Abstract
Hepatozoon canis, causes canine hepatozoonosis, is an important haemoparasite of domestic and wild carnivores, which is increasingly recognized as an important component of veterinary infectious disease because of complex ecological interactions and geographical distribution of its arthropod vectors. This study aimed to investigate the molecular epidemiology of H. canis in domestic (herding and pet) and stray dogs in central Iraq using conventional PCR assay with sequencing and phylogenetic analysis. Furthermore, the study aims to evaluate epidemiological association between molecular data and potential risk factors involving management type, age, sex, geographical distribution, and area to provide a comprehensive understanding for epidemiological and molecular characteristics of H. canis infection in study dogs. Totally, 170 (68 herding, 72 pet, and 30 stray) dogs of various ages and sexes were selected randomly from different rural, sub-urban and urban areas located in Baghdad, Babylon and Wasit provinces during July (2025)-May (2026). Targeting 18S rRNAgene, venous blood of all study dogs were tested molecularly by conventional PCR, and the positive PCR products were sequenced and analysed phylogenetically. Epidemiological data concerned study dogs were reported and subjected to statistical analysis using the GraphPad Prism software. The results revealed that 24.71% of study dogs were having the DNAs of H. canis; in which, molecular epidemiology indicates the presence of identity at 97.70-99.82% and substitution at 0.02%. Subsequently, the study isolates were closely-related to five global isolates / strains at an identity ranged from 98.54% to 100% and mutation 0.02% and distributed as following: 45.24% to the South Korean C21 isolate (ID: PQ326382.1), 33.33% to the Iraqi SUF8 isolate (ID: PZ476549.1), 11.91% to the Thai BK-39 strain (ID: KF621087.1), 7.14% to the Taiwanese 98-TP-DA09-sd02-M strain (ID: MH595900.1), and 2.38% to Italian 4252 isolate. Epidemiological data demonstrated that the prevalence rate of H. canis was significantly higher in stray (46.7%) and herding (36.8%) dogs than pet dogs (4.2%); dogs aged >4 years old (57.9%) than 1-4 (8.8%) and <1 (4.5%) years; females (33%) than males (10.9%); and in Wasit (33.3%) than Babylon (17%) but not (p<0.304) Baghdad (23.3%); however, no remarkable variation (p>0.05) was detected between urban, sub-urban and rural areas. In conclusion, this might represent the first study in studied provinces; in which, local isolates were closely-related to five global isolates/strains. Epidemiological data reported that prevalence rate and risk of H. canis varied significantly according to management type, age, sex and geographical distribution. This suggests that furthermore such studies are greatly needed to provide more information about the incidence of H. canis.phylogenetically. Epidemiological data concerned study dogs were reported and subjected to statistical analysis using the GraphPad Prism software. The results revealed that 24.71% of study dogs were positively infected dogs with H. canis, in which, phylogenetic analysis indicate the presence of identity at 97.70-99.82% and substitution at 0.02%. Subsequently, the study isolates were closely-related to five global isolates / strains at an identity ranged from 98.54% to 100% and mutation 0.02% and distributed as following: 45.24% to the South Korean C21 isolate (ID: PQ326382.1), 33.33% to the Iraqi SUF8 isolate (ID: PZ476549.1), 11.91% to the Thai BK-39 strain (ID: KF621087.1), 7.14% to the Taiwanese 98-TP-DA09-sd02-M strain (ID: MH595900.1), and 2.38% to Italian 4252 isolate. Epidemiological data demonstrated that the prevalence rate of H. canis was significantly higher in stray (46.7%) and herding (36.8%) dogs than pet dogs (4.2%), dogs aged >4 years old (57.9%) than 1-4 (8.8%) and <1 (4.5%) years, females (33%) than males (10.9%), and in Wasit (33.3%) than Babylon (17%) but not (p<0.304) Baghdad (23.3%), however, n

Key words: Apicomplexan haemoparasite, Canine hepatozoonosis, Phylogenetic analysis, Risk factors, Tick-borne pathogens