Sorgenti ambientali (aprile 2003 - aprile 2012)

Legionella pneumophila contamination in a steam towel warmer in a hospital setting

Higa F, Koide M, Haroon A, Haranaga S, Yamashiro T, Tateyama M, Fujita J.

Department of Infectious, Respiratory, and Digestive Medicine, Control and Prevention of Infectious Diseases, Faculty of Medicine, University of Ryukyus, Okinawa, Japan.

J Hosp Infect. 2012 Mar;80(3):259-61.

ABSTRACT: For prevention of nosocomial legionellosis, environmental investigation to identify possible infectious sources is essential. An environmental study in a ward of our hospital revealed that a steam towel warmer was contaminated with legionella whereas no legionella was detected in tap water supplies and shower heads. Water in the apparatus may be a reservoir of legionella. We abandoned the use of all steam towel warmers in our hospital. Based on this finding, we recommend that steam towel warmers in hospital settings be avoided. Otherwise, the apparatus should be drained, cleaned and dried every day.


Electronic-eye faucets: legionella species contamination in healthcare settings

Sydnor ER, Bova G, Gimburg A, Cosgrove SE, Perl TM, Maragakis LL.

Division of Infectious Diseases, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.

Infect Control Hosp Epidemiol. 2012 Mar;33(3):235-40.

ABSTRACT: Objective: To compare heterotrophic plate counts (HPCs) and Legionella species growth from electronic and manual faucet water samples. Design: Proportions of water samples with growth and colony-forming units were compared using Fisher's exact test and the Wilcoxon rank-sum test, respectively. Setting: Two psychiatric units and 1 medical unit in a 1,000-bed university hospital. Methods: Water samples were collected from 20 newly installed electronic faucets and 20 existing manual faucets in 3 hospital units. Manual faucets were located in rooms adjacent to the electronic faucets and received water from the same source. Water samples were collected between December 15, 2008, and January 29, 2009. Four electronic faucets were dismantled, and faucet components were cultured. Legionella species and HPC cultures were performed using standard methods. Results: Nearly all electronic faucets (19/20 [95%]) grew Legionella species from at least 1 water sample, compared with less than half (9/20 [45%]) of manual faucets ([Formula: see text]). Fifty-four (50%) of 108 electronic faucet water cultures grew Legionella species, compared with 11 (15%) of 75 manual faucet water cultures ([Formula: see text]). After chlorine dioxide remediation, 4 (14%) of 28 electronic faucet and 1 (3%) of 30 manual faucet water cultures grew Legionella species ([Formula: see text]), and 8 (29%) electronic faucet and 2 (7%) manual faucet cultures had significant HPC growth ([Formula: see text]). All 12 (100%) of the internal faucet components from 2 electronic faucets grew Legionella species. Conclusions: Electronic faucets were more commonly contaminated with Legionella species and other bacteria and were less likely to be disinfected after chlorine dioxide remediation. Electronic faucet components may provide points of concentrated bacterial growth.


Survey of legionella species found in thai soil

Travis TC, Brown EW, Peruski LF, Siludjai D, Jorakate P, Salika P, Yang G, Kozak NA, Kodani M, Warner AK, Lucas CE, Thurman KA, Winchell JM, Thamthitiwat S, Fields BS.

Respiratory Diseases Branch, Division of Bacterial Diseases, National Center for Immunization and Respiratory Diseases, Centers for Disease Control and Prevention, 1600 Clifton Road, Mailstop G03, Atlanta, GA 30333, USA.

Int J Microbiol. 2012;2012:218791.

ABSTRACT: Members of the Gram-negative genus Legionella are typically found in freshwater environments, with the exception of L. longbeachae, which is present in composts and potting mixes. When contaminated aerosols are inhaled, legionellosis may result, typically as either the more serious pneumonia Legionnaires' disease or the less severe flu-like illness Pontiac fever. It is presumed that all species of the genus Legionella are capable of causing disease in humans. As a followup to a prior clinical study of legionellosis in rural Thailand, indigenous soil samples were collected proximal to cases' homes and workplaces and tested for the presence of legionellae by culture. We obtained 115 isolates from 22/39 soil samples and used sequence-based methods to identify 12 known species of Legionella represented by 87 isolates.


Influence of temperature, chlorine residual and heavy metals on the presence of Legionella pneumophila in hot water distribution systems

Rakić A, Perić J, Foglar L.

Public Health Institute, Split, Croatia.

Ann Agric Environ Med. 2012;19(3):431-6.

ABSTRACT: The microbiological colonisation of buildings and man-made structures often occurs on the walls of plumbing systems; therefore, monitoring of opportunistic pathogens such as Legionellapneumophila (L. pneumophila), both in water distribution mains and in consumers' plumbing systems, is an important issue according to the international and national guidelines that regulate the quality of drinking water. This paper investigates the presence of L. pneumophila in the Dalmatian County of Croatia and the relationship between L. pneumophila presence and heavy metals concentrations, free residual chlorine and water temperature in hot water distribution systems (WDS). Investigations were performed on a large number of hot water samples taken from taps in kitchens and bathrooms in hotels and homes for the elderly and disabled in the Split region. Of the 127 hot water samples examined, 12 (9.4%) were positive for Legionella spp. with median values concentration of 450 cfu × L(-1). Among positive isolates, 10 (83.3%) were L. pneumophila sg 1, and two of them (16.6%) belonged to the genera L. pneumophila sg 2-14. The positive correlation between the water temperature, iron and manganese concentrations, and L. pneumophila contamination was proved by statistical analysis of the experimental data. On the contrary, zinc and free residual chlorine had no observed influence on the presence of L. pneumophila. The presence of heavy metals in water samples confirms the corrosion of distribution system pipes and fittings, and suggests that metal plumbing components and associated corrosion products are important factors in the survival and growth of L. pneumophila in WDS.


The first case of Legionella nagasakiensis isolation from hot spring water

Furuhata K, Edagawa A, Miyamoto H, Goto K, Yoshida S, Fukuyama M.

School of Life and Environmental Science, Azabu University, Sagamihara, Kanagawa 252-5201, Japan.

Biocontrol Sci. 2011 Dec;16(4):171-6.

ABSTRACT: In August, 2010, strain HYMO-6 was isolated from a sample of hot spring water in Aomori, Japan. The 16S rDNA sequences (1,496bp) of this strain (accession number: AB597175) had a similarity of less than 96.6% to other Legionella species, prompting us to hypothesize that this strain might be a novel species belonging to the genus Legionella. However, in March of 2011, it was became clear that the HYMO-6 strain (=JCM 17450 =KCTC 23560 =DSM 24727) was Legionella nagasakiensis CDC-1796-JAP-E(T) (=ATCC BAA-1557(T) =JCM 15315(T)). When this strain was cultured on BCYEα agar at 36°C for 7 d, no long cells were observed. The dominant fatty acids of strain HYMO-6 were 16:1ω7c (32.4%), and the DNA G+C content was 42.0 mol%.


Co-existence of Legionella and other Gram-negative bacteria in potable water from various rural and urban sources

Stojek NM, Dutkiewicz J.

Department of Water and Soil Safety, Institute of Rural Health, Lublin, Poland.

Ann Agric Environ Med. 2011 Dec;18(2):330-4.

ABSTRACT: A total of 320 potable water samples were collected from various rural and urban sources located in the Lublin region of eastern Poland. They comprised: 55 samples of treated (chlorinated) tap water from rural dwellings distributed by the municipal water supply system (MWSS), 111 samples of treated tap water from urban dwellings distributed by the MWSS, 45 samples of untreated well water from household wells and 109 samples from private water supply systems (PWSS) distributing untreated well water. Water samples were examined for the presence and species composition of Legionella, Yersinia, Gram-negative bacteria belonging to family Enterobacteriaceae (GNB-E) and Gram-negative bacteria not belonging to the family Enterobacteriaceae (GNB-NE), by filtering through cellulose filters and culture on respectively GVPC, CIN, EMB and tryptic soya agar media. The occurrence of Legionella in the samples taken from the outlets of the urban MWSS was high (77.5%), and significantly greater compared to frequencies noted in rural MWSS (7.3%), and samples of well water from household wells (28.9%) and PWSS (13.8%) (p<0.001). Strains L. pneumophila serogroups 2-14, L. pneumophila serogroup 1 and Legionella spp. (species other than L. pneumophila) formed respectively 64.3%, 17.5%, and 18.2% of total isolates from urban MWSS, 100%, 0, and 0 of those from rural MWSS, 69.2%, 7.7%, and 23.1% of those from household wells, and 66.7%, 0, and 33.3% of those from PWSS. The concentration of Legionella strains in the positive samples from urban MWSS exceeded the threshold limit value of 100 cfu/100 ml in 86.1%, while in the other sources this value was not exceeded. No Yersinia strains were isolated from the examined water samples. Altogether 8 species or genera of Gram-negative bacteria belonging to Enterobacteriaceae family (GNB-E) and 10 species or genera of Gram-negative bacteria not belonging to the Enterobacteriaceae family (GNB-NE) were found in the examined samples. In the MWSS samples, an inverse relationship was found between Legionella and GNB-E and the numbers of Enterobacter spp. and Serratia spp. strains were significantly more common in the samples without Legionella. By contrast, in the PWSS samples, the numbers of Enterobacter spp., Klebsiella spp. and Salmonella spp. were distinctly and significantly greater (p<0.01-p<0.001) in the samples containing Legionella. Among GNB-NE, Pseudomonas aeruginosa strains occurred significantly more frequently in samples containing Legionella (for MWSS and well water separately p<0.05, for total samples p<0.001). Similarly, strains of Flavobacterium breve and Xanthomonas spp. occurred significantly more often in the samples with Legionella, while the numbers of Aeromonas spp. and Vibrio spp. strains were significantly greater in the samples not containing Legionella. In conclusion, a health risk could be associated with exposure to the water from urban MWSS because of the high prevalence and concentration of Legionella, and with exposure to well water from PWSS because of the correlation of occurrence of Legionella and potentially pathogenic Enterobacteriaceae strains, and the possibility of synergistic eff ects. The adverse eff ects could be also due to the significant correlation of Legionella and Pseudomonas aeruginosa that occured in water from various sources.


Biofilms in drinking water and their role as reservoir for pathogens

Wingender J, Flemming HC.

Biofilm Centre, University of Duisburg-Essen, Universitätsstraße 5, D-45141 Essen, Germany.

Int J Hyg Environ Health. 2011 Nov;214(6):417-23.

ABSTRACT: Most microorganisms on Earth live in various aggregates which are generally termed "biofilms". They are ubiquitous and represent the most successful form of life. They are the active agent in biofiltration and the carriers of the self-cleaning potential in soils, sediments and water. They are also common on surfaces in technical systems where they sometimes cause biofouling. In recent years it has become evident that biofilms in drinking water distribution networks can become transient or long-term habitats for hygienically relevant microorganisms. Important categories of these organisms include faecal indicator bacteria (e.g., Escherichia coli), obligate bacterial pathogens of faecal origin (e.g., Campylobacter spp.) opportunistic bacteria of environmental origin (e.g., Legionella spp., Pseudomonas aeruginosa), enteric viruses (e.g., adenoviruses, rotaviruses, noroviruses) and parasitic protozoa (e.g., Cryptosporidium parvum). These organisms can attach to preexisting biofilms, where they become integrated and survive for days to weeks or even longer, depending on the biology and ecology of the organism and the environmental conditions. There are indications that at least a part of the biofilm populations of pathogenic bacteria persists in a viable but non-culturable (VBNC) state and remains unnoticed by the methods appointed to their detection. Thus, biofilms in drinking water systems can serve as an environmental reservoir for pathogenic microorganisms and represent a potential source of water contamination, resulting in a potential health risk for humans if left unnoticed.


An in-premise model for Legionella exposure during showering events

Schoen ME, Ashbolt NJ.

Office of Research and Development, U.S. Environmental Protection Agency, 26 West Martin Luther King Drive, Cincinnati, OH 45268, USA.

Water Res. 2011 Nov 15;45(18):5826-36

ABSTRACT:An exposure model was constructed to predict the critical Legionella densities in an engineered water system that result in infection from inhalation of aerosols containing the pathogen while showering. The model predicted the Legionella densities in the shower air, water and in-premise plumbing biofilm that might result in a deposited dose of Legionella in the alveolar region of the lungs associated with infection for a routine showering event. Processes modeled included the detachment of biofilm-associated Legionella from the in-premise plumbing biofilm during a showering event, the partitioning of the pathogen from the shower water to the air, and the inhalation and deposition of particles in the lungs. The range of predicted critical Legionella densities in the air and water was compared to the available literature. The predictions were generally within the limited set of observations for air and water, with the exception of Legionella density within in-premise plumbing biofilms, for which there remains a lack of observations for comparison. Sensitivity analysis of the predicted results to possible changes in the uncertain input parameters identified the target deposited dose associated with infections, the pathogen air-water partitioning coefficient, and the quantity of detached biofilm from in-premise pluming surfaces as important parameters for additional data collection. In addition, the critical density of free-living protozoan hosts in the biofilm required to propagate the infectious Legionella was estimated. Together, this evidence can help to identify critical conditions that might lead to infection derived from pathogens within the biofilms of any plumbing system from which humans may be exposed to aerosols.


Enhanced isolation of Legionella species from composted material

McCabe S, Brown A, Edwards GF, Lindsay D.

Scottish Haemophilus Legionella Meningococcus Pneumococcus Reference Laboratory (SHLMPRL), House on the Hill, Stobhill Hospital, Glasgow, UK.

Clin Microbiol Infect. 2011 Oct;17(10):1517-20.

ABSTRACT:Legionella pneumophila and Legionella species were isolated from composted material when freshly prepared buffered charcoal yeast extract (BCYE) was supplemented with glycine (1.5 g/L), polymyxin B sulfate (40 000 IU/L), vancomycin hydrochloride (0.5 mg/L) and cycloheximide (40 mg/L) (GVPC medium) and Modified Wadowsky-Yee (MWY) (Oxoid, Cambridge, UK) plates were used for cultivation, but not with commercially sourced pre-poured GVPC and MWY plates (Oxoid). Legionella cincinnatiensis and pathogenic L. pneumophila serogroup (Sg) 1 Benidorm and France/Allentown were identified, as well as a non-typeable (NT) strain of L. pneumophila. As most laboratories no longer produce their own media, this may contribute to the lack of positive cultures from composted material. The antigenicity of the NT strain is discussed.


Comparison of potentially pathogenic free-living amoeba hosts by Legionella spp. in substrate-associated biofilms and floating biofilms from spring environments

Hsu BM, Huang CC, Chen JS, Chen NH, Huang JT.

Department of Earth and Environmental Sciences, National Chung Cheng University, 168 University Road, Minhsiung Township, Chiayi County 62102, Taiwan, ROC.

Water Res. 2011 Oct 15;45(16):5171-83.

ABSTRACT: This study compares five genera of free-living amoebae (FLA) hosts by Legionella spp. in the fixed and floating biofilm samples from spring environments. Detection rate of Legionella spp. was 26.9% for the floating biofilms and 3.1% for the fixed biofilms. Acanthamoeba spp., Hartmanella vermiformis, and Naegleria spp. were more frequently detected in floating biofilm than in fixed biofilm samples. The percentage of pathogenic Acanthamoeba spp. among all the genus Acanthamoeba detected positive samples was 19.6%. The potential pathogenic Naegleria spp. (for example, Naegleria australiensis, Naegleria philippinensis, and Naegleria italica) was 54.2% to all the Naegleria detected positive samples. In the study, 12 serotypes of possible pneumonia causing Legionella spp. were detected, and their percentage in all the Legionella containing samples was 42.4%. The FLA parasitized by Legionella included unnamed Acanthamoeba genotype, Acanthamoeba griffini, Acanthamoeba jacobsi, H. vermiformis, and N. australiensis. Significant differences were also observed between the presence/absence of H. vermiformis and Legionella parasitism in FLA. Comparisons between the culture-confirmed method and the PCR-based detection method for detecting FLA and Legionella in biofilms showed great variation. Therefore, using these analysis methods together to detect FLA and Legionella is recommended.


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

Valster RM, Wullings BA, van den Berg R, van der Kooij D.

KWR Watercycle Research Institute, Groningenhaven 7, P.O. Box 1072, 3430 BB Nieuwegein, the Netherlands.

Appl Environ Microbiol. 2011 Oct;77(20):7321-8.

ABSTRACT: The study whose results are presented here aimed at identifying free-living protozoa (FLP) and conditions favoring the growth of these organisms and cultivable Legionella spp. in drinking water supplies in a tropical region. Treated and distributed water (±30°C) of the water supplies of three Caribbean islands were sampled and investigated with molecular techniques, based on the 18S rRNA gene. The protozoan host Hartmannella vermiformis and cultivable Legionella pneumophila were observed in all three supplies. Operational taxonomic units (OTUs) with the highest similarity to the potential or candidate hosts Acanthamoeba spp., Echinamoeba exundans, E. thermarum, and an Neoparamoeba sp. were detected as well. In total, 59 OTUs of FLP were identified. The estimated protozoan richness did not differ significantly between the three supplies. In supply CA-1, the concentration of H. vermiformis correlated with the concentration of Legionella spp. and clones related to Amoebozoa predominated (82%) in the protozoan community. These observations, the low turbidity (<0.2 nephelometric turbidity units [NTU]), and the varying ATP concentrations (1 to 12 ng liter(-1)) suggest that biofilms promoted protozoan growth in this supply. Ciliophora represented 25% of the protozoan OTUs in supply CA-2 with elevated ATP concentrations (maximum, 55 ng liter(-1)) correlating with turbidity (maximum, 62 NTU) caused by corroding iron pipes. Cercozoan types represented 70% of the protozoan clones in supply CA-3 with ATP concentrations of <1 ng liter(-1) and turbidity of <0.5 NTU in most samples of distributed water. The absence of H. vermiformis in most samples from supply CA-3 suggests that growth of this protozoan is limited at ATP concentrations of <1 ng liter(-1).


Differential growth of Legionella pneumophila strains within a range of amoebae at various temperatures associated with in-premise plumbing

Buse HY, Ashbolt NJ.

National Exposure Research Laboratory, Office of Research and Development, US Environmental Protection Agency, Cincinnati, OH 45268, USA.

Lett Appl Microbiol. 2011 Aug;53(2):217-24.

