Free-Living Protozoa in Two Unchlorinated Drinking Water Supplies, Identified by Phylogenic Analysis of 18S rRNA Gene Sequences

Abstract

Free-living protozoan communities in water supplies may include hosts for Legionella pneumophila and other undesired bacteria, as well as pathogens. This study aimed at identifying free-living protozoa in two unchlorinated groundwater supplies, using cultivation-independent molecular approaches. For this purpose, samples (<20°C) of treated water, distributed water, and distribution system biofilms were collected from supply A, with a low concentration of natural organic matter (NOM) (<0.5 ppm of C), and from supply B, with a high NOM concentration (7.9 ppm of C). Eukaryotic communities were studied using terminal restriction fragment length polymorphism and clone library analyses of partial 18S rRNA gene fragments and a Hartmannella vermiformis-specific quantitative PCR (qPCR). In both supplies, highly diverse eukaryotic communities were observed, including free-living protozoa, fungi, and metazoa. Sequences of protozoa clustered with Amoebozoa (10 operational taxonomic units [OTUs]), Cercozoa (39 OTUs), Choanozoa (26 OTUs), Ciliophora (29 OTUs), Euglenozoa (13 OTUs), Myzozoa (5 OTUs), and Stramenopiles (5 OTUs). A large variety of protozoa were present in both supplies, but the estimated values for protozoan richness did not differ significantly. H. vermiformis was observed in both supplies but was not a predominant protozoan. One OTU with the highest similarity to Acanthamoeba polyphaga, an opportunistic human pathogen and a host for undesired bacteria, was observed in supply A. The high level of NOM in supply B corresponded with an elevated level of active biomass and with elevated concentrations of H. vermiformis in distributed water. Hence, the application of qPCR may be promising in elucidating the relationship between drinking water quality and the presence of specific protozoa.Free-living protozoa are ubiquitous in natural freshwater environments (7,38,51,71) but also proliferate in engineered water systems, including water treatment systems (3,47,70), distribution systems (6, 75), and tap water installations inside buildings (54, 69). Concentrations of protozoa, determined using cultivation methods and microscopy, range from Ͻ1 to 10 4 cells liter Ϫ1 in treated water (3,47,70,75) and from Ͻ1 to 7 ϫ 10 5 cells liter Ϫ1 in distribution systems (6,61,64,75). Genera of free-living protozoa commonly observed in these systems and in tap water installations include Acanthamoeba,

Pyrosequencing analysis of eukaryotic and bacterial communities in faucet biofilms

“…Modeling of data from a biofilm-batch model indicated that amoebae can only multiply when a threshold concentration of prey bacteria is reached (Kuiper, 2006). Valster et al (2009Valster et al ( , 2011) indicated that the growth of H. vermiformis in drinking water distribution systems is limited at low biofilm concentrations or at ATP concentrations of ≤1 ng L −1 . A number of eukaryotic sequences from the WSW biofilm were related to those of species unlikely to live in biofilm systems, such as Arthropoda and Chordata in Metazoa, and Spermatophyta in Viridiplantae.…”

Relationships between Free-Living Protozoa, Cultivable Legionella spp., and Water Quality Characteristics in Three Drinking Water Supplies in the Caribbean

“…Most information on community composition and abundance of FLP in freshwater environments has been obtained by using cultivation methods and microscopy. Recently, however, the presence and identities of such organisms in drinking water supplies in temperate regions have been studied by using molecular methods for detection and identification (33,51). In two groundwater supplies in the Netherlands a total of 127 operational taxonomic units (OTUs) of FLP were identified based on their 18S rRNA gene sequences.…”

Occurrence of Amoeba and Amoeba-resistent Mycobacteria in Source and Drinking Water

“…detekovali Hartmannellu vermiformis v distribučnom systéme vody, kde bola asociovaná so zvý-šenou koncentráciou organickej hmoty (7,9 ppm uhlí-ku) a aktívnej biomasy meranej ako ATP (10,6 g.l -1 ) (20). Loret a kol.…”

“…Valster a kol. (20) ukázali, že vyššia koncentrácia Hartmannella vermiformis bola získaná v letnom období, ako na jeseň, na tom istom odberovom mieste.…”

“…Dôležité je, že patogénne druhy "amébam rezistentných baktérií" používajú na napadnutie, prežitie a replikáciu v amébach veľmi podobné mechanizmy, aké využívajú pri napádaní buniek pľúcnych makrofágov člo-veka (20). Teda v prípade ich vypustenia z améby do okolia majú už gény virulencie spustené a preto sú porovnateľne virulentnejšie, než pred vstupom do améby.…”