Stomoxys calcitrans, the stable fly, is an obligate hematophagous insect, and both sexes feed on blood for survival and reproduction. It is an ectoparasite of veterinary and economic importance and, due to its painful bite and its role as a mechanical vector of several pathogens, it causes stress, diseases, and significant economic losses to the meat and dairy industries. This scenario is directly associated with residues from the sugarcane industry and excessive organic material such as poultry and cattle manure. Control of this species is mainly based on the use of synthetic insecticides, which continuous and ineffective use has favored the emergence of resistant individuals, in addition to causing environmental impacts and risks to human and animal health. Therefore, natural compounds derived from essential oils, such as Origanum vulgare L. and Thymus vulgaris L., as well as their major compounds carvacrol and thymol, have shown promise as alternatives due to their ectoparasiticidal activities in the current scientific literature. In the present study, a topical application test was performed on adult S. calcitrans, analyzing the longevity and mortality of males and females, and calculating the LD₅₀ and LD₉₀ of each compound. The LD₅₀ values were 4424.65 ppm for O. vulgare L. and 32178.12 ppm for T. vulgaris L. after 24 hours. Their respective major compounds showed LD₅₀ values of 65575.63 ppm for carvacrol and 1823.61 ppm for thymol. In addition, these compounds promoted behavioral alterations associated with impairment of the nervous system. The influence of the compounds on reproductive parameters, such as oviposition, pupation, and adult emergence, was also analyzed. The safety of oregano and thyme essential oils, as well as carvacrol and thymol, was evaluated through cytotoxicity tests on mammalian cells, with thymol being the safest compound after 48 hours of treatment. For this reason, and because it presented the best LD50 and LD90 results, causing the highest mortality rate in adults within 24 hours, thymol was selected for formulation in this study. An EC (emulsifiable concentrate) formulation was developed, and its stability and pH were monitored as validation tests for the formulation. As future perspectives, the thymol-based formulation may and should be tested in outbreak scenarios of S. calcitrans, both in the environment and during adult infestations on animals, to be considered as a potential new bioinsecticide.

Bacteria of the families Anaplasmataceae and Mycoplasmataceae are important infectious agents in veterinary medicine, capable of infecting domestic dogs and cats and causing hematological alterations ranging from subclinical infections to severe clinical manifestations. Diagnosis based exclusively on cytological evaluation has limited sensitivity, making molecular techniques essential for the detection of these microorganisms. This study aimed to investigate the occurrence of Ehrlichia spp., Anaplasma spp., and Mycoplasma spp. in dogs and cats treated at a veterinary teaching hospital in the municipality of Seropédica, Rio de Janeiro, Brazil, using molecular methods, as well as to evaluate possible associations with hematological alterations. A total of 310 blood samples, including 177 from dogs and 133 from cats, were subjected to DNA extraction and molecular detection using conventional PCR, nested PCR, and real-time PCR, targeting regions of the 16S rRNA gene of agents belonging to the genera Mycoplasma, Ehrlichia, and Anaplasma. Molecular analysis revealed positivity for hemoplasmas in 6.45% (20/310) of the samples, with higher frequency observed in male animals, while no samples were positive for Anaplasma phagocytophilum. Cytological evaluation did not allow the identification of hemoparasites, highlighting the low sensitivity of this method. Hematological analysis did not demonstrate a significant association between infection and alterations in hematological parameters, indicating potential underdiagnosis in the absence of molecular methods. The results demonstrate the circulation of hemoplasmas and bacteria of the family Anaplasmataceae in the studied region and reinforce the importance of molecular diagnosis. This study contributes to the understanding of the epidemiology and hematological impact of these agents in domestic animals, emphasizing the need for continuous molecular surveillance and more sensitive diagnostic approaches in veterinary clinical practice.

Ticks are arthropod vectors of several pathogens and can represent a significant risk to public health, especially in interface areas between forest fragments and human flow, such as Conservation Units dedicated to ecotourism. In this context, the objective of this study was to survey the diversity and spatial distribution of ticks on trails within two Conservation Units in the state of Rio de Janeiro, followed by the molecular detection of Rickettsia spp. The methodology involved acarological surveys using flagging, dragging, and walking trap methods on trails located in the Guapiaçu Ecological Reserve (Cachoeiras de Macacu) and the Cicuta Forest Area of Relevant Ecological Interest (Volta Redonda). Adult and nymphal specimens were morphologically identified to the species level, while larvae were identified to the genus level. DNA extraction was performed using the HotSHOT method, with phenol-chloroform re-extraction for samples that failed to amplify. To obtain comprehensive data regarding species diversity, Polymerase Chain Reaction - Restriction Fragment Length Polymorphism (PCR-RFLP) was employed for larval species identification and to confirm nymphal identification. This technique was based on the amplification of the 16S rDNA and COI genes, followed by enzymatic digestion using the restriction endonucleases DraI and VspI, and PstI and MboI, respectively. The investigation of Rickettsia spp. infection and coinfection involved conventional PCR, nested PCR, and PCR-RFLP assays targeting the gltA, ompA, ompB, sca2, and sca9 genes. A total of 582 ticks were collected, distributed across six species: Amblyomma calcaratum, Amblyomma sculptum, Amblyomma dubitatum, Amblyomma varium, Amblyomma ovale, and Ixodes loricatus. Sample positivity varied according to the Rickettsia target gene. For the gltA gene, conducted via conventional PCR, 62 and 41 A. varium samples were positive for the 401 bp and 834 bp fragments, respectively. In the Spotted Fever Group nested PCRs, ompA (360 bp) and ompB (511 bp), positivity was detected in 69 and 60 A. varium samples, respectively, and in one A. ovale specimen. PCR-RFLP and sequencing identified Rickettsia parkeri strain Atlantic Rainforest in A. ovale and Rickettsia amblyommatis in A. varium. Additionally, nested PCRs for sca2 (764 bp) and sca9 (727 bp) detected Rickettsia bellii in the single specimen of I. loricatus and in two A. ovale specimens, revealing a coinfection of R. parkeri strain Atlantic Rainforest and R. bellii in an A. ovale larvae. This study presents the first report of R. amblyommatis in A. varium in the Southeast region and the first reports of R. bellii infection in I. loricatus and coinfection in A. ovale in the state of Rio de Janeiro. It is concluded that the combined use of nested PCR and PCR-RFLP techniques represents an effective and low-cost strategy for larval identification and detection of rickettsial coinfections. The application of these methodologies allows for a more robust epidemiological characterization, revealing pathogen-host interactions that frequently remain underestimated.

The cattle tick Rhipicephalus microplus represents one of the main obstacles to cattle ranching, generating significant economic losses and demanding the intensive use of acaricides, which has led to the selection of multi-resistant populations. Given this scenario, fluralaner, a systemic isoxazoline, emerges as a promising alternative. Considering the central role of hemocytes in ixodid homeostasis, the characterization of their kinetics and viability becomes a fundamental parameter to measure the biological response of these arthropods to exposure to new acaricidal molecules, as outlined in the following assays. The present study aimed to evaluate the effects of fluralaner on the cellular response of engorged R. microplus females, focusing on total count, differential count, and hemocyte viability. Initially, methodological standardization was performed by comparing the total hemocyte count in hemolymph pools and in individual samples. Although no statistical differences were observed in the final cell concentration (4,93 x 106 cél. /mL e 4,73 x 106 cél. /mL, respectively), individual collection showed greater variability. In the differential count, the cell types present in the hemolymph were identified, maintaining the stability of proportions in most treated groups, with the exception of a punctual reduction at the concentration of 0.5 μg/mL at 24 hours. Viability assays using the trypan blue exclusion test demonstrated that fluralaner, at the concentrations and times tested, did not induce immediate cytotoxicity to circulating hemocytes, suggesting that the molecule's mechanism of action on tick reproduction and survival may not be directly linked to primary hemocyte lysis. These results contribute to the cellular understanding of the tick under chemical stress and assist in the validation of experimental protocols for cellular physiology studies in ixodids.

Canine monocytic ehrlichiosis, caused by Ehrlichia canis, represents an important disease of clinical and epidemiological relevance in tropical and subtropical regions, including Brazil. This study aimed to isolate, propagate, and molecularly characterize E. canis strains obtained from naturally infected dogs in the state of Rio de Janeiro, evaluating diagnostic, biological, and phylogenetic aspects of the isolates. Three dogs with clinical signs compatible with infection by tick-borne agents were analyzed, and their blood samples were submitted to hematological examination, microscopic evaluation, molecular detection by PCR, and isolation in Ixodes scapularis cell lines (IDE8 and ISE6). The observation of intracellular morulae in blood smears allowed for initial screening of the samples, later confirmed by PCR targeting the 16S rDNA, dsb, p28, and trp36 genes. The isolation of three new strains (E.canisPretinha, E.canisLyon, and E.canisScooby) was successful in tick embryonic cells, representing the first in vitro isolation of E. canis in Brazil in the IDE8 cell line and propagation in the ISE6 cell line. Descriptive analysis revealed differences in infection intensity among the strains, with the E.canisPretinha strain showing greater in vitro infectivity. Phylogenetic analyses based on the 16S rDNA and dsb genes demonstrated high genetic conservation, confirming the identity of the samples as E. canis. On the other hand, the p28 and trpi36 genes showed genetic variability among the isolates. Two strains clustered with the American genogroup (TEDSVSAPA), while one strain showed a pattern compatible with the Brazilian genogroup, identified by the ASVVPEAE tandem repeat in the trp36 gene. The findings demonstrate the co-circulation of different genogroups of E. canis in the state of Rio de Janeiro and reinforce the importance of the combined use of conserved and variable molecular markers for diagnosis, isolation, and epidemiological characterization of the agent. Furthermore, the use of cell culture of tick-derived strains proved to be an efficient strategy for isolating new Brazilian strains of E. canis, constituting a promising tool for the development of future investigations focused on genetic diversity, biology, and pathogen-vector interaction.

Ehrlichia canis is a Gram-negative, obligate intracellular bacterium responsible for Canine Monocytic Ehrlichiosis (CME), whose transmission occurs through Rhipicephalus linnaei in Brazil. Its distribution has been described in all regions of the country. Studies involving E. canis have identified immunoreactive proteins that present species-specific antigens and are related to host–parasite interaction, providing relevant information about the genetic diversity of the pathogen. In this context, the present study aimed to analyze the prevalence and genetic diversity of E. canis, using the p28 and trp36 genes, in whole blood samples from naturally infected dogs from the microregion of Unaí, Minas Gerais. A total of 386 whole blood samples from dogs were analyzed using molecular techniques based on the p28 and trp36 genes. Among the analyzed samples, 3.1% (12/386) showed amplification for the p28 gene of E. canis. Among the samples positive for p28, 66.7% (8/12) amplified for the trp36 gene. For sequencing and genetic diversity analysis, 11 sequences of the p28 gene and five sequences of the trp36 gene were used. The sequences obtained for the p28 gene showed identity ranging from 99.27% to 100% when compared with E. canis sequences from different geographic regions and, in the amino acid translation analysis, mutation sites were observed. Haplotype network analysis based on the p28 gene revealed nine distinct haplotypes (Hd = 0.9632). For the trp36 gene, simultaneous circulation of American and Brazilian genotypes was observed in the studied region, demonstrating broad genetic diversity and local heterogeneity. It is concluded that E. canis strains circulating in dogs in the microregion of Unaí, Minas Gerais, present genetic diversity in the trp36 and p28 genes. The variation observed in tandem repeat regions and the presence of exclusive haplotypes demonstrate gene flow among strains. The co-circulation of BR and US genogroups reinforces the genetic complexity of the agent in the region and highlights the importance of continuous molecular monitoring.

Stomoxys calcitrans (L.), commonly known as the stable fly, is a hematophagous insect of the Diptera order. Its bite causes intense pain in hosts, which are mainly cattle and horses. When present in large numbers, it leads to significant blood loss and stress, resulting in weight loss and a reduction in meat and dairy production, with a major economic impact on livestock activities. During blood feeding, the insect can mechanically transmit various pathogens to the hosts, causing further losses. The saliva of hematophagous insects contains several pharmacologically active molecules that facilitate feeding and can also influence pathogen infection through immunomodulatory effects on their hosts. This immunomodulatory potential can affect different cells of the immune system, including macrophages, cells of the innate immune system that have the potential to secrete diverse cytokines and pro inflammatory mediators. The genus Herpetomonas is a monophyletic group, present on almost all continents, associated with a wide variety of insect species. Recently, the species Herpetomonas muscarum was isolated from S. calcitrans for the first time. Leishmaniases comprise a group of diseases that present four distinct clinical manifestations, depending on the species of the parasite (cutaneous, diffuse cutaneous, mucocutaneous, and visceral). Visceral leishmaniasis, also known as kala-azar, among other regional names, is a systemic and chronic disease. In Brazil, it is caused by the species Leishmania infantum and transmitted by phlebotomine sandfly insect vectors. Visceral leishmaniasis (VL) is known to affect several animal species, such as marsupials, rodents, edentates, and canids, with dogs being the main reservoir and source of human infection in urban areas. Considering the above, the main objective of the present study was to analyze the immunomodulatory effect of the salivary gland extract (SGE) of S. calcitrans on macrophages, as well as to evaluate its effect on L. infantum promastigote forms and H. muscarum opimastigote forms growth, and to assess the cytotoxicity of the salivary gland extract on macrophages, which are the host cells of L. infantum, besides being curccial to iannate response. Possible mechanisms of action of the salivary gland extract were elucidated through the production of nitric oxide and reactive oxygen species, and by evaluating the parasite load of macrophages infected with L. infantum in the presence and absence of fly saliva. The results demonstrated that the salivary gland extract of S. calcitrans does not have cytotoxic potential toward macrophages. The saliva showed no significant effect on binding or phagocytosis in macrophages infected with L. infantum; however, an increase in NO and ROS was observed in macrophages previously treated with saliva, whether they were stimulated with IFN and LPS or not. When the effect of the extract was evaluated directly on L. infantum promastigotes and H. muscarum opimastigotes, proliferation of H. muscarum induction and a lower percentage of survival of L. infantum were observed compared with untreated controls. Our results show an important role of the saliva of S. calcitrans with pharmacological and biotechnical potential in mammals immune system modulation and in the fly own biology.

Entomopathogenic fungi are a sustainable alternative for the biological control of Rhipicephalus microplus, particularly given the emergence of resistance to chemical acaricides. The production of microparticles by ionic gelation for encapsulating propagules is a promising strategy, as it allows polymers and adjuvants to be combined to protect the fungus and improve its shelf life. This study aimed to develop microparticles containing Metarhizium anisopliae s.l. CG 148 conidia encapsulated with different combinations of sodium alginate (A), pectin (P), glycerol (G), and chitin (Q). The encapsulation efficiency and the influence of chitin addition on conidia production were investigated, and the particles were analyzed for size distribution, morphology, structure, shelf life, UV-B tolerance, thermotolerance, and efficacy in controlling R. microplus in the laboratory and semi-fileld conditions. The ionic gelation efficiency was assessed by quantifying and verifying the viability of the encapsulated conidia. To investigate the effect of chitin on propagule production, the sporulated conidia of the microparticles incubated in culture medium were quantified. Formulations containing chitin (AQ, APQ, AGQ e APGQ) were used in subsequent experiments. Size distribution was analysed using ImageJ, morphology was analysed using stereoscopic and scanning electron microscopy, and infrared spectroscopy (FTIR) was performed. The microparticles and the non-encapsulated conidia (NEC) were stored for one and two months at room temperature and in a refrigerator. For tolerance tests, samples were exposed to UV-B radiation (6.0 kJ m⁻²) and heat (45 °C for four hours). The efficacy of the fungal microparticles in controlling ticks was evaluated in laboratory and semi-field settings. Encapsulation reduced the concentration of conidia by an average of 2.5× without compromising viability. Compared to formulations without chitin, the AQ, APQ, AGQ and APGQ particles showed increases of 60.26%, 84.77%, 37.72% and 54.84%, respectively, in conidia production. The microparticles exhibited spherical shapes with more homogeneous (APQ and APGQ) or heterogeneous (AQ and AGQ) surfaces. The average diameter of the AQ, APQ, AGQ and APGQ microparticles was 1,338 µm, 1,459 µm, 1,244 µm and 1,367 µm, respectively. FTIR analyses suggested physical interactions between the polymeric matrix and the fungus. NEC exhibited greater germination than the fungal microparticles under both storage conditions. Refrigerated storage was more efficient in maintaining microparticle viability than ambient storage. NEC germination decreased after exposure to UV-B radiation (59.08%) and heat (0.19%), compared to AQ (98.59% and 71.74%), APQ (95.48% and 93.87%), AGQ (94.37% and 68.18%) and APGQ (97.95% and 75.5%), respectively. In the laboratory, the microparticles reduced biological parameters and the survival of engorged females compared to the control group, with no difference observed in comparison to the NEC group. In semi-field conditions, AGQ reduced the number of larvae by 59.5% compared to the untreated group, while APQ was ineffective. These findings reinforce the potential of fungal microparticles as an innovative approach to controlling this tick, adding technological value to a final product that offers practical advantages for field use, such as easily application and greater stability.

Aedes aegypti is one of the main vectors of arboviruses of public health relevance, such as dengue, Zika, and chikungunya, and the limitations associated with the continuous use of chemical insecticides reinforce the need for alternative and more sustainable control strategies, including the use of entomopathogenic fungi. In this context, this study aimed to evaluate the response of A. aegypti larvae to infection by the fungus Metarhizium anisopliae, considering both the modulation of the humoral immune response and potential alterations in the intestinal microbiota. Rockefeller strain larvae at the L2 stage were exposed to conidial suspensions of the M. anisopliae CG 153 isolate at a concentration of 1 × 10⁵ conidia mL⁻¹ and maintained under controlled temperature and humidity conditions. In Chapter 1, after 24 and 48 hours of exposure, intestines, fat bodies, and whole larvae were collected for RNA extraction, cDNA synthesis, and gene expression analysis by RT-qPCR, encompassing genes associated with the innate immune pathways Toll, IMD, and JAK-STAT, as well as antimicrobial peptides. In Chapter 2, the intestinal microbiota was analyzed by next-generation sequencing, followed by filtering, quality control, and data processing steps. The results from Chapter 1 showed that the expression of the evaluated genes exhibited variations dependent on the analyzed tissue and post-infection time, without a homogeneous pattern among the components of the immune pathways, while antimicrobial peptides displayed distinct expression profiles across tissues and time points. Microbial ecology analyses indicated maintenance of richness, diversity, and overall composition of the intestinal microbiota between control and infected groups, with no evident separation of microbial profiles at the analyzed time points. Taken together, the results indicate that larval infection by M. anisopliae is associated with an immune response modulated in a tissue- and time-dependent manner, without expressive alterations in the overall structure of the intestinal microbiota under the experimental conditions evaluated, contributing to the understanding of interactions between A. aegypti and entomopathogenic fungi in the context of biological control.

Vector-borne diseases (VBDs) have been a growing concern in feline medicine due to their wide geographic distribution, zoonotic potential, and association with hematological and systemic alterations. In Brazil, studies including cats as hosts and interactions with fleas as protagonists are scarce, especially those correlating findings in both species. The objective of this study was to determine the occurrence of pathogens in C. felis felis and in blood samples from infested and non-infested cats in the Metropolitan Region of Rio de Janeiro, and to relate them to clinical and hematological parameters. Blood samples from 400 cats, 664 fleas, and 14 ticks were evaluated. In the present study, molecular screening for pathogens in cat blood detected Mycoplasma spp. in 77 samples (19.3%), Bartonella spp. in 26 (6.5%), Babesia spp. and R. felis in 3 (0.75%), respectively, followed by Cytauxzoon spp. in 2 (0.5%), Ehrlichia spp. was found in 1 (0.25%), and none of the samples tested positive for Anaplasma spp. Direct examination of blood smears did not reveal structures suggestive of any of the agents sought. In the readings of the buffy coats, structures compatible with morulae and/or inclusions suggestive of the Anaplasmataceae family were observed in 36 (9%) of the slides, of which only 14 were concomitantly positive in the qPCR for any pathogen. Regarding the retroviruses analyzed, 81 (20.3%) were positive for FeLV and 14 (3.5%) for FIV, with 6 (1.5%) animals positive for both retroviruses. Flea infestation was observed in 89 cats, predominantly low-intensity infestations, with most animals presenting fewer than 20 fleas. No statistically significant association was observed between the presence of fleas or the pathogens detected and clinical, hematological, or biochemical changes. When analyzing fleas, R. felis was the most frequently detected microorganism, with a positivity rate of 68.7% (456/664), followed by Bartonella spp. 12.90% (16/124), followed by FeLV, 10.48% (13/124), Mycoplasma spp., 1.61% (2/124), and FIV, 0.81% (1/124). No sample showed detection of Babesia spp., Cytauxzoon spp., Anaplasma spp., or Ehrlichia spp. The results indicate that, although there is circulation of multiple agents in cats and vectors in the studied region, the low parasite load and the predominance of subclinical infections contribute to the absence of measurable clinical impact, highlighting the importance of integrated epidemiological surveillance in urban feline populations.

In Brazil, Cochliomyia hominivorax is the main causative species of primary myiasis in livestock, companion animals, and humans, highlighting its social and economic relevance to animal and public health. Chemical control remains one of the primary management strategies, and insect growth disruptors (IGDs) represent promising alternatives due to their interference with the insect life cycle. Larvae of C. hominivorax obtained from a laboratory colony (CEUA 8634020223) were used to evaluate susceptibility to different IGDs (diflubenzuron, fluazuron, teflubenzuron, triflumuron, and pyriproxyfen). Toxicological analyses were conducted through stage-specific bioassays for each larval instar, considering the physiological and behavioral characteristics of each phase. LC₅₀ values were estimated using Probit analysis in RStudio software. For first-instar larvae (L1), the compounds were incorporated into the larval diet at concentrations of 0.001; 0.005; 0.01; 0.05; 0.1; 0.5; 1; 25; 50; and 100 µg·mL⁻¹, characterizing a bioassay based on combined ingestion and contact exposure. The experimental design consisted of 12 groups with six replicates and ten larvae per replicate (n = 720), in addition to a control group and a solvent group (16% acetone with two drops of Tween 80). Pupation inhibition and adult emergence inhibition were evaluated. LC₅₀ values for pupation inhibition, in ascending order, were 0.01, 0.07, 0.15, 3.82, and 5.14 µg·mL⁻¹ for diflubenzuron, triflumuron, fluazuron, teflubenzuron, and pyriproxyfen, respectively. For emergence inhibition, LC₅₀ values in ascending order were 0.01, 0.03, 0.05, 0.06, and 0.10 µg·mL⁻¹, corresponding to diflubenzuron, triflumuron, teflubenzuron, pyriproxyfen, and fluazuron. For third-instar larvae (L3), at the pre-pupal stage, bioassays were conducted exclusively as contact tests. Whatman No. 1 filter paper discs were impregnated with 470 µL of the active compound solutions and placed in Petri dishes, in sextuplicate, at concentrations of 5; 10; 25; 50; 75; 100; 150; 250; 500; and 1000 µg·mL⁻¹, with ten larvae per replicate (n = 720). For pyriproxyfen, additional concentrations of 0.01; 0.05; 0.1; and 0.5 µg·mL⁻¹ were also evaluated (n = 1020), along with control and solvent (analytical grade acetone) groups. For emergence inhibition in L3, LC₅₀ values were 924.6 µg·mL⁻¹ for diflubenzuron, 329.5 µg·mL⁻¹ for teflubenzuron, 47.2 µg·mL⁻¹ for triflumuron, and 0.4 µg·mL⁻¹ for pyriproxyfen. LC₅₀ estimation for fluazuron was not possible under the bioassay conditions employed. The integration of both methodologies demonstrates that IGD efficacy depends on both the larval instar and the route of exposure, reinforcing their strategic potential in the integrated management of this species.