ABSTRACT: AIMS: The potential effect of in-premise plumbing temperatures (24, 32, 37 and 41°C) on the growth of five different Legionella pneumophila strains within free-living amoebae (Acanthamoeba polyphaga, Hartmannella vermiformis and Naegleria fowleri) was examined.

METHODS AND RESULTS: Compared with controls that actively fed on Escherichia coli prey, when Leg. pneumophila was used as prey, strains Lp02 and Bloomington-2 increased in growth at 30, 32 and 37°C while strains Philadelphia-1 and Chicago 2 did not grow at any temperature within A. polyphaga. Strains Lp02, Bloomington-2 and Dallas 1E did not proliferate in the presence of H. vermiformis nor did strain Philadelphia-1 in the presence of N. fowleri. Yet, strain Bloomington-2 grew at all temperatures examined within N. fowleri, while strain Lp02 proliferated at all temperatures except 41°C. More intriguing, strain Chicago 2 only grew at 32°C within H. vermiformis and N. fowleri suggesting a limited temperature growth range for this strain.

CONCLUSIONS: Identifying the presence of pathogenic legionellae may require the use of multiple host amoebae and incubation temperatures.

SIGNIFICANCE AND IMPACT OF THE STUDY: Temperature conditions and species of amoeba host supported in drinking water appear to be important for the selection of human-pathogenic legionellae and point to future research required to better understand Legionella ecology.


Dispersion of Legionella-containing aerosols from a biological treatment plant, Norway

Blatny JM, Fossum H, Ho J, Tutkun M, Skogan G, Andreassen O, Fykse EM, Waagen V, Reif BA.

Norwegian Defence Research Establishment (FFI), Kjeller, Norway.

Front Biosci (Elite Ed). 2011 Jun 1;3:1300-9.

ABSTRACT: Legionella was detected in aeration ponds (biological treatment plant) at Borregaard Ind. Ltd., Norway, and in air samples harvested directly above these ponds. Since 2005, three outbreaks of legionellosis occurred within a 10 km radius from this plant. This work addresses the dispersion patterns of Legionella-containing particles by characterizing the aerosol plume emitted from these ponds (outbreak source) less than 500 meters using wind-tunnel measurements, CFD simulations, and real-life measurements. The most abundant particles directly over the ponds were less than 6 and more than 15 microm. The results showed that the aerosol plume remained narrow; 180 meters wide at 350 meters downwind of the ponds, and that 2 and 18 microm aerosols were mainly deposited in the vicinity of the ponds ( 150 - 200 meters). Furthermore, the maximum aerosol concentration level appeared 5-10 meters above ground level and the maximum concentration 500 meters downwind was approximately 2 per cent of the concentration level directly above the ponds. Our study demonstrates the strength of combining modeling with real-life aerosol analyses increasing the understanding of dispersion of airborne (pathogenic) microorganisms.


Contamination of the cold water distribution system of health care facilities by Legionella pneumophila: do we know the true dimension?

Arvand M, Jungkind K, Hack A.

Hesse State Health Office, Centre for Health Protection, Dillenburg, Germany.

Euro Surveill. 2011 Apr 21;16(16). pii: 19844.

ABSTRACT: German water guidelines do not recommend routine assessment of cold water for Legionella in healthcare facilities, except if the water temperature at distal sites exceeds 25°C. This study evaluates Legionella contamination in cold and warm water supplies of healthcare facilities in Hesse, Germany, and analyses the relationship between cold water temperature and Legionella contamination. Samples were collected from four facilities, with cases of healthcare-associated Legionnaires' disease or notable contamination of their water supply. Fifty-nine samples were from central lines and 625 from distal sites, comprising 316 cold and 309 warm water samples. Legionella was isolated from central lines in two facilities and from distal sites in four facilities. 17% of all central and 32% of all distal samples were contaminated. At distal sites, cold water samples were more frequently contaminated with Legionella (40% vs 23%, p <0.001) and with higher concentrations of Legionella (≥1,000 colony-forming unit/100 ml) (16% vs 6%, p<0.001) than warm water samples. There was no clear correlation between the cold water temperature at sampling time and the contamination rate. 35% of cold water samples under 20 °C at collection were contaminated. Our data highlight the importance of assessing the cold water supply of healthcare facilities for Legionella in the context of an intensified analysis.


Concentration and Diversity of Uncultured Legionella spp. in Two Unchlorinated Drinking Water Supplies with Different Concentrations of Natural Organic Matter

Wullings BA, Bakker G, van der Kooij D.

KWR Watercycle Research Institute, Groningenhaven 7, P.O. Box 1072, 3430 BB Nieuwegein, Netherlands.

Appl Environ Microbiol. 2011 Jan;77(2):634-41.

ABSTRACT: Two unchlorinated drinking water supplies were investigated to assess the potential of water treatment and distribution systems to support the growth of Legionella spp. The treatment plant for supply A distributed treated groundwater with a low concentration (<0.5 ppm of C) of natural organic matter (NOM), and the treatment plant for supply B distributed treated groundwater with a high NOM concentration (8 ppm of C). In both supplies, the water temperature ranged from about 10°C after treatment to 18°C during distribution. The concentrations of Legionella spp. in distributed water, analyzed with quantitative PCR (Q-PCR), averaged 2.9 (± 1.9) × 10(2) cells liter(-1) in supply A and 2.5 (± 1.6) × 10(3) cells liter(-1) in supply B. No Legionella was observed with the culture method. A total of 346 clones (96 operational taxonomical units [OTUs] with ≥97% sequence similarity) were retrieved from water and biofilms of supply A and 251 (43 OTUs) from supply B. The estimation of the average value of total species richness (Chao1) in supply A (153) was clearly higher than that for supply B (58). In each supply, about 77% of the sequences showed <97% similarity to described species. Sequences related to L. pneumophila were only incidentally observed. The Legionella populations of the two supplies are divided into two distinct clusters based on distances in the phylogenetic tree as fractions of the branch length. Thus, a large variety of mostly yet-undescribed Legionella spp. proliferates in unchlorinated water supplies at temperatures below 18°C. The lowest concentration and greatest diversity were observed in the supply with the low NOM concentration.


High diversity and abundance of Legionella spp. in a pristine river and impact of seasonal and anthropogenic effects

Parthuisot N, West NJ, Lebaron P, Baudart J.

UPMC Univ. Paris 06, UMR 7621, LOMIC, Observatoire Océanologique, F-66650 Banyuls-sur-mer, France.

Appl Environ Microbiol. 2010 Dec;76(24):8201-10.

ABSTRACT: The diversity and dynamics of Legionella species along a French river watershed subject to different thermal and wastewater discharges during an annual cycle were assessed by 16S rRNA gene sequencing and by a fingerprint technique, single-strand conformation polymorphism. A high diversity of Legionella spp. was observed at all the sampling sites, and the dominant Legionella clusters identified were most closely related to uncultured bacteria. The monthly monitoring revealed that Legionella sp. diversity changes were linked only to season at the wastewater site whereas there was some evidence for anthropogenic effects on Legionella sp. diversity downstream of the thermal bath. Quantification of Legionella pneumophila and Legionella spp. by culture and quantitative PCR (qPCR) was performed. Whereas only L. pneumophila was quantified on culture media, the qPCR assay revealed that Legionella spp. were ubiquitous and abundant from the pristine source of the river to the downstream sampling sites. These results suggest that Legionella spp. may be present at significant concentrations in many more freshwater environments than previously thought, highlighting the need for further ecological studies and culturing efforts.


Alternative routes for dissemination of Legionella pneumophila causing three outbreaks in Norway

Olsen JS, Aarskaug T, Thrane I, Pourcel C, Ask E, Johansen G, Waagen V, Blatny JM.

Norwegian Defence Research Establishment (FFI), Norway.

Environ Sci Technol. 2010 Nov 15;44(22):8712-7.

ABSTRACT: Three outbreaks of Legionnaires' disease were reported in the Fredrikstad/Sarpsborg community, Norway, in 2005 and 2008 caused by the L. pneumophila ST15 and ST462 strains determined by sequence based typing. In this retrospective study, we suggest that the aeration ponds, a part of the biological treatment plant at Borregaard Ind. Ltd., are the main amplifiers and primary disseminators of the outbreak L. pneumophila strains. This result is supported by the finding that the ST15 and ST462 strains were not able to survive in air scrubber liquid media more than two days of incubation at the scrubber's operating conditions during the 2005 and 2008 outbreaks. In 2008, >10¹ CFU/L of L. pneumophila ST462 were detected in the aeration ponds. ST15 and ST462 were also detected in the river Glomma in 2005 and 2008, respectively, downstream of the wastewater outlet from the treatment plant (10CFU/L). These findings strongly suggest that the presence of L. pneumophila in the river is due to the release of wastewater from the industrial aeration ponds, demonstrating that the river Glomma may be an additional disseminator of L. pneumophila during the outbreaks. This work emphasizes the need for preventive actions against the release of wastewater containing human pathogens to the environment.


Health risk from the use of roof-harvested rainwater in Southeast Queensland, Australia, as potable or nonpotable water, determined using quantitative microbial risk assessment

Ahmed W, Vieritz A, Goonetilleke A, Gardner T.

Department of Environment and Resource Management, Indooroopilly, Brisbane 4068, Queensland, Australia.

Appl Environ Microbiol. 2010 Nov;76(22):7382-91.

ABSTRACT: A total of 214 rainwater samples from 82 tanks were collected in urban Southeast Queensland (SEQ) in Australia and analyzed for the presence and numbers of zoonotic bacterial and protozoal pathogens using binary PCR and quantitative PCR (qPCR). Quantitative microbial risk assessment (QMRA) analysis was used to quantify the risk of infection associated with the exposure to potential pathogens from roof-harvested rainwater used as potable or nonpotable water. Of the 214 samples tested, 10.7%, 9.8%, 5.6%, and 0.4% were positive for the Salmonella invA, Giardia lamblia β-giardin, Legionella pneumophila mip, and Campylobacter jejuni mapA genes, respectively. Cryptosporidium parvum oocyst wall protein (COWP) could not be detected. The estimated numbers of Salmonella, G. lamblia, and L. pneumophila organisms ranged from 6.5 × 10¹ to 3.8 × 10² cells, 0.6 × 10 to 3.6 × 10 cysts, and 6.0 × 10¹ to 1.7 × 10² cells per 1,000 ml of water, respectively. Six risk scenarios were considered for exposure to Salmonella spp., G. lamblia, and L. pneumophila. For Salmonella spp. and G. lamblia, these scenarios were (i) liquid ingestion due to drinking of rainwater on a daily basis, (ii) accidental liquid ingestion due to hosing twice a week, (iii) aerosol ingestion due to showering on a daily basis, and (iv) aerosol ingestion due to hosing twice a week. For L. pneumophila, these scenarios were (i) aerosol inhalation due to showering on a daily basis and (ii) aerosol inhalation due to hosing twice a week. The risk of infection from Salmonella spp., G. lamblia, and L. pneumophila associated with the use of rainwater for showering and garden hosing was calculated to be well below the threshold value of one extra infection per 10,000 persons per year in urban SEQ. However, the risk of infection from ingesting Salmonella spp. and G. lamblia via drinking exceeded this threshold value and indicated that if undisinfected rainwater is ingested by drinking, then the incidences of the gastrointestinal diseases salmonellosis and giardiasis are expected to range from 9.8 × 10° to 5.4 × 10¹ (with a mean of 1.2 × 10¹ from Monte Carlo analysis) and from 1.0 × 10¹ to 6.5 × 10¹ cases (with a mean of 1.6 × 10¹ from Monte Carlo analysis) per 10,000 persons per year, respectively, in urban SEQ. Since this health risk seems higher than that expected from the reported incidences of gastroenteritis, the assumptions used to estimate these infection risks are critically examined. Nonetheless, it would seem prudent to disinfect rainwater for use as potable water.


Legionellosis in a site with low risk of proliferation

Pañella H, Calzada N, Beneyto V, Valero N, Gracia J, Rodríguez P.

Agència de Salut Pública de Barcelona, Direcció de Serveis de Vigilància Ambiental, Barcelona, España.

Gac Sanit. 2010 Nov-Dec;24(6):498-500.

ABSTRACT: After three cases of legionellosis associated with a spa were identified, an investigation was conducted to confirm the source of infection, determine the risk factors, and establish control measures. Between November 26, 2008 and September 16, 2009, six inspections were carried out, samples were collected for Legionella pneumophila isolation, and water temperature was determined at all the sampling points of the hot water system and at the spa pools. The presence of L. pneumophila serogroup 1 with the same molecular pattern as that found in the clinical isolate was confirmed in the hot water system (with a low proliferation risk according to legislation). The crude attack rate was 0.34% (95% CI: 0.09-0.94). Samples without L. pneumophila were obtained only after structural changes were carried out. Possible factors participating in the cluster were water temperatures between 25 °C and 30 °C, stagnant water and early morning exposure.


Legionella dresdenensis sp. nov., isolated from river water

Lück PC, Jacobs E, Röske I, Schröter-Bobsin U, Dumke R, Gronow S.

Institut für Medizinische Mikrobiologie und Hygiene, TU Dresden, Dresden, Germany.

Int J Syst Evol Microbiol. 2010 Nov;60(Pt 11):2557-62.

ABSTRACT: Legionella-like isolates, strains W03-356(T), W03-357 and W03-359, from three independent water samples from the river Elbe, Germany, were analysed by using a polyphasic approach. Morphological and biochemical characterization revealed that they were Gram-negative, aerobic, non-spore-forming bacilli with a cut glass colony appearance that grew only on L-cysteine-supplemented buffered charcoal yeast extract agar. Phylogenetic analysis based on sequence comparisons of the 16S rRNA, macrophage infectivity potentiator (mip), gyrase subunit A (gyrA), ribosomal polymerase B (rpoB) and RNase P (rnpB) genes confirmed that the three isolates were distinct from recognized species of the genus Legionella. Phenotypic characterization of strain W03-356(T) based on fatty acid profiles confirmed that it was closely related to Legionella rubrilucens ATCC 35304(T) and Legionella pneumophila ATCC 33152(T), but distinct from other species of the genus Legionella. Serotyping of the isolates showed that they were distinct from all recognized species of the genus Legionella. Strains W03-356(T), W03-357 and W03-359 are thus considered to represent a novel species of the genus Legionella, for which the name Legionella dresdenensis sp. nov. is proposed. The type strain is W03-356(T) (=DSM 19488(T)=NCTC 13409(T)).


Isolation and identification of Legionella pneumophila from material reclamation facilities

Ali S, Phillips CA, Phillips PS, Bates M.

School of Health, University of Northampton, Northampton, UK.

Int J Environ Health Res. 2010 Oct;20(5):367-77.

ABSTRACT: Sampling points at a material reclamation facility (MRF) were monitored over three months for the presence of Legionella spp. A number of different Legionellae were isolated and typed to identify L. pneumophila serogroup 1, the serotype which is the most common human pathogen. Phenotypic methods resulted in a total of 61 presumptive isolates of Legionella spp. Using latex agglutination, 26 out of the 61 were identified as L. pneumophila serogroup 1, 23 as L. pneumophila serogroups 2-14, and the remaining 12 were Legionella spp. However, on typing using pulse field gel electrophoresis, the 26 L. pneumophila serotype 1 isolates were a diverse group of 25 PFGE types with none persisting in the environment over time. This diversity suggests that there are a number of contamination sources for this important human pathogen in the MRF environment which constitute a risk to health for operatives in these facilities.


Italian multicentre study on microbial environmental contamination in dental clinics: a pilot study

Pasquarella C, Veronesi L, Castiglia P, Liguori G, Montagna MT, Napoli C, Rizzetto R, Torre I, Masia MD, Di Onofrio V, Colucci ME, Tinteri C, Tanzi M; SItI working group "Hygiene in Dentistry.

Dipartimento di Sanità Pubblica, Università degli Studi di Parma, Italy.

Sci Total Environ. 2010 Sep 1;408(19):4045-51.

ABSTRACT: The dental practice is associated with a high risk of infections, both for patients and healthcare operators, and the environment may play an important role in the transmission of infectious diseases. A microbiological environmental investigation was carried out in six dental clinics as a pilot study for a larger multicentre study that will be performed by the Italian SItI (Society of Hygiene, Preventive Medicine and Public Health) working group "Hygiene in Dentistry". Microbial contamination of water, air and surfaces was assessed in each clinic during the five working days of the week, before and during treatments. Air and surfaces were also examined at the end of the daily activity. A wide variation was found in microbial environmental contamination, both within the participating clinics and relative to the different sampling times. Microbial water contamination in Dental Unit Water Systems (DUWS) reached values of up to 26x10(4)cfu/mL (colony forming units per millilitre). P. aeruginosa was found in 33% of the sampled DUWS and Legionella spp. in 50%. A significant decrease in the Total Viable Count (TVC) was recorded during the activity. Microbial air contamination showed the highest levels during dental treatments and tended to decrease at the end of the working activity (p<0.05). Microbial buildup on surfaces increased significantly during the working hours. As these findings point out, research on microbial environmental contamination and the related risk factors in dental clinics should be expanded and should also be based on larger collections of data, in order to provide the essential knowledge aimed at targeted preventive interventions.


Regrowth of potential opportunistic pathogens and algae in reclaimed-water distribution systems

Jjemba PK, Weinrich LA, Cheng W, Giraldo E, Lechevallier MW.

American Water Research Laboratory, 213 Carriage Lane, Delran, NJ 08075, USA.

Appl Environ Microbiol. 2010 Jul;76(13):4169-78.

ABSTRACT: A study of the quality of reclaimed water in treated effluent, after storage, and at three points in the distribution system of four plants in California, Florida, Massachusetts, and New York was conducted for 1 year. The plants had different treatment processes (conventional versus membrane bioreactor), production capacities, and methods for storage of the water, and the intended end uses of the water were different. The analysis focused on the occurrence of indicator bacteria (heterotrophic bacteria, coliforms, Escherichia coli, and enterococci) and opportunistic pathogens (Aeromonas spp., enteropathogenic E. coli O157:H7, Legionella spp., Mycobacterium spp., and Pseudomonas spp.), as well as algae. Using immunological methods, E. coli O157:H7 was detected in the effluent of only one system, but it was not detected at the sampling points, suggesting that its survival in the system was poor. Although all of the treatment systems effectively reduced the levels of bacteria in the effluent, bacteria regrew in the reservoir and distribution systems because of the loss of residual disinfectant and high assimilable organic carbon levels. In the systems with open reservoirs, algal growth reduced the water quality by increasing the turbidity and accumulating at the end of the distribution system. Opportunistic pathogens, notably Aeromonas, Legionella, Mycobacterium, and Pseudomonas, occurred more frequently than indicator bacteria (enterococci, coliforms, and E. coli). The Mycobacterium spp. were very diverse and occurred most frequently in membrane bioreactor systems, and Mycobacterium cookii was identified more often than the other species. The public health risk associated with these opportunistic pathogens in reclaimed water is unknown. Collectively, our results show the need to develop best management practices for reclaimed water to control bacterial regrowth and degradation of water before it is utilized at the point of use.


