Prevalence and seasonal variation of Acanthamoeba in domestic tap water in greater Sydney, Australia

Background: This study examined the prevalence of free-living Acanthamoeba in domestic tap water in the greater Sydney region, Australia, and determined any seasonal variation in prevalence. Methods: Fifty-four participants were included in this study following approval from an institutional human research ethics committee. Each participant self-collected two samples (one in summer and another in winter) from the surface of the drain of the bathroom sink using an instructional kit. The samples were cultured by inoculating onto a non-nutrient agar plate seeded with Escherichia coli and incubation at 32°C for two weeks. The plates were microscopically examined for the presence of free-living amoeba. DNA was isolated from 20 samples and a polymerase chain reaction (PCR) assay was performed for amplification of the partial sequence of the 18S ribosomal RNA gene. The PCR amplified products were sequenced using Sanger sequencing and genotyping was performed based on the variation in nucleotide sequences. Results: A total of 97 samples were collected over the two collection periods, with 28.6 per cent of samples morphologically classified as Acanthamoeba. The summer period yielded 16 of 54 (29.6 per cent) samples classified as Acanthamoeba, while the winter period yielded 12 of 43 (27.9 per cent) samples classified as Acanthamoeba. There was no statistically significant difference (p = 0.85) between the prevalence of free-living Acanthamoeba in summer compared to winter. Phylogenetic analysis showed that 15 of 20 (75 per cent) isolates belonged to genotype T4, the most frequent genotype isolated in Acanthamoeba keratitis. Conclusion: The prevalence of free-living Acanthamoeba characterised morphologically in domestic tap water of the greater Sydney region was higher than expected, especially considering the low incidence of Acanthamoeba keratitis in Australia. However, this study did not find variation between seasons. As the T4 genotype was most common, Sydney-based practitioners must always consider Acanthamoeba as a possible causative organism in cases of microbial keratitis, regardless of the season.