This study describes the morphology and histology of endoparasitic helminths from marine fish of the families Sciaenidae and Trichiuridae collected at CEASA-RJ, focusing on ecological, sanitary, and taxonomic aspects. Two species of digenean trematodes (Lecithochirium monticelli, L. microstomum) and two nematodes (Anisakis spp., Dichelyne sciaenidicola) were identified. Histological analyses using hematoxylin-eosin, orcein, and Gomori’s trichrome staining revealed structural details of tegument, suckers, musculature, and reproductive organs, showing functional adaptations for attachment and survival in the host’s intestine. Anisakis larvae displayed features consistent with zoonotic L3 stages, while D. sciaenidicola showed traits of chronic infections. The findings highlight the diagnostic value of histology for parasite taxonomy and its relevance to understanding host–parasite coevolution, environmental health monitoring, and the prevention of zoonotic risks related to fish consumption

Rhipicephalus microplus, commonly known as cattle tick, is responsible for significant health and economic challenges, particularly in livestock farming, affecting productivity, increasing farm costs and saving animal welfare. Synthetic acaricides, when used economically, can select for resistant tick situations, leading to the need for integrative alternative solutions. Mycoacaricides offer a promising approach in biological control of ticks. The most trained entomopathogenic fungi for tick control are Beauveria spp. and Metarhizium spp., while Cordyceps species are less explored. The present study aimed to assess the effect of a corn oil formulation based on Cordyceps javanica to control the non-parasitic stages of cattle tick under semi-field conditions. Compatibility tests were performed with C. javanica in different concentrations of corn oil (1, 3 or 5%) and oil silicone (0.01%), the vegetable oil was removed using Solub'Oil®, as germination analyses were performed after 24h and 48h after inoculation in PDA. Laboratory tests were extended with C. javanica formulations topically in larvae and females of R. microplus. The formulation 10⁸ conidia/mL + 3% was selected for the test in semi-field conditions, with three collections being performed for persistence analysis before, 24h and 30 days after treatment, with larval recovery in the pots treated with the formulation reduced by 89% compared to the aqueous control group. The persistence of C. javanica conidia in the treated soil was evaluated one and 30 days after treatment. Germination of C. javanica conidia with oil-in-water formulations was higher than 98% after 24 h, and the highest oil concentrations did not inhibit fungal growth. R. microplus larvae treated under laboratory conditions with 10⁸ conidia/mL + 5% corn oil and oil silicone showed 95.17% mortality five days after treatment. Engorged females exposed to the same conditions experienced a 72.5% reduction in egg mass weight compared to the control. Similar results were presented in formulations with 3% corn oil. Even after 30 days of treatment, soil samples from pots treated with C. javanica showed fungal colonies, evidencing the persistence of the fungus in the soil. Our study supports the integration of oily plant mycoacaricides as part of an integrated management strategy, offering a more sustainable solution that reduces chemical dependence in livestock farming. The successful use of C. javanica as a biological control agent can therefore improve animal health, promote food security and contribute to the sustainability of livestock farming systems.

Changes in society and the urban environment have brought dogs and humans closer together, fostering an affectionate relationship that contributes to the animals' longevity. As a result, there has been an increase in the diagnosis of chronic and malignant diseases such as cancer. Despite advances in veterinary medicine, diseases caused by intestinal parasites, which are still neglected, are one of the main causes of morbidity in dogs, with symptoms such as vomiting, diarrhoea, anorexia and anaemia. Healthy dogs can be asymptomatic, but cancer patients have a compromised immune system, either due to the disease or treatment, which favours the development of clinical signs that affect well-being and can interrupt treatment. Although the rate of endoparasite infection and the incidence of cancer in dogs is high, coproparasitological investigation before treatment is still little practised, and there is no data in the literature on the occurrence of these infections in these patients. The aim of this study was to assess endoparasitoses in dogs with neoplasms and correlate them with the oncological disease (cytological/histopathological type), comparing them with animals free of neoplasms and immunosuppressive diseases. The dogs were divided into two groups: group I (with neoplasms) and group II (free of neoplasms), with 80 individuals in each. For the coproparasitological assessment, the owners were asked to collect faeces from the dogs in jars with MIF for three consecutive days. The material was analysed using three methods: centrifugation-flotation, centrifugation-sedimentation and the Faust technique. The results of the tests and the clinical information obtained from the medical records were tabulated for later analysis. In dogs with cancer, parasites of the genera Ancylostoma spp. and Trichuris spp. were found, while in dogs without neoplasms, Ancylostoma spp., Trichuris spp., Toxocara spp., Giardia sp. and Cystoisospora spp. were observed. The group of patients with neoplasms had the highest number of positive results in the coproparasitological tests, with the dogs diagnosed with skin tumours being the most affected. The age of the positive animals ranged from 4 to 14 years, and males (7/26) had more positive results than females (4/57). The most frequent histological type in the positive males was mastocytoma (4/7), of which 3 had multiple tumours with different histological types. In the females, the tumours varied between mammary neoplasms, ovarian tumours and skin neoplasms, and 2 had more than one histological type. It is concluded that coproparasitological investigations should be carried out during the screening and staging examinations of all dogs with neoplasms, and that dogs with multiple neoplasms appear to be more susceptible to endoparasite infections.

The aim of this study was to evaluate the effectiveness of a paste formulation containing Fluralaner in controlling D. nitens in horses. Fluralaner is a relatively new drug belonging to the isoxazoline class, whose mechanism of action is to block chloride channels controlled by aminobutyric acid (GABA) and chloride channels controlled by glutamate. This drug was administered orally at a dose of 25mg/kg body weight. The study was carried out using a stable test, where 14 Brazilian Pony horses were stabled and divided into a treated group and a control group, animals that received the formulation and animals that did not receive the formulation, respectively. Both groups underwent periodic infestations on days -31, -29, -27, -25, -23, -21, -19, -17, -15, -13, -11, -9, -7, -5, -3, -1. Naturally shed ticks were collected from day -3 to day +29. The average number of ticks shed three days before treatment (-3, -2, -1) was 71.0±48.5 for the animals in the control group and 81.8±66.1 for the animals in the treated group. These data showed no statistically significant differences (p=0.831), which shows that the groups were randomized homogeneously. Comparing the means from D+1 to +29, there was a significant difference (p=0.024) and a high effect size of 1.528 (0.189 - 2.810). The averages for this observation period were 36.4 (4.5-77.9) and 8.3 (3.6 - 20.1) for the control and treated groups, respectively. Disregarding the values obtained for tick drop on day +1, a measure adopted due to zero tick efficacy at this time, the following average tick values were obtained after this time for the period from D+2 to D+29: 33.9 (2.6-71.5) and 1.9 (0.9 to 4.1). There was a significant difference (p<0.0001) and a large effect size of 2.651 (1.009 to 4.229). The tick efficacy for the period from D+1 to +29 was 80.2%. When looking at the tick drop values from day +1 and consequently the efficacy, it can be concluded that it is correlated to the 0 (zero) efficacy of this time. Tick efficacy for the period from D+2 to +29, disregarding D+1, was 95.1%. When looking at the tick drop values on day +1 and, consequently, the efficacy, it can be concluded that it is correlated to the 0 (zero) efficacy of this time. With regard to the results of the action of fluralaner observed in this study on the reproductive aspects of D. nitens, it can be seen that there were significant differences between the average values for the weight of teleogyns, laying weight and percentage of larvae hatching for the two experimental groups.

Theileria equi is an intraerythrocytic protozoan that causes equine piroplasmosis, a disease that affects equids and is responsible for significant health and economic losses to the global equine industry. In Brazil, Rhipicephalus microplus is the only tick species experimentally proven to act as a vector for T. equi. However, the mechanisms involved in this pathogen-vector interaction have not yet been fully elucidated. Therefore, the aim of this study was to analyze the differential expression of genes involved in redox metabolism and the main immune signaling pathways of R. microplus in response to T. equi infection. For this, a chronically infected equine with T. equi (positive control) and a non-infected equine (negative control) were infested with pathogen-free larvae of R. microplus. After the fixation and blood feeding period, whole larvae and nymphs were collected and stored individually, while the females were dissected, and the midgut, ovary, and salivary gland of each specimen were stored separately. RNA was then extracted, and cDNA was synthesized from each sample, followed by relative gene expression assays. In the first chapter, genes related to the redox metabolism of R. microplus in response to T. equi infection were analyzed. A upregulation of both pro-oxidant and antioxidant genes was observed in the midgut and salivary gland of engorged females infected by T. equi, with a particular emphasis on the genes encoding the enzymes dual oxidases (pro-oxidant) and catalase (antioxidant), both being more expressed in the infected group. However, in the salivary gland, the gene encoding the enzyme glutathione-S-transferase was significantly suppressed in the infected group, suggesting that this enzyme may play a key role in the antioxidant response in this tissue. No significant oxidative response was observed in the larval and nymph stages, nor in the ovaries of engorged females. In the second chapter, genes involved in the Toll, IMD, and JAK/STAT immune signaling pathways, as well as genes encoding the antimicrobial peptides microplusin, defensin, and ixodidin, were analyzed. T. equi infection stimulated the activation of the Toll and IMD pathways in the midgut and salivary gland of R. microplus, resulting in a positive regulation of the microplusin and defensin genes in both tissues. However, the JAK/STAT pathway was suppressed in the midgut in response to T. equi infection, which may have led to the non-significant differential expression of the ixodidin gene observed in this tissue. No significant response to T. equi infection through the analyzed pathways was observed in the larval and nymph stages, nor in the ovaries of engorged females. The results obtained in this study provide insights into the molecular mechanisms involved in the interaction between T. equi and R. microplus, contributing to a better understanding of how the tick responds to protozoan infection. Such information aids in identifying key molecules with potential to be used in the development of new strategies for the prevention and control of equine piroplasmosis in the country.

Hemoprotozoan diseases represent a significant challenge to Brazilian cattle farming, mainly due to diagnostic difficulties and the economic losses associated with decreased herd productivity. Among these pathogens, protozoa of the genus Trypanosoma are noteworthy as the causative agents of trypanosomosis, a disease that may present with nonspecific clinical signs such as anemia, weight loss, reduced milk production, and abortions. These manifestations often lead to underdiagnosis and underreporting of the disease in cattle herds. This study aimed to investigate the occurrence of Trypanosoma spp. in cattle from different municipalities in the state of Rio de Janeiro, using Polymerase Chain Reaction (PCR) as the main diagnostic tool. A total of 398 blood samples were collected between 2022 and 2025 from cattle raised on farms located in the municipalities of Vassouras, Miguel Pereira, Seropédica, Teresópolis, Piraí and Volta Redonda. All samples were subjected to PCR using primers targeting the 18S rDNA and cathepsin L (CatL) genes, both widely used markers for the screening and differentiation of Trypanosoma species. Of the animals tested, 13 samples (3.27%) were positive for Trypanosoma spp., all of which originated from a single farm in Volta Redonda. Genetic sequencing confirmed 100% similarity with Trypanosoma vivax strains previously described in Brazil and West Africa. Phylogenetic analysis showed that the sequences clustered with lineages already circulating in the southeastern region of the country. These findings highlight the relevance of PCR as a highly sensitive and specific technique for the detection of bovine trypanosomosis, especially in subclinical infections or cases of low parasitemia where conventional methods tend to fail. Furthermore, the study underscores the importance of incorporating molecular-based epidemiological monitoring strategies to support early diagnosis, sanitary control, and prevention of disease spread in cattle herds across the state of Rio de Janeiro.

Bovine anaplasmosis, caused by Anaplasma marginale, is a hemoparasitic disease of major economic and sanitary importance in global livestock production, transmitted primarily by the tick Rhipicephalus microplus. The disease exhibits high genetic diversity, mainly reflected in the surface proteins MSP1a and MSP4, which influence both antigenic variability and the pathogen’s ability to evade the host immune response. This study aimed to assess the prevalence and genetic diversity of A. marginale in cattle from conventional and organic farms in the state of Rio de Janeiro. The evaluation of the farms sought to determine whether inadequate sanitary management, high vector density, and lack of veterinary supervision contribute to the maintenance and spread of the disease. Blood samples (n = 388) were collected across seven municipalities, followed by DNA extraction, molecular amplification of the GAPDH, msp4, and msp1α genes, and analysis of the msp1α tandem repeats. Results showed a 66.8% prevalence of Anaplasma spp. among the samples analyzed, with the identification of eight circulating strains. Genotype E predominated, with one E1 variant, highlighting the genetic diversity of the pathogen in the region. Regarding sanitary management, the positivity rate did not differ statistically. The findings highlight the need to adopt integrated management and control strategies, including continuous epidemiological surveillance and effective vector-control measures, to reduce the prevalence of anaplasmosis and mitigate its impacts on cattle herds.

Dogs and cats play a fundamental role in today’s society, being considered family members and important companions to humans. These animals can harbor a wide variety of vector-borne agents, among which blood protozoa known as hematozoa, belonging to the genera Babesia, Theileria, Hepatozoon, and Rangelia, stand out. These parasites are part of the phy-lum Apicomplexa, which includes organisms of great veterinary and, in some cases, zoonotic relevance.This study aimed to analyze blood samples and blood smear slides from 338 ani-mals, including dogs and cats, treated at the Small Animal Veterinary Hospital of the Federal Rural University of Rio de Janeiro. The samples were sent to the Veterinary Clinical Patholo-gy Laboratory (LABVET) for hematological and cytological processing and then to the Hemoparasites and Vectors Laboratory (LHV), where DNA extraction, and cytological and molecular analyses were performed. The samples were sent to the Laboratory of Hemopara-sites and Vectors (LHV), where DNA extraction and cytological and molecular analyses were performed. The blood smears, stained using the Giemsa method (1:10) and examined under oil-immersion optical microscopy (1000x), showed no forms compatible with hematozoa. In the molecular analysis, qPCR targeting the hsp70 gene of Babesia vogeli detected a positivity rate of 1.8% (n = 6/338) (five dogs and one cat) while nested PCR targeting a fragment of the 18S rRNA gene revealed 0.6% (2/338) of samples positive for piroplasms. For Hepatozoon spp., 1.8% (n = 6/338) of samples were positive by conventional PCR. The sequences ob-tained showed greater than 99% identity with Babesia vogeli and Hepatozoon canis in Gen-Bank, confirming the molecular identification of the agents. Agreement between the methods used for B. vogeli was evaluated using McNemar’s test and the Kappa coefficient; McNemar’s test indicated no significant difference between techniques (p = 0.125), while the Kappa coefficient demonstrated moderate agreement (0.496; p < 0.001), with discordant re-sults restricted to samples positive only by qPCR, reinforcing its higher diagnostic sensitivity. Hematological evaluations showed anemia, thrombocytopenia, hyperproteinemia, and even unremarkable hemograms in animals positive for B. vogeli, whereas animals positive for H. canis exhibited leukocytosis, anemia, hyperproteinemia, and leukopenia. In conclusion, the detection of hematozoa was low, and molecular methods demonstrated greater diagnostic ef-ficiency than cytology, particularly in infections with low parasitemia. Furthermore, qPCR proved to be significantly more sensitive than nested PCR. These findings highlight the im-portance of applying sensitive diagnostic techniques and underscore the need for additional studies to deepen the understanding of these agents in hospital settings, especially in cats, for which many protozoan infections remain poorly studied.

The control of arthropod vectors and agricultural pests requires sustainable alternatives in light of the limitations and impacts associated with chemical acaricides and insecticides. In this context, entomopathogenic fungi are key elements both in shaping host immune responses and as part of integrated management strategies. This thesis investigated two distinct models: (i) the cattle tick Rhipicephalus microplus, focusing on the interaction of Metarhizium anisopliae with the gut bacteriota in the presence or absence of Anaplasma marginale, and (ii) the moth Noctua pronuba, an important agricultural pest, with emphasis on its humoral immune interactions with Beauveria bassiana and Trichoderma atroviride. In Chapter I, the gut bacterial diversity of engorged R. microplus females was characterized through a metabarcoding approach targeting the V3–V4 region of the bacterial 16S rRNA gene in groups exposed or not to M. anisopliae and infected with A. marginale. Coinfection reshaped the bacteriota, with a higher number of associated taxa and a distinct microbial composition compared to control and singly infected groups. Co-occurrence network analysis revealed that the coinfected microbiome (Metarhizium and Anaplasma) exhibited greater connectivity and robustness, suggesting a compensatory response of the host and bacterial community to dual stress rather than beneficial stability. Functional prediction indicated that these shifts may represent a metabolic adaptation of the bacterial consortium to support tick and/or pathogen survival, supported by the increased abundance of functional genes associated with metabolic pathways in the coinfected group. These findings highlight the modulatory role of the gut bacteriota at the fungus–vector–pathogen interface, with direct implications for the vector competence of R. microplus, and reinforce the potential of fungi to interfere with pathogen transmission cycles. In Chapter II, the immune response of N. pronuba larvae was investigated after exposure to B. bassiana (entomopathogenic) and T. atroviride (non-entomopathogenic). The differential expression of innate immunity-related genes (cecropin1, attacin, and PGRP-A) was quantified by qPCR. B. bassiana induced a stronger early activation of attacin and PGRP-A six hours after treatment, whereas T. atroviride promoted suppression of attacin and PGRP-A at 12 and 24 hours, suggesting an ability to interact with host immune pathways despite lacking evident pathogenicity. These results expand our understanding of recognition and defense mechanisms against fungi and underscore the importance of exploring novel fungal isolates, such as Trichoderma, in biocontrol programs. The two chapters demonstrate that arthropod immune responses and gut bacteriota dynamics are central determinants in the interaction with entomopathogenic fungi and pathogens. Understanding these relationships provides a foundation for developing innovative biocontrol strategies capable of reducing dependence on chemical inputs and contributing to sustainability in animal health and agriculture.

Poultry production is one of the most important agricultural activities for the Brazilian economy and has shown significant growth over the past decade. In 2024, Brazil produced approximately 14.972 million tons of chicken meat, ranking as the second largest global producer and the world leader in exports within this sector. During poultry meat production, it is essential to consider that water consumption ranges from 5,000 to 21,000 liters per ton of meat, generating large volumes of effluents throughout the slaughtering chain. Additionally, poultry farming produces significant amounts of waste, including dead bird carcasses and poultry litter—a mixture of excreta, feathers, skin dander, and food remnants—which can impact water resources through excessive consumption, soil infiltration, and contamination of surface and groundwater.

The use of antimicrobials in animal production has become widespread as growth promoters, helping to control pathogens and enhance animal performance, often through continuous low-dose administration aimed at reducing carcass rejection and productivity losses. Despite their effectiveness, there is growing pressure to replace them due to the alarming rise in antimicrobial resistance. In this context, water becomes a potential route for the dissemination of multidrug-resistant bacteria, increasing the risk of transmission among animals, the environment, and even humans.

Among various classes of antimicrobials, polymyxins have been extensively used in veterinary medicine to control infectious processes and as growth promoters in livestock. Polymyxin resistance was once believed to arise solely from chromosomal regulatory changes and mutations. However, in 2015, the first mobile resistance gene (mcr-1) was identified on a plasmid in Escherichia coli in China. The presence of resistance to polymyxin E (colistin) in mobile genetic elements poses a significant risk to public health, as these elements can spread rapidly via horizontal gene transfer (HGT). The mcr-1 gene has been documented on at least five continents, but its origin, acquisition, emergence, and dissemination mechanisms remain poorly understood.

This study aimed to characterize colistin-resistant Enterobacterales in wastewater samples from the treatment plant of a poultry slaughterhouse located in the municipality of São José do Vale do Rio Preto, in the mountainous region of Rio de Janeiro State, Brazil. Sixteen water samples were collected at different points along the treatment flow. A total of 140 Enterobacterales strains were isolated and identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The isolates were tested for colistin minimum inhibitory concentration (MIC) using broth microdilution; 39 were classified as resistant (MIC > 2 µg/mL). These were screened for the presence of mcr-1 to mcr-10 genes via conventional PCR. Sixteen isolates carried at least one of the targeted genes, with mcr-1 being detected in E. coli, Enterobacter spp., and Klebsiella spp. across all three sampling points, and mcr-10 being identified in Enterobacter bugandensis.

Positive isolates were subjected to clonal typing using Multilocus Sequence Typing (MLST). Additionally, E. coli strains were assigned to phylogenetic groups using the Clermont method. Finally, Enterobacter bugandensis and Klebsiella pneumoniae isolates obtained from treated effluent were selected for whole-genome sequencing. The identification of mcr genes in different Enterobacterales species throughout the treatment chain highlights the role of slaughterhouse wastewater as both a reservoir and a potential source of colistin resistance dissemination. The detection of such genes, even after wastewater treatment, underscores the need for continuous surveillance and the improvement of biosecurity strategies. This study contributes to the understanding of resistance dynamics in environments linked to animal production and supports public policy development aimed at mitigating the risks associated with the spread of resistance genes, particularly under the One Health framework

Apicomplexan protozoa are obligate parasites found in a wide range of vertebrate hosts and hematophagous invertebrates, exhibiting either monoxenous or heteroxenous life cycles depending on the species. Within the suborder Adeleorina, species of the genus Hepatozoon were first recorded infecting small mammals (rodents and marsupials) in Brazil during the last century. In 1915, Hepatozoon muris was detected in the rodent Akodon fuliginosus in São Paulo, and in 1916 another species was reported, described as Haemogregarina didelphydis, in the marsupial Didelphis aurita in the municipality of Rio de Janeiro. More recently, a novel species, Hepatozoon milleri, was described in the rodent Akodon montensis in Botucatu, São Paulo, based on both light microscopy and molecular analyses. In the present study, liver samples and blood smears were collected from 309 non-volant small mammals captured in municipalities of the states of Paraná (Cruz Machado, Lidianópolis, and Ponta Grossa) and Rio de Janeiro (Iguaba Grande and Nova Friburgo), including three species of didelphid marsupials (Didelphis aurita, Monodelphis americana, and Philander quica), 14 species of cricetid rodents, and two species of murid rodents (Mus musculus and Rattus rattus). Blood smears were stained using the Giemsa method, and gamonts were detected by light microscopy in 10% (31/309) of the sampled small mammals (Akodon cursor, Akodon montensis, and Oligoryzomys nigripes). Significant morphometric differences were observed in gamont measurements among Akodon species. Using conventional PCR, 18S rDNA fragments of Hepatozoon spp. were amplified in 23.3% (72/309) of rodents (A. cursor, A. montensis, O. nigripes, Oxymycterus nasutus, Oxymycterus quaestor, Sooretamys angouya, and Mus musculus), whereas no marsupials tested positive for Hepatozoon spp. Phylogenetic analyses showed that sequences obtained from rodents (n = 41) formed a subclade with other sequences from small mammals in Brazil and belonged to four distinct haplotypes. Three long sequences (>1600 bp), representing haplotypes 1, 3, and 4, were subsequently obtained and analyzed. Based on phylogenetic inference, the study reinforces the trophic relationship between rodents and reptiles as a potential transmission route for Hepatozoon in South America. Furthermore, our findings expand the known host range of Hepatozoon spp., reporting Oxymycterus nasutus and Oxymycterus quaestor as new host species and identifying two new haplotypes circulating in rodents from the state of Paraná, where the parasite had not previously been detected in members of this order. Following the qualification examination, the study was expanded to include additional rodent and marsupial samples that also tested positive for Hepatozoon. Additionally, piroplasms were investigated in these populations, with positivity detected in both host groups, significantly enriching the understanding of the parasitic diversity circulating among these mammals.

