Survey of tick-borne pathogens in grazing horses in Kyrgyzstan: phylogenetic analysis, genetic diversity, and prevalence of Theileria equi.

Introduction: Tick-borne pathogens (TBP) are an important group of organisms that can affect animals and humans all over the world. Equine piroplasmosis (EP), caused by Theileria equi and Babesia caballi, is considered one of the most important tick-borne diseases and can cause significant clinical symptoms and mortality in horses. Moreover, EP plays a restrictive role in international horse traditions and transportation. Although these species can cause similar symptoms, there are different 18S rRNA genotypes of T. equi (five genotypes) and B. caballi (three genotypes). Besides piroplasma species, Anaplasma and hemotropic mycoplasmas (HM) are known as other important tick-borne pathogens reported in horses. Methods: In this study, we investigated the presence, prevalence, genetic diversity, and phylogenetic analyses of TBPs using PCRs and DNA sequencing in grazing horses in Kyrgyzstan. For these purposes, a total of 311 blood samples were collected from Chuy, Issyk-Kul, Naryn, Osh, Talas, and Jalal-Abad. Results: DNA amplification of TBP revealed that 23 (7.40%) out of 311 samples were found to be positive for T. equi. However, B. caballi, HM, A. phagocytophilum, and A. capra were not detected in this study. The infection rate of T. equi was higher in males (8.11%) than in females (6.35%) (p=0.2880) and in those older than 5 years (9.02%) than in the 1-4 age group (6.35%) (p=0.1950). Phylogenetic analysis of 18S rRNA revealed that A and E genotypes of T. equi have circulated in grazing horses in Kyrgyzstan. Discussion: Information about the genetic diversity of T. equi is important for understanding the population dynamics of the species and developing effective control strategies against this pathogen. This is the first molecular investigation of A. capra in horses in Kyrgyzstan. Although this pathogen has been detected in different hosts in Kyrgyzstan, it was not detected in this study. However, considering the wide host spectrum of A. capra, it is thought that more large-scale studies are needed to understand the effect of horses on the epidemiology of this pathogen.

First Molecular Evidence of Babesia vogeli, Babesia vulpes, and Theileria ovis in Dogs from Kyrgyzstan.

Tick-borne parasitic diseases cause mild to severe infections among vertebrate hosts, including dogs. Species in the genus Babesia are important tick-borne pathogens and have worldwide distributions. Although there are data on the prevalence and distribution of Babesia species among dogs around the world, there is no information available in Kyrgyzstan, according to a literature review. In this study, 337 dogs were screened by nested PCR for the presence of the 18S small subunit ribosomal RNA (18S SSU rRNA) gene of piroplasm species. Overall prevalence was 6.23% (21/337) for Babesia/Theileria spp. DNA sequencing of positively tested samples revealed that eighteen samples were infected with Babesia vogeli (B. vogeli) (5.34%), two samples with B. vulpes (0.59%), and one sample with Theileria ovis (T. ovis) (0.29%). The phylogenetic analyses and nucleotide sequences in contrast with those present in GenBank revealed that two nucleotide substitutions (594th and 627th) were found between B. vogeli isolates, including ours, indicating that the mutation is relatively rare. The sequences of other pathogens obtained in this study confirmed 100% nucleotide identity with B. vulpes and T. ovis sequences in GenBank. To the best of our knowledge, B. vogeli, B. vulpes, and T. ovis were detected for the first time in dogs from Kyrgyzstan, and it is thought that results will contribute to the understanding of the epidemiology of canine tick-borne pathogens in the country.

The detection and phylogenetic analysis of Anaplasma phagocytophilum-like 1, A. ovis and A. capra in sheep: A. capra divides into two genogroups.

In this study, the presence, prevalence, and genotypes of Anaplasma phagocytophilum, A. ovis, and A. capra in sheep were investigated based on 16 S SSU rRNA, groEL, and gtlA gene-specific polymerase chain reaction (PCR), respectively. The sequences of the genes were used for detection of the phylogenetic position of the species. Additionally, a restriction fragment length polymorphism (RFLP) were carried out for discrimination of A. phagocytophilum and related variants (A. phagocytophilum-like 1 and 2). The prevalence of Anaplasma spp. was found as 25.8% (101/391), while it was found that A. ovis, A. phagocytophilum-like 1, and A. capra are circulating in the sheep herds in Kyrgyzstan, according to the PCRs, RFLP and the partial DNA sequencing results. The positivity rates of A. phagocytophilum-like 1, A. ovis, and A. capra genotype-1 were 6.9, 22.5, and 5.3%, respectively. A total of 32 (8.2%) sheep were found to be mix infected. Moreover, phylogenetic analyses and sequence comparison with those available in the GenBank showed that A. capra formed two distinct genetic groups (A. capra genotype-1 and A. capra genotype-2). Considering the zoonotic potential of these species, it may be necessary to make changes in the interpretation of anaplasmosis cases in animals and there is a need for further studies to determine the pathogenicity of the species/genotypes circulating in animals.

First molecular detection of Anaplasma species in cattle from Kyrgyzstan; molecular identification of human pathogenic novel genotype Anaplasma capra and Anaplasma phagocytophilum related strain.

Anaplasmosis is a rickettsial infection with significant effects on human and animal health, and the discovery of new species or genotypes with zoonotic potential in recent years has increased this importance. The aim of this study was to provide the first assessment of the molecular etiology and prevalence of bovine anaplasmosis in Kyrgyzstan (specifically in the Chuy, Talas, Djalal-Abad, Naryn, and Issyk-Kul regions). The prevalence of bovine anaplasmosis was determined as 1.7% (6/358). PCR and partial DNA sequencing results of the 16S ribosomal RNA (rRNA) gene revealed that Anaplasma centrale, A. phagocytophilum like-1, and the human pathogenic novel genotype A. capra are circulating in cattle herds in Kyrgyzstan. Six DNA nucleotide sequences obtained in this study were deposited in GenBank under the following accession numbers: A. centrale (MW672117, MW672118, MW672119, MW672120), A. phagocytophilum (MW672121), and A. capra (MW672115).

First Molecular Detection of Dirofilaria immitis and D. repens in Dogs from Kyrgyzstan.

BACKGROUND: Dirofilaria immitis and Dirofilaria repens are the causative agents of cardiopulmonary and subcutaneous dirofilariosis, respectively. This neglected disease mainly seen in dogs, cats and wild carnivores is re-emerging recent years. No study was conducted on dirofilariosis in dogs in Kyrgyzstan. PURPOSE: The goal of this study was to investigate Dirofilaria species using PCR and sequencing in dogs from Kyrgyzstan. METHOD: Dirofilaria spp. infection in dogs was screened via convential PCR and sequencing in 337 dogs from Kyrgyzstan. RESULT: The overall prevalence of Dirofilaria spp. was 0.59% (2/337): DNA of D. immitis was detected in one sample and DNA of D. repens in second positive sample. In second sample, parallel co-infection of D. repens with Wolbachia was also found. While D. immitis sequence showed 98.70-100% similarity with previously reported sequences of D. immitis from dog blood, D. repens shared 100% identity with other sequences of D. repens. CONCLUSION: These results provided first evidence for Dirofilaria spp. in Kyrgyzstan and emphasized the veterinary and medical importance.