Molecular prevalence and associated risk factors of Entamoeba spp. in donkeys in Shanxi Province, North China

Background The intestinal protozoa Entamoeba spp. can infect humans and various animals, including donkeys, causing diarrhea and malabsorption and presenting significant risks to animal husbandry and public health. Most Entamoeba species are not pathogenic except for Entamoeba histolytica. China has among the highest rates of donkey farming worldwide. Donkey (Equus asinus) farming is increasingly important in China because of their draft and medicinal value; however, epidemiological data on Entamoeba spp. in donkeys remains limited globally. This study aimed to investigate the prevalence of Entamoeba in donkeys in Shanxi Province, North China, and assess associated risk factors using a molecular approach. Methods Fecal samples of 815 donkeys from three representative geographical locations in Shanxi Province were collected to investigate the presence of Entamoeba spp. A portion of the small-subunit rRNA gene (SSU rRNA) was amplified and sequenced to determine the prevalence and species/genotypes of Entamoeba spp. Statistical analysis of possible risk factors was performed using Statistical Product and Service Solutions (SPSS) 26.0 software. The phylogenetic relationship of Entamoeba spp. was reconstructed using the neighbor-joining (NJ) method in Molecular Evolutionary Genetics Analysis (Mega) 7.0 software. Results The overall prevalence of Entamoeba spp. in donkeys in Shanxi Province was 7.12% (58/815). Two species (Entamoeba sp. RL9 and Entamoeba equi) were identified by sequence analysis; of these, Entamoeba sp. RL9 was the most prevalent species in donkeys in this study. Statistical analysis revealed that the donkeys' sex, region, age, and altitude are the risk factors associated with Entamoeba spp. prevalence (P < 0.05). Phylogenetic analysis indicated that the sequences of Entamoeba sp. RL9 and E. equi isolated from donkeys in this study were clustered with previously reported animal-derived Entamoeba sp. RL9 and E. equi sequences, respectively. Conclusions This study reports the occurrence and prevalence of Entamoeba spp. in donkeys worldwide for the first time to our knowledge. This not only expands the geographical distribution but also broadens the host range of Entamoeba spp., addressing the knowledge gap regarding the prevalence of Entamoeba spp. in donkeys, providing baseline data for carrying out prevention and control of Entamoeba spp. in donkeys in China.