Species of the genus Rickettsia are widely distributed in Brazil, including Rickettsia rickettsii, Rickettsia parkeri, Rickettsia bellii, Rickettsia felis, and Rickettsia amblyommatis, with R. rickettsii being the main causative agent of Brazilian spotted fever. Despite the well-recognized importance of ticks in maintaining these agents, comparative studies across different biomes remain scarce. This study aimed to investigate tick diversity and the occurrence of Rickettsia spp. in free-living ticks and those collected from wild hosts in the Amazon, Atlantic Forest, and Cerrado biomes. A total of 460 ticks were analyzed, with a predominance of the genus Amblyomma (99.7%), collected in the Amazon (n = 303), Cerrado (n = 64), and Atlantic Forest (n = 93). Specimens were morphologically identified and subjected to molecular detection of Rickettsia spp. by conventional PCR targeting the gltA, htrA (17 kDa), and ompA genes, followed by sequencing and phylogenetic analysis. Overall, 9.22% of the samples were positive for Rickettsia spp., with detection restricted to the Amazon and Atlantic Forest biomes. The tick species associated with positive samples were Amblyomma cajennense sensu stricto, Amblyomma sculptum, Amblyomma dubitatum, Amblyomma ovale, Amblyomma geayi, and Ixodes loricatus. In the Amazon, A. cajennense s.s. and A. geayi were detected in Tapirus terrestris, Bradypus variegatus, and in the environment, indicating high ecological plasticity. In the Atlantic Forest, A. sculptum and A. dubitatum parasitizing Hydrochoerus hydrochaeris showed high positivity, reinforcing the role of capybaras as amplifier hosts in enzootic cycles. No Rickettsia spp. was detected in the Cerrado. Positive nymphs were predominantly infected with R. amblyommatis, with a single detection of R. felis in an A. sculptum nymph. No DNA of R. rickettsii was detected in the analyzed samples; however, spotted fever group rickettsiae, such as R. parkeri and R. amblyommatis, were identified. These results demonstrate that the circulation of rickettsiae varies among biomes, reflecting ecological, vector-related, and host-related differences. The study highlights the importance of ticks in maintaining enzootic cycles and emphasizes the need for integrated surveillance at the human–animal–environment interface under a One Health perspective.

Among the main ectoparasites of importance in companion animals, the flies Cochliomyia hominivorax stand out. Several classes of insecticides are used to manage infestations. Fipronil is an insecticide belonging to the phenylpyrazole class, used topically in veterinary medicine. Due to data on human exposure to FIP used through this route of administration, coupled with the trend in the veterinary market towards the development of oral ectoparasiticides, the present study aimed to develop a formulation in the pharmaceutical form of chewable tablets containing fipronil. , with evaluation of the descriptive pharmacokinetics of oral treatment and efficacy in dogs naturally infested with C. hominivorax larvae. The tablets were developed by molding, following the steps of preparing the diluent, binder, mixing the components and incorporating the active ingredient. The pharmacokinetic study was carried out after the oral administration of one unit of fipronil chewable tablet at 72mg, corresponding to 6mg/kg. The efficacy study was conducted through the oral administration of one unit of fipronil chewable tablet at 72mg, according to the natural occurrence of dogs infested by C. hominivorax. The characterization of the tablets was acceptable in relation to quality control criteria, with average weight, dosage and dose uniformity within acceptable limits. When administered orally, it was absorbed and reached systemic concentration quickly, with a Cmax of 3126.53 ng/mL in 2.22h (Tmax). The in vivo clinical efficacy study with the treatment of one unit of the 72mg fiprinil chewable tablet promoted the elimination of 97.76% of C. hominivorax larvae after 24 hours of treatment, with an overall efficacy of 100% in naturally infested dogs.

The study involving hemoparasites from wild and exotic birds, under human and free-living care, directly assists in the conservation of animals ex situ and in situ, by maintaining the health of animals that are kept in institutions and animals that have suffered anthropogenic actions, since Brazil is home to one of the greatest biodiversity on the planet. Studies to determine the occurrence and diversity of these parasites can clarify the parasite-host relationship and the physical conditions that birds find themselves in when harboring the agent. That said, the objective of this study was to investigate the cytological and molecular occurrence of hemoparasites in birds received by the Wild Animal Rehabilitation Center, the commercial breeding facility and the Environmental Education Park, all located in the western zone of the state of Rio de Janeiro. Whole blood was collected from 104 birds, divided into 55 free-ranging birds and 49 birds kept under human care. From this, blood smears were prepared and molecular analyzes were carried out for agents of the genera Borrelia and Ehrlichia, family Trypanosomatidae and the orders Haemosporida and Piroplasmida. The positivity of hemoparasites in the individuals studied was 13.46% (n=14/104) through the molecular technique, being agents of the orders Haemosporida (11.54%; n=12/104), Piroplasmida (0.96%; n=1/104) and for the Trypanosomatidae family (0.96%; n=1/104). Using the blood smear technique, it was possible to observe the presence of trypomastigote forms in a sample compatible with molecular diagnosis targeting the 24Sα-rDNA gene in a sample obtained from Asio clamator. Merozoites suggestive of Piroplasmida were identified in a blood smear and confirmed using PCR targeting the 18S rDNA gene in free-living Sula leucogaster. With sequencing, the Asio clamator agent had 91.60% similarity to the 24Sα-rDNA gene of Trypanosoma theileri, and two samples with 99.57% similarity to the cytB gene fragment of Haemoproteus (Haemoproteus) paramultipigmentatus in specimens of Columbina talpacoti under rescue conditions. Furthermore, Plasmodium sp. was also observed, with 99.9% identity with the fragment of the cytB gene, in a specimen of Ramphastos toco also from free-living. Plasmodium juxtanucleare was found similar (99% to 100%) to other sequences deposited in GenBank in six individuals of Gallus gallus domesticus, as well as P. juxtanucleare in an individual of Ara ararauna, all under captive conditions. Another finding confirmed after sequencing was P. juxtanucleare in a free-living Rupornis magnirostris individual with 99.57% similarity to the cytB gene sequence deposited in GenBank. Studies involving the frequency of hemoparasites in birds assist in the creation of preventive medicine protocols and quarantine actions for animals that will be kept under human care or that will be reintroduced into the wild, reinforcing the need for knowledge of the agent and its host.

Aedes (Stegomyia) aegypti is involved in the dissemination of various arbovirus, representing a challenge for Public Health. Faced with increasing resistance to chemical insecticides and concerns about environmental contamination, it becomes crucial to explore more sustainable control approaches. The aim of this study was to evaluate the persistence of Metarhizium anisopliae CG 153 on A. aegypti larvae under semi-field conditions over 3 weeks, following the weekly introduction of a new batch of larvae, and its impact on the offspring, through the adaptation of the World Health Organization's guidelines for laboratory and field testing of mosquito larvicides. Conidia were extracted from rice medium and suspensions were adjusted to 1×10⁶ and 1×10⁸ conidia/mL. Five types of containers, of different sizes and shapes, were used to simulate artificial breeding sites, each type (N=6) containing 100 to 400 mL of conidial suspension or 0.03% Tween 80, with 25 to 100 larvae (L2). Larval survival was assessed daily for 7 days, with a new batch of larvae introduced every 7 days. Virulence persistence was evaluated for 21 days, with a 10 μl aliquot of suspensions taken from the breeding sites every 7 days for conidial viability monitoring and re-isolation. Dead larvae were removed weekly and subjected to fungal re-isolation and scanning electron microscopy. Surviving larvae were removed after 7 days and pupae transferred to cages, where adults emerged and were fed with a 10% sugar solution and blood for oviposition. Eggs were quantified, offspring counted, and their survival assessed daily for 7 days. Temperature and relative humidity were monitored daily. The concentration of 1×10⁸ conidia/mL was the most virulent over the three weeks, reducing almost 100% of larval survival in all types of containers: large pot, significant reduction in larval survival was observed in the first (χ2= 552.5; P<0.0001), second (χ2= 462.4; P<0.0001), and third weeks (χ2= 313.5; P<0.0001); clay pot, significant reduction in the first week (χ2= 609; P<0.0001), as conidial suspensions and 0.03% Tween solution evaporated; small pot, reduction in larval survival in the first (χ2= 263.8; P<0.0001), second (χ2= 496.8; P<0.0001), and third weeks (χ2= 407.5; P<0.0001); flowerpot and small plate containers were monitored for two weeks as suspensions/solutions evaporated. In both, statistical differences were observed in the first (χ2= 1.362; χ2= 350.9; P<0.0001) and second (χ2= 540.7; χ2= 432.8; P<0.0001) weeks, respectively. The average viability of conidial suspensions decreased over the weeks. Fungal re-isolation in treated larvae showed green colonies, characteristic of M. anisopliae. Scanning electron microscopy revealed M. anisopliae conidia on the larvae's cuticle/natural openings. Therefore, M. anisopliae CG 153 was able to persist in artificial breeding sites and maintain its virulence on A. aegypti larvae under semi-field conditions for 21 days, demonstrating potential for bio-prospecting and field application in the biological control of A. aegypti larvae.

Helminths represent a major public health problem for pets and farm animals, leading to economic losses in the agricultural sector and the risk of zoonoses to their owners. Plant extracts and their components can be an efficient alternative to anthelmintics. The objective of this study was to chemically and biologically characterize the aqueous extract of Eleusine indica and evaluate its antioxidant capacity and anthelmintic action against the nematode Caenorhabditis elegans. To achieve this objective, the extract was characterized in its phenolic components by antioxidant and quantitative tests, and its activity against the adult nematode was performed using concentrations of 50 to 1500 μg/mL-1 in quintuplicate and incubated for 48h. A 10% (w/v) aqueous extract of E. Indica was obtained. After analysis, the product presented 5.50 mg of quercetin/g of dry extract of flavonoids and 8.15 mg of quercetin/g of dry extract for phenolics. When the values of total antioxidants were investigated, the plant extract obtained the value of 63.71 mmol Fe(II)/100 mg of dry extract. Its nematicidal activity indicated LC50 and LC90 of 725.6 and 12926.0 μg/mL-1, respectively. Therefore, the extract of Eleusine indica presented a nematicidal effect in the proposed animal model, and its antioxidant capacity was evaluated.

Dairy cattle farming represents one of the main sources of income and job creation in the Brazilian agricultural sector, being marked by marked heterogeneity both in relation to production systems and the profile of herds and producers. The health status of a dairy herd is essential within the production chain, where the biggest difficulty faced by producers is mastitis. Its etiology is attributed to both environmental and contagious agents, with bacteria of the genus Staphylococcus being the most common cause among all cases and widely distributed throughout the world. The animal production sector becomes crucial in the propagation of both pathogens and resistance genes, with the administration of antimicrobials as growth promoters and prophylactics being the main route of selective pressure on the animal and environmental microbiota. Considering the significant risk posed by infectious conditions and the economic embargoes involved, the WHO declared antimicrobial resistance a substantial threat to global health. The objective of this work was to evaluate the prevalence of species of the genus Staphylococcus and the phenogenotypic profile of antimicrobial resistance against the class of beta-lactams mediated by the mecA and blaZ genes from isolates obtained from samples of bovine milk collected on properties in the south of the state of Rondônia and Baixada Fluminense, as well as the hygienic-sanitary and socioeconomic profile of the producers. The sampling comprised 127 milk samples, which were subjected to phenotypic identification at the genus level in AMVF, tube coagulase test and resistance to bacitracin 0.04UI, and proteomic identification at the species level using the MALDI-TOF technique. The characterization of the phenotypic resistance profile was carried out using the disk diffusion technique given by cefoxitin, oxacillin and penicillin, and the detection of the mecA and blaZ genes through PCR. The survey of the hygienic-sanitary and socioeconomic profile of the properties was carried out by applying a questionnaire during sample collection. The results obtained demonstrated a prevalence of approximately 58% of S. aureus, compared to approximately 42% of CNS species. Phenotypic antimicrobial resistance was observed in only 35% of penicillin-resistant isolates, of which approximately 43% came from the North region and 57% from the Southeast region. Among these, the presence of the mecA gene was confirmed in around 20% of the isolates, with a prevalence of 15% and 25% for the North and Southeast regions, respectively; and blaZ in 7% of the isolates, which were obtained only from samples from the Southeast region. The results obtained provided a comparison of the resistance profile of the isolates and the hygienic-sanitary and socioeconomic profile and properties included in the study. In this way, the need for approaches aimed at meeting the particularities of each region is evident, making deeper studies on the topic essential.

Stomoxys calcitrans is a hematophagous dipteran known as the “stable fly”. The present study aimed to determine under what pressures entomopathogenic nematodes (NEPs) maintain their viability and ability to infect and kill S. calcitrans larvae in by-products of the sugar and alcohol industry.

To obtain the desired pressures, a hydraulic pump with a pressure controller and a sprinkler nozzle were used to reproduce the pressures used in fertigation of sugarcane fields. Groups of 10 third-instar fly larvae were deposited in plastic containers containing two sheets of filter paper and different sugarcane by-products (according to each group). Infectious Juveniles (JIs) of Heterorhabditis bacteriophora HP88, H. baujardi LPP7 and H. indica LPP30 were used, already subjected to pressures of 60, 70 and 80 psi. The total volume was four mL of distilled water per treatment (in bioassays with water) and four mL of 50% vinasse (in bioassays with vinasse), the concentration of NEPs used in all bioassays was 200 JIs/fly larva. The control group had the same concentration of JIs, but without having passed through the pressure system.

There was also a control group without the presence of NEPs. Larval mortality was observed daily. The bioassays were maintained at 27±1° C and 70±10% RH, with six replications. The data were subjected to the Shapiro-Wilk normality and Bartlett homoscedasticity tests. When the assumptions were observed, analysis of variance was applied, followed by the Tukey test (p<0.05) to compare means. It was identified that regardless of the substrate used, nematodes, whether or not subjected to different pressures, impacted S. calcitrans larval mortality. In the bioassay in distilled water without the presence of sugarcane by-products, the species H. bacteriophora HP88 and H. indica LPP30 were the most virulent to S. calcitrans larvae, with mortality rates exceeding 85%. Pressure apparently did not affect the virulence of NEPs, as mortality in groups treated with pressure was higher than in the control group. In filter cake, the species H. bacteriophora HP88 and H. indica LPP30 did not have their virulence affected by the pressures used, whereas H. baujardi LPP7 showed a drop in efficiency after being subjected to 60, 70 and 80 psi. In sugarcane bagasse, only the mortality rates caused by NEPs were variants, with H. bacteriophora HP88 (81.7%) being the most virulent. It was identified that the pressures evaluated did not reduce the virulence of the NEPs, which, after being subjected to 60, 70 and 80 psi, maintained their harmful effect on fly larvae, with an average mortality rate exceeding 70%. In sugarcane straw, H. bacteriophora HP88 had a slight reduction in its virulence when subjected to 80 psi, causing S. calcitrans larval mortality of 72%, however, up to 70 psi, this species promoted mortality of fly larvae in levels above 85%. In the species H. indica LPP30 and H. baujardi LPP7, the application of pressures (60,70 and 80 psi) promoted a reduction in larval mortality by approximately 19% and 17%, respectively, even though the final values were significantly higher. compared to the mortality observed in the control group (4.1%). In vinasse, it was observed that all treatments had mortality averages higher than the control group without NEPs, but equal to each other.

̇Stomoxys calcitrans, commonly known as the stable fly, is a significant mechanical vector that plays a crucial role in the transmission of pathogens in livestock environments, impacting animal health and causing economic losses. Our study focuses on the expression of NADPH oxidases (NOX), enzymes that produce reactive oxygen species (ROS) under certain stimuli, and have various roles in the metabolic activities and redox metabolism of S. calcitrans, under varied nutritional conditions, including fasting, feeding on sucrose, and blood feeding. We also investigated the same parameters following infection with Herpetomonas muscarum, a monoxenous trypanosomatid previously found to naturally infect the fly by our group. The aim of this research is to verify the expression of the NOX enzymes and the modulation of pro- and antioxidant responses in S. calcitrans, particularly in response to different nutritional states and protozoan infection. The adopted methodology encompassed bioinformatic analysis techniques for identification, characterization of NOX isoforms, followed by quantitative gene expression experiments through real-time PCR to measure the levels of NOX in the insect's gut and fat body, as well as redox activities, both pro- and antioxidants. Genes for the NOX5 and DUOX1/2 enzymes were identified from the genome of S. calcitrans, showing homology with those of Drosophila melanogaster and Musca domestica. The results revealed a significant variation in the expression of the NOX enzyme genes in response to the different nutritional states, in addition to a notable increase in NOX5 expression in the fat body after H. muscarum infection. Furthermore, it was observed that the nutritional state directly affects redox activity, modulating both the production of pro-oxidant molecules and the activation of antioxidant responses. Flies in fasting and blood-fed conditions showed an increase in redox activity, indicating a possible adaptive mechanism to deal with oxidative stress. In contrast, sucrose-fed flies exhibited a less intense redox response, suggesting differences in the management of oxidative stress according to the diet. A role of NOX enzymes in regulating the redox metabolism of S. calcitrans is suggested, influencing the insect's ability to respond to nutritional and pathogenic challenges. The modulation of NOX expression and redox activity, investigated for the first time in the present work in S. calcitrans, suggests complex adaptive mechanisms that allow the insect to maintain homeostasis in the face of environmental variations and infections. These findings broaden the understanding of the biology of disease vectors, offering perspectives for the development of vector control strategies that exploit the insect's vulnerability to oxidative stress.

Haematobia irritans is considered a major livestock pest in Brazil and causes economic losses to the livestock industry globally. New control strategies are based on safer compounds, such as the class of insect growth regulators (IGRs), analogous to juvenile hormones that inhibit insect development. The oral efficacy of pyriproxyfen allows the development of formulations that guarantee the safety and extension of the pharmacological action. This study aimed to develop a pyriproxyfen-based Rumino-Reticulum Device (RRD) consisting of films of poly(vinyl)alcohol (PVA) and sodium carboxymethylcellulose (NaCMC) to control the horn fly in cattle. Films were obtained by the solvent casting method by PVA/NaCMC crosslinking. The assays was include physical-chemical characterization, weight variation and pH measurement, swelling degree test (SD), drug content determination, in vitro drug release test, X rays diffraction analysis (XRD), fourier-transform infrared spectroscopy (FTIR), differential scanning calorimetry (DSC), and scanning electron microscopy (SEM). Films were subjected individually to weight variation measurement (n=3), with an average value of 0.446 ± 0.02 mg and presented basic pH values (6.5 ± 0.09), near the ruminal pH (6.5-7.5). The SD predicted by the absorption capacity of the film in the time interval obtained was 331,40% with an absorption capacity. HPLC determined the drug content of films, and the value obtained was 104.81%. The in vitro release results supported a slow-release profile, indicating pseudo-first-order kinetics. FTIR analysis elucidated the characteristic bands of PVA, NaCMC, and pyriproxyfen, and demonstrated the absence of the drug after release. SEM images showed changes in the porosity distribution of the samples after the addition of pyriproxyfen, and XRD analysis showed an increase in crystallinity due to the presence of pyriproxyfen (Xc of 36.59%). Thermal analysis revealed that pyriproxyfen and its delivery altered chain packing, with Tg values of 74 ºC and Tm of 208 °C. The pyriproxyfen-based RRD developed, in addition to fulfilling the characteristics of prolonged release, allows it to be rolled up (compressed form) facilitating swallowing and subsequent conversion to an expanded form that is retained in the rumen throughout the treatment period.

The development of biofilms is a severe health problem, responsible for approximately 80% of microbial infections, including chronic, nosocomial, and medical device-associated infections, which are highly resistant to conventional antibiotics. Essential oils have been evaluated for antimicrobial properties. The study aimed to assess the antimicrobial activity of the essential oils (EOs) of Mimosa verrucosa and Illicium verum against planktonic and sessile cells of Gram-positive and Gram-negative bacteria and evaluate toxicity based on the eukaryotic model of Saccharomyces cerevisiae. Thirty compounds were identified, the main ones being a-pinene (12.6 %), b-pinene (16.7 %) and (E)-caryophyllene (14.2 %) for M. verrucosa, and estragole (4.2 %) and anethole (86.8 %) for I. verum. The results of the minimum inhibitory concentrations (MIC) of the EO of I. verum (29.40 µg/mL) and M. verrucosa (24.89 µg/mL) against planktonic cells showed 99% efficacy against all cells tested (sensitive and resistant S. aureus and E. coli). In sessile cells, the EO of I. verum and M. verrucosa showed respective efficacy against sensitive S. aureus, and all samples were tested. In both analyses, the minimum bactericidal concentration (MBC) test revealed that I. verum caused cell death for sensitive S. aureus and E. coli. However, M. verrucosa only showed bactericidal activity against planktonic cells. The EOs evaluated showed antibacterial and bactericidal activity in the planktonic format. However, only I. verum showed bactericidal activity against sessile cells, indicating its possible potential for treating bacterial biofilms. Considering the expanding resistance to antimicrobials, the EOs tested represent an important therapeutic option, especially against S. aureus and E. coli, which can produce biofilms on various surfaces, becoming a serious public health problem.

The tick Amblyomma sculptum has a wide geographic distribution, high impact on animal health and transmission of zoonotic diseases, which highlight the importance of studying and implementing effective control measures to mitigate the risks associated with this species. Despite this, information on biological aspects of this tick, such as hemocyte dynamics and the possible effects of chemical interference, is still scarce. The objectives of this study were to characterize the ultrastructure of different types of hemocytes in the hemolymph of engorged females of A. sculptum, to evaluate the reproductive efficiency of engorged females against fluazuron and novaluron, and to analyze the challenges of the cellular immune response through the in vitro action of these insect growth disruptors, evaluating quantitatively and morphologically the hemolymph cells. Hemolymph samples were collected by piercing the cuticle in the dorsal region. The identification and ultrastructural analysis of hemocytes was performed using transmission electron microscopy (TEM). The in vitro evaluation of the reproductive efficiency of engorged females of A. sculptum was performed after immersion in different concentrations of fluazuron and novaluron. For the cellular immune response to in vitro challenges, immersion tests were performed with engorged females of A. sculptum in the control and treated groups at concentrations of 7.81, 250 and 4000μg.mL-1 of fluazuron and 1250, 10000 and 40000μg.mL-1 of novaluron. At this stage, the hemolymph was collected at 24h and 48h after immersion. The total hemocyte count was performed in a Neubauer chamber and the determination of the frequency of each cell type was performed by preparing and analyzing hemolymph smears. Four morphologically distinct cell types were identified in females of A. sculptum through TEM: prohemocytes, plasmatocytes, granulocytes and spherulocytes. In the analysis of reproductive efficiency, the values obtained from the concentration of 250 μg.mL-1 of fluazuron achieved satisfactory efficacy, reaching 100% efficacy at the concentration of 4000 μg.mL-1. The results with novaluron did not present significant changes, with the highest concentration achieving 11.4% efficacy. After analyzing the cellular response, it was found that fluazuron promoted a significant decrease in total hemocytes at the concentration of 4000 μg.mL-1 after 48h, as well as increasing the frequency of spherulocytes. Novaluron did not cause changes in total hemocyte counts, however, within 24h, it caused a significant increase in prohemocytes at concentrations of 10,000 and 40,000 μg.mL-1, and within 48h it caused an increase in plasmatocytes and a decrease in granulocytes at concentrations of 10,000 and 40,000 μg.mL-1. In TEM, both insect growth disruptors were able to promote morphological and ultrastructural changes in the phagocytic hemocytes of A. sculptum. It is concluded that from TEM it was possible to characterize four cell types of A. sculptum: prohemocytes, plasmatocytes, granulocytes and spherulocytes. At the highest concentration, fluazuron was able to inhibit larval hatching, while novaluron was not able to act satisfactorily on reproductive efficiency. Furthermore, fluazuron and novaluron were able to promote cellular changes in the hemolymph of engorged female A. sculptum ticks after in vitro exposure. This is the first study to investigate the ultrastructural characteristics of hemocytes from  A. sculptum.

