Genetic diversity of Babesia bovis studied longitudinally under natural transmission conditions in calves in the state of Rio de Janeiro, Brazil

Abstract Serum and DNA samples from 15 naturally infected calves in Seropédica, Brazil, were obtained quarterly from birth to 12 months of age, in order to longitudinally evaluate their humoral immune response against Babesia bovis and the merozoite surface antigen diversity of B. bovis. Anti-B. bovis IgG antibodies were detected by an indirect fluorescent antibody test (IFAT) and enzyme-linked immunosorbent assay (ELISA). Using DNA amplification, sequencing and phylogenetic analysis, the genetic diversity of B. bovis was assessed based on the genes that encode merozoite surface antigens (MSA-1, MSA-2b and MSA-2c). The serological results demonstrated that up to six months of age, all the calves developed active immunity against B. bovis. Among the 75 DNA samples evaluated, 0, 3 and 5 sequences of the msa-1, msa-2b and msa-2c genes were obtained, respectively. The present study demonstrated that the msa-2b and msa-2c gene sequences amplified from blood DNA of B. bovis-positive calves were genetically diversified. These data emphasize the importance of conducting deeper studies on the genetic diversity of B. bovis in Brazil, in order to design diagnostic antigens and vaccines in the future.摘要：本研究从巴西塞罗佩迪卡（Seropédica）的15头自然感染牛巴贝斯虫（Babesia bovis）的犊牛中，于出生至12月龄期间每季度采集血清与DNA样本，以纵向评估其针对牛巴贝斯虫的体液免疫应答，以及牛巴贝斯虫的裂殖子表面抗原多样性。本研究采用间接荧光抗体试验（indirect fluorescent antibody test, IFAT）与酶联免疫吸附试验（enzyme-linked immunosorbent assay, ELISA）检测抗牛巴贝斯虫IgG抗体；通过DNA扩增、测序及系统发育分析，基于编码裂殖子表面抗原（merozoite surface antigen）的MSA-1、MSA-2b与MSA-2c基因评估牛巴贝斯虫的遗传多样性。血清学结果显示，至6月龄时，所有犊牛均已产生针对牛巴贝斯虫的主动免疫。在所检测的75份DNA样本中，分别获得msa-1、msa-2b与msa-2c基因序列0条、3条和5条。本研究证实，从牛巴贝斯虫阳性犊牛血液DNA中扩增得到的msa-2b与msa-2c基因序列存在遗传多样性。上述数据凸显了在巴西开展牛巴贝斯虫遗传多样性深入研究的重要意义，以期未来能够开发针对性的诊断抗原与疫苗。