Molecular survey and genetic characterization of Anaplasma marginale isolates in cattle from two regions of Russia.

Anaplasma marginale is an intraerythrocytic tick-borne rickettsial pathogen that causes bovine anaplasmosis, an economically important disease of cattle worldwide. Major surface protein MSP1α has been used as a stable marker in identifying geographical strains of A. marginale. The genetic diversity of A. marginale based on MSP1α has been reported in several countries all over the world. Only a few molecular surveys of A. marginale strains have been conducted in Russia. The aim of this study was molecular detection and characterization of A. marginale isolates in cattle from two regions of Russia. Blood samples from 62 cattle were collected and screened for the presence of A. marginale by real-time PCR targeting the msp4 gene. Anaplasma marginale DNA was detected in 26 cattle (42%). The partial msp1α gene containing tandem repeat sequences and msp4 gene were amplified from msp4-positive samples, cloned and sequenced. Sequence analysis revealed that two msp4 genotypes were found. The genetic diversity of A. marginale strains was analyzed based on the MSP1α tandem repeats structure and 5'-UTR microsatellite. Sixteen new genotypes of A. marginale were found in 17 animals. Seven animals (41%) were infected by more than one genotype. Eight new tandem repeats are described for the first time. The number of repeats differed between 1 and 6 across the isolates. The msp1α microsatellite analysis revealed that six genotypes were identified; one of them was not previously described. Phylogenetic analysis revealed that Russian isolates formed four separate clades. The tandem repeat and microsatellite analyses of the msp1α gene showed a high genetic diversity among the isolates. The present study provided the first evidence of genetic diversity of A. marginale in cattle in Russia.