Sarcocystis species are intracellular protozoan parasites with an intermediate-definitive host life cycle based on a predator-prey relationship. Most Sarcocystis species infect specific hosts or closely related host species, causing various clinical signs to the hosts and in some cases rendering the meat unusable for human consumption as it can still be a risk factor for human infection, as it occurs through the ingestion of raw or undercooked meat. A total of 27 cervids were submitted for post-mortem examination from 2017 to 2023 at the Pathological Anatomy Sector of the Universidade Federal do Rio de Janeiro. Four cervids in total were diagnosed with meningoencephalitis, representing 14.8% of the causes of death in the Cervidae family. Three Rusa timorensis from that group were diagnosed with meningoencephalitis caused by Sarcocystis sp. infection and presented compatible histological characteristics, according to the established criteria. Brain tissue samples from three deer with neurological signs were analyzed and encephalitis was observed microscopic with the presence of structures compatible with merozoites of Sarcoscystis. In addition, immunohistochemistry of these tissues was performed and protozoan research was carried out in deer blood from the BioParque blood sample bank through molecular analysis. This study involves recording the appearance of Sarcocystis sp. in histopathological lesions associated with neurological signs in captive cervids from the City of Rio de Janeiro; in addition to their diagnosis, their macro and microscopic aspects and their clinical presentation and epidemiology where described.

Numerous species of domestic and wild animals act as reservoirs of hemoparasites transmitted by ticks, causing hemoparasites of great importance to public health, such as Brazilian Spotted Fever. Known to cause high fever, this disease is a zoonosis with worldwide distribution and is caused by bacteria of the Rickettsiaceae family, presenting a complex diagnosis. The agent's detection has been facilitated due to access to molecular techniques, such as the polymerase chain reaction (PCR). Military personnel in training or operations, due to frequent activities and prolonged exposure, are more predisposed to infestations and at greater risk of infection by bacteria of the Rickettsiaceae family. Therefore, the objective of the present study was to carry out the molecular diagnosis of Rickettsia spp. in ticks parasitizing humans, domestic and wild animals circulating in areas belonging to Academia das Agulhas Negras, Resende, Rio de Janeiro, Brazil. A sampling of ticks infesting different species of vertebrate hosts (humans, wild and domestic animals) collected and identified by Prado (2022) during the years 2019 to 2021 were used. A stratified random sample of 494 ticks were selected to be molecularly tested for the presence of Rickettsia spp. corresponding to 52 tick samples on domestic animals, 138 tick samples on captured wild animals, 37 tick samples on small mammals, 200 tick samples on collared peccaries and 67 tick samples on military personnel. The samples were subjected to DNA extraction, followed by a prior amplification of the partial sequence of the nuclear ITS-2 gene of ticks (family Ixodidae) for quality control of the extraction and conventional PCR for the detection of target DNA corresponding to the gltA gene of ticks Rickettsia spp.. A sample of A. dubitatum was positive during molecular reactions, amplifying the gltA gene fragment of Rickettsia spp.. The corresponding sample belongs to Hydrochoerus hydrochaeris. This finding reveals that wild animals can harbor rickettsial agents, and raises the alarm about animals as sentinels and even humans who are in areas of vulnerability. It is necessary to deepen the study, in addition to carrying out the sequencing.

Responsible for affecting livestock activity with economic losses due to its parasitism, the horn fly, Haematobia irritans, is literally annoying to cattle, because the incessant bites interfere on weight gain and leather quality, which directly harms the milk and meat production. These factors affect health and welfare of these animals, and the control of this ectoparasite is indispensable, done for a long time indiscriminately with synthetic insecticides, causing flies’ resistance to some of these drugs. To reduce the use of these synthetic molecules and then reduce damage to animals, humans and the environment, an alternative is essential oils (EOs) with a large spectrum of insecticidal action, however, few EOs have been tested against H. irritans. Therefore, the objective of this work was to evaluate the insecticidal activity of Eugenia caryophyllus (clove), Cinnamomum cassia (cassia cinnamon), Thymus vulgaris (white thyme) and Illicium verum (star anise) EOs on adult flies of H. irritans, and to determinate its lethal concentration (LC) values: LC₅₀ and LC₉₀. The EOs were obtained commercially, and their constituents were determined through gas chromatography. The major ones being respectively: eugenol, (E)-cinnamaldehyde, thymol and ocymene, and (E)-anethole. The specimens from naturally infested cattle from the field area of the Laboratory of Experimental Chemotherapy in Veterinary Parasitology (LQEPV) at the Universidade Federal Rural do Rio de Janeiro (UFRRJ) were exposed to filter paper discs (63.62cm²) impregnated with different concentrations of the EOs in Petri dishes (90x15mm). The evaluation of mortality were made 2 and 4 hours after exposure. The concentrations that showed 100% of mortality at the 2h evaluation were 11.79 and 15.72 µg/cm² for clove, 39.30 µg/cm² for cinnamon, 47.15 µg/cm² for thyme and 235.77 µg/cm² for anise. And at the 4h evaluation of 15.72 µg/cm² for clove, 15.72 and 39.30 µg/cm² for cinnamon, 31.44 and 47.15 µg/cm² for thyme, and 157.18 and 235.77 µg /cm² for anise. The LC₅₀ and LC₉₀ were statistically calculated using Probit analysis using the RStudio Team Software program with a 95% confidence interval (p≤0.05). The LC₅₀ of the EOs at 2 and 4h were: 5.04 and 5.27 µg/cm² for clove, 8.57 and 5.03 µg/cm² for cinnamon, 18.57 and 14.08 µg/cm² for thyme and 83 .91 and 71.88 µg/cm² for anise. The LC₉₀ at 2 and 4h were: 11.71 and 10.03 µg/cm² for clove, 19.26 and 11.22 µg/cm² for cinnamon, 27.41 and 18.80 µg/cm² for thyme and 132, 78 and 101.30 µg/cm² for anise. Except LC₅₀ of clove EO, all LCs after 4h of exposure reduced. Despite this, the oil that showed the greatest performance was clove EO, followed by cinnamon, thyme and anise. Concluding that all evaluated oils showed in vitro insecticidal activity against the horn fly.

Periodontal disease (PD) arises from the formation of bacterial biofilm on the surface of teeth and their adjacent tissues. It is characterized as the most prevalent oral condition in humans and animals, being present in approximately 70% of adult dogs. In veterinary medicine, its prevention and treatment are challenging since establishing a hygiene routine is not straightforward, and the persistence of conventional formulations in the oral cavity is hindered by saliva action and mechanical mouth activity. Thus, innovation in this field is essential, especially in the face of bacterial resistance that may be involved in different stages of the disease. In this context, the use of products derived from natural sources becomes an alternative to conventional products, such as flavonoids, which stand out due to their antimicrobial properties and their regenerative action on soft and hard tissues, which make up the oral mucosa. Among them, there are quercetin (QRC) and rutin (RTN), which already have reports showing their antimicrobial activity against sensitive and resistant strains. Therefore, this study aimed to estimate the minimum inhibitory and bacteriostatic concentrations of these two flavonoids against nine strains, both sensitive and resistant, of importance for PD. After the assays, QRC proved to be more promising and was selected as the active molecule for pharmaceutical development tests. The following were developed: a hydrogel (HG) and a carboxymethyl cellulose sodium-based film (CMC-Na) and a chitosan-based HG (CTS). As a result, the formulations maintained a pH range of 6.5 to 7.5, ensuring safety during application, except for the CTS HG, which showed a slightly lower pH than ideal. The content of CMC-Na and CTS HG, as well as the film, was 98.2%, 95.0%, and 97%, respectively. The formulations showed extended residence times, exceeding 5 hours. Regarding in vitro release, the CMC-Na HG exhibited zero-order kinetics and a release rate of 32.1% in 24 hours, while the CTS HG and the polymeric film exhibited pseudo-first-order kinetics with release rates of 18.6% and 26% in 24 hours, respectively. Morphological analyses of the polymeric film revealed pores in the formulations containing QRC and greater compactness in the placebo film. Structural analyses showed that the presence of QRC does not alter the structure of the components present in the film but results in a more intense C=C stretching. The combination of all results indicated that the developed formulations have innovative potential for preventing and treating canine PD.

Stomoxys calcitrans (Diptera: Muscidae) is an obligatory hematophagous fly, an ectoparasite
of various mammal species, with bovines and equines being the most affected due to its
aggressive and persistent feeding habits. Population outbreaks, related to increased sugarcane
p roduction in livestock areas, and the increased resistance to insecticides, intensify concerns
about this species, as it is responsible for the mechanical transmission of various pathogens
among parasitized animals, as well as causing significant losses in cattle farming. Therefore,
studies that enable the development of insect control strategies are of great importance. The
generalist feeding habit of S. calcitrans provides an interesting point of investigation. For this
reason, this study analyzed the influence of different diets (sucrose solution and bovine, equine,
ovine, rabbit, and human blood) and developmental substrates (sugarcane based and alfalfa
hay based ) on the fly's biology. Analyses of feeding behavior showed a greater attraction of the
fly t o bovine and equine blood and sucrose solution, consistent with the fly's preferred hosts in
nature and its habit of consuming sugary foods in its natural habitat. Longevity experiments
demonstrated higher survival rates in flies fed with ovine blood, foll owed by bovine blood,
developed on sugarcane based substrate. Furthermore, feeding on ovine blood, in both
substrates, resulted in the highest values of reproductive potential for the species. Contrary to
our expectations, feeding on equine blood showed ne gative impacts on the fly's survival and
reproduction, despite horses being considered one of its preferred hosts. When observing
ovarian development, delays in organ development were observed as a result of feeding on
equine and rabbit blood, while feedin g on bovine and ovine blood showed superior patterns of
development, demonstrating relevant results for the analyzed oviposition rates. The different
diets and developmental substrates did not show significantly different results among the
analyzed groups. The observed changes are possibly based on the nutritional differences of each
condition, as well as direct impacts on digestion and the microbiota. A better knowledge of S.
calcitrans biology contributes to the development of novels control strategies at the same time
that we intend to optimize the parameters of colony flies production.

The objective of the present work was the molecular detection, characterization, and morphological analysis of bacteria of the families Anaplasmataceae and Mycoplasmataceae in populations of small mammals in areas of the Amazon forest, in the state of Acre, Brazil. The importance of this investigation is based on obligate intracellular bacteria that participate in wild cycles as zoonotic agents, and therefore of interest for human health, in addition to parasites of importance for domestic animals, and the animal's own ecology and life cycle. host. The study areas are concentrated in forested areas of the Amazon rainforest, in the city of Rio Branco, State of Acre, Brazil. For direct diagnosis, 186 samples of blood smear slides from previously captured rodents, marsupials and bats were used to detect, characterize, and measure inclusions indicative of bacteria from the families Anaplasmataceae and Mycoplasmataceae. Liver samples stored in RNA Later® with DNA extraction using DNeasy Blood & Tissue Kit (Qiagen®) were used for molecular diagnosis. Of the liver tissue samples analyzed, 14.52% (n=27/186) were considered positive for Mycoplasma sp. by PCR using primers Myco 322s and Myco 938as which amplify 620 base pairs of the 16S rDNA fragment. The diagnosis of Anaplasmataceae was hampered by the non-specificity of the PCR reactions with the tested primer pairs, and when the technique was specific, it resulted in a 16S rDNA fragment from Wolbachia sp. Therefore, these positive data were not considered for analysis. The positivity in blood smear slides for bacteria from the Anaplasmataceae family was 6.99% (n=13/186), while for hemotropic mycoplasmas it was 1.61% (n=3/186). Regarding the location, there was a significant statistical difference between the areas of São Raimundo and Floresta Piracema and the areas of Santa Cecília, EMBRAPA, APA do Amapá, and Colégio Agróciola, where São Raimundo and Floresta Piracema presented a higher percentage of positivity for Mycoplasma sp. compared to the others. Regarding the sequencing result of the 16S rDNA fragment of Mycoplasma sp., three samples from bats of the genera Glossophaga (22052), Artibeus (22108), Carollia (22152) had their sequences well read in the chromatogram, with approximately 600bp. The highest percentage of blast similarity for 22052 was 98.41% with a sample of Glossophaga commissarisi (LR699022), from Switzerland; for 22108 it was 99.82% with a sample of Platyrrhinus lineatus (MZ048291), from Mato Grosso do Sul, Brazil; and for 22152 it was 98.28% with a sample of Desmodus rotundus from Belize (KY932722). This study revealed different genotypes of Mycoplasma sp. in bats and demonstrated the presence of morulae in leukocytes and platelets, characteristic of bacteria of the Anaplasmataceae family in rodents, marsupials, and bats. These results suggest that hemotropic mycoplasmas are endemic in bats from Acre and that much remains to be discovered about the nature of obligate intracellular bacteria in these animals.

The Atlantic Rainforest is one of the largest centers of biodiversity and species endemism in the world. Relief is one of the elements responsible for producing so much diversity. The environmental conditions in the mountains are very different from those in lowland areas because they have a greater temperature range, water restrictions and greater geographical isolation. As a result, species experience adaptations that reflect environmental variations and can disrupt biotic exchanges with neighboring plains. Hematozoan parasites such as Trypanosoma sp. are transmitted by a series of dipteran vectors abundantly present in this environment, affecting most local hosts and exerting selective pressure on avifauna, causing deleterious effects on health, reproductive success, bbehavior,and community structure. The aim of this study was to correlate the elevation gradient of Itatiaia National Park with the prevalence and diversity of Trypanosoma sp. in the blood of wild birds using blood smears and PCR screening to verify the predominance of this hematozoan. Of the 230 slide smears observed under microscopy, the infective evolutionary form of Trypanosoma was found in 17 and 42 samples were PCR positive. Prevalence in the lower elevation gradient, below 1,000, was higher than in the higher elevation gradient, above 2,000, which suggests that temperature, environmental conditions, and vector diversity are more abundant in lower elevation areas, favoring transmission of the hematozoan. Phylogenetic analyses revealed 17 new sequences very close to Trypanosoma Polygranularis found in Peliperdix lathami in Cameroon, Africa. A single sequence clustered close to the Trypanosoma everetti clade, a branch close to the Trypanosoma everetti-like species found in the host Dumetella carolinensis, USA. Our study demonstrated the diversity and prevalence of trypanosome species in different altitudinal gradients in the PNI. In Itatiaia National Park, the genetic diversity among Trypanosoma lineages in wild birds appears to be quite limited. Out of the 18 sequences obtained during this study, an impressive 17 are closely related to Trypanosoma polygranularis according to their phylogenetic analysis. Furthermore, the Trypanosoma sp. strains identified in this research exhibit a broad host range, as they have been observed in no fewer than 12 different bird species.

Brazil is the largest exporter and producer of chicken meat in the world. These birds naturally have the presence of bacteria such as Escherichia coli in the gastrointestinal tract that can cause illness or even death of these animals. In order to avoid this, antimicrobials have been added to water and feed as a prophylactic measure and acting as a performance enhancer. However, this may favor the selection of bacteria resistant to existing antimicrobials. Among the mechanisms of resistance in Gram-negative bacteria, the most important is the production of enzymes β-lactamases, especially those of the extended spectrum (ESBL) and AmpC. Knowing that during this process of poultry meat production there is consumption around 5,000 to 21,000 L of water per ton of meat, generating a large amount of effluents along the slaughter chain, it is understood that water becomesis another way of spreading the multidrug-resistant bacteria present in this environment, enhancing transmission between animals, the environment and humans. Thus, the present study aimed to evaluate the quality of circulating water of an avian slaughterhouse and to draw a resistance profile and to detect the presence of isolates of Escherichia coli producing ESBL and AmpC through water samples. The samples were collected in the municipality of São José do Vale do Rio Preto, in the mountainous region of the State of Rio de Janeiro, where the Rio Preto drains the river, at different points of water passage from the poultry slaughterhouse, totaling 16 samples. The Most Probable Number (MPN) technique was used to determine the number of total coliforms and thermotolerant coliforms. The water from the entrance of the slaughterhouse was within the standard required by the legislation, while the treated effluents were ejecting a load that exceeds the maximum quality limit. They were isolated and identified by MALDI-TOF, 76 Escherichia coli. Ampicillin was the least effective antibiotic and Meropenem the most. In addition, 61.8% (47/76) presented resistance to at least one antibiotic, being among these 6.38% (3/47) considered multiresistant (MDR). In relation to the different collection points, it is observed that the entrance has a lower diversity of antibiotic resistance when compared to the exit. Among the isolates analyzed,19.7% (15/76) indicated possible production of ESBL and 6.6% (5/76) of AmpC in the Diffusion Disc Test. In the Double Disc Sensitivity Test (DDST), 10.5% (8/76) were positive 11.8% (9/76) for the Approximation Test with CAZ/CZC and 15.8% (12/76) with CTX/CTC. Only 3 were positive in the three confirmatory tests. Among the 76 isolates, 21% (16/76) were positive for the bla_CTX-M-1 gene, and 17.1% (13/76) for the bla_CMY-2 gene, where 6.5% (5/76) presented co-production of the gene. When comparing these data with the phenotypic, 68.7% (11/16) had the bla_CTX-M-1 gene but did not show ESBL phenotype and 84.6% (11/13) had the bla_CMY-2 gene without resistance to cefotaxime. Thus, this study makes it possible to understand the impact of these production facilities as contributors to the phenomenon of antimicrobial resistance through water in a One Health perspective.

Anaplasma platys is a gram-negative, obligatory intracellular bacterium, with tropism for platelets and the etiological agent of Canine Infectious Cyclic Thrombocytopenia (TCIC), a disease distributed worldwide, transmitted by the ixodid tick Rhipicephalus sanguineus sensu lato and commonly associated, as the name implies, with thrombocytopenia in dogs. It is predominant in regions with a subtropical and tropical climate, including Brazil, where it occurs with prevalence rates ranging from 2 to 55%, with reports in all regions of the country. It is known to be a zoonotic agent and its vector has already been described as parasitizing humans in Brazil. Despite its importance, several aspects of the infection are still unknown in many parts of the country. Therefore, this study aimed to determine the prevalence, using hematological and molecular methods, analyze the epidemiological aspects, and evaluate the genetic diversity of A. platys, using the gltA gene, in whole blood samples from naturally infected thrombocytopenic dogs from two mesoregions of the Rio de Janeiro state. Four hundred and four whole blood samples from thrombocytopenic dogs, from the Metropolitana and Sul Fluminense mesoregions, in the state of Rio de Janeiro, were used to perform a complete blood count, blood smear search for tick-borne parasites, molecular diagnosis, genetic sequencing and phylogenetic analysis. Furthermore, the correlation of the variables sex, breed and age of the dogs with positivity for A. platys was verified. Of the samples analyzed, 4.7% (19/404) amplified for the A. platys gltA gene. Of these, 15 were subjected to sequencing and phylogenetic analysis. The sequences showed 99-100% identity with isolados from countries on several continents and formed a unique clade. Anemia, hypochromia, hypoproteinemia and the variable sex (male dogs) were associated with A. platys positivity. The results demonstrated that there is a single genotype of A. platys circulating in the state of Rio de Janeiro and that male dogs are the most infected by the pathogen.

The selection of an ideal DNA extraction method must take into account factors such as sensitivity, consistency, speed and ease of execution. The literature lacks sufficient evidence to support the choice of a specific method to extract DNA from avian whole blood samples. The objective of this work was to standardize a genomic DNA extraction method. Five protocols were tested, including the DNeasy Blood & Tissue Kit (Qiagen), PureLink® Genomic DNA Mini Kit (Invitrogen), Salting Out, Hotshot and Phenolchloroform. And the quantification and verification of the quality of the extracted genetic material was carried out. The samples extracted from the “PureLink Genomic DNA Mini” Commercial Kit showed visibly better characteristics in terms of the evaluated parameters compared to the others, obtaining better results and greater efficiency. In the inhibition test, the Invitrogen kit and the Salting out method stood out, which presented lower mean Cq (p-value 0.0568), however the kit presented a lower standard deviation value. This study provides a scientific basis for extracting high-quality genomic DNA from quantities as small as blood when samples are stored on FTA® cards. This is particularly beneficial for bird research, given the wide variation in sizes and weights between species. It is concluded that the standardization of protocols plays a fundamental role in molecular biology, since the efficient extraction of genetic material is essential for the success of subsequent analyses. The Atlantic Forest is one of the richest biomes in biodiversity in the world and is home to an impressive variety of wild birds. Parasites belonging to the genus Haemoproteus are classified in the family Haemoproteidae and have as vectors hematophagous insects from the families Ceratopogonidae and Hippoboscidae. The objective of this study was to evaluate the genetic diversity of parasites of the genus Haemoproteus in the Atlantic Forest biome, in the Caparaó (MG), Itatiaia and Serra dos Órgãos (RJ) National Parks, between 700 and 2500 meters of altitude. Among the 586 birds sampled, 94 were positive for Plasmodium/Haemoproteus, obtaining an overall prevalence of 16.04% (n=94/586). In the phylogenetic analysis, the sequences obtained in this study were grouped into 6 well-supported clades. Clade A was composed of 10 samples that were grouped into an exclusive clade, with Haemoproteuserythrogravidus as its sister group. In clade B, four samples grouped into the Haemoproteuserythrogravidus group. In clade C, four samples formed an exclusive group, with Haemoproteuszosteropis as the sister group. A total of 13 samples grouped into the Haemoproteusnucleocentralis clade (clade D), 5 samples into the Haemoproteusparaortalidum clade (clade E). The lineages grouped in clade B were only found in Zonotrichiacapensis. Clade C was only found at 1500 meters altitude. Regarding the locations sampled, H. erythrogravidus was only found in the CaparaóNP. And there was no occurrence of Haemoproteusparaortalidum in the Serra dos Órgãos NP. A relatively high prevalence and genetic diversity in wild birds was observed in the studied region. Through the results obtained in this study, we enriched knowledge about the global diversity of parasites of the genus Haemoproteus in endemic birds of different species that occur in highland areas of the Atlantic Forest of Brazil. 

The Aedes aegypti mosquito is of great importance in the epidemiological chain of arboviruses that affect Public Health. Thinking about resistance to chemical insecticides and more ecological control solutions, the use of entomopathogenic fungi is increasingly being studied as an alternative to control A. aegypti. This work investigated the effects of Metarhizium anisopliae and Beauveria bassiana on oviposition, ovarian morphology and lipid profile in the fat body and ovaries of females exposed to the fungi, in addition to investigating the antimicrobial activity in A. aegypti larvae challenged by these fungi. To evaluate the effect of fungal exposure on oviposition and ovary morphology, females 6 to 8 days post-emergence were fed with mouse blood and exposed or not to M. anisopliae CG 153 at a concentration of 107 conidia/mL for 24h. , with the following groups being formed: G1= exposed to the fungus minutes before feeding; G2= fed 24h before fungal exposure; G3= fed 24h after fungal exposure; CTR= exposed to 0.03% Tween 80 minutes before feeding. Ovaries were dissected at 0, 24, 48 and 72h and investigation was carried out by histopathology. No statistical difference was observed in the following parameters: number of engorged females in each group (P=0.6358); quantity of total eggs produced (P~0.5948); comparison between the quantity of eggs per female fed (P=0.3191); or the number of live mosquitoes at the end of the experiments (P=0.7507). The ovarian follicles of mosquitoes exposed to M. anisopliae CG153 showed loss of tissue architecture after 48 hours of treatment, with rupture of the follicular epithelium, reduction in the number of oocytes and changes in the morphology of nurse cells. To evaluate the effect of fungal exposure on the lipid profile, females (6-8 days post-emergence; N=45) were exposed to M. anisopliae CG 153 or B. bassiana CG 479 at a concentration of 107 conidia/mL or Tween 80 at 0.03%. After 24, 48 or 72 hours of exposure, the ovaries and fat body of the females were dissected for lipid extraction and subsequent analysis of neutral lipids by one-dimensional thin layer chromatography. There was no statistical difference in the fatty acid and cholesterol profile in the fat body (P=0.7705 and P~0.8365) and ovaries (P=0.9798 and P>0.9999) at the times observed after exposure. Groups of 700 second stage larvae were immersed in 700 mL of B. bassiana CG 479 suspension at a concentration of 106 conidia/mL or 0.03% Tween 80. After 16, 24, 40 and 48h of exposure, a pool of 60 larvae (N=240) were macerated to evaluate antimicrobial activity by performing an antibiogram on strains of Escherichia coli, Staphylococcus aureus and Pseudomonas sp. resistant and sensitive to beta-lactams. However, no antimicrobial activity was identified in larvae macerate even after 48 hours of exposure to B. bassiana CG 479. Therefore, further studies on the effects of M. anisopliae CG 153 and B. bassiana CG 479 isolates on larvae and females of A. aegypti must be carried out to better understand the fungus-host dynamics.