First isolation of Legionella species, including L. pneumophila serogroup 1, in Greek potting soils: possible importance for public health

Velonakis EN, Kiousi IM, Koutis C, Papadogiannakis E, Babatsikou F, Vatopoulos A.

Department of Microbiology, National School of Public Health and Central Public Health Laboratory, Athens/Hellenic Centre for Infectious Disease Control (KEELPNO), Ministry of Health, Athens, Greece.

Clin Microbiol Infect. 2010 Jun;16(6):763-6.

ABSTRACT: A total of 21 Legionella isolates were recovered from six out of 22 samples of potting soil from the Athens area, Greece. Legionella pneumophila (serogroups 1 and 2-15) and species and serotypes included in the group of L. longbeachae serogroups 1 and 2, L. bozemanii serogroups 1 and 2, L. dumoffii, L. gormanii, L. jordanis, L. micdadei and L. anisa were isolated on BCYEalpha agar containing cysteine, GVPC and natamycin and on BCYEalpha agar containing cysteine, Wadowsky Yee supplement and natamycin. The bacterial load was 4000-120 000 CFU/g of potting soil. The isolation of L. pneumophila serogroup 1 from Greek potting soils is reported here for the first time.


Is domestic tap water a risk for infections in neutropenic patients?

von Baum H, Bommer M, Forke A, Holz J, Frenz P, Wellinghausen N.

Section Infection Control and Hospital Epidemiology, Institute of Medical Microbiology and Hygiene, University Hospital of Ulm, Steinhoevelstr. 9, 89075, Ulm, Germany.

Infection. 2010 Jun;38(3):181-6.

ABSTRACT: BACKGROUND: Home care has become popular in the management of hemato-oncologic patients. Therefore, we conducted a prospective study to assess whether tap water from the domestic environment of neutropenic patients poses a risk for infections from the waterborne pathogens nontuberculous mycobacteria (NTM), Legionella spp., and Pseudomonas aeruginosa. MATERIALS AND METHODS: Tap water samples were taken in the homes of 65 hemato-oncologic patients who were discharged from the hospital whilst neutropenic and had a suspected period of neutropenia of a minimum of 10 days. Selective culture for Legionella, P. aeruginosa, and NTM was performed. Patients who required hospital readmission were monitored for infection with the aforementioned pathogens over the following 3 months. RESULTS: NTM were cultured in 62 (95.4%) households in concentrations from 1 to 1,000 CFU/500 ml. The facultative pathogenic species Mycobacterium chelonae (58.5% of taps) and M. mucogenicum (38.5% of taps) were most frequently detected. Legionella spp. was cultured from six households (9.2%), including five households with L. pneumophila in concentrations from 25 to 2,500 CFU/500 ml. P. aeruginosa was found in seven households (10.8%) in concentrations from 5 to 2,500 CFU/500 ml. While clinical infection with Legionella spp. was not detected in any patients, infection with M. chelonae and P. aeruginosa occurred in one and seven patients, respectively. However, transmission from household water could not be confirmed. CONCLUSION: Although the risk of infection from household water-borne pathogens appears low, preventive measures may be considered on an individual basis in patients with long-term immunosuppression as well as in patients with long-term central-vascular catheterization.


Drinking water quality in household supply infrastructure--A survey of the current situation in Germany

Völker S, Schreiber C, Kistemann T.

Institute for Hygiene and Public Health, University of Bonn, Sigmund-Freud-Str. 25, 53105 Bonn, Germany.

Int J Hyg Environ Health. 2010 Jun;213(3):204-9.

ABSTRACT: As a result of the amendment to the German Drinking Water Ordinance in 2001, local public health authorities are obliged to monitor the water supply in installations providing water for public use (Section 18 German Drinking Water Ordinance). With a systematic and nationwide survey of locally available data relating to hygienic drinking water quality and the existing drinking water infrastructure in buildings, the extent of microbial contamination of in-building distribution systems in Germany is intended to be assessed. To gain an overview of the microbial contamination of drinking water in public buildings all 419 local public health authorities in Germany were contacted in 2007. In a detailed study with a representative cooperation level of 5% of these local public health authorities, the available data relating to microbiological, chemical, physical and technical parameters gained from in-building distribution systems were collected. Drinking water parameters were combined with regard to the total number of analyses and the absolute number as well as the percentage of limit compliance failures (n=108,288). Limits exceeded were classified as the failure to comply with the German Drinking Water Ordinance, DVGW technical regulations and Federal Environment Agency recommended limits. The highest rates of samples exceeding these limits were found for the parameter Legionella sp. which contaminated 12.8% of all samples (n=22,786; limit: 100 CFU/100ml), followed by heterotrophic plate count at 36 degrees C (3.5%, n=10,928; limit: 100 CFU/1 ml) and Pseudomonas sp. (2.9%, n=3468; limit: 0 CFU/100ml). Legionella sp. and Pseudomonas sp. pose a direct health risk to immunosuppressed users. Additionally, for some chemical parameters, such as nickel, iron and lead, a potential risk for the health of consumers was detected. Further data analysis may reveal whether this contamination is related to stagnation where there is only sporadic use or whether other factors are involved in the process of microbial growth in installation systems. Copyright 2010 Elsevier GmbH. All rights reserved.


Water birthing: retrospective review of 2625 water births. Contamination of birth pool water and risk of microbial cross-infection

Thöni A, Mussner K, Ploner F.

Reparto di Ginecologia e Ostetricia, Ospedale di Vipiteno, Bolzano, Italia.

Minerva Ginecol. 2010 Jun;62(3):203-211.

ABSTRACT: The aim of this study was to document the practice of 2625 water births at Vipiteno over the period 1997-2009 and compare outcome and safety with normal vaginal delivery. The microbial load of the birth pool water was analyzed, and neonatal infection rates after water birth and after land delivery were compared. Methods. The variables analyzed in the 1152 primiparae were: length of labor; incidence of episiotomies and tears; arterial cord blood pH and base excess values; percentage of pH<7.10 and base excess values >/=12 mmol/L. In all 2625 water births, the variables were: analgesic requirements; shoulder dystocia/ neonatal complications; and deliveries after a previous caesarean section. Bacterial cultures of water samples obtained from the bath after filling (sample A) and after delivery (sample B) were analyzed in 300 cases. The pediatricians recorded signs of suspected neonatal infection after water birth and after conventional vaginal delivery. Results. There was a marked reduction in labor duration in the primiparae who birthed in water; the episiotomy rate was 0.46%. Owing to the pain relieving effect of the warm birth pool water, pain relievers (oppiates) were required in only 12.9% of water births. Arterial cord blood pH and base excess values were comparable in both groups. Shoulder dystocia/neonatal complications were managed in 4 water births; 105 women with a previous caesarean section had a water birth. In sample A, the isolated micro-organisms were Legionella spp. and Pseudomonas aeruginosa; in sample B, there was elevated colonization of birth pool water by total coliform bacilli and Escherichia coli. Despite microbial contamination of birth pool water during delivery, antibiotic prophylaxis, as indicated by clinical and laboratory suspicion of infection, was administered to only 0.98% of babies after water birth versus 1.64% of those after land delivery. Conclusions. Results suggest clear medical advantages of water birthing: significantly shorter labor duration among the primiparae; a net reduction in episiotomy rates; and a marked drop in requests for pain relievers. During expulsion of the fetus at delivery, fecal matter is released into the birth pool water, contaminating it with micro-organisms. Despite this, water birthing was found to be safe for the neonate and did not carry a higher risk of neonatal infection when compared with conventional vaginal delivery.


Evaluation of Legionella pneumophila contamination in Italian hotel water systems by quantitative real-time PCR and culture methods

Bonetta S, Bonetta S, Ferretti E, Balocco F, Carraro E.

Dipartimento di Scienze dell'Ambiente e della Vita, University of Piemonte Orientale A. Avogadro, Alessandria, Italy.

J Appl Microbiol. 2010 May;108(5):1576-83.

ABSTRACT: AIMS: This study was designed to define the extent of water contamination by Legionella pneumophila of certain Italian hotels and to compare quantitative real-time PCR with the conventional culture method. METHODS AND RESULTS: Nineteen Italian hotels of different sizes were investigated. In each hotel three hot water samples (boiler, room showers, recycling) and one cold water sample (inlet) were collected. Physico-chemical parameters were also analysed. Legionella pneumophila was detected in 42% and 74% of the hotels investigated by the culture method and by real-time PCR, respectively. In 21% of samples analysed by the culture method, a concentration of >10(4) CFU l(-1) was found, and Leg. pneumophila serogroup 1 was isolated from 10.5% of the hotels. The presence of Leg. pneumophila was significantly influenced by water sample temperature, while no association with water hardness or residual-free chlorine was found. CONCLUSIONS: This study showed a high percentage of buildings colonized by Leg. pneumophila. Moreover, real-time PCR proved to be sensitive enough to detect lower levels of contamination than the culture method. SIGNIFICANCE AND IMPACT OF THE STUDY: This study indicates that the Italian hotels represent a possible source of risk for Legionnaires' disease and confirms the sensitivity of the molecular method. To our knowledge, this is the first report to demonstrate Legionella contamination in Italian hotels using real-time PCR and culture methods.


Free-living amoebae, Legionella and Mycobacterium in tap water supplied by a municipal drinking water utility in the USA

Marciano-Cabral F, Jamerson M, Kaneshiro ES.

Department of Microbiology and Immunology, Virginia Commonwealth University School of Medicine, Richmond VA, 23298, USA.

J Water Health. 2010 Mar;8(1):71-82.

ABSTRACT: Legionella and Mycobacterium can proliferate within free-living amoebae (FLA) where they are protected from disinfectants at concentrations that can kill bacteria but not protozoa. Despite effective treatment of drinking water, microbes can enter water utility distribution systems (DS) and hence the plumbing within building premises. Additionally, biofilm formation may account for the persistence of microbes in the DS. In the present study a domestic water tap in north-central United States (USA) was sampled in March and September 2007 and analysed for FLA, Legionella and Mycobacterium. Identification of organisms was determined by growth on specific culture media, light and electron microscopy, and amplification of DNA probes specific for each organism. In both the spring and fall samples, amoebae, Legionella and Mycobacterium were detected. However, Acanthamoeba was prominent in the spring sample whereas Vahlkampfia and Naegleria were the amoebae detected in the autumn. Bacterial proliferation in laboratory cultures was noticeably enhanced in the presence of amoebae and biofilms rapidly formed in mixed amoebae and bacteria cultures. It is hypothesized that temperature affected the dynamics of FLA species population structure within the DS and that pathogenic bacteria that proliferate within FLA, which are themselves opportunistic pathogens, pose dual public health risks.


Legionella in industrial cooling towers: monitoring and control strategies

Carducci A, Verani M, Battistini R.

Biology Department, University of Pisa, Pisa, Italy.

Lett Appl Microbiol. 2010 Jan;50(1):24-9.

ABSTRACT: AIMS: Legionella contamination of industrial cooling towers has been identified as the cause of sporadic cases and outbreaks of legionellosis among people living nearby. To evaluate and control Legionella contamination in industrial cooling tower water, microbiological monitoring was carried out to determine the effectiveness of the following different disinfection treatments: (i) continuous chlorine concentration of 0.01 ppm and monthly chlorine shock dosing (5 ppm) on a single cooling tower; (ii) continuous chlorine concentration of 0.4 ppm and monthly shock of biocide P3 FERROCID 8580 (BKG Water Solution) on seven towers. METHODS AND RESULTS: Legionella spp. and total bacterial count (TBC) were determined 3 days before and after each shock dose. Both strategies demonstrated that when chlorine was maintained at low levels, the Legionella count grew to levels above 10(4) CFU l(-1) while TBC still remained above 10(8 )CFU l(-1). Chlorine shock dosing was able to eliminate bacterial contamination, but only for 10-15 days. Biocide shock dosing was also insufficient to control the problem when the disinfectant concentration was administered at only one point in the plant and at the concentration of 30 ppm. On the other hand, when at a biocide concentration of 30 or 50 ppm was distributed throughout a number of points, depending on the plant hydrodynamics, Legionella counts decreased significantly and often remained below the warning limit. Moreover, the contamination of water entering the plant and the presence of sediment were also important factors for Legionella growth. CONCLUSIONS: For effective decontamination of outdoor industrial cooling towers, disinfectants should be distributed in a targeted way, taking into account the possible sources of contamination. SIGNIFICANCE AND IMPACT OF THE STUDY: The data of the research permitted to modify the procedure of disinfection for better reduce the water and aerosol contamination and consequently the exposure risk.


An outbreak of Pontiac fever due to Legionella longbeachae serogroup 2 found in potting mix in a horticultural nursery in New Zealand

Cramp GJ, Harte D, Douglas NM, Graham F, Schousboe M, Sykes K.

Te Puna Waiora, Tairawhiti District Health, Gisborne, New Zealand 4040.

Epidemiol Infect. 2010 Jan;138(1):15-20.

ABSTRACT: Previous outbreaks of Pontiac fever have invariably been associated with water droplet spread of Legionella spp. In January 2007 three workers from a horticultural nursery were admitted to hospital with non-pneumonic legionellosis. Investigations showed that a working party of ten people had been exposed to aerosolized potting mix; nine of these workers met the case definition for Pontiac fever. The presence of genetically indistinguishable Legionella longbeachae serogroup 2 was demonstrated in clinical specimens from two hospitalized workers and in the potting mix to which they had been exposed. A further seven cases were diagnosed by serological tests. This is the first documented outbreak of Pontiac fever from L. longbeachae serogroup 2 confirmed from inhalation of potting mix. Pontiac fever is likely to be under-diagnosed. We advocate the introduction of an industry standard that ensures the use of face masks when handling potting mix and attaching masks and warning labels to potting mix bags sold to the public.


Detection of potentially pathogenic bacteria in the drinking water distribution system of a hospital in Hungary

Felföldi T, Heéger Z, Vargha M, Márialigeti K.

Department of Microbiology, Faculty of Science, Eötvös Loránd University, Budapest.

Clin Microbiol Infect. 2010 Jan;16(1):89-92.

ABSTRACT: The drinking water distribution system of a hospital was investigated using standard cultivation techniques, taxon-specific PCRs targeting pathogenic bacteria, denaturing gradient gel electrophoresis, cloning and sequencing. The results obtained verify the higher sensitivity of PCR compared to cultivation for detecting Legionella and Pseudomonas aeruginosa. Moreover, several other opportunistic pathogenic bacteria, such as Escherichia albertii, Acinetobacter lwoffi and Corynebacterium tuberculostrearicum, were detected, emphasizing that drinking water systems, especially those with stagnant water sections, could be the source of nosocomial infections.


Water quality in dental chair units. A random sample in the canton of St. Gallen

Barben J, Kuehni CE, Schmid J.

Division of Paediatric Pulmonology, Children's Hospital, St. Gallen, Switzerland.

Schweiz Monatsschr Zahnmed. 2009;119(10):976-85.

ABSTRACT:This study aimed to identify the microbial contamination of water from dental chair units (DCUs) using the prevalence of Pseudomonas aeruginosa, Legionella species and heterotrophic bacteria as a marker of pollution in water in the area of St. Gallen, Switzerland. Water (250 ml) from 76 DCUs was collected twice (early on a morning before using all the instruments and after using the DCUs for at least two hours) either from the high-speed handpiece tube, the 3 in 1 syringe or the micromotor for water quality testing. An increased bacterial count (>300 CFU/ml) was found in 46 (61%) samples taken before use of the DCU, but only in 29 (38%) samples taken two hours after use. Pseudomonas aeruginosa was found in both water samples in 6/76 (8%) of the DCUs. Legionella were found in both samples in 15 (20%) of the DCUs tested. Legionella anisa was identified in seven samples and Legionella pneumophila was found in eight. DCUs which were less than five years old were contaminated less often than older units (25% und 77%, p<0.001). This difference remained significant (0=0.0004) when adjusted for manufacturer and sampling location in a multivariable logistic regression. A large proportion of the DCUs tested did not comply with the Swiss drinking water standards nor with the recommendations of the American Centers for Disease Control and Prevention (CDC). 


Variable bacterial load of Legionella spp. in a hospital water system

Napoli C, Iatta R, Fasano F, Marsico T, Montagna MT.

Department of Biomedical Science - Hygiene Section, University of Bari, Bari, Italy.

Sci Total Environ. 2009 Dec 20;408(2):242-4.

ABSTRACT: Several approved protocols for the prevention of Legionella pneumonia base the type of intervention (to disinfect or not) on the level of contamination found (cfu/L). However, if the level of contamination by Legionella spp. of a water system fluctuates in a short period of time, inadequate sampling could lead to different decisions being made. To determine if there are significant variations in the bacterial count of Legionella spp., water samples were taken at different times from the same sites. Eight wards were selected from a large hospital in Southern Italy and a water sample was taken from 21 taps in each ward at the same time each day for 5 consecutive days. A Freidman test detected statistically significant differences in average Legionella spp. load over the 5 sampling days (p value<0.001). This fluctuating load can have practical implications: the Italian Guidelines recommend disinfection only for a Legionella count>10,000 cfu/L in hospitals without documented cases of disease. In the present study, the daily average loads varied, during the 5-day sampling period, above and below this cut-off (10,000 cfu/L). This means that the decision to disinfect or not would be different depending on which day the sampling was carried out. Our data suggest that, especially in health-care facilities, a single sampling would not give a realistic estimation of risk; therefore, even at lower levels of bacterial load, measures should be taken to reduce it further.


Pseudo-outbreak of legionnaires disease among patients undergoing bronchoscopy - Arizona, 2008

Centers for Disease Control and Prevention (CDC).

MMWR Morb Mortal Wkly Rep 2009 Aug 14;58(31):849-54MMWR.

