Filogenetska analiza i molekularna karakterizacija terenskih izolata bakterije Anaplasma spp. izoliranih iz goveda u Indiji

In India, the economic losses incurred due to ticks and tick-borne diseases are estimated around 498.7 million US dollars. Among these diseases, anaplasmosis causes significant mortality and morbidity in animals, leading to significant production losses. However, only scant information on the molecular characterization, phylogenetic and matrix analysis of Anaplasma spp in cattle is available. Therefore, the objective of the present study was to perform phylogenetic and molecular characterization of field isolates of Anaplasma spp. infecting dairy animals in Uttar Pradesh, India. Blood samples from 200 cattle showing the clinical signs of bovine anaplasmosis were collected from the western part of Uttar Pradesh, India. To detect Anaplasma spp., screening of blood smears and molecular confirmation by PCR was performed. The molecular characterization was done by sequencing the 16S rRNA gene and its bioinformatic analysis using MEGA version X. On the basis of microscopic examination, Anaplasma spp. were detected in 106 (53%) samples, while the 16S rRNA gene-based polymerase chain reactions revealed positive results in 176 (88%). The sequencing of the 16S rRNA gene and the bioinformatic analysis of sequences revealed the existence of three different populations of Anaplasma spp., viz., Anaplasma marginale and two other Anaplasma spp. genetically related to Anaplasma capra and Anaplasma ovis, circulating in the blood of infected cattle. All the field isolates of A. marginale and A. ovis from Uttar Pradesh, India, were clustered in a single clade with others isolated from Iran, Brazil, Thailand and Israel, while A. capra isolates from India, established in same clade of A. capra, have been reported from Japan, China and South Korea. To the best of the authors’ knowledge, this is the first report of Anaplasma capra and Anaplasma ovis from bovine in India, and indicates the potential of cattle to serve as reservoirs of these pathogens, leading to the requirement for further studies of these emerging zoonotic pathogens and their possible zoonotic potential.Ekonomski gubici koje u Indiji uzrokuju krpelji i bolesti koje se prenose krpeljima procjenjuju se na oko 498,7 milijuna američkih dolara. Među tim je bolestima i anaplazmoza, koja uzrokuje znatnu smrtnost i pobol među životinjama, što vodi do velikih gubitaka u proizvodnji. S obzirom na oskudne podatke o molekularnoj karakterizaciji, filogenetskoj i matriksnoj analizi bakterije Anaplasma spp. u goveda, cilj je ovog rada bio dobiti znanstvene informacije iz navedenih područja istraživanja na terenskim izolatima Anaplasma spp. koja inficira mliječna goveda u zapadnom dijelu savezne države Uttar Pradesh u Indiji. Prikupljeni su uzorci krvi od 200 goveda koja su pokazivala kliničke znakove anaplozmoze. Za detekciju bakterije učinjena je analiza razmaza krvi i molekularna potvrda PCR-om. Karakterizacija izolata na molekularnoj razini provedena je sekvenciranjem gena 16S rRNA i njegovom bioinformatičkom analizom upotrebom MEGA verzije X. Mikroskopskim je pregledom Anaplasma spp. otkrivena u 106 uzoraka (53%), dok je PCR analizom gena 16S rRNA pozitivno bilo 176 uzoraka (88%). Sekvenciranje gena 16S rRNA i bioinformatička analiza slijedova otkrili su postojanje triju različitih populacija bakterije Anaplasma spp., i to Anaplasma marginale i dviju drugih anaplazmi genetski povezanih s vrstama Anaplasma capra i Anaplasma ovis koje cirkuliraju u krvi zaraženih goveda. Svi su terenski izolati bakterija A. marginale i A. ovis iz države Uttar Pradesh svrstani u jednu filogenetsku kladu zajedno s izolatima iz Irana, Brazila, Tajlanda i Izraela, dok su izolati A. capra iz Indije svrstani u kladu s A. capra iz Japana, Kine i Južne Koreje. Prema autorovim je saznanjima ovo prvo izvješće o vrsti Anaplasma capra i Anaplasma ovis u goveda u Indiji što upućuje na potencijal goveda da budu rezervoari ovih patogena. To pak upućuje na potrebu za daljnjim istraživanjima bakterije Anaplasma spp. s obzirom na njezinu emergenciju i zoonotski potencijal