Well water sources simultaneous contamination with Cryptosporidium and Acanthamoeba in East-Southeast Asia and Acanthamoeba spp. in biofilms in the Philippines.

Cryptosporidium is the leading agent of waterborne parasitic protozoan outbreaks and is the second leading cause of infant mortality due to diarrhoea worldwide. Acanthamoeba spp. causes Acanthamoeba keratitis (AK) and a life-threatening condition known as granulomatous amoebic encephalitis (GAE). The present study aimed to assess the water quality of an indigenous and a rural community for waterborne parasitic protozoan contamination. Aquatic samples (n = 22) were processed by filtration of 500 mL portion through a 1.2 µm pore size glass microfiber filter and eluted for light microscopy, culture in non-nutrient agar, and PCR analysis. Overall, 36% (8/22) of the investigated aquatic samples were positive for either Cryptosporidium spp. oocysts (13%; 3/22) or Acanthamoeba spp., (36%; 8/22) or both (13%; 3/22). Cryptosporidium spp. oocysts were detected in 27% (3/11) of wet season samples only while Acanthamoeba spp. were detected in 18% (2/11) and 55% (6/11) of wet and dry season samples, respectively. Subsequently, molecular detection for Acanthamoeba species identified A. lenticulata and A. hatchetti with 98-99% BLAST similarity. This is the first report on the simultaneous contamination of Cryptosporidium and Acanthamoeba in well water sources in East-Southeast Asia, the first detection of Acanthamoeba spp. in biofilms in the Philippines, and the longest viability demonstrated for A. lenticulata in two-year-old water samples stored at room temperature.

Acanthamoeba species isolated from Philippine freshwater systems: epidemiological and molecular aspects.

Free-living amoeba (FLA) research in the Philippines is still in its infancy but has, by far, demonstrated the presence of potentially pathogenic species. Acanthamoeba may cause sight-threatening and central nervous system infections to humans, yet its epidemiologic distribution from local environmental sources is yet to be defined. The present study aimed to provide a baseline epidemiologic distribution of Acanthamoeba spp. in freshwater systems in the Philippines and establish potential pathogenicity of isolates through thermo-tolerance assay. A total of 63 water samples were collected from 13 freshwater systems all over the Philippine archipelago. The low-volume (50 ml) water samples were processed and cultured on non-nutrient agar lawned with Escherichia coli and observed for amoebic growth using light microscopy. Amoebic culture demonstrated 14.28% (9/63) positivity while further molecular testing of culture-positive plates using Acanthamoeba-specific primers demonstrated 100% (9/9) confirmation of Acanthamoeba species. Genotyping of Acanthamoeba isolates revealed T1, T3, T4, T5, T7, T11, and T15 genotypes. Thermo-tolerance assay demonstrated that T5 and T7 genotypes were potentially pathogenic strains. The evidence of environmental distribution of Acanthamoeba spp. in the freshwater systems in the Philippines and thermo-tolerance profile of isolates are significant aspects of amoeba study in public health and calls for initiatives in the dissemination of relevant information and the expansion of knowledge, awareness, and policies on pathogenic waterborne amoeba to mitigate, prevent, detect, and report cases of human infections.