ABSTRACT: Legionnaires disease (LD) is a potentially fatal form of pneumonia acquired by inhalation of aerosolized water containing Legionella bacteria. Legionella is a common cause of health-care--associated pneumonia, particularly in settings with hematopoietic stem-cell or solid-organ transplant recipients. On July 25, 2008, the Arizona Department of Health Services (ADHS) notified CDC of four patients who had Legionella cultured from specimens obtained during bronchoscopies performed at a medical center in Arizona. To characterize transmission and identify the source, ADHS and CDC began an investigation on August 1. This report summarizes the results of that investigation, which determined that the patients did not have LD and that nonsterile ice used to cool saline-filled syringes for bronchoalveolar lavage was the likely source of Legionella contamination of these clinical specimens. Ice was supplied by two ice machines, which became contaminated by heavy Legionella colonization within the center's potable water supply during a 6-month period (February--July 2008). Findings from the investigation underscore the importance of adherence to recommended infection control practices and surveillance for LD in health-care settings. Clinicians and endoscopy technicians should ensure that nonsterile items are not introduced during bronchoscopy procedures.


The role of biofilms and protozoa in Legionella pathogenesis: implications for drinking water

Lau HY, Ashbolt NJ.

National Exposure Research Laboratory, U.S. Environmental Protection Agency, Cincinnati, OH, USA.

J Appl Microbiol. 2009 Aug;107(2):368-78.

ABSTRACT: Summary Current models to study Legionella pathogenesis include the use of primary macrophages and monocyte cell lines, various free-living protozoan species and murine models of pneumonia. However, there are very few studies of Legionella spp. pathogenesis aimed at associating the role of biofilm colonization and parasitization of biofilm microbiota and release of virulent bacterial cell/vacuoles in drinking water distribution systems. Moreover, the implications of these environmental niches for drinking water exposure to pathogenic legionellae are poorly understood. This review summarizes the known mechanisms of Legionella spp. proliferation within Acanthamoeba and mammalian cells and advocates the use of the amoeba model to study Legionella pathogenicity because of their close association with Legionella spp. in the aquatic environment. The putative role of biofilms and amoebae in the proliferation, development and dissemination of potentially pathogenic Legionella spp. is also discussed. Elucidating the mechanisms of Legionella pathogenicity development in our drinking water systems will aid in elimination strategies and procedural designs for drinking water systems and in controlling exposure to Legionella spp. and similar pathogens.


The colonization of hot water systems by Legionella

Hruba L.

Center of Hygienic Laboratories, Vrchlickeho 57, 587 25 Jihlava, Czech Republic,

Ann Agric Environ Med. 2009 Jun;16(1):115-9.

ABSTRACT: This study surveyed Legionella in 805 samples taken from 18 hot water systems under operating conditions. The results were analyzed and discussed in relation to water temperature, legislative requirements and optimization of the systems. The temperature of most samples (71%) ranged from 45-60 degrees C. The highest levels of colonization by L. pneumophila were found at water temperatures from 30-35 degrees C. At temperatures above 50 degrees C there was a large decrease in the number of positive samples, as well as the number of Legionella in individual samples. However, L. pneumophila was found in some samples having a temperature of 55-60 degrees C. These results indicate that the legislative requisite temperature of 50 degrees C for hot water systems is insufficient. A system operating temperature of 55 degrees C might be a better optimum, given the economical and safety limitations of temperatures as high as 60 degrees C. If it is impossible or ineffective to use classical method superheating (70-80 degrees C) then it is necessary take into account the chemical decontamination of water.


Potentially pathogenic amoeba-associated microorganisms in cooling towers and their control

Pagnier I, Merchat M, La Scola B.

Unité de recherche sur les maladies infectieuses et tropicales émergentes (URMITE) CNRS UMR 6236, Faculté de Médecine de Marseille, 13385 Marseille Cedex 05, France.

Future Microbiol. 2009 Jun;4:615-29.

ABSTRACT: Cooling towers provide a favorable environment for the proliferation of microorganisms. Cooling towers generate a biofilm and often aerosolize contaminated water, thereby increasing the risk of microorganism dissemination by human inhalation. This pathogen dissemination was first revealed by the epidemics of Legionnaires' disease that were directly related to the presence of cooling towers, and since then, the ecology of Legionella pneumophila has been well studied. Each country has specific standards regarding the acceptable amount of microorganisms in cooling tower systems. However, those standards typically only concern L. pneumophila, even though many other microorganisms can also be isolated from cooling towers, including protozoa, bacteria and viruses. Microbiological control of the cooling tower system can be principally achieved by chemical treatments and also by improving the system's construction. Several new treatments are being studied to improve the efficiency of disinfection. However, as most of these treatments continue to focus solely on L. pneumophila, reports of other types of pathogens continue to increase. Therefore, how their dissemination affects the human populous health should be addressed now.


Survey of pathogenic free-living amoebae and Legionella spp. in mud spring recreation area

Hsu BM, Lin CL, Shih FC.

Department of Earth and Environmental Sciences, National Chung Cheng University, Chiayi, Taiwan, ROC.

Water Res. 2009 Jun;43(11):2817-28.

ABSTRACT: Acanthamoeba, Hartmannella, and Naegleria are free-living amoebae, ubiquitous in aquatic environments. Several species within these genera are recognized as potential human pathogens. These free-living amoebae may facilitate the proliferation of their parasitical bacteria, such as Legionella. In this study, we identified Acanthamoeba, Hartmannella, Naegleria, and Legionella using various analytical procedures and investigated their occurrence at a mud spring recreation area in Taiwan. We investigated factors potentially associated with the prevalence of the pathogens, including various water types, and physical and microbiological water quality parameters. Spring water was collected from 34 sites and Acanthamoeba, Hartmannella, Naegleria, and Legionella were detected in 8.8%, 35.3%, 14.7%, and 47.1%, respectively. The identified species of Acanthamoeba included Acanthamoeba castellanii and Acanthamoeba polyphaga. Nearly all the Hartmannella isolates are identified as Hartmannella vermiformis. The Naegleria species included Naegleria australiensis and its sister groups, and two other isolates referred to a new clade of Naegleria genotypes. The Legionella species identified included unnamed Legionella genotypes, Legionella pneumophila serotype 6, uncultured Legionella spp., Legionella lytica, Legionella drancourtii, and Legionella waltersii. Significant differences (Mann-Whitney U test, P<0.05) were observed between the presence/absence of Hartmannella and total coliforms, between the presence/absence of Naegleria and heterotrophic plate counts, and between the presence/absence of Legionella and heterotrophic plate counts. This survey confirms that pathogenic free-living amoebae and Legionella are prevalent in this Taiwanese mud spring recreation area. The presence of pathogens should be considered a potential health threat when associated with human activities in spring water.


Pseudo-outbreak of Legionella pneumophila serogroup 8 infection associated with a contaminated ice machine in a bronchoscopy suite

Schuetz AN, Hughes RL, Howard RM, Williams TC, Nolte FS, Jackson D, Ribner BS.

Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, Georgia, USA.

Infect Control Hosp Epidemiol. 2009 May;30(5):461-6.

ABSTRACT: OBJECTIVE: To investigate the marked increase noted over an 8-month period in the number of Legionella pneumophila isolates recovered from bronchoalveolar lavage fluid specimens obtained during bronchoscopy in our healthcare system. SETTING: Bronchoscopy suite that serves a 580-bed tertiary care center and a large, multisite, faculty practice plan with approximately 2 million outpatient visits per year. METHODS: Cultures of environmental specimens from the bronchoscopy suite were performed, including samples from the air and water filters, bronchoscopes, and the ice machine, with the aim of identifying Legionella species. Specimens were filtered and acid-treated and then inoculated on buffered charcoal yeast extract agar. Serogrouping was performed on all isolates recovered from patient and environmental samples. RESULTS: All L. pneumophila isolates recovered from patients were serogroup 8, a serogroup that is not usually recovered in our facility. An epidemiologic investigation of the bronchoscopy suite revealed the ice machine to be contaminated with L. pneumophila serogroup 8. Patients were exposed to the organism as a result of a recently adopted practice in the bronchoscopy suite that involved directly immersing uncapped syringes of sterile saline in contaminated ice baths during the procedures. At least 1 patient was ill as a result of the pseudo-outbreak. Molecular typing of isolates recovered from patient and environmental samples revealed that the isolates were indistinguishable. CONCLUSIONS: Extensive cleaning of the ice machine and replacement of the machine's water filter ended the pseudo-outbreak. This episode emphasizes the importance of using aseptic technique when performing invasive procedures, such as bronchoscopies. It also demonstrates the importance of reviewing procedures in all patient areas to ensure compliance with facility policies for providing a safe patient environment.


Survey of wastewater indicators and human pathogen genomes in biosolids produced by class a and class B stabilization treatments

Viau E, Peccia J.

Department of Chemical Engineering, Environmental Engineering Program, Yale University, New Haven, CT 06520, USA.

Appl Environ Microbiol. 2009 Jan;75(1):164-74.

ABSTRACT: Accurate modeling of the infectious aerosol risk associated with the land application of biosolids requires an in-depth knowledge of the magnitudes and changes in pathogen concentrations for a variety of class A and class B stabilization methods. The following survey used quantitative PCR (qPCR) and culture assays to detect environmentally resistant bacterial and viral pathogens and biosolid indicator organisms for 36 biosolid grab samples. Biosolids were collected from 14 U.S. states and included 16 class B mesophilic anaerobic digestion (MAD) samples and 20 class A biosolid samples from temperature-phased anaerobic digestion (TPAD), MAD plus composting (COM), and MAD plus heat pelletization processes. The indicator concentrations of fecal coliforms and male-specific coliphages as well as pathogen genome concentrations for human adenovirus species, Legionella pneumophila, Staphylococcus aureus, and Clostridium difficile were significantly lower in the class A samples, and a multivariate analysis of variance ranked the stabilization processes from the lowest pathogen/indicator load to the highest as (i) class A COM, (ii) class A TPAD, and (iii) class B MAD. Human adenovirus genomes were found in 88% of the class B samples and 70 to 100% of the class A samples. L. pneumophila, S. aureus, and C. difficile genomes were detected at the qPCR assay detection limits in 19 to 50% of the class B and class A anaerobic digestion samples, while L. pneumophila was detected in 50% of the class A compost samples. When considering all the stabilization methods, both the fecal coliform and the male-specific coliphage concentrations show a significant linear correlation with the pathogen genome concentrations. This survey provides the necessary pathogen concentrations to add to biosolid aerosol risk and pathogen exposure analyses and clarifies the effectiveness of class A stabilization methods with the pathogen and indicator loads in biosolids.


Effect of bacterial interference on biofilm development by Legionella pneumophila

Guerrieri E, Bondi M, Sabia C, de Niederhäusern S, Borella P, Messi P.

Department of Biomedical Science, University of Modena and Reggio Emilia, Via Campi 287, Modena 41100, Italy.

Curr Microbiol. 2008 Dec;57(6):532-6.

ABSTRACT: In the ecology of Legionella pneumophila a crucial role may be played by its relationship with the natural flora; thus we investigated the interactions between Legionella and other aquatic bacteria, particularly within biofilms. Among 80 aquatic bacteria screened for the production of bacteriocin-like substances (BLSs), 66.2% of them were active against L. pneumophila. The possible effect of some of these aquatic bacteria on the development and stability of L. pneumophila biofilms was studied. Pseudomonas fluorescens, the best BLS producer, showed the greatest negative effect on biofilm formation and strongly enhanced the detachment of Legionella. Pseudomonas aeruginosa, Burkholderia cepacia, Pseudomonas putida, Aeromonas hydrophila, and Stenotrophomonas maltophilia, although producing BLSs at different levels, were less active in the biofilm experiments. Acinetobacter lwoffii did not produce any antagonistic compound and was the only one able to strongly enhance L. pneumophila biofilm. Our results highlight that BLS production may contribute to determining the fate of L. pneumophila within ecological niches. The interactions observed in this study are important features of L. pneumophila ecology, which knowledge may lead to more effective measures to control the persistence of the germ in the environment.


Microbial contamination of dental unit waterlines in Istanbul, Turkey

Göksay D, Cotuk A, Zeybek Z.

Department of Biology, Faculty of Science, Istanbul University, Vezneciler-Eminönü, Istanbul, 34120, Turkey.

Environ Monit Assess. 2008 Dec;147(1-3):265-9.

ABSTRACT: The water used in dental unit waterlines (DUWLs) acts as a coolant for the high-speed equipment and as an irrigant during dental treatments. There are kind of water tanks. DUWLs provide a favorable environment for microbial biofilm and multiplation primarily due to the high surface in the tubing and the character of fluid dynamics in narrow, smooth-walled waterlines. Biofilms can harbour opportunist pathogens such as Legionella sp., Pseudomonas sp. Several studies have shown that DUWLs have high levels of microbial contamination. Presence of high level of microbial contamination is an important problem for dentists and dental patients who are immunocompromised. We collected water samples from DUWLs of 20 private dental offices. We have determined that only 2 (3.4%) out of 59 dental unit water samples were found to meet the standard (<200 for DUWLs water quality by American Dental Association (ADA). Of the 59 water samples examined, 14 (24%) were positive for Pseudomonas sp. and 18 (30.5%) were positive for fungi. The most common 14 bacterial strains and seven fungi were isolated. Of bacterial strains, 57.1% were identified: Majority of the bacterial species isolated from our samples was identified as Pseudomonas fluorescens, Pasteurella haemolytica, Photobacterium damsela, Ochrobacter anthropi, Moraxella sp., Aspergillus flavus, Penicillium expansum. Legionella sp. were not detected in all water samples.


Legionella species colonization of water distribution systems, pools and air conditioning systems in cruise ships and ferries

Goutziana G, Mouchtouri VA, Karanika M, Kavagias A, Stathakis NE, Gourgoulianis K, Kremastinou J, Hadjichristodoulou C.

Department of Hygiene and Epidemiology, Faculty of Medicine, University of Thessaly, Larissa, Greece.

BMC Public Health. 2008 Nov 24;8:390.

ABSTRACT: BACKGROUND: Legionnaires' disease continues to be a public health concern in passenger ships. This study was scheduled in order to investigate Legionella spp. colonization of water distribution systems (WDS), recreational pools, and air-conditioning systems on board ferries and cruise ships in an attempt to identify risk factors for Legionella spp. colonization associated with ship water systems and water characteristics. METHODS: Water systems of 21 ferries and 10 cruise ships including WDS, air conditioning systems and pools were investigated for the presence of Legionella spp. RESULTS: The 133 samples collected from the 10 cruise ships WDS, air conditioning systems and pools were negative for Legionella spp. Of the 21 ferries WDS examined, 14 (66.7%) were legionellae-positive. A total of 276 samples were collected from WDS and air conditioning systems. Legionella spp. was isolated from 37.8% of the hot water samples and 17.5% of the cold water samples. Of the total 96 positive isolates, 87 (90.6%) were L. pneumophila. Legionella spp. colonization was positively associated with ship age. The temperature of the hot water samples was negatively associated with colonization of L. pneumophila serogroup (sg) 1 and that of L. pneumophila sg 2 to 14. Increases in pH >/=7.8 and total plate count > or =400 CFU/L, correlated positively with the counts of L. pneumophila sg 2 to 14 and Legionella spp. respectively. Free chlorine of > or =0.2 mg/L inhibited colonization of Legionella spp. CONCLUSION: WDS of ferries can be heavily colonized by Legionella spp. and may present a risk of Legionnaires' disease for passengers and crew members. Guidelines and advising of Legionnaires' disease prevention regarding ferries are needed, in particular for operators and crew members.


Biodiversity of amoebae and amoebae-resisting bacteria in a drinking water treatment plant

Thomas V, Loret JF, Jousset M, Greub G.

Center for Research on Intracellular Bacteria, Institute of Microbiology, Faculty of Biology and Medicine, University of Lausanne, Lausanne, Switzerland.

Environ Microbiol. 2008 Oct;10(10):2728-45.

ABSTRACT: The complex ecology of free-living amoebae (FLA) and their role in spreading pathogenic microorganisms through water systems have recently raised considerable interest. In this study, we investigated the presence of FLA and amoebae-resisting bacteria (ARB) at various stages of a drinking water plant fed with river water. We isolated various amoebal species from the river and from several points within the plant, mostly at early steps of water treatment. Echinamoeba- and Hartmannella-related amoebae were mainly recovered in the drinking water plant whereas Acanthamoeba- and Naegleria-related amoebae were recovered from the river water and the sand filtration units. Some FLA isolates were recovered immediately after the ozonation step, thus suggesting resistance of these microorganisms to this disinfection procedure. A bacterial isolate related to Mycobacterium mucogenicum was recovered from an Echinamoeba-related amoeba isolated from ozone-treated water. Various other ARB were recovered using co-culture with axenic Acanthamoeba castellanii, including mycobacteria, legionella, Chlamydia-like organisms and various proteobacteria. Noteworthy, a new Parachlamydia acanthamoebae strain was recovered from river water and from granular activated carbon (GAC) biofilm. As amoebae mainly multiply in sand and GAC filters, optimization of filter backwash procedures probably offers a possibility to better control these protists and the risk associated with their intracellular hosts. 


Tracking airborne Legionella and Legionella pneumophila at a biological treatment plant

Blatny JM, Reif BA, Skogan G, Andreassen O, Høiby EA, Ask E, Waagen V, Aanonsen D, Aaberge IS, Caugant DA.

Norwegian Defence Research Establishment (FFI), P.O. Box 25, N-2027 Kjeller, Norway.

Environ Sci Technol. 2008 Oct 1;42(19):7360-7.

ABSTRACT: Biological treatment plants are frequently used to degrade organic substances in wastewater from wood refinement processes. Aeration ponds in such plants provide an optimal growth environment for many microorganisms, including Legionella species. To investigate whether legionellae could be dispersed as aerosols from the ponds and transported by the wind, the wetted-wall cyclone SASS 2000(PLUS) and the impactors MAS-100 and STA-204 were used to collect air samples directly above, upwind, and downwind of aeration ponds during a 4-month period. Computational fluid dynamics was used a priori to estimate the aerosol paths and to determine suitable air-sampling locations. Several Legionella species, including Legionella pneumophila, were identified in air samples at the biological treatment plant using microbiological and molecular methods. L. pneumophila was identified up to distances of 200 m downwind from the ponds, but, in general, not upwind nor outside the predicted aerosol paths. The highest concentration level of viable legionellae was identified directly above the aeration ponds (3300 CFU/m3). This level decreased as the distance from the aeration ponds increased. Molecular typing indicated that a single clone of L. pneumophila was dispersed from the ponds during the period of the study. Thus, our study demonstrated that aerosols generated at aeration ponds of biological treatment facilities may contain L. pneumophila, which then can be transported by the wind to the surroundings. The methods used in this study may be generically applied to trace biological aerosols that may pose a challenge to environmental occupational health.


