Effect of thermal treatment on free-living amoeba inactivation

AbstractAims: To evaluate the effect of temperature on two amoeba strains of thegenera Acanthamoeba and two amoeba strains of the genera Hartmannellaseparately treated depending on their life stage, trophozoite or cyst, when cellsare directly exposed under controlled conditions.Methods and Results: For thermal treatments, three temperatures wereselected 50, 60 and 70°C, and a microcosm was designed using dialysis bags.The inactivation of each strain was determined using a method based on themost probable number quantification on agar plates. The results showed thatfor all amoeba strains, thermal treatment was more effective againsttrophozoites compared with cyst stages. The inactivation patterns showedstatistical differences between the two genera analysed at temperatures above50°C. The effectiveness of the thermal treatments at 60 and 70°C was higherfor both life stages of Hartmannella vermiformis strains compared withAcanthamoeba strains, being the most resistant Acanthamoeba cysts.Conclusions: Free-living amoebae have been isolated in a wide range ofenvironments worldwide due to their capacity to survive under harshconditions. This capacity is mainly based on the formation of resistant forms,such as double-walled cysts, which confers a high level of resistance as shownhere for thermal treatments.Significance and Impact of Study: Free-living amoebae survival can promote arapid recolonization of drinking water systems and is a likely source ofemerging opportunistic pathogens such as Legionella. Because of that a betterunderstanding of the factors that affect micro-organism inactivation in watersystems would allow more efficient application of disinfection treatments.IntroductionFree-living amoebae (FLA) are widespread protozoa andnormal inhabitants of soil, and in particular, fresh-watersources (Rodriguez-Zaragoza 1994). These aquatic sourcesinclude environmental water systems such as lakes, ponds(Barbeau and Buhler 2001) and man-made water systems,including water supplies, hospital water networks,cooling towers, swimming pools and hydrotherapy baths(Schmitz-Esser et al. 2008; Thomas et al. 2010).Amoebae are a group of unicellular micro-organismsthat have a two-stage life cycle: a replicating and feedingtrophozoite stage and a latent cyst stage. In the trophozoitestage, FLA are important predators of prokaryotic andeukaryotic micro-organisms with much influence on thecomposition of the microbial community feeding on bio-films and detritus (Rodriguez-Zaragoza 1994; Ramaleyet al. 2001; Schmitz-Esser et al. 2008). Specifically, theyestablish symbiotic relationships with other microbesincluding bacteria of the genera Legionella, Mycobacte-rium, Campylobacter and Listeria, which, apart frombeing a nutritional source, are able to survive amoebadigestion and multiply within them (Greub and Raoult2004; Schmitz-Esser et al. 2008). Moreover, some FLAspecies can infect humans and other mammals, causingserious illnesses like ocular and central nervous system