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The current study included the microscopic diagnosis of Entamoeba histolytic parasite, (140) stool samples were collected from children ranging in age from one to fourteen years with diarrhea from the Children's Hospital in Kirkuk from September 2020 to August 2021 using direct examination methods. The percentage of total infection was (23.57%). The current study also included the use of the polymerase chain reaction (PCR) method to diagnose amoeba infection, and the genetic sequencing method was used to determine the type of amoeba. Where 28 positive samples were subjected (through microscopy) to molecular examination by diagnosing the 18s ribosomal RNA gene, and the results showed that 19 out of 28 samples were (positive for amoeba) and 17 of 19 samples were (positive for Entamoeba histolytica). Molecular characterization and genetic analysis of E.histolytica was carried out and it was found that the closest recorded isolates belong to isolates from Baghdad, Diwaniyah and Dhi Qar (from Iraq) and to South Africa, Egypt, India and Iran. Also, (11) samples were sent to the International Gene Bank (NCBI) and approval was obtained to register the study samples that were subjected to molecular characterization and given them the serial numbers, OL771805, OL771806, OL771807, OL771808, OL771809, OL771810, OL771811, OL771812, OL771813 and OL77.