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Recent outbreaks of enterically transmitted infections, including acute hepatitis A and shigellosis, have raised the concerns of increasing Entamoeba histolytica infection (EHI) among people living with HIV (PLWH) in Taiwan. This study investigated the prevalence of EHI, its temporal trends, and associated factors among newly diagnosed PLWH in Taiwan. Medical records of newly diagnosed PLWH at six medical centers in Taiwan between 2009 and 2018 were reviewed. The annual prevalence of invasive amoebiasis and seroprevalence of E. histolytica were determined and examined by the Cochran-Armitage test. The clinical characteristics associated with invasive amoebiasis and seropositivity for E. histolytica were analyzed in multivariable regression models. Among 5362 patients seeking HIV care at six medical centers in Taiwan during the 10-year study period, 119 (2.2%) had invasive amoebiasis at the time or within six months of their HIV diagnosis. Among 3499 who had indirect hemagglutination antibody (IHA) determined, 284 (8.1%) had positive IHA (≥1:32) and 205 (5.9%) had high-titre IHA (≥1:128). The prevalence of invasive amoebiasis increased from 1.3% in 2012 to 3.3% in 2018 (p = 0.024). Invasive amoebiasis was independently associated with a greater age, men who have sex with men, rapid plasma reagin titre ≥1:4, and concurrent shigellosis and giardiasis. Increasing prevalence of invasive amoebiasis among newly diagnosed PLWH in Taiwan calls for strategies to prevent ongoing transmission in this population. Routine screening of EHI for early diagnosis and treatment is recommended, especially among men who have sex with men and those who present with other sexually or enterically transmitted infections., Author summary Outbreaks of enterically transmitted infection, including acute hepatitis A and shigellosis, among men who have sex with men and people living with HIV have been reported in Taiwan and in many developed countries in recent years. This study reveals that the prevalence of invasive amoebiasis among newly diagnosed people living with HIV increased in Taiwan since 2012, accompanied by increasing seroprevalence of syphilis and hepatitis A virus infection. This study also shows that concurrent infections with shigellosis and giardiasis and history of syphilis were independently associated with invasive amoebiasis, which indicates that transmission of Entamoeba histolytica might have occurred through sexual behaviours that increased faecal-oral contact in this population. In the era of improved access to HIV prevention and treatment, emerging and re-emerging enterically transmitted infections, including amoebiasis, pose an ongoing health threat to at-risk individuals and the public as a whole.