Detec??o de Tritrichomonas foetus em s?men bovino criopreservado atrav?s de isolamento e rea??o em cadeia da polimerase

• Autor(a) principal: Ribeiro, Laura
• Data de Publicação: 2020
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: Biblioteca Digital de Teses e Dissertações da UFRRJ
• Texto Completo: https://tede.ufrrj.br/jspui/handle/jspui/6260
• topic: Tricomon?ase; biologia molecular; s?men congelado; Trichomoniasis; molecular biology; frozen semen; Medicina Veterin?ria

The Bovine Genital Trichomoniasis (BGT) is an infectious disease triggered by a protozoan called Tritrichomonas foetus, which is transmitted by venereal route and by contaminated fomites. The disease is economically relevant, since its main consequence is abortion, besides producing temporary infertility. In males the disease is asymptomatic and permanent, characterizing the bulls as silent nurses and emphasizing the direction of diagnosis in this category of animals. The identification and detection of the protozoan by microbiological culture are considered as "gold standard" among diagnostic techniques in Brazilian artificial insemination centers. However, the possibility of a false negative is a reality and is needed more sensitive and specific techniques of diagnose, since the Artificial Insemination (AI) is the current way to prevent BGT. T. foetus has already been found in semen samples and is also able to remain viable when frozen, raising AI to a risk factor when sanitary measures are not applied or the diagnostic method is insufficient Thus, this study is aimed to detect the presence of T. foetus in samples of cryopreserved bovine semen, through culture and Polymerase Chain Reaction (PCR). The semen straws will be purchased from artificial insemination centers. A total of 27 samples were analyzed. Once in the laboratory, they were thawed in water at 37?C, part was inoculated in a test tube containing Diamond medium, incubated at 35?C with consequent growth evaluation and evaluated every 24 hours via direct microscope examination and the other part diluted in PBS for subsequent molecular analysis. After 10 days of cultivation, all samples were negative. For molecular detection, the Quick-DNA Miniprep Kit (Zymo Research) without proteinase K was used for DNA extraction. The primers used in PCR were TRF3 and TRF4. The PCR result was negative for all samples, corroborating with that obtained in the culture. It was concluded that the samples used were negative for the presence of T. foetus in both diagnostic methods, proving the innocuousness of the commercial semen tested.