РАЗРАБОТКА И ПРЕДВАРИТЕЛЬНАЯ ОЦЕНКА НЕПРЯМОГО ИФА НА ОСНОВЕ РЕКОМБИНАНТНОГО AMA-1 ДЛЯ ОБНАРУЖЕНИЯ АНТИТЕЛ К BABESIA BIGEMINA У КРУПНОГО РОГАТОГО СКОТА

Bovine babesiosis, caused by Babesia bigemina, remains one of the most significant vector-borne animal diseases. However, existing laboratory diagnostic methods have several limitations, particularly when testing asymptomatic carriers. The aim of this study was to develop an indirect enzyme-linked immunosorbent assay (ELISA) based on the recombinant AMA-1 antigen of B. bigemina, to preliminarily evaluate its diagnostic properties, and to use it to detect specific antibodies in cattle. The study was conducted on 500 blood samples collected in the Atyrau and Zhambyl regions of Kazakhstan. A control panel was formed based on the results of PCR and sequencing of the 18S rRNA gene fragment of Babesia spp. Optimization of the indirect ELISA conditions was performed using a checkerboard titration method using the recombinant AMA-1 protein. After optimization, the assay was used to test the entire sample. The optimal antigen concentration was found to be 5 μg/ml with a final plasma dilution of 1:40. On a preliminary panel of 70 samples, the developed ELISA demonstrated a relative sensitivity of 100.0%, a relative specificity of 98.0%, and a high level of agreement with PCR (κ=0.97). Among the 500 animals tested, antibodies to recombinant AMA-1 were detected in 43 (8.6%) animals. In the Atyrau region, seropositivity was 29.0%, while in the Zhambyl region it was 3.5% (p<0.001). These results demonstrate the potential of using recombinant AMA-1 as a diagnostic antigen in the development of serological tests for the detection of antibodies to B. bigemina. The developed laboratory prototype requires further independent validation on an expanded panel of samples before practical implementation.