Spatiotemporal distribution of thermophilic free-living amoebae in recreational waters: A 5-year survey in Guadeloupe (French West Indies)

Abstract

Free-living amoebae (FLA) such as Acanthamoeba, Balamuthia mandrillaris, Naegleria fowleri and Sappinia pedata are naturally widespread in freshwater, causing rare but fatal and debilitating infections in humans. Although recent studies have shown an increase in infection rates, there is a paucity of epidemiological studies regarding the presence of these emerging pathogens in water. Herein, we studied the diversity and relative abundance of thermophilic FLA in different recreational baths in a tropical climate for 5 years.From 2018 to 2022, a total of 96 water samples were collected from 7 recreational baths (natural, tiled, regularly cleaned or not, and with temperatures ranging from 27 to 40 °C). DNA was extracted from FLA cultivated at 37 °C to detect thermophilic culturable FLA. Metabarcoding studies were conducted through FLA 18S rRNA gene amplicons sequencing; amplicon sequence variants (ASV) were extracted from each sample and taxonomy assigned against PR2 database using dada2 and phyloseq tools. We also searched for Naegleria sp. and N. fowleri using PCR targeting ITS and NFITS genes (respectively) and we quantified them using an optimized most probable number (MPN) method for FLA. Our results showed that differences in FLA diversity and abundance were observed amongst the 7 baths, but without a clear seasonal distribution. Naegleria, Vermamoeba and Stenamoeba were the most represented genera, while the genera Acanthamoeba and Vahlkampfia were mainly found in 2 baths. The MPN values for Naegleria sp. (NT/l) increased between 2018 and 2022, but the MPN values for N. fowleri (NF/l) seemed to decrease.Globally, our results showed that since we cannot establish a seasonal distribution of FLA, the regular presence of FLA (namely Naegleria and Acanthamoeba) in recreational waters can pose a potential threat in terms of neuroinfections as well as Acanthamoeba keratitis. It is thus imperious to perform the regular control of these baths as a preventive health measure.