Canine visceral leishmaniasis (CVL) is a chronic and fatal disease caused by the protozoan Leishmania infantum, which also infects humans and other animal species. Its transmission takes place by infected sandflies of the Lutzomyia longipalpis species, with dogs being the main domestic reservoir. Brazil currently does not have a real-time surveillance system for CVL notification and information on numbers of infected animals and injuries is flawed and undersized, not having the same centrality and similar mechanisms used in human health surveillance systems. CVL has major impacts on animal and human health, which, due to the lack of an integrated notification system, may not represent the trend curves of this disease and are often restricted to publications by educational and research institutions and technical information. A total of 477 records were made in the system based on training trips in surveillance and diagnosis of canine visceral leishmaniasis and data on confirmed cases of visceral leishmaniasis from the Ministry of Health. The inclusion of data allowed the construction and visualization of maps from the system to evaluate the distribution and georeferencing of cases, the inclusion of photos, videos and images from Google Earth. The application worked smoothly and even in areas where there was no signal, it allowed location from the mobile device's GPS data. All data collected from the application were imported into the surveillance and control platform without loss of data or information collected. The data obtained by the project allowed the construction of a spatial distribution map of CVL, which from the spatial analysis verified a higher concentration of CVL cases in the Northeast, North, Midwest and Southeast regions, coinciding with the same distribution of occurrence in human cases of human visceral leishmaniasis. The application allowed the construction of a form for registering cases and including data, allowing exportation in different formats for later analysis and identification. The use of the surveillance and control application (viconSaga mobile) proved to be a viable, low-cost, customizable tool, being an alternative for registering CVL cases, contributing to obtaining centralized data on CVL that will allow a set of actions for the knowledge and detection of the determinants and conditions of this disease in order to recommend prevention and control measures for CVL, suggesting a methodology that can be used nationwide by official surveillance services. The general objective of this study was to verify the effectiveness and usability of the ViconSaga mobile application as a data collection tool for the registration of CVL cases, and support for surveillance and control of canine visceral leishmaniasis. The study aimed to register cases of leishmaniasis in dogs from immunochromatographic serological tests, from training and qualification trips and epidemiological surveillance, in all regions of Brazil, performing georeferencing and a spatial analysis of CVL cases from of the application and platform records, Evaluate the efficiency of the mobile notification system through the platform for epidemiological studies., Evaluate the results obtained by the notification and inserted in the platform in relation to the official data of the health surveillance system and contribute to the perception of the LVC in Brazil

Leishmaniasis are among the most prevalent and neglected endemic diseases worldwide, being of great importance in public health. Visceral leishmaniasis is an infectious zoonotic disease caused by the protozoan Leishmania infantum, an obligate intracellular parasite of the cells of the mononuclear phagocytic system of vertebrate hosts. The transmission of the parasite to humans and animals occurs primarily through the bite of infected female sandflies, popularly known as “straw mosquitoes”. In the Southeast region of Brazil, the municipality of Volta Redonda, in the interior of the state of Rio de Janeiro, has been presenting cases of canine visceral leishmaniasis (CVL), in addition to reported human cases. Chapter I of the thesis aimed to carry out a survey of cases of canine visceral leishmaniasis (LVC) at the Centro de Controle de Zoonoses (CCZ) in the municipality of Volta Redonda (VR), in the interior of the state of Rio de Janeiro (RJ), in the period from January 2016 to December 2020, using the VICON SAGA Platform for georeferencing the cases. It was observed that the search/demand for diagnosis in the CCZ of the city of Volta Redonda has been increasing over the years; the number of animals diagnosed with CVL increased by 286% in the research period; skin lesions are the main clinical alterations found in patients positive for CVL; the large number of neighborhoods presenting positive cases of CVL alerts to the occurrence of the disease, mainly in urban areas; the VICON SAGA platform made it possible to identify areas with the highest occurrence of CVL and, consequently, the areas at greatest risk; the profile of the animals treated at the CCZ of VR are SRD animals, with short hair, medium size and showing nonspecific clinical signs. Chapter II aimed to carry out a systematic review to elucidate the main epidemiological factors associated with CVL in South America from 2000 to 2020, using PubMed, Lilacs, Web of Science and Scopus platforms as databases. Of the 3076 articles found, 20 epidemiological studies were selected. It is concluded that factors associated with the environment are the most studied in all epidemiological studies and the closer the shelters for dogs are to forest regions, the greater the chance of infection by L. infantum; it is not possible to establish an association between race and a higher occurrence of L. infantum infection; articles that were able to establish correlation with age, reported that adult animals are the most exposed.

Coccidians are obligate intracellular protozoans that infect most invertebrates, all classes of vertebrates, including domestic and wild animals and humans. They present intestinal life cycles, although extra-intestinal life stages are observed in some species. They are usually identified from fecal samples. Brazil has a diverse avifauna currently represented by 1,971 species of different orders. Passeriformes is the most representative order that is often positive for different species of coccidian parasites, mainly of the genera Isospora Schneider, 1881 and Eimeria Schneider, 1875. The study of the interactions between these parasites and hosts allows us to understand several ecological, evolutionary and behavioral processes, including sexual selection, reproductive success, migration, competitiveness, among others. Currently, there are several techniques and methodologies that can be applied to identify a coccidian species in addition to morphological analysis, from confirmation of host susceptibility, determination of parasite density, degree of pathogenicity and identification of inter- and intra-specific differences that provide relevant taxonomic adjustments. Finally, the advent of Molecular Biology, a recognized and established practice, mainly in the study of coccidian parasites in production animals, offers complementary information to microscopy and immunological methods, for example, since the sequencing of the microorganism's DNA serves as an additional study to taxonomy and phylogeny. Based on these facts, this work, in chapters I, II, III and IV, aimed to genetically identify coccidians from wild birds captured in the Atlantic Forest regions of southeastern Brazil, demonstrating the isolation of coccidia oocysts from fecal samples of wild birds, with subsequent DNA extraction in order to amplify and sequence different genic regions of the cox1, cox3 genes and fragments of the small and large subunit of the rDNA of the mitochondrial DNA, evaluating the applicability of these new sequences in the differentiation of species and phylogenetic studies.

Macrophages are phagocytic cells of the innate immune system present in several organs, composing a cell population involved in homeostasis and tissue protection. Macrophages can differentiate, according to the microenvironment, into cells capable of eliminating pathogenic microorganisms or into cells capable of restoring the balance of damaged tissue. Their plasticity highlighted them as targets of several drugs capable of orientate their response to numerous diseases. Leishmaniasis is a set of clinical manifestations that afflict 1 million people annually in lack of financial and health conditions. Macrophages are the host cells of the parasite Leishmania sp. harboring amastigotes that multiply until they disrupt the cell, thus establishing the infection. The treatment for this disease are toxic and expensive, making consumption and distribution on a large scale unfeasible. Natural products are an easily accessible, low-cost and safe option with pharmacological activities and their chemical structure allows modifications for their improvement. We evaluated the immunomodulatory activity of 3 natural products (-)-Guaiol, Piperine (AF1), 1,8-cineole and a synthetic piperine derivative: N4-cyclohexyl-1,2,4-triazol-3- thione (AF2) in different macrophages strains of murine P388D1 and RAW 264.7 and canine DH82. The compounds demonstrated a dose-dependent leishmanicidal activity in vitro for Leishmania amazonensis promastigotes with IC50 of 56.24 µM, 9.36 µM and 8.73 µM for (-)-Guaiol, AF1 and AF2, respectively. The cell lines showed viability above 70% in the treatments with the compounds (-)-Guaiol, AF1 and AF2. Preliminary results demonstrate that AF2 and 1,8-cineole at a concentration of 50 µM decreased the phagocytic activity of the DH82 after incubation with promastigotes. In RAW 264.7 cells stimulated with LPS, treatments with the compounds increased the production of nitric oxide (NO). In DH82, it is possible to observe a decrease in NO production in nonstimulated cells treated with (-)-Guaiol, different from the treatment with AF2 and 1,8- cineole, which increased in relation to the control. The murine strain P388D1 stimulated with LPS, where treatments with (-)-Guaiol and 1,8-cineole decreased NO production, while AF2 led to an increase in NO. ROS production in RAW 264.7 cultures without stimulus and treated with AF1, AF2 and 1,8-cineole increased ROS production in relation to control. In RAW 264.7 cultures stimulated with LPS, ROS production decreased after AF2 treatment. In LPS-stimulated P388D1, treatment with (-)-Guaiol and AF1 decreased ROS production. The DH82 cells decreased its ROS production in AF1 and increase in AF2. In stimulated cells, treatment with AF1 and 1,8-cineole led to a decrease while with (-)-Guaiol there was an increase in ROS production. The analysis of MHC and costimulatory molecules of RAW 264.7 murine macrophages stimulated or not with LPS+INF-γ and treated, did not show alteration in the expression of MHC II, CD80 and CD86. Preliminary results demonstrate that the compounds tend to decrease MHC II expression in unstimulated and LPS-stimulated DH82 cells. In P388D1 there was a decrease in the population of MHC I + cells treated with 60 µM of AF2 and AF2+LPS and an increase in the population of MHC II+ cells treated with AF2, AF2+LPS and infected with L.a and treated with AF2. Finally, real-time PCR results demonstrate that murine RAW 264.7 and canine DH82 macrophages increased mRNA expression of the cytokine IL-10 after treatment with (-)-Guaiol, AF1 and 1,8-cineole. Expression of IL10, IL-12, TLR4 and TLR9 in RAW 264.7 and DH82 cells decreased after treatment with the compounds. The unstimulated P388D1 cell line treated with (-)-Guaiol and AF1 led to a reduction of TLR4 expression in the cells. The results imply that the effect of the compounds may be different depending on the cell type and that they can modulate the production of NO, ROS and the expression of inflammatory and anti-inflammatory cytokines as well as altering the expression of MHC and Toll like receptors (TLR) in cells.

During the last 20 years, Brazilian researchers have played a leading role in efforts to improve our knowledge of the bioecology, the presence and importance of pathogens and the dynamics of tick populations associated with wild animals in South America. Many of these findings were provided in the form of molecular data, principally sequencing of molecular markers amplified using the polymerase chain reaction (PCR) technique. However, despite the advances achieved so far, there are still a number of barriers that prevent molecular techniques from becoming the standard approach for identifying ticks associated with wild animals in Brazil. Research carried out by our group within the PPGCV/UFRRJ, between 2018-2019, resulted in the development of a robust and low-cost system (which serves as an alternative to sequencing) for the identification, at the species level, of ticks of the Ixodidae family. The system is based on the Polymerase Chain Reaction-Restriction Fragment Length Polymorphism technique (PCR-RFLP), using a fragment of the mitochondrial gene that encodes 16S ribosomal RNA as target. To facilitate access to this system and to promote its use by other Brazilian researchers, an online tool called “TickCutter” was developed. The present project aimed to expand and improve this tool on two fronts. Firstly, a new tool called “COIsearcher” was developed using an alternative molecular marker, specifically Cytochrome C Oxidase I (COI), in order to address some of the limitations associated with the use of a single molecular marker. Secondly, through the inclusion of PCR-RFLP data (band patterns determined by in silico digestion of virtual “amplicons” (~460 nucleotides) of the 16S rDNA gene with the enzymes DraI and VspI), derived from tick species of the Argasidae family and/or from novel representatives of the Ixodidae family. A total of 35 new banding patterns were identified among 363 previously unpublished 16S rDNA sequences derived from ticks of the Ixodidae family (deposited in the GenBank between 06/09/2019 to 08/03/2021. The inclusion of these data served to increase the number of species identified by the tool. Regarding the Argasidae family of ticks, 31 banding profiles were identified among the 75 sequences obtained from GenBank (which represented 22/25 of the Argasidae species recognized in Brazil). However, it was observed that patterns derived from six species (Antricola guglielmonei, Ornithodoros capensis, Ornithodoros guaporensis, Ornithodoros kohlsi, Ornithodoros marinkellei and Ornithodoros mimon) generated conflicting identifications with banding profiles derived from the following species of the Ixodidae family: Amblyomma coelebs, A. geayi, A. tigrinum, Dermacentor nitens, Haemaphysalis leporispalustris, Ixodes auritulus and Rhipicephalus microplus. The new “COIsearcher” tool was developed in a very similar manner to the “TickCutter 16S” tool, and took advantage of some of the solutions developed to solve problems encountered during the development of the 16S module. However, some modifications were necessary due to the differences between the markers, particularly in terms of the size of the amplicons. The database of the new module was established based on the in silico digestion of virtual “amplicons” (709 nucleotides) of the COI gene, derived from tick species of the Ixodidae family, with the enzymes AluI and MboI. A total of 65 banding profiles were recorded among a total of 281 sequences derived from 27 of the 33 species of the genus Amblyomma. In addition, 23 additional banding patterns were identified from within 570 sequences deposited for the species Dermacentor nitens, Haemaphysalis juxtakochi, H. leporispalustris, I. auritulus, R. microplus and Rhipicephalus sanguineus. The interspecific discriminatory power of the “COIsearcher” tool was high. However, some conflicting identifications were detected, in a manner similar to what was previously observed with the “TickCutter 16S” tool. The solution found to resolve these conflicts was the identification of a third enzyme capable of generating discriminatory patterns at the species level. The application of this strategy allowed the differential identification of 86/88 of the banding patterns present within the “COIsearcher” database, a result considered to be highly satisfactory. It was concluded that the modifications introduced to the “TickCutter” platform will provide greater flexibility and discriminatory power to the current identification system and will offer a solution to most of the limitations associated with the use of a single molecular marker.

The nematode Caenorhabditis elegans has been used as an experimental model for studies in several sectors of biology and medicine due to the low economic costs for maintenance, cultivation and execution of experiments. The use of cryopreservation technique for this species can generate a source of genetically stable living tissues and cells for a variety of purposes. Currently, standard preservation techniques for C. elegans have shown difficulties to obtain results with high success rates in terms of survival. Thus, it is necessary to constantly develop new techniques and tests for new substances that can guarantee greater viability and stability in the conservation of cultures. The present study sought to define a cryopreservation methodology using different substances and associations in order to improve the maintenance and transport of the C. elegans culture. Three determining factors for survival and culture maintenance capacity of C. elegans during freezing and thawing were considered, namely the use of a preservative agent, the association with a gelling agent and the storage time in liquid nitrogen, using freezing in two steps. The use of glycerol and polyvinylpyrrolidone (PVP) showed satisfactory results in the association of gelling agents such as agar, carbopol and carboxymethylcellulose (CMC), respectively. Polyethylene glycol (PEG) and polysorbate 80 (T80) compared to the other substances tested, did not obtain significant efficiency, despite confirming their cryoprotective character between repetitions. There were no results demonstrating that these components evaluated individually brought benefits to the cryopreservation of C. elegans. Considering the unfolding of the factors among themselves for the survival of C. elegans, it was identified that the maintenance of individuals alive after thawing from seven days of storage is essentially dependent on the use of preservatives associated with some type of gelling agent.

The objective of the present study was to carry out an epidemiological analysis based on the detection of Anaplasma platys through the Real Time PCR technique – also known as qPCR – in blood samples from dogs domiciled in municipalities in the mountain region and low altitude regions located in the Rio de Janeiro state. After detection, the positive samples were submitted to statistical analysis in order to verify the association of epidemiological factors with the presence of A. platys DNA in the blood of dogs from these regions, as well as to elaborate statistical models that can predict variables related to the A. platys infection. Samples were collected from 456 dogs from four municipalities in different regions of the state – Petrópolis and Teresópolis, located in the mountain region – and Barra do Piraí and Paracambi – located in the southern Fluminense region and metropolitan region, respectively. From the blood collection, the genetic material was extracted and submitted to the detection of a specific fragment of the gltA gene, through the application of the Polymerase Chain Reaction – quantitative variation (qPCR) technique. An Epidemiological Questionnaire was developed and applied to dog owners, addressing factors that could be associated with the occurrence of Anaplasma platys infection in animals from the regions selected for the study. Factors such as sex, race, score, size, fur length, age, altitude, zone, climatic period, region of the household, health status of the animal, number of dogs in the household, presence of cats, presence of other domestic animals, presence of rodents, presence of wild animals, presence of cattle, presence of vegetation, street access, use of parasiticides, presence of yard, veterinary assistance, health assessment, housing, castration, flea infestation, infestation by ticks of the Rhipicephalus sanguineus species, infestation by ticks of the genus Amblyomma. Of the 456 samples tested by qPCR, 6.14% were considered positive for Anaplasma platys. Among the 28 animals in which A. platys was detected, 28.6% belonged to Petrópolis and 14.3% to Teresópolis – totaling 42.9% of cases in the mountain region – while 39.3% belonged to Paracambi and 17.8 % to Barra do Piraí – low altitude municipalities, representing 57.1% of occurrences in the regions in question. When analyzing the variables submitted to Logistic Regression, it was observed that dogs residing in low-altitude regions are 3.08 times more likely to be infected by A. platys than animals domiciled in the mountain region. As for the number of dogs in the household, it was found that, in households that have more than one dog, these animals are 3.40 times more likely to be infected with A. platys than a single dog in a household. This study showed that altitude and number of dogs in the home are epidemiological factors of relevance and interest in studies associated with the infection of dogs by A. platys in the municipalities of Petrópolis and Teresópolis – mountain region – and Paracambi and Barra do Piraí – low altitude regions.

Livestock productivity can be significantly diminished by the effects of gastrointestinal parasitism. As well, it can impact dogs and cats and, consequently, humans due to its zoonotic potential. The need for food without pesticide residues, better working conditions and an increase in cases of anthelmintic resistance opens the way for sustainable alternatives in the control of helminthiases, such as essential oils. In this context, Caenorhabditis elegans is used to research phytoinputs with anthelmintic activity at different stages of development through toxicological evaluation in different strains of C. elegans. The aim of the present study was to evaluate the activities of essential oils against C.elegans and thus to assess their potential for use as an anthelmintic product. The following EOs were used: Cymbopogon flexuosus, Eugenia caryophyllus, Illicium verum, Mentha piperita, Mentha spicata, Pelargonium graveolens and Thymus vulgaris. These were solubilized in DMSO. In vitro lethality tests were carried out on adults of wild strain (N2) and strains resistant to ivermectin, albendazole and levamisole, egg hatchability and egg laying. The essential oil of Illicium verum showed better nematicidal activity in N2 adults and inhibition of egg laying. On the other hand, the essential oil of Pelargonium graveolens exhibited better inhibition of egg hatching. In general, compared to the N2 strain, the essential oils of I. verum, P. graveolens and T. vulgaris showed sensitivity at different degrees of concentrations. However, in ivermectin-resistant strains and exposed to high concentrations in the levamisole-resistant strain, P. graveolens essential oil did not exhibit efficacy.

Fleas are hematophagous ectoparasites that feed blood on warm-blooded animals. They have great importance in parasitology as vectors of different etiological agents capable of causing diseases in different vertebrate hosts. The use of entomopathogenic nematodes (EPNs) has been studied as an alternative in the biological control of arthropods, in an attempt to reduce, for example, environmental contamination and resistance to chemical substances. The present study aims to evaluate the susceptibility and mortality of seven-day-old larvae of Ctenocephalides felis felis by entomopathogenic nematodes of the species Heterorhabditis amazonensis, strain NEPET11, under different experimental conditions. The experiment was divided into two stages, in which 70 seven-day-old larvae of C. felis felis were used, divided into seven plates, six of which were exposed to the solution containing the NEPs and a control. The Petri dishes submitted to infection contained 10 flea larvae and 600 μL of solution with infective juveniles (IJs) of entomopathogenic nematodes (120 NEPs/larva), whereas the control contained 10 flea larvae and 600 μL of distilled water, both were kept in climatized chamber at a temperature of 25±1ºC, 70-80UR, being observed for a period of 48 hours to assess larval mortality. The infection was confirmed through the dissection of a flea larva from each Petri dish and the observation of the presence of juveniles and/or adults inside it. In the second stage, 10 Petri dishes containing seven-day-old larvae of C. felis felis infected by IJs of H. amazonensis NEPET11 at a concentration of 120 IJs/flea larva were used. Five Petri dishes were kept in BOD at a temperature of 25±1°C, 70-80UR and the other five were kept at room temperature. In addition to these, there were two control Petri dishes containing 10 flea larvae and 600 μL of distilled water, kept under the same conditions. The percentage of flea larvae mortality was evaluated by analysis of variance (ANOVA), followed by Tukey's test with a significance level of 5% (p≤0.05) using the SISVAR statistical program. In the first experimental stage, it was possible to verify the infection of flea larvae by H. amazonensis, with an average mortality percentage equal to 98.33%. In the second experimental stage, the infection was also successful in the two conditions tested. The mean percentage of mortality after 48 hours of observation was 96% for plates kept in BOD and 98% for those kept in ambient conditions, with no significant difference between them. In both stages it was not possible to recover the infective juveniles in White's trap. The present study suggests that C. felis felis flea larvae are susceptible to infection by H. amazonensis NEPET11, and that EPN has a high virulence for the stage of the evaluated life cycle of this flea species, being efficient when used under experimental conditions.The EPNs can be a promising tool for the biological control of fleas, however, more studies should be carried out using both the host species and the nematode species, since this was the first study evaluating the infection of C. felis felis larvae by H. amazonensis.

The emergence and spread of antimicrobial resistance is one of the three main threats to Public Health in the 21st century and must be analyzed in an integrated One Health approach, as it is a health risk shared by people, animals and the environment. Despite understanding the multifactorial origin of antimicrobial resistance, little is known about the contribution of environments aimed at the production, maintenance and care of animals in disseminating this phenomenon. Among these, the necropsy space represents a point of cohesion, being a place of extreme relevance for research and understanding of the circulation of bacterial microbiota and its resistance genes. The present study evaluated the occurrence of superbugs in samples of animals necropsied at the Federal Rural University of Rio de Janeiro, considering the priority criteria established by the World Health Organization (WHO), where Pseudomonas aeruginosa and Acinetobacter baumannii resistant to carbapenems and bacteria of the order Enterobacterales resistant to carbapenems and producing ESBL were classified as level 1 or critical; vancomycin-resistant Enterococcus faecium, methicillin-resistant Staphylococcus aureus and with resistance or intermediate resistance to vancomycin, and Salmonella spp. resistant to fluoroquinolones as level 2 or high priority; and Shigella resistant to fluoroquinolones and S. pneumoniae not susceptible to penicillin as level 3 or medium priority.

Of the 198 samples collected from 45 animals, being 20 companion animals, 20 production animals, and three wild ones, 325 strains were isolated, of which 51,38% (167/325) were Enterobacterales, 31,69% (103/325) Staphylococcus spp., 12,62% (41/325) Enterococcus spp., 2,46% (8/325) Streptococcus spp. and 1,85% (6/325) BGNNF. MALDI-TOF proved to be an efficient tool for bacterial identification, especially in Enterococcus spp. and Enterobacterales. The agreement between biochemical, proteomic and genotypic techniques in identifying Staphylococcus spp. was 80,58%, which confirms the importance of the association between different diagnostic methods for the characterization of this genus, leading to the correct direction of the resistance analysis. 8,74% (9/103) of Staphylococcus spp. showed phenotypic resistance indicative of PBP2a production, with detection of the mecA gene in all strains. Phenotypic resistance to vancomycin was evidenced in E. faecalis, with detection of the vanB gene. In 29,13% (30/103) of Staphylococcus spp. there was detection of the blaZ gene. The ESBL phenotypic research was performed through screening and confirmatory antibiograms recommended by the CLSI. 11,98% (20/167) of enterobacteria showed resistance to beta-lactams in the screening antibiogram and 80% (16/20) of them were positive in the confirmatory test. The search for genes encoding ESBL revealed the presence of bla_SHV in 10,18% (17/167), bla_TEM in 6,59% (11/167) and bla_CTX-M in 4,19% (7/167). There was no detection of carbapenemase-producing strains. No mcr genes were detected. These results reveal species characterized as critical superbugs in the necropsy environment and reinforce the need to monitor these strains in the veterinary environment, not only for the adoption of adequate control and treatment measures for the animals but also for the implementation of safe protocols for the disposal of their carcasses.

