Ticks and tick-borne diseases (TTBDs) are considered major causes of economic loss in the livestock sector which incur an annual control cost estimated at US$ 498.7 million in India. Among these diseases, babesiosis, theileriosis and anaplasmosis are listed among the top ten livestock diseases in India and cause significant mortality and morbidity among cattle. However, molecular characterization of bovine Babesia and Anaplasma species are scant; thus, the aim of this study is to perform molecular characterization of field isolates of Babesia spp. and Anaplasma spp. infecting bovines in Kerala, South India. Blood smears and whole blood samples were collected from a total of 199 apparently healthy adult female cattle in Kerala. Based on microscopy, Babesia spp., Theileria orientalis and Anaplasma spp. organisms were detected in 9 (4.5%), 40 (20%) and 6 (3%) samples, respectively. Genus-specific polymerase chain reactions for amplification of 18S rRNA of Babesia spp. and 16S rRNA of Anaplasma spp. revealed positive results with 18 (9%) and 14 (7%) samples. The phylogenetic analysis of 18S rRNA gene sequences of Babesia spp. confirmed the existence of two different populations of Babesia spp. circulating in the blood of infected cattle viz., Babesia bigemina and a Babesia sp. genetically related to Babesia ovata. Further phylogenetic analysis using rap-1a sequences of isolates of B. bigemina revealed higher levels of genetic heterogeneity. However, the field isolates of B. bigemina displayed only slight heterogeneity when the rap-1c gene was examined. Polymerase chain reaction followed by sequencing and phylogenetic analysis of 16S rRNA gene of Anaplasma spp. revealed the existence of Anaplasma marginale, Anaplasma bovis and Anaplasma platys in bovines in South India. Based on msp4 gene sequences, all the field isolates of A. marginale from Kerala were clustered in a single clade with others isolated from around the world. To our knowledge, this study forms the first report on occurrence of B. ovata-like parasites and A. platys in cattle from India.

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In the present study, 111 blood samples were collected from apparently healthy cats belonging to four districts of Kerala, southern India, and they were investigated for the presence of hemoparasites and hemoplasmas by light microscopic examination and polymerase chain reaction (PCR). The microscopic examination of the Giemsa-stained blood smears did not reveal any parasites/organisms. However, PCR followed by nucleotide sequencing could detect 10 (9.01%) out of 111 samples infected with Hepatozoon felis, 3 (2.70%) with Cytauxzoon spp., and 10 (9.01%) with Mycoplasma spp. None of the samples revealed amplicons specific for the Babesia spp. and Trypanosoma evansi. The phylogenetic analysis of 18S ribosomal RNA (rRNA) gene sequences of H. felis revealed the existence of two different populations of H. felis circulating in the blood of infected cats. The phylogenetic tree was constructed based on 18S rRNA gene sequences of Cytauxzoon spp. and revealed that these isolates formed a distinct clade and do not cluster with any of the isolates from other countries. Among the 10 samples positive for Mycoplasma spp. infections, 7 were detected positive for Candidatus Mycoplasma haemominutum, two for Mycoplasma haemofelis, and one for Candidatus Mycoplasma turicensis. Phylogenetic analysis of 16S rRNA gene sequences of Mycoplasma spp. showed no distinct geographical grouping of the sequences. The sequences of M. haemofelis, Candidatus M. haemominutum, and Candidatus M. turicensis identified in the study clustered along with their respective isolates from around the world. To the best of our knowledge, this study forms the first report of molecular detection of Cytauxzoon spp. and Candidatus M. turicensis in cats from India.

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