Occurrence and diversity of legionellaceae in polar lakes of the antarctic peninsula

Carvalho FR, Nastasi FR, Gamba RC, Foronda AS, Pellizari VH.

Laboratory of Environmental Microbiology, Department of Microbiology, Institute of Biomedical Science, São Paulo, SP, Brazil.

Curr Microbiol. 2008 Oct;57(4):294-300.

ABSTRACT: Legionellaceae is a family of Gram-negative, mesophilic, and facultative intracellular parasitic bacteria that inhabits freshwater environments. In this article, the Legionella population of water samples from the North and South Lake, located close to the Brazilian Scientific Station on King George Island, Keller Peninsula, Antarctica has been characterized. Culture onto selective medium and a independent-culture method were applied to the samples. In our attempt to isolate Legionella species from Antarctic lakes, we were able to obtain one L. pneumophila colony by an amoebic coculture procedure followed by plate culture onto a selective medium. In addition, results obtained from phylogenetic inference showed the presence of noncharacterized specimens of Legionella spp. These findings indicated the presence of legionellae in Antarctica and suggest that these bacteria can adapt to extreme conditions and open new possibilities for understanding the survival strategies of mesophilic Legionellaceae living in Antarctic environments. Furthermore, the isolation of these symbiotic bacteria in Antarctic lakes will allow future studies on cold-resistant mechanisms of legionellae in polar environments.


 Legionella bacteria in shower aerosols increase the risk of Pontiac fever among older people in retirement homes

Bauer M, Mathieu L, Deloge-Abarkan M, Remen T, Tossa P, Hartemann P, Zmirou-Navier D.

Laboratoire d'Hydro-Climatologie Médicale Santé Environnement, School of Medicine, 9 Avenue de la Forêt de Haye, 54500 Vandoeuvre les Nancy, France.

J Epidemiol Community Health. 2008 Oct;62(10):913-20.

ABSTRACT: BACKGROUND: 828 elderly subjects residing in nursing homes were followed up during 4 months to ascertain incidence of symptoms associated with Pontiac fever (PF) in a non-epidemic setting. METHODS: The exposure situation was inhalation of Legionella bacteria while showering. An audit of the hot water system in all institutions allowed ascribing each subject to a water quality area wherefrom one shower was sampled for Legionella assays at the end of the follow-up period. Legionella were detected in water and aerosols using the culture (CFU, colony forming units) and in situ hybridization (FISH) techniques. RESULTS: Among 32 Pontiac-like episodes, 29 cases complied with the operational definition of PF elaborated for this study. Incidence density was 0.11 case/person-year (95% CI 0.07 to 0.15). Water concentrations greater than 105 Legionella FISH/l and 104 Legionella CFU/l were associated with an increased risk of PF (respectively RR 2.23, p = 0.05 and RR 2.39, p = 0.11, with significant dose-response patterns: p for trend <0.04). The condition also seems associated with aerosol concentrations above 103 Legionella FISH/l of air. A significantly higher risk of Pontiac-like episodes (RR 6.24, 95% CI 2.12 to 18.38) was seen for elderly subjects receiving corticosteroid therapy. CONCLUSION: The water and threshold values identified in this research could be used to inform guidance measures aimed at protecting institutionalised older people from Legionnaires' disease. Immunosuppressive therapy in the same population group can significantly enhance susceptibility to Legionella bacteria.


 Surveillance for waterborne disease and outbreaks associated with drinking water and water not intended for drinking--United States, 2005-2006

Yoder J, Roberts V, Craun GF, Hill V, Hicks LA, Alexander NT, Radke V, Calderon RL, Hlavsa MC, Beach MJ, Roy SL; Centers for Disease Control and Prevention (CDC).

Division of Parasitic Diseases, National Center for Zoonotic, Vector-Borne, and Enteric Diseases, CDC, Atlanta, GA 30341, USA.

MMWR Surveill Summ. 2008 Sep 12;57(9):39-62.

ABSTRACT: PROBLEM/CONDITION: Since 1971, CDC, the U.S. Environmental Protection Agency (EPA), and the Council of State and Territorial Epidemiologists have maintained a collaborative Waterborne Disease and Outbreak Surveillance System (WBDOSS) for collecting and reporting data related to occurrences and causes of waterborne-disease outbreaks (WBDOs) and cases of waterborne disease. This surveillance system is the primary source of data concerning the scope and effects of waterborne disease in the United States. REPORTING PERIOD: Data presented summarize 28 WBDOs that occurred during January 2005--December 2006 and four previously unreported WBDOs that occurred during 1979--2002. DESCRIPTION OF SYSTEM: The surveillance system includes data on WBDOs associated with recreational water, drinking water, water not intended for drinking (WNID) (excluding recreational water), and water use of unknown intent. Public health departments in the states, territories, localities, and Freely Associated States (FAS) (i.e., the Republic of the Marshall Islands, the Federated States of Micronesia, and the Republic of Palau, formerly parts of the U.S.-administered Trust Territory of the Pacific Islands) are primarily responsible for detecting and investigating WBDOs and voluntarily reporting them to CDC by a standard form. Only cases and outbreaks associated with drinking water, WNID (excluding recreational water), and water of unknown intent (WUI) are summarized in this report. Cases and outbreaks associated with recreational water are reported in a separate Surveillance Summary. RESULTS: Fourteen states reported 28 WBDOs that occurred during 2005--2006: a total of 20 were associated with drinking water, six were associated with WNID, and two were associated with WUI. The 20 drinking water-associated WBDOs caused illness among an estimated 612 persons and were linked to four deaths. Etiologic agents were identified in 18 (90.0%) of the drinking water-associated WBDOs. Among the 18 WBDOs with identified pathogens, 12 (66.7%) were associated with bacteria, three (16.7%) with viruses, two (11.1%) with parasites, and one (5.6%) mixed WBDO with both bacteria and viruses. In both WBDOs where the etiology was not determined, norovirus was the suspected etiology. Of the 20 drinking water WBDOs, 10 (50) were outbreaks of acute respiratory illness (ARI), nine (45%) were outbreaks of acute gastrointestinal illness (AGI), and one (5.0%) was an outbreak of hepatitis. All WBDOs of ARI were caused by Legionella, and this is the first reporting period in which the proportion of ARI WBDOs has surpassed that of AGI WBDOs since the reporting of Legionella WBDOs was initiated in 2001. A total of 23 deficiencies were cited in the 20 WBDOs associated with drinking water: 12 (52.2%) deficiencies fell under the classification NWU/POU (deficiencies occurred at points not under the jurisdiction of a water utility or at the point-of-use), 10 (43.5%) deficiencies fell under the classification SWTDs (contamination at or in the source water, treatment facility, or distribution system), and for one (4.3%) deficiency, classification was unknown. Among the 12 NWU/POU deficiencies, 10 (83.3%) involved Legionella spp. in the drinking water system. The most frequently cited SWTD deficiencies were associated with a treatment deficiency (n = four [40.0%]) and untreated ground water (n = four [40.0%]). Three of the four WBDOs with treatment deficiencies used ground water sources. INTERPRETATION: Approximately half (52.2%) of the drinking water deficiencies occurred outside the jurisdiction of a water utility. The majority of these WBDOs were associated with Legionella spp, which suggests that increased attention should be targeted towards reducing illness risks associated with Legionella spp. Nearly all of WBDOs associated with SWTD deficiencies occurred in systems using ground water. EPA's new Ground Water Rule might prevent similar outbreaks in the future in public water systems. PUBLIC HEALTH ACTIONS: CDC and EPA use surveillance data to identify the types of water systems, deficiencies, and etiologic agents associated with WBDOs and to evaluate the adequacy of current technologies and practices for providing safe drinking water. Surveillance data also are used to establish research priorities, which can lead to improved water-quality regulation development. The majority of drinking water deficiencies are now associated with contamination at points outside the jurisdiction of public water systems (e.g., regrowth of Legionella spp. in hot water systems) and water contamination that might not be regulated by EPA (e.g., contamination of tap water at the POU). Improved education of consumers and plumbers might help address these risk factors.


Multiresistant waterborne pathogens isolated from water reservoirs and cooling systems

Blasco MD, Esteve C, Alcaide E.

Departamento de Microbiología y Ecología, Universitat de València, Burjassot, Valencia, Spain.

J Appl Microbiol. 2008 Aug;105(2):469-75.

ABSTRACT: AIMS: To determine the incidence of multiple antibiotic-resistant strains of the emergent human pathogens Legionella pneumophila, Pseudomonas aeruginosa and mesophilic Aeromonas species among those isolated from water reservoirs and industrial cooling systems. METHODS AND RESULTS: Water from four natural water reservoirs and four industrial cooling towers was sampled for 1 year period. The total heterotrophs, mesophilic Aeromonas, Pseudomonas spp. and Legionella spp. counts were performed as recommended by standard procedures, and the sensitivity of the isolates to 27 antibiotics was tested. A total of 117 Aeromonas, 60 P. aeruginosa and 15 L. pneumophila strains were isolated and identified by means of biochemical tests and DNA probes. 46.4% of Aeromonas, and 100% of P. aeruginosa isolates presented multiple resistance. Legionella pneumophila strains were generally sensitive to the drugs used. CONCLUSIONS: Antibiotic-resistant pathogenic bacteria belonging to P. aeruginosa and mesophilic Aeromonas species are common in natural aquatic environments. Thus, the risk of waterborne diseases owing to domestic and industrial uses of freshwater should be re-examined from the increase of bacterial resistance point of view. SIGNIFICANCE AND IMPACT OF THE STUDY: These data confirm the emergence of bacteria resistant to antibiotics in aquatic environments.


Protection of waterborne pathogens by higher organisms in drinking water: a review

Bichai F, Payment P, Barbeau B.

Ecole Polytechnique de Montreal, Department of Civil, Geologic and Mining Engineering, P.O. Box 6079, Succ. Centre Ville, Montreal, QC H3C3A7, Canada.

Can J Microbiol. 2008 Jul;54(7):509-24.

ABSTRACT: Higher organisms are ubiquitous in surface waters, and some species can proliferate in granular filters of water treatment plants and colonize distribution systems. Meanwhile, some waterborne pathogens are known to maintain viability inside amoebae or nematodes. The well-documented case of Legionella replication within amoebae is only one example of a bacterial pathogen that can be amplified inside the vacuoles of protozoa and then benefit from the protection of a resistant structure that favours its transport and persistence through water systems. Yet the role of most zooplankton organisms (rotifers, copepods, cladocerans) in pathogen transmission through drinking water remains poorly understood, since their capacity to digest waterborne pathogens has not been well characterized to date. This review aims at (i) evaluating the scientific observations of diverse associations between superior organisms and pathogenic microorganisms in a drinking water perspective and (ii) identifying the missing data that impede the establishment of cause-and-effect relationships that would permit a better appreciation of the sanitary risk arising from such associations. Additional studies are needed to (i) document the occurrence of invertebrate-associated pathogens in relevant field conditions, such as distribution systems; (ii) assess the fate of microorganisms ingested by higher organisms in terms of viability and (or) infectivity; and (iii) study the impact of internalization by zooplankton on pathogen resistance to water disinfection processes, including advanced treatments such as UV disinfection.


Prevalence study of Legionella spp. contamination in Greek hospitals

Mavridou A, Smeti E, Mandilara G, Pappa O, Plakadonaki S, Grispou E, Polemis M.

Technological Educational Institution of Athens Athens, Greece.

Int J Environ Health Res. 2008 Aug;18(4):295-304.

ABSTRACT: Water and swab samples were collected from 13 hospitals and analyzed for Legionella counts. Legionella was detected in eight out of 13 hospitals and in 22 of 130 water and swab-collected samples. A total of 72.7% of the strains were L. pneumophila ser. 1, 22.7% were L. pneumophila ser. 2-14, and 4.5% did not belong to any of these groups. AFLP typing of the L. pneumophila ser. 1 strains generated two distinguishable AFLP types. There was no significant correlation to the sample type with Legionella recovery. Legionella isolation was more likely to occur in the cooling towers than the water system. Water temperatures of 30-40 degrees C seem to favor Legionella growth. Of the 265 serum samples taken from the medical and technical staff for the control of IgG titre, 89.4% were negative, 7.2% were positive, and for 3.4% the result was doubtful. No association between IgG titre and maximum observed level of Legionella occurrence was detected.


Hospital-acquired legionellosis originating from a cooling tower during a period of thermal inversion

Engelhart S, Pleischl S, Lück C, Marklein G, Fischnaller E, Martin S, Simon A, Exner M.

Institute of Hygiene and Public Health, University of Bonn, Sigmund-Freud-Str. 25, 53105 Bonn, Germany.

Int J Hyg Environ Health. 2008 Jul;211(3-4):235-40.

ABSTRACT: A case of hospital-acquired legionellosis occurred in a 75-year-old male patient who underwent surgery due to malignant melanoma. Legionellosis was proven by culture of Legionella pneumophila serogroup 1 from bronchoalveolar lavage (BAL) fluid. Being a chronic smoker the patient used to visit the sickroom balcony that was located about 90 m to the west of a hospital cooling tower. Routine cooling tower water samples drawn during the presumed incubation period revealed 1.0x10(4) CFU/100 ml (L. pneumophila serogroup 1). One of three isolates from the cooling tower water matched the patient's isolate by monoclonal antibody (mab)- and genotyping (sequence-based typing). Horizontal transport of cooling tower aerosols probably was favoured by meteorological conditions with thermal inversion. The case report stresses the importance of routine maintenance and microbiological control of hospital cooling towers.


The high prevalence of Legionella pneumophila contamination in hospital potable water systems in Taiwan: implications for hospital infection control in Asia

Yu PY, Lin YE, Lin WR, Shih HY, Chuang YC, Ben RJ, Huang WK, Chen YS, Liu YC, Chang FY, Yen MY, Liu CC, Ko WC, Lin HH, Shi ZY.

Center for Environmental Laboratory Services, National Kaohsiung Normal University, 62 Shen-chong Rd, Yanchao, Kaohsiung, 824, Taiwan.

Int J Infect Dis. 2008 Jul;12(4):416-20.

ABSTRACT: BACKGROUND: The major sources of Legionnaires' disease (LD) are the potable water systems of large buildings including hospitals, nursing homes, and hotels. Culturing the hospital water system for Legionella allows a preventive approach for hospital-acquired LD. However, hospital-acquired LD is rarely reported in Taiwan, and environmental cultures of Legionella in hospital water systems in Taiwan have never been systematically performed. OBJECTIVE: The objective of this study was to determine if Legionella is present in hospital water systems in Taiwan. Water quality analysis was also performed to determine if geographic differences in water quality result in different Legionella positivity rates. METHOD: The water systems of 16 hospitals throughout Taiwan were tested for Legionella by culture. Standardized culture procedures were followed. RESULTS: Legionella pneumophila was isolated from 63% (10/16) of the hospital water systems; 19% (3/16) of the hospitals had an L. pneumophila positive rate greater than 30%. L. pneumophila serogroups 1 and 6 (strains that are most responsible for Legionella infections) were isolated from 80% (8/10) and 60% (6/10), respectively, of the hospitals that yielded L. pneumophila in their water distribution systems. CONCLUSION: As was shown in epidemiological studies in the USA and Spain, hospital-acquired legionellosis may be prevalent but underdiagnosed in Taiwan.


Gram-negative bacteria in water distribution systems of hospitals

Stojek NM, Szymanska J, Dutkiewicz J.

Department of Occupational Biohazards, Institute of Agricultural Medicine, Jaczewskiego 2, 20-090 Lublin, Poland.

Ann Agric Environ Med. 2008 Jun;15(1):135-42.

ABSTRACT: A total of 67 samples of tap water were collected from faucets and showerheads in 6 hospitals located in the Lublin province (eastern Poland). The samples were examined for the presence and species composition of Legionella, Gram-negative bacteria belonging to family Enterobacteriaceae (GNB-E) and Gram-negative bacteria not belonging to family Enterobacteriaceae (GNB-NE), by filtering through cellulose filters and culture on respectively GVPC, EMB and tryptic soya agar media. On average, Legionella was isolated from 65.7% of the water samples taken in hospitals. Strains of the Legionella pneumophila types 2-14 predominated, forming 74.6% of total Legionella isolates. Legionella pneumophila type 1 strains constituted 13.5% of the total count, while other species of Legionella (referred to as Legionella spp.) formed 11.9% of the total. The concentrations of Legionella in positive water samples ranged from 3-350 cfu/100 ml. GNB-E were not found in the examined water samples. GNB-NE were isolated from 79.1% of the water samples taken in hospitals in the concentrations 11-300 cfu/100 ml. Species of the family Pseudomonadaceae predominated among GNB-NE strains isolated from the examined water samples, forming on average 71.5% of the total count. Altogether, 20 GNB-NE species were identified in the examined samples, out of which 12 were potentially pathogenic. In conclusion, Gram-negative flora of water samples taken in the examined hospitals complies with potable water sanitary standards by the lack of Enterobacteriaceae species, but creates a moderate health risk because of mediocre concentrations of Legionella and the presence of potentially pathogenic non-enterobacterial species.


Italian multicenter study on infection hazards during dental practice: control of environmental microbial contamination in public dental surgeries

Castiglia P, Liguori G, Montagna MT, Napoli C, Pasquarella C, Bergomi M, Fabiani L, Monarca S, Petti S; SItI Working Group Hygiene in Dentistry.

Istituto di Igiene e Medicina Preventiva, Università degli Studi di Sassari, Via P, Manzella 4, 07100 Sassari, Italy.

BMC Public Health. 2008 May 29;8:187.

