Genetic diversity of Entamoeba Species among Children Under 5 years in the Vhembe District, Limpopo Province, South Africa

Background: Our understanding of the disease caused by the various Entamoeba species and its epidemiology is changing with time, and very little is known about their genetic diversity. Therefore, this study sought to investigate the prevalence and genetic diversity of Entamoeba species among children under 5 years in the Vhembe District, South Africa. Methods: : A total of 534 stool samples from 313 children (both males and females) aged under 5 were collected from 12 villages in Vhembe District, Limpopo. The prevalence of Entamoeba infections was examined by microscope and PCR, followed by Sanger sequencing for specific regions of 18S rRNA gene to identify and differentiate the circulating species of Entamoeba in the study population. Results: Of the 313 children recruited in the study, 163 were females and 150 were males, the ages ranged from 1 to 3 years. Of the 534 samples, 130/534 (24.3%) were microscopically positive for Entamoeba cysts. However, Entamoeba genus-specific DNA amplification using PCR identified 43/534 (8%) of Entamoeba species. Twenty positive amplicons were sequenced by Sanger sequencing technologies. Out of the twenty samples, twelve (60%) were confirmed to be Entamoeba species. The Entamoeba species identified in the study as evidenced by BLAST calls in the NCBI database and phylogenetic tree after narrowing the search using the option Entamoeba taxid (5758) from the NCBI database included 4 E. polecki (33.3%), 6 E. coli (50%), one E. muris (8.3%) and one E. hartmanni (8.3%). The phylogenetic tree showed the close relationship between isolated species and the ones in the GenBank. Conclusion: The current study has shown for the first time the presence of E. polecki in humans and the existence of possibly two types of E. coli infecting humans. Our findings further emphasize the need for the re-evaluation of the pathogenicity of species such as E. polecki which are quite common in the study population and might be responsible for some of the health complications.