Fleas are among the ectoparasites that most affect companion animals. They produce and transmit diseases that pose a danger to the health of animals and humans. The control of these insects involves the use of insecticides. Insect growth disruptors are a group of insecticides that can act as an alternative in flea control. The objective of this work was to evaluate the activity of novaluron and pyriproxyfen on Ctenocephalides felis felis larvae in in vitro tests during cycle interruption, determining the lethal concentration (LC) of the two compounds. Initially, the larval substrate impregnation method to be used in the experiment was evaluated. For this, a 400 ppm pyriproxyfen solution was prepared and an impregnation volume of 200 µL was used for two grams of substrate. The result was satisfactory, and the methodology did not interfere with the result, with 100% inhibition of the flea development cycle and there was no interference in the development of the cycle for control. To carry out the experiment with the disruptors, the aforementioned methodology was used. Different concentrations of pyriproxyfen (0.049 to 25 ppm) and of novaluron (0.001 to 5.0 ppm) were prepared. After substrate impregnation, the first instar larvae were exposed to the treated larval substrate. The material was kept under controlled conditions for a period of 21 days. Qualitative and quantitative assessments were performed and lethal concentrations were determined through Probit analysis. In the evaluation of the test with pyriproxyfen, an efficiency of 100% was verified in concentrations greater than 1,563 ppm. From the pupae, live and dead fleas were recovered, the dead ones showed changes in morphology, color and size. It was observed that there was a significant difference between the amount of live fleas recovered from the control and treated groups, in addition to a greater number of females compared to males. The calculated lethal concentration LC₅₀ and LC₉₀ of pyriproxyfen was 0.07 and 0.27 ppm, respectively. In the novaluron test there was 100% efficacy at the highest concentration (5.0 ppm). Some dead fleas were recovered from the pupae, they showed changes in the exoskeleton, color and limbs, but without visible changes in size. The amount of live fleas recovered was discontinuous between the concentrations, the control had a significant difference with the treated ones from the concentration 1.0 ppm. Regarding sex, they were mostly females, except in the concentration of 1.75 ppm. The calculated lethal concentration LC₅₀ and LC₉₀ of novaluron were 0.25 and 2.29 ppm, respectively. Based on the results of this work, it can be stated that both pyriproxyfen and novaluron have shown activity on Ctenocephalides felis felis larvae, interrupting the normal developmental cycle of the flea. In addition, novalorun was tested for the first time in a flea laboratory colony in Brazil, proving to be an alternative for the flea control.

Hemoparasites are responsible for causing great damage to the health of animals, whether they are production, companion or wild. Many pathogens have zoonotic potential and represent an important public health topic. In this sense, the objective of the present study was to detect the infection by bacteria of the Anaplasmataceae family in wild animals from the Clínica de Reabilitação de Animais Silvestres (CRAS) of the Universidade Estácio de Sá, municipality of Rio de Janeiro, state of Rio de Janeiro, Brazil. Blood samples were collected from 148 birds, 52 mammals and 20 reptiles, totaling 220 animals, from January 2019 to August 2021. The samples were identified e conditioned to posterior processing at the Laboratory for Cultivation of Cells and Hemoparasites of the Federal Rural University of Rio de Janeiro (UFRRJ), where they were subjected to DNA extraction. The samples were subjected to nested-PCR and conventional PCR techniques for the detection of the target DNA corresponding to 16S rDNA, gltA e GroEL genes of Anaplasmataceae, as well as specific targets to the genus Anaplasma spp. (16 rDNA and rpoB) and Ehrlichia spp. (Dsb). After all assays, the Anaplasma sp. DNA was detected only in one specimen of Coendou spinosus by the amplification of the 16S rDNA and GroEL genes. At sequencing of the amplifying products, the corresponding sample positive to Anaplasma sp. presenting the follow identity: 99,4% with Anaplasma sp. (830/835) decribed in dromedary in 16S rDNA gene and 81,11% with Anaplasma platys at GroEL gene. In this way, it demonstrates that wild animals can be infected by these bacteria, can act as a reservoir and maintainers of the epidemiological cycle of important pathogens for animal and human health.

The present study evaluated the compatibility between the isolates of Beauveria bassiana (LCM S19 and LCM S20) and Metarhizium anisopliae (LCM S01) with the essential oil of Illicium verum, the effectiveness of the isolates and the oil separately in the control of larvae, pupae and adults. of Aedes aegypti and also the in silico analysis of the oil. For compatibility between fungi and oil, the germination of the isolates and the growth of colony diameter were evaluated. For the germination percentage, 10 μL of fungal suspension associated or not with oil were inoculated into plates containing BDA + 0.5% chloramphenicol. After 16 hours of incubation, the percentage of germination was verified. To evaluate radial colony growth, 10 μL of each suspension were inoculated in the center of plates containing Oat medium and measured for 9 days. For larvicidal and pupicidal activity, groups containing 10 larvae or pupae of A. aegypti were kept in disposable cups containing 15 mL of essential oil solution at 40 ppm, 60 ppm, 80 ppm and 100 ppm or fungal suspension at concentrations of 104, 105, 106, 107 and 108 propagule/ml. The survival of larvae and pupae was monitored daily. In the biological assay with adults, 10 mosquitoes were transferred to disposable cups containing filter paper, previously impregnated with 1mL of essential oil concentrations or fungal concentrations, covering the entire inner surface of the cup. The control group of all experiments contained dechlorinated water with 0.03% Tween 80. The data were submitted to the normality test, and after submitted to the Analysis of Variance using Tukey's test. The Kaplan-Meier and Log-rank tests were used to analyze the survival curve and the mean survival time (S50). The significance level was 95% (P≤0.05). The concentrations of I. verum OE did not interfere with conidia germination and did not change the colony growth of the isolates LCM S01 (Metarhizium anisopliae), LCM S19 and LCM S20 (Beauveria bassiana). In the biological assay with the O.E., it showed larvicidal activity at all concentrations studied and pupicidal activity at concentrations of 60 and 100 ppm. In silico analysis showed that 79.96% of the oil is composed of (E)-anethole that can affect 89 possible targets in the arthropod. In the biological assay with the fungal isolates, all, regardless of the propagule used, were able to reduce the survival rate of the larvae at a concentration of 108 propagule/mL. Pupicidal activity
VII
was demonstrated by all isolates at different concentrations. The LCM S01 isolate was able to reduce adult survival at three different concentrations. The isolates LCM S19 and LCM S20 were also virulent at the highest concentrations for adults. It is concluded that the fungal isolates and the essential oil are compatible and good options for controlling larvae, pupae and adults of A. aegypti.

Dopamine (DA) is a biogenic monoamine that modulates ticks and insect hemocytes related to the immune system of these arthropods. However, the detailed role of DA in the immune response of ticks is yet to be elucidated. The present study analyzed the effect of a DA receptor antagonist when ticks were challenged with the entomopathogenic fungus Metarhizium anisopliae. Our study evaluated the survival and biologic parameters of Rhipicephalus microplus, its hemolymph’s phenoloxidase activity, the hemocytes’ phagocytic index, DA detection in the hemocytes, and hemocyte quantification. Seven groups were formed as follow: control ticks (CTR), ticks inoculated with phosphate buffer (PBS), inoculated with antagonist 1ɳM (SCH A), 1μM (SCH B), M. anisopliae conidia (MA) and associations (SCH A + MA and SCH B + MA). There was a reduction in survival comparing the means of MA (7 days), with SCH A + MA (5 ½ days; P=0.0253) and SCH B + MA (4 ½ days; P=0.0291). The egg production index (IPO) was lower in the SCH A + MA (9.6%) and SCH B + MA (8.9%) compared to CTR (49.7%; P<0.0001) and the MA (25.1%) (P=0.0098; P=<0.01). The phagocytic index observed in ticks treated with Metarhizium alone was 57.3% in contrast to 18.1% (SCH A+MA) (P<0.0001) and 25.3% (SCH B+MA) (P<0. 0001). No changes in phenoloxidase activity and DA levels in R. microplushemocytes were detected in the presence of the antagonist. The quantification of hemocytes treated with antagonist associated or not with the fungus was similar. This result supports the hypothesis that DA is crucial in the tick defense process, changing the phagocytic capacity of hemocytes and the susceptibility of ticks to infection by entomopathogenic fungi.

The aim of this survey is to investigate the interaction between domestic dogs and the forested areas around the Palmares Environmental Protection Area (APA Palmares), a community located in the Atlantic Forest in the State of Rio de Janeiro. The APA area is located at altitudes ranging from 831 to 985 meters above sea level. Dogs were classified into three categories, domiciled, semi-domiciled and stray dogs. It was observed that the domiciled dogs, although kept in fenced yards, come across with some wild animals, which visit the homes, probably looking for food or even shelter as in the case of skunks and hedgehogs. Semi-domiciled and stray dogs frequently visit the forested area located close to the residences, therefore they are in contact with wild animals. According to information from owners, some dogs were treated with ectoparasiticides whenever owners detect infestations by ectoparasites.  Dogs were examined as sentinel animals to diagnose the presence of common parasitic ticks of wild animals. Tick collections took place monthly from January to December, 2019. Overall, 60 (33.9%) positive dogs were diagnosed among 177 examined dogs. Three species of ticks were identified: Rhipicephalus sanguineus, Amblyomma aureolatum and Amblyomma ovale. Overall, 279 adult ticks were collected, distributed as follows: 143 (51.3 %) R.sanguineus, 135 (48.4 %) A. aureolatum and a single female of A. ovale. Tick/dog interaction was evaluated in terms of housing, treatment and host sex using the Chi-square test for R.sanguineus and A. aureolatum. The Chi square result indicated dependence of both species for the housing and sex and independence for the treatment.

Dermatopathies are commonly observed in rabbits caused by ectoparasites, mainly Psoroptes ovis and Leporacarus gibbus, responsible for developing otitis externa and the presence of black spots on the fur, respectively. Although several treatments have already been described for the control of parasites in rabbits, new compounds and routes of administration are objectives of studies, with the aim of developing safer and more practical products for use. However, the present study aimed to evaluate the efficacy of a group of molecules, isoxazolines (afoxolaner, fluralaner, lotilaner and sarolaner), in controlling psoroptic mange and infestation by L. gibbus in naturally infested rabbits. To determine the effectiveness, clinical (determined by the score of the lesions: 1 to 4) and parasitological (through the evaluation of crusts and fur: on days 0, +3, +7, +14, +21, +28 and + 35). A total of 36 rabbits infested with both mites will be used throughout the experiment. Divided into 5 experimental groups and 1 control group. So far, 2 groups have been executed. The animals were clinically evaluated for lesional score and subjected to parasitological examination on days 0, +3, +7, +21, +28 and +35 post medication, with 1 group receiving fluralaner therapy at a dose of 25mg / kg and another group was medicated with lotilaner at a dose of 20mg / kg, both were treated orally in a single dose. The control group was submitted to the same evaluation and examination procedures, using placebo medication. Thus, it will occur with all other groups to be treated. Those treated with fluralaner after 72 hours were found to be negative in the parasitological examination for both mites. Those treated with fluralaner after 72 hours were found to be negative in the parasitological examination for both mites. The rabbits treated with lotilaner, after 72 hours, only 2 animals presented live mites in the parasitological exam for P. ovis and 1 positive rabbit for L. gibbus. In both groups, no adverse clinical reactions were observed. It is expected to prove that this new class can be used in rabbits, being an alternative for the treatment of these mites in this species.

 Over the last decades, multiple factors such as the strengthening of the relationship between humans and pets, environmental changes resulting from the urbanization of peri-urban and rural areas, the significant intensification of animal production have led to a significant change in the human-animal dynamics with consequently increased circulation of bacterial pathogens between humans and animals and the emergence and re-emergence of diseases. Identifying emerging bacteria in the routine of veterinary diagnosis has been presented as a great challenge. Generally, these pathogens do not have reference studies from samples from the animal environment, and sometimes methodologies developed for samples of human origin are not effective for its identification. Another aspect associated with this emergence is the circulation of resistance genes, pointing to the importance of characterizing the antimicrobial resistance of bacterial species from different animal environments. This work characterized bacterial species and their respective resistance profile from wild animals, poultry production, and clinical laboratory samples of different species. Samples from different wild animal species, such as maritacas, jabutism, cachorro do mato and mão pelada, provided 103 isolates of the Enterobacterales family, 28 Staphylococcus spp, 2 Streptococcus spp and 1 Enterococcus spp. A strain of Pantoea dispersa isolated from maritaca was detected harboring the colistin resistance gene mcr-9. From the tortoise samples, 2 samples were detected presenting the blaTEM resistance gene, 1 presented the blaCTX gene and 1 presented both genes. Among the samples of poultry production, Enteobacterales strains revealed 45.45% (20/44) of ESBL-producing strains, with 35% (9/20) blaSHV, 20% (4/20) blaCTX-M, 15% ( 3/20) blaTEM, 10% (2/20) showing the blaSHV and blaCTX genes, and 10% (2/20) showing blaSHV and blaTEM, simultaneously. Among the 51 strains of Enterococcus spp (51 strains), 1 strain of E. faecium from chick cloaca was identified with the vanB gene and in 1 strain of E. faecalis from adult chicken, vanA and vanB were identified simultaneously. Of these same 51 strains, 23.53% (12/51) showed resistance to streptomycin. Among the strains of Staphylococcus spp. 1 isolate with the mecA gene was detected. The poultry litter was also evaluated and 01 positive sample for the blaVIM gene was detected. The 35 samples of Acinetobacter spp. were identified by proteomic, genotypic and sequencing analysis, to assess the difference between the techniques, the Kappa test was used to compare MALDI-TOF and PCR, MALDI-TOF and rpoB, PCR and rpoB, and between the three techniques. The resistance profile of these samples was also evaluated, with 54.28% (19/35) classified as MDR, 51.42% (18/35) had 01 or more ESBL genes, with 04 strains with the blaCTX gene, 01 strain with blaSHV, 09 strains with blaTEM, 03 strains with blaCTX and blaTEM and 01 strain with blaSHV and blaTEM. The present work sought to identify bacteria and detect resistance genes in different veterinary environments. This is a big challenge mainly because there are still few works and phenotypic analysis standards on animal themes. The role of wild, production, or companion animal has great participation in disseminating resistance gene in the environment, emphasizing One Health and generating an alert as to the focus of new studies to understand and evaluate veterinary environments.

Sandflies are the main vectors of leishmaniasis, a neglected zoonotic disease
of great importance for public health, caused by the protozoan of the genus Leishmania.
The aim of this study was to robustly assess the dynamics, the spatial distribution, the
seasonality, food predilection and infection rate of sandflies captured in
Seropédica, in Rio de Janeiro. Collections were carried out in five districts of the city: Valão
das Louças, São Miguel, Santa Sofia, Campo Lindo and Fazenda Caxias. the sand flies
were captured in the period from August 2016 to July 2018 and the number of specimens
was correlated to climatic variables (temperature, humidity and precipitation) for the study
of seasonality. Males were identified by morphology and females were
for molecular analysis. For carrying out the DNA extractions of the
females, four extraction protocols were tested: Phenolchloroform, HotShot, Salting out
and Blood & Tissue QIAGEN® commercial kit. To compare the data, a
amplification intensity scale obtained in conventional PCR, with gene target
constitutive coxI. Analysis of food source identification and infection rate of
sandflies by Leishmania spp. were carried out with all females fed.
captured. For the identification of food sources, the multiplex PCR technique was applied
real-time targeting on the cytochrome b gene of several animals. The determination of the rate of
infection of sandflies was performed by real-time PCR technique with target on
mkDNA gene of the protozoan. 

The positive sandfly species were identified by
gene sequencing by the Sanger method. Infection rate data and species of
positive sandflies were interpolated with the geographic coordinates of each point of
capture in QGIS software 3.14.15. The two main species identified by morphology
were Nyssomyia intermedia and Migonemyia migonei. A positive correlation was observed and
moderate between the number of sandflies and the climatic variables of temperature and
rainfall index. The most cost-effective extraction protocol for
sandflies was phenolchloroform, with the highest amplification rate and lowest mean of
inhibition. Of the total of 360 fed females, 6.11% were positive for the gender
Leishmania and according to genetic sequencing, belonging to species N.
intermedia Nyssomyia whitmani and M. migonei. According to the source detection analysis
food, the three main species of preference for sandflies were humans, the
chickens and dogs. According to the Kernel map, it was possible to observe positive points of
greater intensity in rural areas of the municipality. According to the study, it was possible
identify that rural areas have a higher risk of infection for the disease and that
main species of vectors related to LTA transmission in Rio de Janeiro, then
present in the municipality of Seropédica, with seasonal variation under the influence of temperature and
precipitation, it is essential to adopt a control measure to prevent
effective, the emergence of new cases of the disease in the region.

Theileria equi is a protozoan that infects horses, persists throughout the animal's life, and is challenging to eliminate from the organism through the use of drugs indicated to treat the disease, making it evident that preventive measures are of paramount importance in equine breeding. The tick Rhipicephalus microplus is the only tick species known as a biological vector of T. equi for horses in Brazil, proven by experimental studies. The vector competence of ticks is closely linked to the ability of pathogens to evade tick defense mechanisms. Thus, understanding the molecular and cellular interactions at the tick-pathogen interface may provide new targets for blocking the transmission of T. equi by R. microplus and other fundamental metabolic pathways for the infection, multiplication, and persistence of T. equi in the tick. Due to the lack of information on the mechanisms involved in the tick-pathogen interaction, the present study aimed to seek greater knowledge about: the transcriptional profile of the salivary gland of R. microplus in response to T. equi infection; and also the differential expression of participants of signaling pathways that act in the immune defense of the tick gut when challenged with T. equi under experimental conditions. The data generated significantly contribute to the advancement of scientific knowledge, as they provide genetic and molecular information of high technological value for exploration in many other studies for the parasite mentioned above.

The small animal veterinary clinic plays an important role in the diagnosis, treatment, control and prophylaxis of the main diseases that affect dogs. In this context, hemoparasitosis represent the most common parasitic disease, regardless of gender, age and race, being considered endemic in the urban environment. Despite efforts to combat this disease, many animals still suffer from the disease, as many of the methods to control its vectors have not been effective. Due to this reality, the first chapter aimed to analyze the laboratory and epidemiological aspects involved in the prevalence of blood parasites in the urban environment. From a laboratory point of view, animals with thrombocytopenia, activated and anemic monocytes were the ones that presented the highest risk of being infected by hemoparasites. From an epidemiological point of view, the most relevant aspects were the size of the animal and the presence of ticks. The second chapter presented as objective to analyze the seroprevalence of the main endemic hemoparasites in the small animal clinic in 5 veterinary clinics using a commercial kit ELISA test. Of the 1132 selected exams, the seroprevalence of hemoparasites was 44.3% for at least one agent, being Ehrlichia spp. (33.3%), the most prevalent gender, followed by Dirofilaira immitis (7.1%), Anaplasma spp. (3.7%) and B. burgdorferi (0.3%). Regarding co-infection cases, the seroprevalence in the study population was 16.2% for at least two bioagents analyzed, with the highest seroprevalence of Ehrlichia spp. + Anaplasma spp. (7.9%), followed by Ehrlichia spp. + D. immitis (6.4%). The third chapter aimed to evaluate the accuracy of three diagnostic techniques for the agents, Ehrlichia spp. Anaplasma spp. and Babesia spp. in the routine of the small animal clinic. Samples from 70 domiciled dogs were collected for convenience. Comparisons were made between positive and negative animals in serological and molecular tests regarding the main signs and symptoms. All animals included presented at least 3 clinical signs and symptoms and laboratory alterations suggestive of hemoparasitosis. Blood samples were analyzed by smear, serology and qPCR. Sensitivity, specificity, positive and negative predictive values were calculated. The prevalence of Ehrlichia spp was 40% and 37.1% Anaplasma spp. was 34.3% and 14.3% and Babesia spp. was 8.6% and 14.3% in the ELISA and qPCR tests, respectively. The general parasitism of the 3 agents for the 3 techniques ranged from 1.43% to 82.86%. The ELISA test showed the highest sensitivity for the diagnosis of Ehrlichia spp. (Se= 37.93%) and the lowest sensitivity for the diagnosis of Anaplasma spp. (Se= 8.33%). The dot-ELISA test showed the sensitivity (Se=33.33%) and the highest PPV (93.33%) for the diagnosis of Babesia spp. Animals with epistaxis were 100% positive for Ehrlichia spp. in ELISA and 80% in PCR. Real time PCR was considered the fundamental technique for the diagnosis of Ehrlichia spp. Anaplasma spp. and Babesia spp. being possible to determine to select animals with subclinical, acute and chronic disease.

Tick-borne diseases are among the most significant occupational health problems, and military personnel are at particularly high risk due to exposure. This study aimed to analyze the diversity of ticks in military training areas across different municipalities and biomes in the Southeast region of Brazil; to analyze the diversity of ticks collected from domestic and wild animals and human beings in a single military training area; to molecularly detect Rickettsia spp. in free-living tick samples; and to carry out a “survey” of behaviors, attitudes, and practices in the military community regarding the issue of ticks. The research on tick diversity was carried out by means of flannel trawling, flagging, and/or collection from operators’ clothing in 66 areas used for training by six military institutions. A total of 9,374 ticks were collected, representing five genera and 10 species, namely Amblyomma sculptum, A. dubitatum, A. brasiliense, A. longirostre, A. aureolatum, Amblyomma spp., Dermacentor nitens, Riphicephalus spp., Ixodes spp., and Haemaphysalis spp. The study of tick species diversity infesting multiple hosts was carried out in the military area of Resende Municipality, Rio de Janeiro State, in which a total of 1,331 ticks from 111 animals (29 domestic and 82 wild) and 67 ticks parasitizing 31 human military volunteers were collected. The ixodids comprised 14 species distributed in four genera: A. aureolatum, A. auricularium, A. brasiliense, A. calcaratum, A. dubitatum, A. incisum, A. longirostre, A. ovale, A. sculptum, Amblyomma spp., Ixodes loricatus, Ixodes schulzei, D. nitens and R. sanguineus. Molecular detection of Rickettsia spp. in samples of free-living ticks collected in military training areas yielded 11 positive samples, ten with similarity to Rickettsia bellii (in nine nymphs of A. dubitatum and one nymph of A. sculptum) and two similar to Rickettsia sp. (strains 463 and 464, in two nymphs of A. dubitatum). The detection of ticks positive for Rickettsia spp. suggests an occupational hazard for the military, increasing the risk of human cases. An exploratory survey on the subject was also developed and administered to 655 military personnel from five municipalities in the Southeast region, verifying that ticks were the type of “pest” that most affected military personnel in the field, with most military personnel having already experienced infestation at least once in the region where they serve, some recently. The present work reveals a high frequency of tick species in most military training areas, associated with the traces and presence of wild animals (most often capybaras), suggesting a predisposition to the occurrence of human cases of spotted fever. It is the first record of infestation by A. sculptum, A. brasiliense, and A. aureolatum in military personnel in the country, warns of the hazard of exposing military personnel to ticks infected with Rickettsia spp. in military training areas, and establishes the first base on the knowledge, perceptions, attitudes, and practices of the military in relation to the subject in Brazil. This One Health research endeavor will contribute to a better understanding of these vectors, their epidemiological network, and the related occupational hazards in military institutions, thereby facilitating sanitary management and prevention measures in these areas.

