Entamoeba gingivalis is present in the oral cavity of humans and is associated with periodontal disease. Consequently, this study aimed to comprehensively investigate the E. gingivalis infection and the associated risk factors among individuals suffering from periodontal conditions. A cross-sectional descriptive study was carried out within a cohort of periodontal patients. Dental plaque specimens were meticulously collected and subsequently subjected to thorough examination using the polymerase chain reaction (PCR)-based technique targeting the small subunit ribosomal RNA (SrRNA) gene of the organism. The occurrence of risk factors for E. gingivalis infection was analyzed by the chi-square test and binary logistic regression. Out of the 230 participants, 60 were clinically diagnosed with periodontitis, while 170 were afflicted with gingivitis. Out of the 230 patients, 25 (10.9%) tested positive for E. gingivalis infections. An in-depth analysis unveiled that a significant majority of infections were recorded within subgroups characterized by a marital status (15.45%), manifestation of periodontitis (25.00%), and concomitant presence of underlying disease (20.83%). Furthermore, the high risk factor associated with E. gingivalis infection was the female (OR adj = 13.65, 95% CI = 1.08-173.21), followed by periodontitis (OR adj = 3.30, 95% CI = 1.21-9.00), respectively. The study employs a molecular diagnostic approach to screen for E. gingivalis enrichment within a subset of periodontal patients with advancing disease. The findings emphasize the necessity for further research to elucidate the pathogenesis of E. gingivalis and advocate for vigilant surveillance within a substantial population of periodontal patients.