ABSTRACT: BACKGROUND: The present study assessed microbial contamination in Italian dental surgeries. METHODS: An evaluation of water, air and surface microbial contamination in 102 dental units was carried out in eight Italian cities. RESULTS: The findings showed water microbial contamination in all the dental surgeries; the proportion of water samples with microbial levels above those recommended decreased during working. With regard to Legionella spp., the proportion of positive samples was 33.3%. During work activity, the index of microbial air contamination (IMA) increased. The level of microbial accumulation on examined surfaces did not change over time. CONCLUSION: These findings confirm that some Italian dental surgeries show high biocontamination, as in other European Countries, which highlights the risk of occupational exposure and the need to apply effective measures to reduce microbial loads.


Dynamics of Legionella spp. and bacterial populations during the proliferation of L. pneumophila in a cooling tower facility

Wéry N, Bru-Adan V, Minervini C, Delgénes JP, Garrelly L, Godon JJ.

INRA, UR50, Laboratoire de Biotechnologie de l'Environnement, Avenue des Etangs, Narbonne F-11100, France.

Appl Environ Microbiol. 2008 May;74(10):3030-7.

ABSTRACT: The dynamics of Legionella spp. and of dominant bacteria were investigated in water from a cooling tower plant over a 9-month period which included several weeks when Legionella pneumophila proliferated. The structural diversity of both the bacteria and the Legionella spp. was monitored by a fingerprint technique, single-strand conformation polymorphism, and Legionella spp. and L. pneumophila were quantified by real-time quantitative PCR. The structure of the bacterial community did not change over time, but it was perturbed periodically by chemical treatment or biofilm detachment. In contrast, the structure of the Legionella sp. population changed in different periods, its dynamics at times showing stability but also a rapid major shift during the proliferation of L. pneumophila in July. The dynamics of the Legionella spp. and of dominant bacteria were not correlated. In particular, no change in the bacterial community structure was observed during the proliferation of L. pneumophila. Legionella spp. present in the cooling tower system were identified by cloning and sequencing of 16S rRNA genes. A high diversity of Legionella spp. was observed before proliferation, including L. lytica, L. fallonii, and other Legionella-like amoebal pathogen types, along with as-yet-undescribed species. During the proliferation of L. pneumophila, Legionella sp. diversity decreased significantly, L. fallonii and L. pneumophila being the main species recovered.


Isolation and identification of amoeba-resisting bacteria from water in human environment by using an Acanthamoeba polyphaga co-culture procedure

Pagnier I, Raoult D, La Scola B.

Unité des Rickettsies, CNRS UMR 6020, Faculté de Médecine de Marseille, 13385 Marseille Cedex 05, France.

Environ Microbiol. 2008 May;10(5):1135-44.

ABSTRACT: Amoeba-resisting bacteria (ARB) such as Legionella spp. are currently regarded as potential human pathogens living in the environment. To detect ARB from both human and environmental samples, co-culture with amoebae has been demonstrated as an efficient tool. However, using this procedure, mostly water from cooling towers and hospital water supplies have been investigated as the possible reservoir of ARB. In the present study, we studied ARB population in 77 environmental water samples including rivers, fountains, lakes and domestic wells in the south of France. As a result, a total of 244 isolates corresponding to 89 different species of ARB, but not Legionella spp., were identified. Ability to grow within and/or to be lytic for amoebae was revealed for the first time for several human pathogens. Six isolates are likely to be the members of a new or uncharacterized genus/species. An anaerobic bacterium, Clostridium frigidicarnis was demonstrated to be lytic for amoebae. This preliminary work demonstrates that the water environment in the vicinity of humans is a reservoir of ARB, including well-known pathogens for which amoebae and/or water was not recognized earlier as a possible reservoir.


A Legionnaires' disease outbreak: a water blaster and roof-collected rainwater systems

Simmons G, Jury S, Thornley C, Harte D, Mohiuddin J, Taylor M.

Auckland Regional Public Health Service, Auckland 1035, New Zealand.

Water Res. 2008 Mar;42(6-7):1449-58.

ABSTRACT: In February 2006, an outbreak of Legionnaires' disease (LD) was identified in Beachlands, a small, isolated east Auckland suburb. It was investigated through case finding, a case-control study, sampling potential sources of infection and by molecular typing (using sequence-based typing (SBT) of all Legionella pneumophila serogroup 1 (Lp1) isolates). Lp1 was isolated from the respiratory tract of one case, the roof-collected rainwater systems of five households (three associated with cases) and from a water blaster at a nearby marina. All isolates were indistinguishable, exhibiting the same SBT allele pattern. Three LD cases lived within 500m of the water blaster (the fourth case within 1250m) and downwind in prevailing conditions. Another domestic roof-collected rainwater supply contaminated by Lp1 (identical SBT pattern) was incidentally identified in another suburb 4km east of Beachlands. This is the first outbreak of LD linked to roof-collected rainwater supplies and the first isolation of Legionella from these systems in New Zealand. Aerosols containing Legionella discharged to air by the marina water blaster may have infected some cases directly or may have seeded roof-collected rainwater systems. Some cases may have been exposed by contaminated bathroom showers. Roof-collected rainwater systems need appropriate design, careful cleaning and the maintenance of hot water temperatures at a minimum of 60 degrees C to reduce the chances of Legionella multiplying. Further research into the ecology of Legionella in roof-collected rain water systems is indicated.


Genetic diversity of Legionella pneumophila in hospital water systems

Oberdorfer K, Müssigbrodt G, Wendt C.

Institute of Hygiene, University of Heidelberg, Im Neuenheimer Feld 324, 69120 Heidelberg, Germany.

Int J Hyg Environ Health. 2008 Mar;211(1-2):172-8.

ABSTRACT: It has been shown that different patients who had acquired legionellosis in a hospital setting were infected with the same strain even years apart. However, there are no longitudinal data describing the molecular epidemiology of Legionella pneumophila strains that contaminate a water system. This raised the question if there are any shifts of L. pneumophila strains over time, or after carrying out control measures. Using genotyping on a large collection of isolates, we investigated in a retrospective study the distribution of L. pneumophila serogroups and PFGE types in six different hospitals of the University of Heidelberg between 1991 and 2001. A total of 2012 water samples were drawn for routine testing and for evaluation of control measures, 747 samples were positive for L. pneumophila. Serogroups were determined by latex agglutination or by direct fluorescence assay; and 515 L. pneumophila isolates from water systems and six from patients underwent PFGE typing after SfiI-restriction. We identified seven serogroups and 19 genotypes among the water isolates. Each hospital had one to four predominating PFGE types that were stable over the investigation period. The oldest buildings in hospitals 4 and 5 (built 1876 and 1907) had more types than the newest one (built 1986). In all hospitals PFGE types were identified that could be found only sporadically. Although each hospital had its own warm water supply, we identified types that could be found in more than one hospital. However, there was no overlap of types in buildings that were fed from different wells. Infrequently occurring nosocomial legionellosis (n=3) were only caused by predominant strains. Contamination of water supplies seemed to be dominated by stable genotypes, even after various control measures. Additional genotypes could be isolated sporadically, however, their pathogenetic relevance seemed to be questionable.


Occurrence of Legionella in hot water systems of single-family residences in suburbs of two German cities with special reference to solar and district heating

Mathys W, Stanke J, Harmuth M, Junge-Mathys E.

Institute for Hygiene, University of Muenster, Robert-Koch-Str. 41, 48129 Muenster, Germany.

Int J Hyg Environ Health. 2008 Mar;211(1-2):179-85.

ABSTRACT: A total of 452 samples from hot water systems of randomly selected single family residences in the suburbs of two German cities were analysed for the occurrence of Legionella. Technical data were documented using a standardized questionnaire to evaluate possible factors promoting the growth of the bacterium in these small plumbing systems. All houses were supplied with treated groundwater from public water works. Drinking water quality was within the limits specified in the German regulations for drinking water and the water was not chlorinated. The results showed that plumbing systems in private houses that provided hot water from instantaneous water heaters were free of Legionella compared with a prevalence of 12% in houses with storage tanks and recirculating hot water where maximum counts of Legionella reached 100,000 CFU/100ml. The presence of L. pneumophila accounted for 93.9% of all Legionella positive specimens of which 71.8% belonged to serogroup 1. The volume of the storage tank, interrupting circulation for several hours daily and intermittently raising hot water temperatures to >60 degrees C had no influence on Legionella counts. Plumbing systems with copper pipes were more frequently contaminated than those made of synthetic materials or galvanized steel. An inhibitory effect due to copper was not present. Newly constructed systems (<2 years) were not colonized. The type of hot water preparation had a marked influence. More than 50% of all houses using district heating systems were colonized by Legionella. Their significantly lower hot water temperature is thought to be the key factor leading to intensified growth of Legionella. Although hot water systems using solar energy to supplement conventional hot water supplies operate at temperatures 3 degrees C lower than conventional systems, this technique does not seem to promote proliferation of the bacterium. Our data show convincingly that the temperature of the hot water is probably the most important or perhaps the only determinant factor for multiplication of Legionella. Water with a temperature below 46 degrees C was most frequently colonized and contained the highest concentrations of legionellae. It is evident that the same factors affecting colonization by Legionella in large buildings also exist in small residential water systems. If temperatures are low there is no difference between large and small systems and Legionella counts are high in both. Since private residences are an important source of community-acquired legionellosis, these findings emphasize the need for preventive control measures in small residential buildings. In some situations it may be necessary to install filtration devices at the point-of-use.


Genotypic variability and persistence of Legionella pneumophila PFGE patterns in 34 cooling towers from two different areas

Sanchez I, Garcia-Nuñez M, Ragull S, Sopena N, Pedro-Botet ML, Esteve M, Rey-Joly C, Sabria M.

Infectious Diseases Unit, Hospital Universitari Germans Trias i Pujol, Autonomous University of Barcelona, Badalona, Spain.

Environ Microbiol 2008 Feb;10(2):395-9.

ABSTRACT: Genotypic variability and clonal persistence are important concepts in molecular epidemiology as they facilitate the search for the source of sporadic cases or outbreaks of legionellosis. We studied the genotypic variability and persistence of Legionella pulsed-field gel electrophoresis (PFGE) patterns over time (period > 6 months) in 34 positive cooling towers from two different areas. In area A, radius of 70 km, 52 indistinguishable PFGE patterns were differentiated among the 27 cooling towers. In 13 cooling towers we observed >or= 2 PFGE patterns. Each cooling tower had its own indistinguishable Legionella PFGE pattern which was not shared with any other cooling tower. In area B, radius of 1 km, 10 indistinguishable PFGE patterns were obtained from the seven cooling towers. In four, we observed >or= 2 PFGE patterns. Three of these 10 indistinguishable PFGE patterns were shared by more than one cooling tower. In 27 of 34 cooling towers the same PFGE pattern was recovered after 6 months to up to 5 years of follow-up. The large genotypic diversity of Legionella observed in the cooling towers aids in the investigation of community outbreaks of Legionnaires' disease. However, shared patterns in small areas may confound the epidemiological investigation. The persistence of some PFGE patterns in cooling towers makes the recovery of the Legionella isolate causing the outbreak possible over time.


An outbreak of legionnaires disease caused by long-distance spread from an industrial air scrubber in Sarpsborg, Norway

Nygård K, Werner-Johansen Ø, Rønsen S, Caugant DA, Simonsen Ø, Kanestrøm A, Ask E, Ringstad J, Ødegård R, Jensen T, Krogh T, Høiby EA, Ragnhildstveit E, Aaberge IS, Aavitsland P.

Norwegian Institute of Public Health, Oslo, Norway.

Clin Infect Dis. 2008 Jan 1;46(1):61-9.

ABSTRACT: On 21 May 2005, the Norwegian health authorities were alerted by officials from a local hospital that several recent patients had received the diagnosis of legionnaires disease; all patients resided in 2 neighboring municipalities. We investigated the outbreak to identify the source and to implement control measures. METHODS: We interviewed all surviving case patients and investigated and harvested samples from 23 businesses with cooling towers and other potential infection sources. The locations of the businesses and the patients' residences and movements were mapped. We calculated attack rates and risk ratios among people living within various radii of each potential source. Isolates of Legionella pneumophila were compared using molecular methods. RESULTS: Among 56 case patients, 10 died. The case patients became ill 12-25 May, resided up to 20 km apart, and had not visited places in common. Those living up to 1 km from a particular air scrubber had the highest risk ratio, and only for this source did the risk ratio decrease as the radius widened. Genetically identical L. pneumophila serogroup 1 isolates were recovered from patients and the air scrubber. The air scrubber is an industrial pollution-control device that cleans air for dust particles by spraying with water. The circulating water had a high organic content, pH of 8-9, and temperature of 40 degrees C. The air was expelled at 20 m/s and contained a high amount of aerosolized water. CONCLUSIONS: The high velocity, large drift, and high humidity in the air scrubber may have contributed to the wide spread of Legionella species, probably for >10 km. The risk of Legionella spread from air scrubbers should be assessed.


Transcriptional profiling of Legionella pneumophila biofilm cells and the influence of iron on biofilm formation

Hindré T, Brüggemann H, Buchrieser C, Héchard Y.

Laboratoire de Chimie de l'Eau et de l'Environnement, UMR 6008, Université de Poitiers, 40 Avenue du Recteur Pineau, 86022 Poitiers Cedex, France.

Microbiology. 2008 Jan;154(Pt 1):30-41.

ABSTRACT: In aquatic environments, biofilms constitute an ecological niche where Legionella pneumophila persists as sessile cells. However, very little information on the sessile mode of life of L. pneumophila is currently available. We report here the development of a model biofilm of L. pneumophila strain Lens and the first transcriptome analysis of L. pneumophila biofilm cells. Global gene expression analysis of sessile cells as compared to two distinct populations of planktonic cells revealed that a substantial proportion of L. pneumophila genes is differentially expressed, as 2.3 % of the 2932 predicted genes exhibited at least a twofold change in gene expression. Comparison with previous results defining the gene expression profile of replicative- and transmissive-phase Legionella suggests that sessile cells resemble bacteria in the replicative phase. Further analysis of the most strongly regulated genes in sessile cells identified two induced gene clusters. One contains genes that encode alkyl hydroperoxide reductases known to act against oxidative stress. The second encodes proteins similar to PvcA and PvcB that are involved in siderophore biosynthesis in Pseudomonas aeruginosa. Since iron has been reported to modify biofilm formation in other species, we further focused on iron control of gene expression and biofilm formation. Among the genes showing the greatest differences in expression between planktonic cells and biofilm, only pvcA and pvcB were regulated by iron concentration. A DeltapvcA L. pneumophila mutant showed no changes in biofilm formation compared to the wild-type, suggesting that the pvcA product is not mandatory for biofilm formation. However, biofilm formation by L. pneumophila wild-type and a DeltapvcA strain was clearly inhibited in iron-rich conditions.

Aerosolization of mycobacteria and legionellae during dental treatment: low exposure despite dental unit contamination

Dutil S, Veillette M, Mériaux A, Lazure L, Barbeau J, Duchaine C.

Institut universitaire de cardiologie et de pneumologie de l'Université Laval, Hôpital Laval, Québec, Canada.

Environ Microbiol. 2007 Nov;9(11):2836-43.

ABSTRACT: Dental unit waterlines (DUWL) support growth of a dense microbial population that includes pathogens and hypersensitivity-inducing bacteria, such as Legionella spp. and non-tuberculous mycobacteria (NTM). Dynamic dental instruments connected to DUWL generate aerosols in the work environment, which could allow waterborne pathogens to be aerosolized. The use of the real-time quantitative polymerase chain reaction (qPCR) provides a more accurate estimation of exposure levels compared with the traditional culture approach. Bioaerosol sampling was performed 13 times in an isolated dental treatment room according to a standardized protocol that included four dental prophylaxis treatments. Inhalable dust samples were taken at the breathing zone of both the hygienist and patient and outside the treatment room (control). Total bacteria as well as Legionella spp. and NTM were quantified by qPCR in bioaerosol and DUWL water samples. Dental staff and patients are exposed to bacteria generated during dental treatments (up to 4.3 E + 05 bacteria per m(3) of air). Because DUWL water studied was weakly contaminated by Legionella spp. and NTM, their aerosolization during dental treatment was not significant. As a result, infectious and sensitization risks associated with legionellae and NTM should be minimal.


Colonization of legionella species in hotel water systems in Turkey

Erdogan H, Arslan H.

Department of Infectious Diseases and Clinical Microbiology, Baskent University Alanya Hospital, Antalya, Turkey.

J Travel Med. 2007 Nov-Dec;14(6):369-73.

ABSTRACT: Background. The goal of this study was to evaluate the prevalence of Legionella species in hotel water distribution systems in Alanya, Turkey, which is an important tourism center. Methods. Water and swab samples were obtained from 52 Turkish hotels from August 2003 to September 2005. Water samples were collected in 100 mL sterile containers and were concentrated by membrane filters with a pore size of 0.45 mum. Heat treatment was used to eliminate other microorganisms from the samples, which were then spread on buffered charcoal yeast extract alpha agar plates and glycine, vancomycin, polymyxin, cycloheximide agar plates. Cysteine-dependent colonies were identified by latex agglutination. Results. In all, 491 water and swab samples were analyzed. The results of all samples were negative for Legionella in 16 (30.8%) hotels. Legionella species (92.5% of which were Legionella pneumophila) were detected in 93 (18.9%) of the samples. The most frequently isolated species were L pneumophila serogroups 6 (63.5%) and 1 (21.5%). Conclusions. Legionella pneumophila serogroup 6 was the most common isolate detected in Turkish hotel water systems in our study. The result of Legionella urinary antigen tests, which are the diagnostic tests most often used to identify legionnaires' disease, may be negative in people infected with L pneumophila serogroup 6. We suggest that clinicians should apply the whole spectrum of laboratory methods for the detection of legionnaires' disease in patients with pneumonia of unknown origin and history of travel to Alanya, Turkey.


Phylogenetic study of legionella species in pristine and polluted aquatic samples from a tropical atlantic forest ecosystem

Carvalho FR, Vazoller RF, Foronda AS, Pellizari VH.

Laboratory of Environmental Microbiology, Department of Microbiology, Institute of Biomedical Science, University of São Paulo, Room 148 Lineu Prestes avenue, 1374, Cidade Universitária São Paulo, SP, Brazil.

Curr Microbiol. 2007 Oct;55(4):288-93.