Stomoxys calcitrans is a cosmopolitan and hematophagous dipteran. Its feeding habit is defined as aggressive and persistent, having different warm-blooded species as hosts. It is considered a pest of economic importance to cattle and other animals, causing reduction of the weight and milk production in livestock. In rearing environments, members of the genus Stomoxys play an important role in the epidemiology of transmissible diseases, being mechanical vectors of several pathogens. Knowing and understanding the cellular components of the fly's immune system and the relationship between the S. calcitrans organism and the pathogens it transmits is essential to clarify the pathways involved in these interactions, identifying the best research targets for the development of control techniques for this livestock pest. Thus, in this work we isolated and quantified the number of circulating hemocytes in different evolutionary phases of the fly, identifying larger populations in the evaluated field individuals. From the isolation, we were able to identify and characterize these hemocytes in four types, namely: prohemocytes, granulocytes, plasmatocytes and oenocytoids. Prohemocytes were in the lower number of cells in the assays, and plasmatocytes were the most abundant cells. Responding to an experimental infection by Herpetomonas muscarum, the time of greatest hemocytic response ocurred at  two hours after the injection of trypanosomatid. In general, the populations of different types of hemocytes increased after infection, with the exception of prohemocytes, which decreased. The production of extracellualr traps (ETs) by S. calcitrans stimulated with LPS was also evaluated, being possible to identify the release of DNA after stimulation. We identified the presence of a parasite in the hemolymph of S. calcitrans in individuals from the field, larvae in the laboratory and adults generated in a colony. It was isolated and identified at the species level by DNA sequencing, being the trypanosomatid called H. muscarum. To evaluate its interaction aspects with its host, we initially performed its growth curve to understand its development and identify the best times for laboratory tests. The interaction of this parasite was analyzed at the intestinal level, by in vivo and in vitro experiments, demonstrating that it is able to interact and settle in the intestine even after a few hours of interaction. Finally, we evaluated developmental aspects of the stable fly infected orally with H. muscarum. The longevity of individuals was not significantly altered, as well as their oviposition. However, the viability of the eggs was affected, thus reducing their hatching and, consequently, impairing the development of larvae in pupae and pupae in adult individuals. Thus, the reproduction of S. calcitrans is affected when it is infected by H. muscarum. More evaluations should be carried out, for a better understanding of the hemocytic reactions of S. calcitrans, but this work already elucidates the types of hemocytes present in its circulation and methods of their extraction. The interactions of the fly with the trypanosomatid reveal a potential use in two aspects, namely: (1) use of the trypanosomatid for controlling the population of S. calcitrans; (2) use of these organisms as an experimental model for understanding the relationship between dipterans and the trypanosomatids that infect them, through laboratory tests.

Canine auricular cholesteatoma, a disease unknown to many veterinarians, is an epidermoid cyst that forms in the middle ear cavity, often as a complication of otitis media, but the etiopathogenesis remains controversial and unknown in veterinary medicine. There are few reports in the world and in Brazil; possibly because it is underdiagnosed. The objectives of the study were: (i) to report the epidemiological data of animals with the disease treated between the years 2017 to 2022 with cholesteatoma; (ii) propose the best diagnostic method for cholesteatoma; (iii) correlate the prognosis of the disease with the therapy adopted. A total of 105 records were evaluated, 100 of the canine species and 5 of the feline species, obtaining information on the history, anamnesis, clinical evolution and signs present. As well as the results of computed tomography, otoendoscopy and histopathology with confirmation of the diagnosis of cholesteatoma. In general, histopathology is the most used method described for confirmation of cholesteatoma, however the most known material collection is done by surgical procedure. It is concluded that there is a growing number of cases of cholesteatoma in dogs and otoendoscopy in association with computed tomography is effective methods for early diagnosis.

Periodontal disease is the most common oral disease affecting both men and animals, predominantly occurring in dogs over three years old. It’s result of the progressive inflammatory response due to the accumulation of bacterial plaque and the signs and symptoms include halitosis, pain, accumulation of dental calculus, enlarged of the gingival pouch, severe inflammation and infections, destruction of the gums, teeth and bones, bacteremia with progression to septicemia. Plants extracts like the one from Psidium guajava L have antimicrobial activity and the potential for use in the prevention and treatment of oral diseases. Popularly known as guava tree [goiabeira] in Brazil, the Psidium guajava L. is used worldwide in traditional medicine and in the food industry. The objective was to characterize the antimicrobial and antioxidant activity of the aqueous extract of the guava leaf and the development and quality control of a modified oral mucoadhesive release formulation containing extract, for the treatment and prophylaxis of periodontal disease in dogs. Microbiological methods such as disk diffusion and broth microdilution, chemical methods such as thin layer chromatography, antioxidant analysis (DPPH), total phenolics were used. In addition, uniformity of weight, pH, swelling index, disintegration time, infrared and sterility of the films. The aqueous extract obtained yield of 1.2%, found in thin layer chromatography the presence of flavonoids, antioxidant capacity of EC50 = 140 µg.mL^-1 and total phenolics of 17.02 ± 6.87 mg equivalent of gallic acid per g of dry extract. In the evaluation of microbial susceptibility, maximum inhibition halos (mm) were obtained for resistant S. aureus as 18 ± 3.46, for sensitive S. aureus 16 ± 0, for resistant S. pseudintermedius 17.3 ± 1.15, for sensitive S. pseudintermedius 18 ± 0 and for sensitive beta-hemolytic Streptococcus 21 ± 2.00, with a concentration of 3.6 mg / cm² at medium. In the broth microdilution test, the minimum inhibitory concentration (MIC) for all strains of 8.8 mg.mL-1 and the minimum bactericidal concentration (CBM), only for S. pseudintermedius (resistant) was 8.8 mg. mL-1, for the other strains it was greater than 11.4 mg.mL-1. The buccal film information presents homogeneity and malleability, having a pH compatible with the buccal one (from 6.35 ± 0.07 to 6.45 ± 0.18), degree of swelling from 27.0 ± 62.8% to 369.8 ± 28.0%, and the disintegration time in approximately 4 minutes. The physical infrared test evidenced that there were no interactions between the components and the base itself. The results demonstrated that it was possible to develop a polymeric film of oral dissolution with release in melting having antioxidant and antimicrobial capacity.

Allergic flea bite dermatitis (DAPP) is a common dermatological disease in Brazil, which brings discomfort both to the animal and to the guardian for seeing the animal with intense itching. The diagnosis is usually made through anamnesis and physical examination. These methods are questioned by the owner, now more enlightened and demanding, who seeks a conclusive, objective and quick diagnosis. Some studies have been carried out with the intradermal test (TID) to help diagnose this pathology, but the results differ. An alternative would be the use of the prick test (PT), which has been widely accepted in human medicine as a fast, safe, reliable and low cost method. The aim of the study was to evaluate the prick test as an auxiliary method in the diagnosis of flea allergy dermatitis in dogs. 24 dogs from the Experimental Chemotherapy Laboratory in Veterinary Parasitology (LQEPV) of the beagle breed, male and female, aged one year or more were selected. These animals were divided into two groups: group 1 containing 12 beagles known to have no DAPP and group 2, composed of 12 dogs with clinical diagnosis of DAPP. The animals were physically restrained, placed in lateral decubitus and had the chest area trichotomized and cleaned. Then, the test was carried out. One drop of each solution was used: for the negative control, a glycerin saline solution (50% glycerol - protein stabilizer) and phenolate (0.45% phenol - preservative), positive control (histamine) and the extract of total flea, all in duplicate, and this was inoculated into the dermis with the aid of a Duotip-Test II puntor. The readings were taken after 15 minutes and those papules that presented mean orthogonal diameter equal to or greater than 3 mm of the mean diameter of the negative control were considered positive reactions. Reading 24 and 48 later was also done to assess late reactions. The reactions observed to the flea extract in both groups were weak and considered to be non-reactive. The results suggest that the PT performed in this study had no sensitivity for the diagnosis of DAPP. Therefore, it is necessary to conduct a further study with patients seen in the clinical routine or with other extracts to improve the sensitivity of the technique.

Due to the problems generated by the indiscriminate use of chemical acaricides to control ticks, alternative methods have been developed, such as the use of entomopathogenic fungi. However, these entomopathogens have their viability compromised when applied under natural conditions, being indispensable the development of formulations. In this context, microencapsulation technologies for biocontrol agents are promising; since it provides protection against environmental factors as well as increased the encapsulated microorganism shelf life. Ionic gelation is a simple and financially viable technique when employed for the entomopathogenic fungi encapsulation. Thus, the aim of the present study was encapsulate the Metarhizium anisopliae conidia (LCM S01) in 2 and 3% sodium alginate, using the ionic gelation technique and, evaluate the external morphology, concentration, viability, shelf life, tolerance of UV-B irradiation, thermotolerance and efficacy of these microparticles in the control of Rhipicephalus microplus engorged females. The external morphology was characterized by ScanningElectron Microscopy (SEM) and the encapsulated conidia (EC) viability was determined by the germination percentage. For shelf life, conidia encapsulated in 2 (EC 2%) or 3% (EC 3%) of sodium alginate and unencapsulated (NEC) were stored for 1, 3, 5, 7, 9 and 11 months at room temperature and in a freezer.   For UV-B tolerance assay, EC and NEC were exposed to 6.0 or 8.0 kJ m-2, while for thermotolerance, they were submitted to 42 ºC in a water bath for 2, 4 and 6 hours. Furthermore, the biological parameters of engorged females exposed to 30, 60 or 90 mg of microparticles under laboratory conditions wereevaluated. 2 and 3% sodium alginate fungal particles were spherical with a more homogeneous and heterogeneous surface, respectively. Encapsulation did not cause severe conidia losses and did not affect their viability. Encapsulation increased the conidia shelf life stored at room temperature for 1, 3 and 5 months compared to NEC. In the freezer, NEC germinated more than EC. The EC and NEC showed greater viability in the freezer when compared to room temperature. Heat exposure for 6 hours reduced the NEC germination compared to EC 2 and 3%. Different UV-B doses exposure also significantly reduced the NEC germination compared to EC. The particles were able to reduce significantly the engorged females’ biological parameters when compared to NEC and the control group, however, there was no significant difference between the particles evaluated. In general, no significant differences were observed between the sodium alginate concentrations used. Thus, the formulations developed in the present study increased M. anisopliae (LCM S01) shelf life, thermotolerance and UV-B tolerance, and were effective in R. microplus engorged females control, showing promising potential to  control this tick.

Antimicrobial resistance is a problem in infections caused by enterobacteria, mainly by strains that produce beta-lactamases, which provide resistance to a wide range of antibiotics. Escherichia coli is a commensal gastrointestinal microbiota bacteria associated with infections in the urinary tract, and strains that produce these enzymes are important pathogens in Veterinary and Human Medicine. Despite the detection of beta-lactamase production being standardized by some committees, such as CLSI (Clinical Laboratory Standard Institute) and BRCast (Brazilian Committee on Antimicrobial Susceptibility Testing), the divergences between them generate extreme difficulty in interlaboratory standardization, generating an crucial barrier to the knowledge of the genuine epidemiology of these enzymes. Thus, the present study aimed to evaluate the resistance profile to β-lactams in E.coli isolated from urine samples from dogs, cats, and humans with urinary tract infections, in addition to researching - through the disc diffusion technique with interpretive reading, the production of ESBL (Extended Spectrum Beta-lactamase), AmpC and Carbapenemases, comparing the detection methods recommended by the normative documents. The study was carried out with 100 strains of E. coli from 273 urine samples from humans, and 97 from 103 samples of dogs and cats, totaling 197 isolates. All isolates had a resistance profile of less than 50% compared to the tested antimicrobials, ampicillin being the least effective, considering both the cutoff points of CLSI and BRCast. A total of 10% (10/100) and 11.16% (22/197) of E.coli from human and animal samples, respectively, were suspected of producing ESBL in the screening test. Among them, 56.25% (18/32) presented suspicion considering the cutoff points of both recommended documents. Of the total of suspected isolates, 81.25% (26/32) were confirmed in the tests - considering at least one normative document. Only 1% (1/100) and 5.15% (5/97) of human and animal samples were AmpC producers, respectively, and among these, four (66.7%, 4/6) were co-producers of ESBL . Finally, two isolates of human origin showed resistance to ertapenem, indicating possible production of carbapenemase. The emergence of multidrug-resistant bacterial strains is worrying due to the aspect of One Health since domestic animals can be reservoirs and transmitting agents for humans and vice versa, where the environment also acts in a potentiating way, contributing significantly to this problem.

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease that affects dogs, cats, horses, humans and primates. The pathogenesis is multifactorial, being associated with genetic, hormonal and environmental factors. SLE leads to deposition of immune complexes in joints and various organs, resulting in clinical manifestations. The participation of T and B lymphocytes is crucial for the pathogenesis of the disease. B-1 lymphocytes, a subpopulation of B cells, have characteristics that may contribute to the pathogenesis of autoimmune diseases because they are capable of secreting cytokines such as IL-10, exerting a modulating action on both the acute and chronic inflammatory response, they present antigens on T cells, they participate in innate and adaptive immunity and are the main producers of natural antibodies. Understanding the role of B-1 cells in the immunopathogenesis of SLE may open up fronts for studies on immunotherapeutic strategies that enable the control of this complex and multifactorial disease. The present work evaluated the role of B-1 cells in the immunopathogenesis of SLE using the pristane induction model. SLE was induced in BALB/c, XID (B-1 deficient) and XID female mice repopulated with B-1. The animals were evaluated for six months. We found that BALB/c pristane and XID repopulated with B-1 showed characteristic signs of the disease, such as lipogranuloma formation and splenomegaly. Notably, BALB/c pristane also presented ascites, joint edema, arthritis, kidney damage with immune complex deposition. In the spleen, animals with B-1 had a higher percentage of B cells (CD19+, IgM+), BALB/c pristane than CD8 T (CD3+, CD8+), while XID pristane had increased TCD4 (CD3+, CD4+). In the peritoneal lavage, after induction with pristane, there was a decrease in B cells (CD19+, IgM+) and an increase in B1a (CD19+ IgM+ CD5+) in BALB/c pristane. XID pristane showed an increase in TCD4 (CD3+, CD4+) and BALB/c pristane than TCD8 (CD3+, CD8+). In peripheral blood, BALB/c had a higher number of lymphocytes, XID had a neutrophilic profile, and in all SLE groups there was an increase in monocytes. The cytokines IL-10, IL-6 and IFN-gama were increased in repopulated BALB/c and XID. Based on these results, we suggest that the presence of B-1 cells may contribute to the development of SLE.

Rhipicephalus microplus tick has a significant impact on Brazilian cattle raising, being responsible for economic losses estimated at 3.24 billion dollars per year. Excessive use of chemical acaricides leads to tick resistance, in addition to affecting non-target organisms and the environment. Entomopathogenic fungi and essential oils have been shown to be effective in controlling bovine ticks. Here, the effect of the isolate of Beauveria bassiana CG 479 and the plant compound eugenol, alone or combined at different concentrations, against bovine ticks were evaluated. Initially, the germination rate of B. bassiana was evaluated at 1 × 10⁶ con/mL combined or not with eugenol at 0.18 mg / mL or 0.75 mg / mL. Regardless of the dose, eugenol was not able to reduce fungus germination. As well, the impact of both bioagents on the biological and reproductive parameters of engorged females of R. microplus were analyzed. Bioassays were conducted according to Bennett (1974), as a result, B. bassiana at 1 × 10⁸ con/mL combined with eugenol at 0.18 mg / mL or 0.75 mg / mL demonstrated the best control potential as well, how it reduced all reproductive parameters by about 50%. For evaluation by light microscopy, the ovaries of female ticks (24 and 72 hours after exposure to the fungus and/or eugenol) were collected and stained with hematoxylin and eosin, and histopathological analysis was performed to identify the morphological changes in the tissues ovaries. In general, regardless of time, oocyte vacuolization, pedicel hypertrophy and disorganization of oocyte membranes were observed in ovaries of R. microplus females exposed to B. bassiana at 1 × 10⁸ con/mL combined with eugenol at 0.18mg/mL or 0.75 mg/ml. Ovarian lipid profile was evaluated by thin layer chromatography according to Bligh & Dyer (1959) and Angelo (2013). The results obtained demonstrate that fatty acid and cholesterol were only affected by B. bassiana combined with eugenol at 72 hours post-exposure. The changes caused in reproductive parameters, such as the 47.5% reduction in total posture, which led to a control percentage of up to 56.48%, can be explained by the changes observed in the ovaries and lipid profile of treated females, and demonstrate the potential of the entomopathogenic fungus combined with eugenol as an alternative to control bovine ticks and reduce the reproductive efficiency of R. microplus.

The fungal genus Metarhizium is commonly found in soil and plant roots, and it is one of the main agents used in the biocontrol of agricultural pests. In this context, pasture is a favorable environment for the development of this microorganism, where the plant's rhizosphere and the presence of hosts such as the tick Rhipicephalus microplus make the best ecological niche for the fungus. However, biotic and abiotic factors interfere on its persistence in the environment; it is essential to select fungal isolates, consider ecological interactions with other microorganisms, and formulate them. Based on this, the present study addresses some tests, which consider the applicability of the genus Metarhizium spp. in the control of pests that inhabit the soil. Therefore, the following were evaluated: heat tolerance and the ability of different isolates of the fungus to colonize tissues of the Vigna radiata plant; the antimicrobial and acaricidal effect of fungal exudate; and the effectiveness of granular formulations of M. robertsii microesclerotia and blastospores to control cattle ticks. In the heat tolerance test, conidial suspensions of the fungus were exposed to 25 ° C, 40 ° C and 45 ° C, for 2h, 4h, 8h and 12h, and the germination was evaluated after incubation for 48h at 28 ± 1 ° C, based on that, heat-tolerant isolates were selected. For the study of endophytic colonization, seeds, root, stem, and leaf of V. radiata were treated with fungus suspensions, and the colonization of the plant tissues by different isolates was evaluated. To assess the effect of fungal exudate on ticks and bacteria, drops of this metabolite secreted by the colonies of Metarhizium spp. were collected. Then, from the disk diffusion test, the effect on strains of two bacteria isolated from the soil and on resistant and sensitive strains of Staphylococcus and Escherichia coli was evaluated. For the action on ticks, the fungal exudate was inoculated in tick females and the biological parameters, mortality, and immune response of the arthropod were evaluated. Finally, for the test with microsclerotia and blastospore granular formulations, the soil of pots containing Brachiaria decumbens were treated or not with granular formulations. Subsequently, females of R. microplus were exposed to this environment and mortality, the number of larvae obtained in each group, as well as the persistence of the fungus in the soil were evaluated. As regards to the results, in the heat tolerance test, all Metarhizium spp. isolates tested germinated well after 12h at 40 ° C and were considered good isolates, those that did not suffer drastic variation in germination over the time of exposure. When evaluated the ability to colonize V. radiata, all tested isolates were able to colonize roots, stems, and leaves, with variation in the colonization of different tissues. The fungal exudate was also able to inhibit the growth of the resistant and sensitive strains of Staphylococcus spp., and it affected more the strains of bacteria isolated from the soil than the others. The exudate also directly affected engorged R. microplus females, reducing the cellular immune response, causing death, and decreased egg production. The treatment with different concentrations of granular formulations of M. robertsii proved to be effective in controlling the cattle tick under the conditions tested. This study provides data about the application of Metarhizium spp. in the soil and contributes to future research, showing the bioprospecting of this microorganism.

Fipronil (FIP) is an ectoparasiticide of the phenylpyrazole class, used in veterinary medicine in topical form. Supported by evidence of uncontrolled human exposure to FIP and environmental damage caused by commercially available formulations, its use as a medicine for oral administration has become promising. The effectiveness of FIP against the flea Ctenocephalides felis felis and the tick Rhipicephalus sanguineus and its pharmacokinetics and its main active metabolite, fipronil sulfone (SULF), were evaluated after oral administration of tablets in three different doses (2, 4 and 6 mg/kg) in Beagle dogs, in a single treatment. Through the plasma concentration curves it was possible to observe that the FIP showed rapid absorption and metabolization and slow elimination. The values of Cmax (β = 0.7653) and AUC0-t (β = 0.3209) did not increase proportionally with increasing dose. At 48h after treatment, doses of 4 mg/kg (AUC0-t = 442.39 ± 137.35 µg/mL*h) and 6 mg/kg (AUC0-t = 421.32 ± 102.84 µg/mL*h) provided 100% and 99% efficacy for fleas and 95% and 98% for ticks, respectively. The estimated EC₉₀ for FIP + SULF was 1.30 µg/mL for C. felis felis and 2.16 µg/mL for R. sanguineus. The correlation of the FIP pharmacokinetic and efficacy data demonstrated its potential to be administered orally in the form of tablets for the control of ectoparasites in dogs, as a safer alternative for animals, humans and environment, aligned with the One Health concept.

During a survey conducted from March to June 2016, 356 dogs and 69 horses were sampled for the presence of ticks and possible pathogens transmitted by them from rural areas located in the Atlantic Forest biome, municipality of Ilhéus, state of Bahia, Both the ticks collected and the blood of the hosts were molecularly tested for the presence of hemoparasites DNA and later analyzed using the PCR technique. Five species of ticks, Rhipicephalus sanguineus (s.l.), Amblyomma ovale, A. sculptum, A.naponense and Rhipicephalus microplus were collected on dogs, while A. sculptum and Dermacentor nitens were collected on horses. Overall, 242 ticks from dogs and 62 from horses and blood from all dogs and horses were analyzed. R. parkeri Atlantic Forest strain was detected in A. ovale collected on dogs. Erlichia canis and Babesia vogeli were detected in R. sanguineus (s.l) and in the blood of the examined dogs. This is the first record of an A. naponense nymph in dogs in the state of Bahia. The multivariate analysis performed using the logistic regression method revealed that dogs with access to forest areas and/or have contact with wild animals are more likely to be positive in blood samples for E.canis and B.vogeli. 

The body's physiological response to an offending agent is inflammation and can be triggered by an infection and / or injury. The acute phase of this process is a rapid response and occurs within the first minutes and hours after pathogen recognition. Their resolution usually results in the elimination of infectious agents and repair of normal tissue architecture and function. The chronic phase begins when attempts to restore homeostasis are unsuccessful. Inflammation control through immune system components is a promising alternative to the use of conventional drugs that often do not meet patients' demands due to their multiple side effects. B-1 cells are capable of producing natural antibodies, may have T lymphocyte antigens and produce various cytokines, including the anti-inflammatory cytokine IL-10. Thus, they have the potential to modulate the inflammatory response. In this paper, we evaluated the population dynamics of leukocytes in the blood and peritoneal cavity of BALB / c and XID mice during the acute inflammatory response triggered by LPS. For this, we use XID mice, whose peritoneum is an environment devoid of B-1 cells. Our results revealed that XID animals spontaneously have a high number of peripheral blood neutrophils and this population becomes even larger after LPS inoculation. Concomitantly, high levels of IL-6 were detected. In addition, the peritoneal cavity of these animals also has a larger amount of neutrophils compared to BALB / c mice. This data does not change after stimulation in our experimental conditions. In the phagocytosis assays, we observed that the number of macrophages capable of phagocytosis is statistically equal between BALB / c and XID, but the number of internalized yeasts is lower in the LID and IFN-γ stimulated XID group. This suggests greater microbicidal power of the macrophages of these mice. This result is corroborated by the nitrite dosage in the culture supernatant, in which the stimulated XID macrophages produced more nitric oxide than the control group. The body of facts points to the development of a more intense inflammatory response in XID mice compared to BALB / c mice, probably due to impaired production of anti-inflammatory cytokine IL-10. Thus, our analysis reports the unprecedented finding of increased neutrophil population in the blood and peritoneal cavity of XID mice, confirms the importance of B-1 lymphocytes in modulating the inflammatory response, and suggests that these may be future targets for investigations into immunotherapy strategies.

