Free-living amoeba diversity in river waters of western Brazil and their in vitro and in vivo pathogenic potential

Free-living amoebae (FLA) are key components of aquatic and terrestrial ecosystems, but their diversity and distribution in central-western Brazil remain poorly characterized. Some FLA are pathogenic and may harbor bacteria of public health relevance. We aimed to identify and to characterize the FLA biodiversity in the Cuiabá River basin and to infer their pathogenic potential. Water samples were collected at eight sites across two hydrological periods, and physicochemical and microbiological parameters were quantified. FLA were isolated on non-nutrient agar seeded with heat-inactivated Escherichia coli and identified by using morphology and molecular markers. We obtained 39 isolates (six genera and 14 species); Acanthamoeba (51.3%), Vannella (23.1%), and Vermamoeba (12.8%) were predominated. Only V. vermiformis was detected during both the high-water (when four exclusive species were detected) and low-water (when nine exclusive species were detected) periods. Seventeen FLA strains ( n = 35) killed zebrafish larvae ( Danio rerio ); 11 of these strains (64.7%) were Acanthamoeba species, and the remaining strains belonged to the genera Flamella , Naegleria , Ptolemeba , Vannella , and Vermamoeba . Thermo- and osmotolerance did not correlate with zebrafish larval mortality. These findings expand current knowledge of FLA biodiversity in Brazil and reinforce that integrated FLA monitoring in the Cuiabá River basin is needed. • This is the first time Flamella balnearia and Ptolemeba bulliensis have been reported in Brazil. • Acanthamoeba hatchetti, A. jacobsi, A. quina, A. stevensoni , and Vannella planctonica are newly reported in Brazil. • Osmotolerance and thermotolerance varied among and within species. • Challenging zebrafish larvae with FLA yielded strain-dependent larval mortality. • Osmotolerance and thermotolerance did not correlate with zebrafish larval mortality.