First study on molecular identification of Anaplasma ovis in sheep in southern Kazakhstan

Background and Aim: Anaplasmosis in small ruminants is a tick-borne infection caused mainly by the obligate intraerythrocytic bacterium Anaplasma ovis. It is usually subclinical, with persistent infection in affected animals, but acute disease can occur, particularly in young animals. The pathogen is widespread in Central Asia and neighboring regions. In Kazakhstan, the infection was first detected in 1929. However, until now, diagnosis in the country has been based on traditional microscopic examination of blood smears. There were no reliable data on the prevalence and genetic diversity of Anaplasma spp. in sheep in Kazakhstan. This study aimed to determine the occurrence of Anaplasma spp. infection in sheep in southern Kazakhstan, a high-risk region for tick-borne diseases, using PCR and to identify the species by sequencing. Materials and Methods: A cross-sectional study was conducted on apparently healthy adult ewes from 77 settlements in 34 districts of Kyzylorda, Turkistan, Zhambyl, Almaty, and Jetisu, southern Kazakhstan. A total of 2553 whole blood samples collected in midsummer 2022 and 2023 were analyzed for Anaplasma spp. using polymerase chain reaction targeting the 404 bp groEL gene fragment. The amplification products from the 441 positive samples were sequenced using the Sanger sequencing method. Phylogenetic analysis of the obtained sequences was performed using the maximum likelihood model. Results: Overall, 1017/2553 (39.8%; 95% confidence interval: 37.9%-41.7%) ewes tested were positive for Anaplasma spp. Positive animals were found in 68/77 (88%) of the settlements from which samples were taken. The percentage of Anaplasma spp.-positive ewes varied significantly from 21.3% to 50.1% in the provinces. Altitude <500 m above sea level was identified as a risk factor for Anaplasma infection. All amplification products were identified as A. ovis through sequencing. Phylogenetic analysis of the groEL gene fragment sequences revealed the presence of two A. ovis genotypes; one was 100% identical to sequences from isolates from China and the other was >99.5% identical to isolates from Africa, Cyprus, and China. Conclusion: This first molecular study revealed a widespread of A. ovis infection in adult ewes in southern Kazakhstan. Altitude <500 m was identified as a risk factor. Therefore, clinical cases associated with A. ovis are expected in this region, especially in young animals. Future studies are needed to determine the clinical and economic impact of anaplasmosis on sheep production in the country, to investigate seasonal patterns of infection, and to identify tick species or other arthropods that act as local vectors. This information is useful for developing possible control measures and evaluating their effectiveness.