The flea (Ctenocephalides felis) is the principal ectoparasite of dogs and cats worldwide acting as a vector of zoonotic pathogens including Rickettsia felis and Bartonella. The diversity of host associated microbes and their interactions within their hosts, including arthropods, are fundamental to the ecological functions within those communities and may contribute to host evolution. Moreover, symbiotic interactions may influence the transmissions of zoonotic pathogens, via effects upon vectoral competence. Knowledge of the microbiota of vectors has been used to develop novel approaches for control based on the concept of microbiota manipulation. A key component in this strategy is the presence of a stable/core microbiota. The present study characterized the stability, over seven years, of the cultivable microbiota of a laboratory colony of C. felis. Bacteria associated with different life stages were isolated by culture on plates of nutrient agar. Bacteria were identified, to the genus or species level, by polymerase chain reaction (PCR) amplification of a 500 base pair (bp) fragment of the gene encoding prokaryotic 16S ribosomal RNA, followed by nucleotide sequencing. Cultures were characterized for susceptibility to seven antimicrobial compounds (ampicillin, chloramphenicol, mercury chloride, nitrofurantoin, oxytetracycline, rifampicin and streptomycin), using a microdilution method. Each of the different life stages presented a unique microbiota, however a core component of all samples were members of the genus Staphylococcus, with some demonstrating multiple-drug resistant phenotypes. The most prevalent species were S. saprophyticus, S. nepalensis, S. lentus and S. cohnii all of which,  have been reported as opportunistic pathogens with zoonotic potential. The constant presence of the same species of Staphylococcus, in multiple life stages, suggests they are essential components of the microbiota and by implication of the biology of the fleas. Future research will examine the effects of manipulating the core microbiome as the first step in the development of novel strategies for infestation control. 

Fleas of the species Ctenocefalides felis felis, also known as cat fleas, are hematophagous ectoparasites of the order Siphonaptera. Several animal species can be parasitized, including man. The fleas are widely distributed and are of great importance in Veterinary Medicine, which can cause severe itching and allergic dermatitis in pets, in addition to being vectors of various pathogens such as rickettsiae, bacteria, worm cestodes, nematodes and protozoa, both in animals and in humans. The control is done through integrated management of the environment and animals infested through chemical products. Current studies demonstrate that the inappropriate use of such chemicals to control this insect has become an item of great concern due to the occurrence of resistant populations in some regions. Thus, the search for natural products that act in biological control, such as fungi, bacteria, viruses, protozoa and entomopathogenic nematodes has become important in insect control. In recent years, the study on entomopathogenic nematodes (EPNs) has increased, making this an alternative in the control of insects of veterinary importance, as they present several advantages in relation to their use, and can therefore be used alone or in conjunction with Chemicals. The present study aimed to standardize and evaluate the susceptibility of seven-day-old flea larvae of the species Ctenocephalides felis felis by entomopathogenic nematodes of the genus Heterorhabiditis. The study was divided into two stages, where the first three bioassays aimed to standardize the infection technique of flea larvae of C. felis felis, verifying the amount of liquid necessary for the nematodes to move and infect the larvae (400, 600 and 1000 µl); to evaluate the susceptibility of infection and mortality of flea larvae by entomopathogenic nematodes Heterohabiditis bacteriophora (strain HP88) in three different concentrations (120, 160 and 200 EPNs/flea larvae). The second bioassay aimed to evaluate the susceptibility to infection and mortality of C. felis felis larvae by the entomopathogenic nematode Heterohabiditis indica (strain LPP30) in three different concentrations (120, 160 and 200 EPNs/flea larvae). All bioassays were divided into two groups, one containing a specific diet for larvae and the other without a diet. Parametric data were evaluated by analysis of variance (ANOVA) followed by the Tukey test (p <0.05). In the first three bioassays with H. bacteriophora (HP88), it was possible to standardize the infection of larvae of C. felis felis, in Petri dishes with 6 cm in diameter, where the most adequate concentration of nematodes containing liquid was 600 µl, showing significant difference when compared to the bioassay that used 400 µl. In the bioassay in which the 1000 µl volume was used, mortality was 100%, including in the control groups, suggesting that the amount of liquid is not suitable for infection. Still for the species H. bacteriophora (HP88), in the volume of 600µl, there was no significant difference between the groups with and without diet, nor in the concentration of EPNs used for the infection. In the second stage of the study, infection of the larvae of C. felis felis by entomopathogenic nematodes H. indica (LPP30) was proven, both in the group with diet and in the group without diet, as well as in the three concentrations of entomopathogenic nematodes, with no significant difference between them. Through these results, it can be suggested that in vitro infection of seven-day flea larvae of the species Ctenocephalides felis felis by the species Heterorhabditis bacteriophora (HP88) and Heterorhabditis indica (LPP30) is possible, which is a tool to be studied for the use in the biological control of these insects.

Belonging to the Amblyomma cajennense complex, the tick Ambyomma sculptum can be found in the various regions of Brazil. It is known as star tick or horse tick and parasite domestic, wild and human animals, being the main transmitter of Rickettsia rickettsii etiological agent of Brazilian Spotted Fever. It has great importance in veterinary medicine because of several damages to its hosts, in addition to the suspicion of transmitting Theileria equi to horses. There are few active ingredients registered for the control of the A. sculptum tick. Fluazuron is an insect growth inhibitor used in the control of ticks that interferes with the arthropod's chitin production, preventing the phase change. The resistance of the tick population to antiparasitic agents represents an obstacle to control programs. Studies on the tick's immune system seek to assess changes in the hemolymphatic cellular response that make the immunology and physiology involved in the control and resistance mechanisms understand. The aim of this work was to characterize quantitatively and morphologically the hemocytes present in the hemolymph of engorged females of the A. sculptum tick and to evaluate the cellular immune response of this tick after in vitro exposure to fluazuron. For hemolymphatic cell characterization of A. sculptum. hemolymph was collected through a cuticle perforation in the dorsal part of the tick. The hemolymph was placed directly on a glass slide to make blood smears that were stained with Giemsa's solution. Differential hemocyte counting was performed based on the morphology observed in an optical microscope in 1000-fold magnification, by identifying the first 100 cells found in the stained smear. The evaluation of cell sizes and their components was performed by reading the slides made for differential counting of hemocytes with the aid of an Olympus optical microscope model BX51 with polarized light, coupled to a camera of the same brand model UC30. The total hemocyte count was performed with the aid of an optical microscope and a Neubauer Chamber. To evaluate the cellular response after in vitro exposure to fluazuron, immersion tests were performed with the engorged females of A. sculptum in the control (with diluents only) and treated (7.81, 250 and 4000 ppm) groups. Hemolymph was collected for total and differential hemocyte counts at 24h and 48h after immersion. The cellular characterization revealed that the A. sculptum tick has an average of 1024 cells / μL and six cell types: prohemocytes, plasmatocytes, granulocytes, spherulocytes, adipohemocytes and oenocytoids that are distributed differently in the hemolymph, where granulocyte was the most frequent type. Fluazuron was able to make cellular changes in the tick A. sculptum promoting a decrease in total hemocytes at a concentration of 4000ppm after 48h as well as reducing the frequency of granulocytes and increasing the frequency of spherulocytes in the differential count.

The use of mechanized harvesting and the processing of sugar cane nowadays, generate by-products that help in the proliferation of Stomoxys calcitrans (Linnaeus, 1758), which affects various species of domestic animals and man, causing economic losses due to the occurrence of outbreaks . With the constant use of chemicals to control the fly in the stables, resistance to them is observed and this has led to further studies in biological control, where entomopathogenic nematodes (NEP) appear as organisms with potential use for biological control . The objective of the present study was to verify the efficiency of Heterorhabditis bacteriophora - HP88 and H. baujardi - LPP7, maintained at different temperatures of vinasse on S. calcitrans larvae in the laboratory. Each experimental unit consisted of ten third instar larvae placed in plastic pots containing four milliliters of the 50% vinasse solution. In each treatment, 300 infectious juveniles of H. bacteriophora - HP88 line or H. baujardi - LPP7 line were added per larva in the vinasse solution described above, heated for ten minutes at temperatures of 25, 28, 31, 34, 37 and 40 ° C, the treated groups and the control groups of the experiment were kept in an air-conditioned BOD chamber at 25 ± 1 ° C and 70 ± 10% RH. In total, 6 repetitions were performed for each treatment, with mortality observed daily for a period of 15 days. The results were evaluated, and in the statistical analysis, using the GLIMMIX procedure at 5% probability, it was found that there was no statistical difference between the treated groups, control groups and temperatures for larval mortality rate (P = 0, 8573), percentage of dead pupae (P = 0.1782) and adult emergence (P = 0.4386). When only the groups with presence and absence of NEPs were evaluated, larval mortality rates of 30% and 14.17% were observed for H. bacteriophora – HP88 line and H. baujardi - LPP7 line respectively, while the control groups presented 3,89% in H. bacteriophora – HP88 strain and 8.61% in H. baujardi - LPP7 strain. In the evaluation of temperatures, significant statistical differences were observed only in the temperatures 37 and 40 ºC of H. baujardi - strain LPP7 in the larval mortality rate. In pupal mortality rates, the group with H. bacteriophora - HP88 strain showed averages of 32.5, 34.17 and 27.5% at temperatures of 25, 31 and 34 ºC, respectively. In the group with H. baujardi - lineage LPP7, temperatures of 37 and 40 ºC presented 40% and 37.5% of pupal mortality. In the analysis of the adult emergence rate, there was a lesser emergence at temperatures of 25, 28, 31 and 34 ºC in the group with the presence of H. bacteriophora – strain HP88 and the group with H. baujardi - strain LPP7 with the lowest rates of emergence of adults at temperatures of 37 and 40 ºC. The results of the study show that there was a decrease in the mortality of the immature stages of S. calcitrans caused by NEPs maintained at different temperatures of vinasse, when compared to other studies without heating it. There is a need for further studies to adapt the use of NEP together with vinasse, heated or not.

In Brazil, the importance of Visceral Leishmaniasis constitutes a serious public health problem, with great incidence, wide geographical distribution and the possibility of taking on serious and lethal forms. In Latin America, Leishmania infantum causes canine visceral leishmaniasis (CVL). A total of 399 medical records and their respective serum samples were submitted to the tests recommended by the Ministry of Health. From the dogs attended in the routine system at the Veterinary Clinical Care, the animals which were submitted to the tests were seroreactive for LVC, as followed: TR-DPP® with 45 (11,28%) of animals were positive for CVL and EIE®-LVC with 11 (2.76%) and with a final result of 4 (1.00%) seropositive for CVL, due to the concomitance of the final result of TR-DPP® + EIE ®-LVC compatible with the recommendations of the Ministry of Health. One of the animals examined was negative for CVL in both tests, characterized as asymptomatic for being positive for the bone marrow puncture cytological test, which prompted this study. There was no characterization of CVL between gender differences, but the predominance of infected animals was consisted of female in both used tests. Regarding the age group, most animals were characterized as middle aged, the longer the life span; greater was the probability of becoming infected with the studied etiological agent. The predominance of seroreagent to CVL was consisted of small animals, predominant in the studied region. Most of the clinical manifestations observed here were similar to those related to other etiologies observed in most animals during their veterinary clinical care. Even so, suspicious animals should be examined for having clinical manifestations compatible with CVL, even if the final diagnosis is characterized by involvement by another etiology, even if it comes from areas considered free of cases favorable to the appearance of CVL in domestic dogs. Furthermore, putting the tutors themselves at risk for being the etiological agent of CVL, Leishmania (Leishmania) infantum chagasi considered as a zoonosis of compulsory notification.

The families of the order Passeriformes can be parasitized by several coccidian species, mainly of the genera Isospora Schneider, 1881 and Eimeria Schneider, 1875. In this context, the current work aimed to identify, characterize and quantify species of coccidian parasites of the families Thamnophilidae, Tyrannidae, Conopophagidae and Icteridae in Itatiaia National Park (PNI) mainly. Nineteen expeditions were carried out at the PNI, and three other expeditions were carried out in two different locations: Cacaria and Barra Mansa. Birds were captured with mist nets. From all 204 bird specimens were captured: 100 taminophilidae, 89 tyrannids, 7 conopophagids and 8 icterids, after processing the samples, observed to be coccidians of the genera Isospora or Eimeria from four families addressed in this study. Thus, the current work presents a review of the scientific literature on the themes of ecology and taxonomy of coccidia of Thamnophilidae, Tyrannidae, Conopophagidae and Icteridae, and, as a result, five scientific articles in chronological order describing a new host Cacicus haemorrhous (Linnaeus, 1766) and locality (PNI) for Isospora bellicosa Upton, Stamper and Whitaker, 1995, and a new species Isospora guaxi Silva & Berto, 2017 parasitizing C. haemorrhous in the family Icteridae; a new species Isospora lopesi Silva-Carvalho & Berto, 2018, from Platyrinchus mystaceus Vieillot, 1818 at PNI in the Tyrannidae family; a new host Pyriglena leucoptera (Vieillot, 1818) and locality (PNI) for Isospora sagittulae McQuistion and Capparella, 1992 in the Thamnophilidae family; a new species Isospora borbai Silva-Carvalho et Berto, 2019 parasitizing two distinct hosts, Conopophaga melanops (Vieillot, 1818), and Conopophaga lineata (Wied, 1831) in the Conopophagidae family; and finally a new host Dysithamnus mentalis (Temminck, 1823) for Isospora parnaitatiaiensis Silva, Rodrigues, Lopes, Berto, Luz, Ferreira, Lopes, 2015 in the Thamnophilidae family in PNI.

Several approaches to stable fly control have already been considered and/or applied; passing through chemical control, with low viability due to possible environmental damage, and generation of populations resistant to various chemicals; waste management (organic matter), which depends on manpower training and awareness of the actors involved in the process; to the current paradigm of microbiological control, either by itself or in an integrated control system. The biological characteristics and other information attributed to entomopathogenic nematodes (EPN) indicate that these agents are promising in agricultural systems in the control of pests that develop partially or totally in the soil, with the possibility of low cost implementation for farmers; limiting financial losses caused by weight loss and/or animal productivity. It should be considered, however, that following the expansion of sugarcane cultivation areas and the increase in the Brazilian cattle herd, the financial loss caused by S. calcitrans tends to be much greater than these published estimates. The experiments described for this study dealt with the factors with the greatest potential for interference with the implementation of a EPN-based biological control system for Stomoxys calcitrans – the maintenance of nematode pathogenicity among the most favorable sugarcane by-products available to fly development, and the resistance to temperatures similar to those of fertigation in sugarcane fields. The present study aimed to verify the pathogenic potential of Heterorhabditis nematodes on S. calcitrans larvae when exposed to vinasse, filter cake, and sugarcane bagasse; observe the effect of different temperatures on the maintenance of EPN virulence; to evaluate the influence of S. calcitrans larvae age on EPN virulence, and to consider the viability of EPN application on S. calcitrans pupae. Furthermore, this study aimed to offer tools that will help mitigate the problems caused by S. calcitrans in the relationship between sugarcane industries, livestock zones and public health.

Digenetic trematode infection can cause changes in the reproductive activity of snails that are intermediate hosts. However, these changes vary according to the species of snail and parasite involved, age of the host, infective parasite load, abiotic and biotic factors. Our aim was verify the reproductive alterations in Subulina octona after infection by Paratanaisia bragai. The infected snails were individually exposed for 24 hours to 20 parasite eggs and four groups were formed (10, 20, 30 and 40 d.p.i.), along with control groups. Every 10 days, the number of eggs in the reproductive tract, number of eggs hatched, galactogen content and histopathological changes were evaluated. The regression analysis of the reproductive parameters indicated the inversion of the observed patterns for the number of eggs per snail and of eggs hatched during the experimental period between the control and infected snails. However, in relation to the amount of galactogen, the two groups followed the same pattern of variation. In the histology, we observed the presence of male and female gametes with marked reduction in the number of oocytes. The results indicate that the intra-snail development of the parasite affects the reproductive biology of the host.

Stomoxys calcitrans (Diptera), known as stable fly presents great veterinary importance. Males and females feed on blood, a parasitic habit that results in damage to livestock. The consequences of blood feeding include bite stress, which leads to lower weight gain and milk production, and infection with pathogens that S. calcitrans mechanically transmits. The annual damage caused by the stable fly in Brazil is estimated at US $ 340 million. Maintaining this organism under laboratory conditions is a crucial step in establishing research lines under controlled conditions. Insect colonies establishment in the laboratory is important for several studies, ranging from behavior observations to control strategies. This work presents data from S. calcitrans colony that we established in our lab over five months and strategies for breeding this fly in the laboratory. Additionally, we intend to define different actions to better understand the physiological aspects of the vector, as well as its interaction with pathogens in order to better understand the dynamics of mechanical transmission. The saliva composition of hematophagous insects is a crucial evolutionary factor for hematophagy and pathogens have been benefiting from its properties in this relationship. Salivary glands are organs of great importance, both for blood feeding and for the dynamics of pathogen transmission. Therefore, we characterized the salivary glands of S. calcitrans morphologically. The analyzes performed in this work revealed that this organ has merocrine gland characteristic, with polygonal cells presenting large nuclei, besides exhibiting cytoplasm rich in mitochondrial profiles and rough endoplasmic reticulum, characteristics of high protein synthesis activity. Understanding how S. calcitrans interacts with pathogens is an important step to clarify the dynamics of transmission by this fly, and the chosen model was Leishmania, etiological agent of leishmaniasis, diseases with anthropozoonotic characteristics. Leishmania sp. has been able to interact with the S. calcitrans gut, but the mechanisms by which this binding occurs must still be clarified.

Triatomines are responsible for the vector transmission of Chagas disease, which affects between 6 and 8 million people in continental Latin America, with an incidence of 28 thousand cases per year. In the state of Ceará, the predominance of Caatinga, besides a rural area with precarious human habitations, provides several types of shelters for these insects. In the present study, we evaluated the geographical distribution, the dispersion rates and the natural Trypanosoma cruzi infection in triatomines collected in the Cariri region, southern Ceará, and their association with dwelling, socioeconomic and environmental characteristics. The highest dispersion rates were registered in Antonina do Norte and Potengi. We found significative correlation between triatomine dispersion and income, work and environmental variables in the municipalities. The species T. pseudomaculata represented most of the specimens captured during the study period. In the domestic environment, most of the collected triatomines were identified as T. brasiliensis or T. pseudomaculata. The municipalities of Farias Brito and Potengi registered the highest triatomine rates for each 10 thousand inhabitants. The species P. megistus and R. nasutus presented the highest mean indexes for T. cruzi natural infection. In Farias Brito and Potengi, most of the dwellings where T. cruzi-positive triatomines were captured had external bathroom, perch, cats, dogs and hen houses. However, we did not find significative correlation between the characteristics of the domiciles and the
positivity for T. cruzi in triatomines. Our results reinforce the importance of the entomological surveillance for Chagas disease vectors in endemic areas in the Brazilian northeast.

The Atlantic Forest is considered one of the 34 biodiversity hotspots on the planet due to its high species richness and endemism, however the anthropic actions that occur in this biome have contributing to the faunal decline and particularly, hemoparasite infections may be further threat to the birds of this ecosystem. Thus, understanding the diversity, distribution patterns and evolutionary aspects of these parasites in wild bird hosts is important to design actions in conservation programs. In this study, blood samples from 188 wild birds were subjected to conventional PCR genetic analysis using 18S rDNA sequence to identify Trypanosoma spp resulting in a total of 15 positive samples; seven specimens from the Itatiaia National Park (RJ / MG), two in Turdus flavipes and five in T. albicollis, as well as in the capture areas belonging to Zona da Mata Mineira for six individuals of the species Tachyphonus coronatus, one species of Thamnophilus caerulescens and one of Synallaxis spixi. Five clones from each PCR reaction were submitted to sequencing, totaling 15 sequences from each bird specimen evaluated. All 15 sequences obtained from T. albicollis and T. flavipes were identical to each other. In the six specimens of T. coronatus, 5 distinct sequences were observed. All strings of T.caerulescens and S. spixi were identical to each other. Using a strategy of cloning different PCR sequences, it was possible to observe one of the examples of T. coronatus two sequences of different trypanosomatics, or with probable evidence of co-infection. Phylogenetic reconstruction showed the grouping of the seven new strains with parasites of the genus Trypanosoma. The new strains were grouped into two clades. The lines JB03, JB04, JB05, ITA01 and ITA02 are grouped together with the species Trypanosoma bennetti. As the lines JB01 and JB02 are grouped externally to the groups of species T. avium, T. cullicavium and T. corvi. As the new trypanosome lines found in this study are grouped with occurrence lines recorded in Europe, Asia and Africa. According to the evolutionary distance data selected in the present study, we can observe the same genetic strain of Trypanosoma sp. is capable of infecting different species of birds, just as the same bird species can be infected by another strain of Trypanosoma sp. This study, in an unprecedented way, is the first to access a molecular diversity of parasites of the genus Trypanosoma in birds in Brazil. Until then, all studies related to avian trypanosomes were based mainly on reports of occurrence in blood smears of birds in studies aimed at hemoparasites in general. For Brazil, this pioneering study selected possible methods to research a diversity of avian trypanosomes and shows the great potential that exists to be explored in this group of parasites. Due to the results of this study accredited in Brazil, due to its large territorial dimensions and the great biodiversity of hosted birds and insect vectors, it may be one of the richest species of avian trypanosomes in the world.

Compounds of plant origin have been identified as promising in the control of ticks. In chapter 1, the present work aimed at the phytochemical study of essential oils (OEs) from the bark of Cinnamomum zeylanicum (cinnamon) and from the stem of Eremanthus erythropappus (candeia) and evaluation of the tick activity of these OE (s), together with their major isolated compounds and acetylated analog on Rhipicephalus microplus. The results of gas chromatography and mass spectrometry showed the compound (E)-cinnamaldehyde as the major component of the cinnamon OE (86.93%), and α-bisabolol as the main compound of the cinnamon OE (78.71%). The bioassays with the non-fed larvae were performed using the larval pack test at concentrations from 0.31 to 10.0 mg / mL. For engorged females, the immersion test was performed at concentrations of 2.5 to 60.0 mg / mL. With the exception of cinnamyl acetate, which showed low activity for engorged females, it was possible to observe the potential of these compounds of plant origin on R. microplus. In chapter 2, the activity of (E)-cinnamaldehyde and α-bisabolol was investigated on fifty-one populations of R. microplus, with different profiles of resistance to the different active principles that exist commercially. In order to characterize the degree of resistance, ticks based on deltamethrin (Pyrethroid - Butox®), amitraz (Amidine - Triatox®) and chlorfenvinfos (Organophosphate - Supokill®) were used in their commercial concentration in the female immersion test. For the calculation of the LC₅₀ of the larvae translated with the isolated compounds, the larval pack test was performed at concentrations from 0.31 to 10.0 mg / mL. In this stage, tests were also performed with the Porto Alegre strain (POA), a sensitive strain for reference in the calculation of the Resistance Ratio (RR). Based on the results generated, it was not possible to infer a correlation between the efficiency of ticks and the LC₅₀ values of the isolated compounds tested, thus not being observed cross-resistance, indicating that the different results found may be related to existing phenotypic variations between populations. In chapter 3, the lipid profile of the fatty body and eggs of R. microplus engorged females exposed to a concentration of 10.0 mg / mL of (E)-cinnamaldehyde and α-bisabolol were evaluated. To characterize the lipid profile, the techniques of thin layer chromatography and gas chromatography coupled to a mass spectrometer were applied. In addition, an in silico study was carried out in order to know the possible molecular targets of these plant compounds. The results demonstrated changes in the profile of lipids present in the fatty body and eggs of females treated with (E)-cinnamaldehyde and α-bisabolol, showing a possible mechanism of action of these compounds.