Molecular detection and phylogenetic characterization of Anaplasma phagocytophilum and the first evidence of Anaplasma platys in dogs from Iran: epidemiological and hematological findings

Abstract

Anaplasma phagocytophilum and Anaplasma platys are tick-borne pathogens of dogs, with zoonotic and veterinary importance. Data on their prevalence and hematological effects in Iran are scarce, and A. platys has not been molecularly reported in dogs in the country. This study investigated the molecular prevalence, risk factors, clinical presentation, phylogenetic position, and hematological alterations associated with these pathogens in dogs from Ahvaz, southwestern Iran. Blood samples were collected from 146 dogs between May 2024 and May 2025. Dogs with concurrent infections caused by pathogens other than A. phagocytophilum and A. platys were excluded. Hemi-nested PCR targeting groEL ( A. phagocytophilum ) and two-step PCR approach targeting 16 S rRNA ( A. platys ) were performed, followed by sequencing and maximum likelihood phylogenetic analysis. Hematological parameters were compared between infected and non-infected dogs, and risk factors were analyzed using logistic regression. Microscopic examination detected Anaplasma morulae in only two symptomatic dogs (1.4%), whereas PCR revealed substantially higher prevalence. Among 144 clinically healthy dogs, 54 (37.5%) were PCR-positive for at least one target Anaplasma spp.: 35 (24.3%) for A. phagocytophilum , 30 (20.8%) for A. platys , including 11 (7.6%) dual infections. Phylogenetic analysis confirmed species identity: the three A. phagocytophilum groEL sequences (PZ518362–364) clustered within the A. phagocytophilum clade and demonstrated closest affinity with globally distributed mammalian isolates, strongly suggesting affiliation with Ecotype I. The three A. platys 16 S rRNA sequences (PZ489281–283) formed a well-supported A. platys clade, clearly separated from other Anaplasma spp., providing the first molecular evidence of A. platys in Iran. Lack of tick control was the sole independent predictor of infection (adjusted OR ≈ 15; P < 0.001). Despite absent clinical signs, infected dogs presented significant hematological alterations. Subclinical A. phagocytophilum infection was associated with marked thrombocytopenia (mean platelets 98 × 10³/µL vs. 337 × 10³/µL in controls) and reduced erythrocyte, leukocyte, and neutrophil counts. A. platys -infected dogs exhibited milder but significant decreases in erythrocyte and platelet counts. Co-infected dogs had the most pronounced abnormalities. Only two dogs developed overt clinical disease (fever, lameness, diarrhea, cytopenias). This study provides the first molecular evidence and phylogenetic confirmation of A. platys infection in dogs in Iran. The A. phagocytophilum isolates clustered with strains commonly assigned to Ecotype I and therefore appear most consistent with this ecotype. Subclinical infections are common and associated with significant hematological changes, particularly thrombocytopenia. The strong association with lack of tick control underscores prevention as key. Infected dogs may serve as sentinels for human exposure risk, supporting One Health surveillance.