Tristeza parasitária bovina: diagnóstico molecular, efeito sobre a fertilidade e desenvolvimento de um produto para prevenção

Abstract

Anaplasma marginale and Babesia spp. are parasites that cause hemolytic anemia in cattle and make up the cattle tick fever complex (TF). Tick fever is endemic in most of the Brazilian territory, causing significant economic losses. The most sensitive diagnosis is obtained through molecular techniques. Therefore, the objective of this thesis was to determine the occurrence of A. marginale and Babesia spp., using molecular diagnostic, in Brazil, to verify the influence of these parasites on hematological parameters and conception rate of Angus heifers, and to develop a slow and prolonged release vehicle for TF prevention. Initially, a systematic review was carried out, with a search for studies that used molecular techniques to detect A. marginale and Babesia spp. in cattle in the Brazilian territory. Overall, 38 studies were included in the review, with the highest occurrence of A. marginale and B. bovis found in the state of Roraima (98.78% and 95.47%, respectively) and B. bigemina in the state of São Paulo (86.25%). To verify the impact of infection by these hemoparasites on the conception rate, 94 Angus heifers were inserted into a timed artificial insemination (TAI) protocol. Blood samples were collected at the beginning of the protocol (D-10) and on the day of pregnancy diagnosis (D+34) to determine hematological variables. An aliquot of D-10 blood was used for total genomic DNA (gDNA) extraction and all samples were subjected to quantitative PCR (qPCR) to investigate target genes for A. marginale and B. bovis. All positive samples for B. bovis were submitted to absolute quantification, determining copy number (CN) of B. bigemina and B. bovis. Comparisons were made between pregnant (P) and non-pregnant (NP) groups and correlation between CN and hematological parameters. The conception rate at the first service was 47.87% (45/94), only on D-10 the NP had higher hemoglobin than the P. All heifers were positive for A. marginale and B. bovis in qPCR, while in absolute quantification, six heifers were negative for B. bigemina (four P and two NP) and two negatives for B. bovis, one from each group. There was no difference with CN ratio and significant correlation was found only between hemoglobin/erythrocyte (P<0.0001; r=0.8082) and hemoglobin/hematocrit (P=0,0049; r = 0,3009), respectively. For an innovative strategy for the prevention of TF, a slow and sustained release emulsion was developed, performing in vitro release tests, and comparing it with a commercial formulation, and testing to verify pain and/or reactions at the application site. In in vitro tests, emulsions releases occurred more slowly when compared to the commercial product. In the test with animals, an increase in volume at the application site was noted, without manifestations of significant pain. It is concluded that the occurrence of A. marginale and Babesia spp. ranged from 7.32% to 98.78% and, despite the amountof data found on the occurrence and use of molecular diagnosis, there are still gaps and research opportunities, mainly on the quantification of losses in the herds with higher occurrence of TF, and for the development of formulations for the prevention of these diseases.