ABSTRACT: Legionella species are ubiquitous bacteria in aquatic environments. To examine the effect of anthropogenic impacts and physicochemical characteristics on the Legionellaceae population, we collected water from two sites in the Itanhaém River system in the Atlantic Forest of Brazil. One sample was collected from an upstream pristine region, the other from a downstream estuarine region moderately affected by untreated domestic sewage. Cultures on a selective medium failed to isolate Legionella species. Culture-independent methods showed that water from the estuarine aquatic habitat contained DNA sequences homologous to the 16S ribosomal DNA gene of Legionella pneumophila and non-pneumophila species. In pristine water, only two sequences related to L. pneumophila were detected. The results suggest that salinity and anthropogenic factors, such as wastewater discharge, favor a diversity of Legionella species, whereas pristine freshwater selects for Legionella pneumophila.


Persistence of the same strain of Legionella pneumophila in the water system of an Italian hospital for 15 years

Scaturro M, Dell'eva I, Helfer F, Ricci ML.

Department of Infectious, Parasitic, and Immune-mediated Diseases, Istituto Superiore di Sanita, Viale Regina Elena 299, 00161-Rome, Italy.

Infect Control Hosp Epidemiol. 2007 Sep;28(9):1089-92.

ABSTRACT: In 2004, an outbreak of legionnaires disease occurred in a hospital in northern Italy with a water system that had been disinfected multiple times since 1990 and equipped with a continuous disinfecting system. Molecular typing linked the outbreak to contamination of the hospital water system and demonstrated the persistence of a predominant strain of Legionella pneumophila for 15 years.


Do contaminated dental unit waterlines pose a risk of infection?

Pankhurst CL, Coulter WA.

Department of Oral Medicine, King's College London Dental Institute, Bessemer Road, London, United Kingdom.

J Dent. 2007 Sep;35(9):712-20.

ABSTRACT: OBJECTIVES: To review the evidence that the dental unit waterlines are a source of occupational and healthcare acquired infection in the dental surgery. DATA: Transmission of infection from contaminated dental unit waterlines (DUWL) is by aerosol droplet inhalation or rarely imbibing or wound contamination in susceptible individuals. Most of the organisms isolated from DUWL are of low pathogenicity. However, data from a small number of studies described infection or colonisation in susceptible hosts with Legionella spp., Pseudomonas spp. and environmental mycobacteria isolated from DUWL. The reported prevalence of legionellae in DUWL varies widely from 0 to 68%. The risk from prolonged occupational exposure to legionellae has been evaluated. Earlier studies measuring surrogate evidence of exposure to legionellae in dental personnel found a significant increase in legionella antibody levels but in recent multicentre studies undertaken in primary dental care legionellae were isolated at very low rate and the corresponding serological titres were not above background levels. Whereas, a case of fatal Legionellosis in a dental surgeon concluded that the DUWL was the likely source of the infection. The dominant species isolated from dental unit waterlines (DUWL) are Gram-negative bacteria, which are a potent source of cell wall endotoxin. A consequence of indoor endotoxin exposure is the triggering or exacerbation of asthma. Data from a single large practice-based cross-sectional study reported a temporal association between occupational exposure to contaminated DUWL with aerobic counts of >200cfu/mL at 37 degrees C and development of asthma in the sub-group of dentists in whom asthma arose following the commencement of dental training. SOURCES: Medline 1966 to February 2007 was used to identify studies for this paper. STUDY SELECTION: Design criteria included randomised control trials, cohort, and observational studies in English. CONCLUSIONS: Although the number of published cases of infection or respiratory symptoms resulting from exposure to water from contaminated DUWL is limited, there is a medico-legal requirement to comply with potable water standards and to conform to public perceptions on water safety.


Molecular evidence for the ubiquitous presence of Legionella species in Dutch tap water installations

Diederen BM, de Jong CM, Aarts I, Peeters MF, van der Zee A.

Laboratory of Medical Microbiology and Immunology, St Elisabeth Hospital, Tilburg, The Netherlands.

J Water Health. 2007 Sep;5(3):375-83.

ABSTRACT: Our aim was to investigate the occurrence and identity of Legionella spp. in Dutch tap water installations using culture, real-time PCR and sequence analysis. The PCR assays used were a 16S rRNA gene based PCR with both a Legionella species specific probe and a L. pneumophila specific probe and a L. pneumophila-specific PCR based on the sequence of the mip gene. A total of 357 water samples from 250 locations in The Netherlands was investigated. The detection rates of Legionella spp. were 2,2% (8 of 357) by culture, and 87,1% (311 of 357) by PCR. The majority of samples was found to contain Legionella species other than L. pneumophila. These comprised of Legionella Like Amoebal Pathogens (LLAPs), L. busanensis, L. worsliensis and others. Fourteen (3,9%) samples were positive for L. pneumophila by either culture, 16S rRNA based PCR and/or mip based PCR. It is apparent from this study that Legionella spp. DNA is ubiquitous in Dutch potable water samples. Our findings further suggest that LLAPs and viable but nonculturable (VBNC) Legionella represent a large proportion of the population in man-made environments.


Legionella contamination in the water system of hospital dental settings

Veronesi L, Capobianco E, Affanni P, Pizzi S, Vitali P, Tanzi ML.

Public Health Department, University of Parma, Italy.

Acta Biomed. 2007 Aug;78(2):117-22.

ABSTRACT: BACKGROUND AND AIM OF THE WORK: Among hospital facilities the dental unit is an environment that is at major risk of Legionella due to equipment such as the air/water syringe, the turbine, the micromotor and the scaler which generate potentially harmful aerosols that may to be a source of exposure to Legionella spp. particularly in immunodeficient patients, and those affected by chronic diseases, and also in dental personnel. Therefore, an examination of the extent of Legionella spp. contamination in the dental chairs waterlines and the incoming water supply of some public dental units is the subject of the present study. METHODS: From February 2002 to March 2004, a total of 208 water samples were collected: 160 samples from the water supply of 4 dental chair and 48 samples from the cold incoming tap water of 2 units. RESULTS: Legionella spp. was detected in 46 samples (22.1% ): 19 of them (41.3% of Legionella spp.; 9.1% of the total) were Legionella pneumophila; Pseudomonas aeruginosa was detected in 86 samples (41.4%) and both microorganisms were detected in 2 samples (0.96%). CONCLUSIONS: Our results show a microbiological condition in dental settings, that is not at all satisfactory due to the presence of Legionella in concentrations that are considered to be a health hazard (> or = 10(3)) in certain cases. Given the extent of the health risk in these surroundings, the difficulty in its assessment, and also considering the wide diffusion of general dental care, our investigation has confirmed the need to regularly monitor the microbiological condition of water in dental units.


Legionella pneumophila in cooling towers: fluctuations in counts, determination of genetic variability by pulsed-field gel electrophoresis (PFGE), and persistence of PFGE patterns

Ragull S, Garcia-Nuñez M, Pedro-Botet ML, Sopena N, Esteve M, Montenegro R, Sabrià M.

Infectious Diseases Unit, Germans Trias i Pujol University Hospital, Cta Canyet s/n, 08916 Badalona (Barcelona), Spain. msabria.germanstrias@gencat.met

Appl Environ Microbiol. 2007 Aug;73(16):5382-4.

ABSTRACT: The concentrations of Legionella pneumophila in cooling towers may vary considerably over short periods of time, producing significant fluctuations throughout the year. Despite genetic variability, in small geographical areas the same indistinguishable pulsed-field gel electrophoresis patterns may be shared among different cooling towers and persist over time.


Increased rainfall is associated with increased risk for legionellosis

Hicks LA, Rose CE Jr, Fields BS, Drees ML, Engel JP, Jenkins PR, Rouse BS, Blythe D, Khalifah AP, Feikin DR, Whitney CG.

Epidemic Intelligence Service, Office of Workforce and Career Development, Centers for Disease Control and Prevention, Atlanta, GA, USA.

Epidemiol Infect. 2007 Jul;135(5):811-7.

ABSTRACT: Legionnaires' disease (LD) is caused by Legionella species, most of which live in water. The Mid-Atlantic region experienced a sharp rise in LD in 2003 coinciding with a period of record-breaking rainfall. To investigate a possible relationship, we analysed the association between monthly legionellosis incidence and monthly rainfall totals from January 1990 to December 2003 in five Mid-Atlantic states. Using negative binomial model a 1-cm increase in rainfall was associated with a 2.6% (RR 1.026, 95% CI 1.012-1.040) increase in legionellosis incidence. The average monthly rainfall from May to September 1990-2002 was 10.4 cm compared to 15.7 cm from May to September 2003. This change in rainfall corresponds to an increased risk for legionellosis of approximately 14.6% (RR 1.146, 95% CI 1.067-1.231). Legionellosis incidence increased during periods of increased rainfall; identification of mechanisms that increase exposure and transmission of Legionella during rainfall might lead to opportunities for prevention.


Influence of aquatic microorganisms on Legionella pneumophila survival

Guerrieri E, Bondi M, Borella P, Messi P.

Department of Biomedical Sciences, University of Modena and Reggio Emilia, Modena, Italy.

New Microbiol. 2007 Jul;30(3):247-51.

ABSTRACT: The ability of aquatic bacteria Pseudomonas fluorescens SSD (Ps-D) and Pseudomonas putida SSC (Ps-C) to support the persistence of Legionella pneumophila (Lp-1) in an artificial water microcosm was investigated for 42 day, at two different incubation temperatures. At 4 degrees C, individually suspended Lp-1 was no longer detectable just after 24 hours, while in co-cultures with Pseudomonas, Lp1 showed a better survival capability. At 30 degrees C, Lp-1 alone displayed high survival rates over the entire period of observation. When Lp-1 was inoculated with Ps-C and Ps-D, its count showed a marked decrease, followed by a gradual and costant decline.


Detection of Legionella spp. and some of their amoeba hosts in floating biofilms from anthropogenic and natural aquatic environments

Declerck P, Behets J, van Hoef V, Ollevier F.

Laboratory of Aquatic Ecology, Department of Biology, Zoological Institute, Katholieke Universiteit Leuven, Charles Deberiotstraat 32, 3000 Leuven, Belgium.

Water Res. 2007 Jul;41(14):3159-67.

ABSTRACT: Floating biofilms develop at the water-air interface and harbor numerous microorganisms, some of which are human pathogens like Legionella pneumophila. The presence of Legionella spp. and especially L. pneumophila in such biofilms was investigated. In parallel, the occurrence of Naegleria spp., Acanthamoeba spp., Willaertia spp., Vahlkampfia spp. and Hartmanella spp. was determined and it was examined whether Acanthamoeba spp. isolates were naturally infected with L. pneumophila bacteria. Eight anthropogenic and 37 natural aquatic environments were sampled between June and August 2005. Both Legionella spp. and L. pneumophila were present in 100% of the floating biofilms of the anthropogenic aquatic systems. Eighty-one percent of all natural floating biofilm samples were positive for Legionella spp. and 70% of these samples were positive for L. pneumophila. Legionella concentrations were in the range of 10(1)-10(2)cells/cm(2). Naegleria spp. and Acanthamoeba spp., two well-known L. pneumophila amoeba hosts, were present in 50-92% and 67-72% of floating biofilm samples, respectively. Acanthamoeba spp. isolates appeared to be naturally infected with L. pneumophila bacteria as proved by fluorescent in situ hybridization.


A controlled study of Legionella concentrations in water from faucets with aerators or laminar water flow devices

Huang WK, Lin YE.

Graduate Institute of Environmental Education, National Kaohsiung Normal University, 62 Shen-chong Rd., Yanchao, Kaohsiung, Taiwan 824.

Infect Control Hosp Epidemiol 2007;28:765-766.



Legionella impletisoli sp. nov. and Legionella yabuuchiae sp. nov., isolated from soils contaminated with industrial wastes in Japan

Kuroki H, Miyamoto H, Fukuda K, Iihara H, Kawamura Y, Ogawa M, Wang Y, Ezaki T, Taniguchi H.

Department of Microbiology, School of Medicine, University of Occupational and Environmental Health, Fukuoka 807-8555, Japan.

Syst Appl Microbiol. 2007 Jun;30(4):273-9.

ABSTRACT: In this study, we tried to isolate legionellae from nine Legionella DNA-positive soil samples collected from four different sites contaminated with industrial wastes in Japan. Using culture methods with or without Acanthamoeba culbertsoni, a total of 22 isolates of legionellae were obtained from five of the nine samples. Identification of species and/or serogroups (SGs), performed by DNA-DNA hybridization and agglutination tests, revealed that the 22 isolates consisted of ten isolates of Legionella pneumophila including five SGs, five Legionella feeleii, and one each of Legionella dumoffii, Legionella longbeachae, and Legionella jamestownensis. The species of the remaining four isolates (strains OA1-1, -2, -3, and -4) could not be determined, suggesting that these isolates may belong to new species. The 16S rDNA sequences (1476-1488bp) of the isolates had similarities of less than 95.0% compared to other Legionella species. A phylogenetic tree created by analysis of the 16S rRNA (1270bp) genes demonstrated that the isolates formed distinct clusters within the genus Legionella. Quantitative DNA-DNA hybridization tests on the OA1 strains indicated that OA1-1 should be categorized as a new taxon, whereas OA1-2, -3, and -4 were also genetically independent in another taxon. Based on the evaluated phenotypic and phylogenetic characteristics, it is proposed that one of these isolates from the soils, OA1-1, be classified as a novel species, Legionella impletisoli sp. nov.; the type strain is strain OA1-1(T) (=JCM 13919(T)=DSMZ 18493(T)). The remaining three isolates belong to another novel Legionella species, Legionella yabuuchiae sp. nov.; the type strain is strain OA1-2(T) (=JCM 14148(T)=DSMZ 18492(T)). This is the first report on the isolation of legionellae from soils contaminated with industrial wastes.


Legionella in the dental office

Bodrumlu E, Alaçam T, Bayraktar A.

Operative Dentistry and Endodontics Department, Faculty of Dentistry, Ondokuz Mayis University, Samsun, Turkey.

Int J Dent Hyg. 2007 May;5(2):116-21. 

ABSTRACT: OBJECTIVE: This study aimed to assess the microbiology of dental unit water and municipal water in terms of Legionella species and total bacteria levels. METHODS: The presence of Legionella species was investigated using the culture method, direct fluorescent antibody and polymerase chain reaction techniques in collected dental unit water and municipal water samples from 71 dental offices in Ankara, Turkey. In addition, total bacterial counts were assessed using the culture method. RESULTS: In 27% of the dental unit water samples and in 13% of municipal water samples, the number of colony-forming units (cfu ml(-1)) significantly exceeded acceptable values for high-risk group patients. No Legionella spp. was found in the dental unit water samples. Legionella SG3 was found in only one municipal water sample. CONCLUSION: The dental unit water systems examined in this study did not include Legionella spp., but other bacteria at high numbers were determined. This is a potential threat, especially for elderly people, the medically compromised patients receiving regular dental treatment and the dental clinic staff.


Risk factors for contamination of hotel water distribution systems by Legionella species

Mouchtouri V, Velonakis E, Tsakalof A, Kapoula C, Goutziana G, Vatopoulos A, Kremastinou J, Hadjichristodoulou C.

Department of Hygiene and Epidemiology, Faculty of Medicine, University of Thessaly, 22 Papakiriazi Str., Larissa 41222, Greece.

Appl Environ Microbiol. 2007 Mar;73(5):1489-92.

ABSTRACT: The Legionella colonization frequency at 385 Greek hotel hot and cold water distribution systems was 20.8%. Legionella contamination was associated with the presence of an oil heater (odds ratio [OR]=2.04, 95% confidence interval [CI]=1.12 to 3.70), with the sample temperature (OR=0.26, 95% CI=0.1 to 0.5), with seasonal operation (OR=3.23, 95% CI=1.52 to 6.87), and with the presence of an independent disinfection system (OR=0.30, 95% CI=0.15 to 0.62). The same water temperatures, free-chlorine levels, and pHs differently affect the survival of various Legionella spp.


 Does using potting mix make you sick? Results from a Legionella longbeachae case-control study in South Australia

O'Connor BA, Carman J, Eckert K, Tucker G, Givney R, Cameron S.

Communicable Disease Control Branch, Department of Health South Australia, Australia.

Epidemiol Infect. 2007 Jan;135(1):34-9.

ABSTRACT: A case-control study was performed in South Australia to determine if L. longbeachae infection was associated with recent handling of commercial potting mix and to examine possible modes of transmission. Twenty-five laboratory-confirmed cases and 75 matched controls were enrolled between April 1997 and March 1999. Information on underlying illness, smoking, gardening exposures and behaviours was obtained by telephone interviews. Recent use of potting mix was associated with illness (OR 4.74, 95% CI 1.65-13.55, P=0.004) in bivariate analysis only. Better predictors of illness in multivariate analysis included poor hand-washing practices after gardening, long-term smoking and being near dripping hanging flower pots. Awareness of a possible health risk with potting mix protected against illness. Results are consistent with inhalation and ingestion as possible modes of transmission. Exposure to aerosolized organisms and poor gardening hygiene may be important predisposing factors to L. longbeachae infection.


Environmental survey of Legionella pneumophila in hot springs in Taiwan

Lin YE, Lu WM, Huang HI, Huang WK.

Graduate Institute of Environmental Education and Center for Environmental Laboratory Services, National Kaohsiung Normal University, Kaohsiung, Taiwan.

J Toxicol Environ Health A. 2007 Jan;70(1):84-7.

ABSTRACT: Acquisition of sporadic community-acquired legionnaires' disease has been linked to hot springs and whirlpool baths. Outbreaks of hot spring-associated legionnaires' disease were reported in Japan in the last few years. Although the mode of transmission is unclear, the presence of Legionella in hot springs may discourage hot springs resort visits by the general public. An environmental survey was conducted to determine the presence of Legionella in hot springs in Taiwan. In total, 55 water samples were collected from 19 hot springs resorts; 21% (4/19) of the hot spring resorts sampled yielded L. pneumophila in the public hot springs bath. Legionella pneumophila serogroups 1 and 6, L. pneumophila serogroup 3, L. pneumophila serogroup 5, and L. pneumophila serogroup 7 were isolated from four different resort spas, respectively. The total sample positivity rate for L. pneumophila was 11% (6/55). The risk of occurrences of legionnaires' disease outbreaks associated with hot springs water in general public is unknown, and epidemiologic investigations should be conducted for locating the potential sources of Legionella for those cases of community-acquired legionnaires' disease. Disinfection of hot springs for Legionella may be necessary if the risk of contracting legionnaires' disease from hot springs can be validated by an evidence-based approach.


