Banca de DEFESA: NELSON MEIRELES DA SILVA

Uma banca de DEFESA de MESTRADO foi cadastrada pelo programa.
STUDENT : NELSON MEIRELES DA SILVA
DATE: 07/11/2023
TIME: 14:00
LOCAL: Plataforma Google Meet
TITLE:

Standardization of a protocol for extracting genomic DNA stored on FTA® cards and genetic diversity of parasites of the genus Haemoproteus in wild birds from the Montane Atlantic Forest.

KEY WORDS:

Haemosporidian;birdlife; phylogenetic analysis

PAGES: 119
BIG AREA: Ciências Agrárias
AREA: Medicina Veterinária
SUBÁREA: Medicina Veterinária Preventiva
SPECIALTY: Doenças Parasitárias de Animais
SUMMARY:

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. 

COMMITTEE MEMBERS:
Presidente - 3701492 - HUARRISSON AZEVEDO SANTOS
Interna - 3103528 - CLAUDIA BEZERRA DA SILVA
Interna - 3103478 - MARISTELA PECKLE PEIXOTO
Externa à Instituição - ALINE MOREIRA DE SOUZA - UFF
Externo à Instituição - DANIEL DA SILVA GUEDES JUNIOR - FIOCRUZ