title: Biofilm Composition and Threshold Concentration for Growth of Legionella pneumophila on Surfaces Exposed to Flowing Warm Tap Water without Disinfectant.
abstract: Legionella pneumophilain potable water installations poses a potential health risk, but quantitative information about its replication in biofilms in relation to water quality is scarce. Therefore, biofilm formation on the surfaces of glass and chlorinated polyvinyl chloride (CPVC) in contact with tap water at 34 to 39°C was investigated under controlled hydraulic conditions in a model system inoculated with biofilm-grownL. pneumophilaThe biofilm on glass (average steady-state concentration, 23 ± 9 pg ATP cm-2) exposed to treated aerobic groundwater (0.3 mg C liter-1; 1 μg assimilable organic carbon [AOC] liter-1) did not support growth of the organism, which also disappeared from the biofilm on CPVC (49 ± 9 pg ATP cm-2) after initial growth.L. pneumophilaattained a level of 4.3 log CFU cm-2in the biofilms on glass (1,055 ± 225 pg ATP cm-2) and CPVC (2,755 ± 460 pg ATP cm-2) exposed to treated anaerobic groundwater (7.9 mg C liter-1; 10 μg AOC liter-1). An elevated biofilm concentration and growth ofL. pneumophilawere also observed with tap water from the laboratory. TheBetaproteobacteria PiscinibacterandMethyloversatilisand amoeba-resistingAlphaproteobacteriapredominated in the clones and isolates retrieved from the biofilms. In the biofilms, theLegionellacolony count correlated significantly with the total cell count (TCC), heterotrophic plate count, ATP concentration, and presence ofVermamoeba vermiformisThis amoeba was rarely detected at biofilm concentrations of <100 pg ATP cm-2A threshold concentration of approximately 50 pg ATP cm-2(TCC = 1 × 106to 2 × 106cells cm-2) was derived for growth ofL. pneumophilain biofilms.IMPORTANCELegionella pneumophilais the etiologic agent in more than 10,000 cases of Legionnaires' disease that are reported annually worldwide and in most of the drinking water-associated disease outbreaks reported in the United States. The organism proliferates in biofilms on surfaces exposed to warm water in engineered freshwater installations. An investigation with a test system supplied with different types of warm drinking water without disinfectant under controlled hydraulic conditions showed that treated aerobic groundwater (0.3 mg liter-1of organic carbon) induced a low biofilm concentration that supported no or very limited growth ofL. pneumophilaElevated biofilm concentrations andL. pneumophilacolony counts were observed on surfaces exposed to two types of extensively treated groundwater, containing 1.8 and 7.9 mg C liter-1and complying with the microbial water quality criteria during distribution. Control measures in warm tap water installations are therefore essential for preventing growth ofL. pneumophila.
authors.affiliation: KWR Watercycle Research Institute, Nieuwegein, the Netherlands Dick.van.der.Kooij@kwrwater.nl.
authors.forename: Dick
authors.initials: D
authors.lastname: van der Kooij
authors.affiliation: Vitens NV, Zwolle, the Netherlands.
authors.forename: Geo L
authors.initials: GL
authors.lastname: Bakker
authors.affiliation: KWR Watercycle Research Institute, Nieuwegein, the Netherlands.
authors.forename: Ronald
authors.initials: R
authors.lastname: Italiaander
authors.affiliation: KWR Watercycle Research Institute, Nieuwegein, the Netherlands.
authors.forename: Harm R
authors.initials: HR
authors.lastname: Veenendaal
authors.affiliation: KWR Watercycle Research Institute, Nieuwegein, the Netherlands.
authors.forename: Bart A
authors.initials: BA
authors.lastname: Wullings
mesh_terms: D000656:Amoeba / Q000254:growth & development; D001419:Bacteria / Q000254:growth & development; D001422:Bacterial Adhesion; D018441:Biofilms / Q000254:growth & development*; D018533:Biomass; D015169:Colony Count, Microbial; D003300:Copper; D004202:Disinfectants; D060766:Drinking Water / Q000382:microbiology*; D005618:Fresh Water / Q000737:chemistry / Q000382:microbiology*; D005898:Glass; D006358:Hot Temperature; D016952:Legionella pneumophila / Q000254:growth & development* / Q000302:isolation & purification / Q000502:physiology; D007877:Legionnaires' Disease / Q000517:prevention & control; D009532:Nickel; D013499:Surface Properties; D014871:Water Microbiology; D014881:Water Supply
chemical_list: D004202:Disinfectants; D060766:Drinking Water; D003300:Copper; D009532:Nickel
keywords: Legionella pneumophila; predominating biofilm bacteria; threshold biofilm concentration; warm tap water