Molecular Characterisation of Anaplasma marginale and Theileria orientalis in Slaughtered Bovines of Aizawl District, Mizoram

Abstract

Background: Bovines are domesticated ungulates mainly raised as livestock for milk, meat and leather and as draught animals. A slaughterhouse study helps to assess the disease status of herds and contains valuable information about the incidence and epidemiology of animal diseases. Haemoprotozoan diseases are one of the major problems which adversely affect the health and productivity of cattle. The diverse climatic zones of India are highly conducive to the survival and propagation of vectors and vector-borne pathogens. However, there is no data on proper slaughterhouse study of bovines in Mizoram even in India for determination of the prevalence of haemoprotozoan infections. Methods: The blood samples were collected in an EDTA vials. The blood smear was prepared, stained with Giemsa stain and examined for the presence of haemoprotozoan. The DNA was isolated from positive samples by use of a DNeasy Blood and Tissue Kit. Then PCR was performed for all the isolated DNA collected from positive samples. After confirmation through PCR, the representative samples for Anaplasma sp. and Theileria sp. were sent for sequencing. Then the sequencing results were annotated using BLAST, ClustalW multiple alignment program of MEGA9 software. The phylogenetic tree was generated by using the Neighbour-Joining method, keeping bootstrap consensus from 1000 replicates. Result: After the blood smear examination, PCR was performed for confirmation. Molecular detection of Anaplasma marginale, Theileria orientalis was done. Molecular characterization of 2 isolates from Anaplasma marginale and Theileria orientalis was performed by sequencing. The sequence analysis of rpoB gene of isolates of Anaplasma revealed both the isolates were placed in the same clade in the phylogenetic tree. The phylogenetic analysis of Thileria isolates revealed that isolate Miz-86 was closely placed with the Type-N3 clade. Whereas the other isolate i.e. Miz-87, belonged to the Type-5 clade.