Legionella contamination in the hospital environment: monitoring of the hot water distribution systems of three hospitals in Catania (Italy)

Pignato S, Coniglio MA, Faro G, Cantaro P, Carini SA, Mangano G, Cunsolo R, Coco G, Giammanco G.

Dipartimento G.F. Ingrassia Igiene e Sanità Pubblica, Laboratorio di Riferimento Regionale per le Legionellosi, Università di Catania.

Ig Sanita Pubbl. 2006 Nov-Dec;62(6):635-52.

ABSTRACT: This study evaluated the presence and extent of contamination with Legionella spp. in the hot water distribution systems of three hospitals in Catania (Italy). In total, 291 hot water samples were collected between September 2002 and August 2005 and these were examined in order to monitor the hospital distribution systems and evaluate the efficacy of decontamination measures. L. pneumophila was detected at variable concentrations up to over 10000 UFC/L at several collection sites in some hospital buildings and branches of the water distribution system while other buildings/branches were found to be free of contamination. The most frequently isolated serogroup was L. pneumophila serogroup 3, occasionally associated with serogroups 4, 5 and 6. Molecular typing of Legionella strains by pulsed-field gel electrophoresis of genomic DNA restriction fragments identified four different genotypes, each recovered from a different branch of the distribution system. Decontamination procedures, including shock hyperchloration and two different thermal shock methods, performed between October 2003 and August 2005, led to only temporary reductions in contamination. In fact, previous concentrations of the same L. pneumophila serogroup were found within 3 to 8 months of decontamination. In order to prevent and monitor Legionella infections, sterilizing filters were installed in water taps of all wards with high-risk patients and urinary antigen testing was performed on all patients diagnosed with nosocomial pneumonia. No cases of Legionella pneumonia were identified in 2005.


Influence of amoebae and physical and chemical characteristics of water on presence and proliferation of Legionella species in hospital water systems

Lasheras A, Boulestreau H, Rogues AM, Ohayon-Courtes C, Labadie JC, Gachie JP.

Service Hygiène Hospitalière, Hôpital Pellegrin, CHU Bordeaux, Bordeaux cedex, France.

Am J Infect Control. 2006 Oct;34(8):520-5.

ABSTRACT: The reservoir for hospital-acquired Legionnaires' disease has been shown to be the potable water distribution system. The objectives of the present study were as follows: (1) to examine the possible relationship between physical-chemical characteristics of water such as temperature, pH, hardness, conductivity, and residual chlorine and the presence of amoebae as growth-promoting factors for Legionella species and (2) to determine eradication measures for water distribution systems to seek ways of reducing the risk of legionellosis. Ten hospitals in southwest France took part in this study. Water samples were collected from 106 hot water faucets, showers, hot water tanks, and cooling towers. Two analyses were performed to analyze the association between water characteristics and (1) the presence of Legionella species and (2) the proliferation of Legionella species. Of the 106 water samples examined, 67 (63.2%) were positive for Legionella species. Amoebae were detected in 73 of 106 (68.9%) samples and in 56 of 67 (86.6%) Legionella species-positive samples (P < 10(-6)). In these positive samples, conductivity was lower than 500 microOmega(-1).cm(-1) in 58.2% (P = .026), temperature was below 50 degrees C in 80.6% (P = .004), and hardness was significantly higher (P = 002) than in Legionella species-negative samples. Neither Legionella species nor amoebae were isolated from any sampling point in which the water temperature was above 58.8 degrees C. Multivariate analysis shows that high hardness and presence of amoebae were strongly correlated statistically with the presence of Legionella when showers, tanks, pH, and temperature promoted their proliferation. This study shows the importance of water quality evaluation in assessing environmental risk factors and in selecting the most appropriate prevention and control measures in hospital water systems.


Pilot study on the presence of Legionella spp in 6 Italian cities' dental units

Montagna MT, Tatò D, Napoli C, Castiglia P, Guidetti L, Liguori G, Petti S, Tanzi ML; Gruppo di Lavoro SItI L'Igiene in Odontoiatria.

Dip. di Scienze Biomediche ed Oncologia Umana, Sezione di Igiene, Università degli Studi di Bari.

Ann Ig. 2006 Jul-Aug;18(4):297-303.

ABSTRACT: Among the microorganism involved in environmental contamination, Legionella spp is actually considered an important infectious hazard. The aim of this study is to evaluate the incidence of Legionella spp in water samples collected from 138 dental unit selected from public outpatient clinics of 6 Italian cities. The samples were taken from oral rinsing cup, air-water syringe, ultrasonic scaler and the turbine to investigate Legionella spp, Pseudomonas aeruginosa and the total heterotrophic count at 36 degrees and 22 degrees. Legionella spp was present in 33,3% dental unit water; but a significant difference was shown among the enrolled cities. In 43,5% of water sample Legionella concentration was 1.000-10.000 CFU/L and in 30,4% was >10.000 CFU/L. L. pneumophila 1 was found in 23,9% of water samples. The results demonstrate that the concentration of Legionella spp in dental unit water lines could be high and this suggests that the exposure to these micoorganism during the dental practise could be a potential health risk both for dental personnel and for the patients too, especially when immunocompromised.


Correlation between Legionella contamination in water and surrounding air

Crimi P, Macrina G, Grieco A, Tinteri C, Copello L, Rebora D, Galli A, Rizzetto R.

Department of Health Sciences, University of Genoa, Genoa, Italy.

Infect Control Hosp Epidemiol. 2006 Jul;27(7):771-3.

ABSTRACT: We evaluated Legionella pneumophila contamination of water and surrounding air in a burn care department equipped with bathtubs. In water, the bacterium always aerosolized, but in surrounding air, it diluted itself to such a point that it became undetectable at 1 m or more from the source of emission, which indicated that patients were at low risk of inhaling a dangerous quantity of the microbe.

A legionellosis case due to contaminated spa water and confirmed by genomic identification in Taiwan

Su HP, Tseng LR, Tzeng SC, Chou CY, Chung TC.

Center for Disease Control, Department of Health Taiwan.

Microbiol Immunol. 2006;50(5):371-7.

ABSTRACT: Tracing the source of a legionellosis (LG) case revealed that the Legionella pneumophila (LP) strain isolated from patient's sputum shared the same serogroup (SG) and PFGE-type with 4 LP strains obtained from a spa center. With a high LP-contamination rate (81.2%, 13/16) in all of its 16 basins, this spa center was also found to have a multi-genotypic distribution among its 13 LP isolates, which can be categorized into 5 PFGE-types. Despite such a serious contamination in the spa center, which usually had ca. 100 visitors per day, this male patient, bearing LG-risk factors of long-term heavy smoking and alcoholism, was the only case identifiable after an active investigation. To explore the possible reason for this sporadic infection, all 5 PFGE-types of LP isolated were assayed for their presence of two important virulent genes (lvh and rtx A) and were identified as either less-virulent (lvh (+) , rtx A(+)) or non-virulent (lvh (-), rtx A (-)) types. The strong virulent type (lvh (+), rtx A (+)) usually seen in clinical strains elsewhere was not found here. Moreover, the LG-causative type in this infection was the only one to be classified as the less-virulent type, with the presence of lvh gene indicating its relatively more virulent potential than other 4 PFGE-types. Accordingly, mutual interaction between LP's virulent potential and patient's health-status was suggested to be the force directing the opportunistic infection of this sporadic case. This is the first spa-associated infection caused by SG 2 of LP.


Legionella pneumophila in commercial bottled mineral water

Klont RR, Rijs AJ, Warris A, Sturm PD, Melchers WJ, Verweij PE.

Department of Medical Microbiology, Radboud University Nijmegen Medical Center, Nijmegen, The Netherlands.

FEMS Immunol Med Microbiol. 2006 Jun;47(1):42-4.

Sixty-eight commercial bottled mineral waters (64 brands, 68 different 'best-before dates') were tested for the presence of bacteria and fungi. Six samples were Legionella antigen positive and six were Legionella pneumophila PCR positive. Two samples were both Legionella antigen and L. pneumophila PCR positive. Legionella cultures were negative. Although the PCR might have detected only dead Legionella cells, the PCR has been described to detect specifically viable but not culturable (VBNC) L. pneumophila cells as well. Whether VBNC bacteria may be present in bottled mineral waters and the risk for infection this may pose for severely immunocompromised patients should be investigated.


Microbial ecology of drinking water distribution systems

Berry D, Xi C, Raskin L.

Department of Civil and Environmental Engineering, University of Michigan, Ann Arbor, 1351 Beal Avenue, Ann Arbor, MI 48109-2125, USA.

Curr Opin Biotechnol. 2006 Jun;17(3):297-302.

The supply of clean drinking water is a major, and relatively recent, public health milestone. Control of microbial growth in drinking water distribution systems, often achieved through the addition of disinfectants, is essential to limiting waterborne illness, particularly in immunocompromised subpopulations. Recent inquiries into the microbial ecology of distribution systems have found that pathogen resistance to chlorination is affected by microbial community diversity and interspecies relationships. Research indicates that multispecies biofilms are generally more resistant to disinfection than single-species biofilms. Other recent findings are the increased survival of the bacterial pathogen Legionella pneumophila when present inside its protozoan host Hartmannella vermiformis and the depletion of chloramine disinfectant residuals by nitrifying bacteria, leading to increased overall microbial growth. Interactions such as these are unaccounted for in current disinfection models. An understanding of the microbial ecology of distribution systems is necessary to design innovative and effective control strategies that will ensure safe and high-quality drinking water.


Isolation and characterization of Legionella spp. and Pseudomonas spp. from greenhouse misting systems

Zietz BP, Dunkelberg H, Ebert J, Narbe M.

Medical Institute of General Hygiene and Environmental Health, University of Gottingen, Gottingen, Germany.

J Appl Microbiol. 2006 Jun;100(6):1239-50.

Abstract Aims: Greenhouse misting systems used for watering plants produce fine aerosols. They are a possible cause for bacterial infections. This study investigates the colonization of greenhouse misting systems with Legionella spp. and Pseudomonas spp. and evaluates a possible health hazard. Methods and Results: Between June and September 2003, a total of 80 water samples were collected in 20 different greenhouse systems in Germany, each tested on two different occasions. Each time, water was drawn at a central tap and at the outlet of spray nozzles. Sampled greenhouses were used to cultivate various plants and trees for commercial, recreational or scientific reasons, some of them in tropical conditions. Legionella spp. were detected in 10% of the systems (two systems), but only in low numbers. On the contrary, Pseudomonas spp. were recovered from 70% of the greenhouse watering systems (14 systems), occasionally at counts greater than 10 000 CFU per 100 ml. A random amplified polymorphic DNA polymerase chain reaction typing method was used to demonstrate that each colonized greenhouse had one or several individual strains of Legionella and Pseudomonas that could not be detected in any other system. Conclusions: This study demonstrates that aerosolizing greenhouse watering systems may be contaminated with Legionella or Pseudomonas which under certain circumstances could become a potential source of infection for workers and visitors. Significance and Impact of the Study: The study results indicate that greenhouse misting systems should be included in Legionella and Pseudomonas monitoring and control programs.


Detection of Legionella pneumophila serogroup 7 strain from bathwater samples in a Japanese hospital

Fujimura S, Oka T, Tooi O, Meguro M, Chiba M, Kawamura M, Maki F, Takeda H, Watanabe A.

Department of Microbiology, Miyagi University, 1 Gakuen, Taiwa-cho, Miyagi 981-3298, Japan.

J Infect Chemother. 2006 Apr;12(2):105-8.

Hospital-acquired legionellosis is one of the serious problems in nosocomial infection. For risk assessment of nosocomial Legionella infection, we surveyed samples from bathrooms for public use in three hospitals and two nursing homes to determine whether Legionella pneumophila was present. A total of 70 hot bathwater samples and samples wiped from bathtubs were collected at 1-h intervals. Fifteen shower-water and 15 inner-head samples were obtained at the start of a bath. Water samples were cultured using the Legionella spp. selective medium, and discrimination between L. pneumophila and other Legionella spp. was performed by PCR analysis. L. pneumophila serogroup 7 was detected in 1 bathwater and 1 wiped sample, both of which were collected 1 h after daily use from the same bathtub in a hospital. However, L. pneumophila SG7 was not detected in any other samples. Furthermore, the concentrations of free residual chlorine in most bath- and shower-water samples were lower than 0.1 mg/l. These results suggest that L. pneumophila has become a potential pathogen for nosocomial infections in public-type hospital baths. From the point of view of an infection-control program, it might be advisable to hold the concentration of free residual chlorine at 0.2-0.4 mg/l, which is generally required for public baths in Japan.


Legionella anisa, a Possible Indicator of Water Contamination by Legionella pneumophila

van der Mee-Marquet N, Domelier AS, Arnault L, Bloc D, Laudat P, Hartemann P, Quentin R.

Service de Bacteriologie et Hygiene, CHRU Trousseau, 37044 Tours Cedex 9, France.

J Clin Microbiol. 2006 Jan;44(1):56-9.

ABSTRACT: Legionella anisa is one of the most frequent species of Legionella other than Legionella pneumophila in the environment and may be hospital acquired in rare cases. We found that L. anisa may mask water contamination by L. pneumophila, suggesting that there is a risk of L. pneumophila infection in immunocompromised patients if water is found to be contaminated with Legionella species other than L. pneumophila.


Soil as a source of Legionella pneumophila serogroup 1 (Lp1)

Wallis L, Robinson P.

School of Public Health, Faculty of Health Sciences, La Trobe University, Victoria, Bundoora.

Aust N Z J Public Health. 2005 Dec;29(6):518-20.

ABSTRACT: OBJECTIVE: To investigate the potential source of a case of Legionnaires' disease caused by an unusual serotype of Legionella pneumophila serogroup 1 (Lp1) in regional Victoria in May 2001. METHOD: Epidemiological and environmental investigation of the source of infection of a case of Legionnaires' disease in regional Victoria in May 2001. RESULTS: Extensive environmental investigations did not reveal any cooling water tower systems close to the residence or the shopping centre that the case visited prior to illness. The sputum culture and a soil sample from the field at the plant nursery where the case worked prior to illness were both positive for Legionella pneumophilia serogroup 1, MDU pulsovar 97:103. CONCLUSION: Legionella pneumophila has been found in soil and was further found to be associated with a case of Legionella pneumophila. IMPLICATIONS: Public health authorities should consider exposures to soil and potting mixes when investigating cases of Legionella pneumophila where the case has no apparent association with cooling towers. Safe gardening practices should be promoted among the community.


Prevalence of Legionella pneumophila serogroup 1 in water distribution systems in Izmir province of Turkey

Uzel A, Ucar F, Esin Hames-Kocabas E.

Department of Biology, Basic and Industrial Microbiology Section, Faculty of Sciences, Ege University, Bornova-Izmir, Turkey.

APMIS. 2005 Oct;113(10):664-669.

ABSTRACT: Legionella pneumophila serogroup 1 occurrence has been investigated in 168 hot water samples from 24 hotels, situated in 6 counties in Izmir province of Turkey, from 15 June to 30 September of the year 2000. Sampling was carried out at 15-day intervals and seven samples were taken from each of the hotels' hot water reservoirs and hot water networks. The samples were (1 L) concentrated using polycarbonate filters (mesh size 0.22 mum). Isolation was achieved using selective medium, GVPC agar. The samples were concentrated by membrane filtration, divided into three portions and cultured without pretreatment, after acid treatment, and after heat treatment, on GVPC agar. One hundred and ten isolates were identified as L.pneumophila sg 1 using the Legionella Latex Test (Oxoid). Arbitrarily primed PCR (AP PCR) was employed to assess the clonal relationship between Legionella pneumophila sg 1 isolates from the hot water samples of the hotels. Three genotypes of L. pneumophila sg 1 isolates were identified. With a high prevalence of type A, 22 hotels were found to be colonized with L. pneumophila serogroup 1, while only 2 were free from the bacteria.


Isolation of pathogenic Legionella species and legionella-laden amoebae in dental unit waterlines

Singh T, Coogan MM.

Department of Immunology and Microbiology, National Institute for Occupational Health, P.O. Box 4788, Johannesburg 2000, South Africa.

J Hosp Infect. 2005 Nov;61(3):257-62.

ABSTRACT: Legionella released into the air during treatment are a potential source of infection. Water stagnation in dental unit waterlines (DUWLs) creates biofilms and promotes the proliferation of these micro-organisms. This study investigated the presence of amoeba infected with legionella, L. pneumophila and other pathogenic Legionella species in a dental teaching hospital. Water samples were collected in the morning and afternoon from 99 dental units and 16 taps connected to the municipal water supply. Samples were plated on selective media and tested for legionella using the direct immunofluorescent antibody technique and the latex agglutination test. Legionella were found in 33% of the DUWLs and in 47% of the mains taps supplying these units. Legionella-laden amoebae occurred in one mains tap sample and in 20% of DUWLs in a clinic of the teaching hospital. L. micdadei was the predominant species isolated from this clinic. L. pneumophila serogroups 2-14 predominated in the mains water, whereas L. pneumophila serogroup 1 was found in approximately half of the contaminated DUWLs and mains taps irrespective of the time of sampling. Pathogenic Legionella species seeded by municipal water into DUWLs is a potential source of legionella infection for both dental personnel and patients during prolonged dental treatment. This problem is compounded by the presence of legionella-laden amoebae which may contain levels of organism well within the infective dose. The interaction of legionella with amoebae is an important ecological factor that may significantly increase the risk of legionellosis, and thus should be given further consideration in the refinement of risk assessment models.


Water ecology of Legionella and protozoan: environmental and public health perspectives

Borella P, Guerrieri E, Marchesi I, Bondi M, Messi P.

Department of Hygiene and Microbiology, University of Modena and Reggio Emilia, Via Campi 287, 41100 Modena, Italy.

Biotechnol Annu Rev. 2005;11:355-80.

ABSTRACT: Ecological studies on Legionella spp. are essential to better understand their sources in the natural environments, the mechanism of their entry into man-made water systems and the factors enabling their survival and growth in aquatic habitats. Legionella spp. exhibits peculiar and multiple strategies to adapt to stressful environment conditions which normally impair other germ survival. These strategies include the ability to enter in a viable but non-cultivable (VBNC) state, to multiply intracellularly within a variety of protozoa, such as amoebae, to survive as free organisms within biofilms and to be enhanced/inhibited by the presence of other aquatic bacteria. The host-parasite interaction has been shown to be central in the pathogenesis and ecology of L. pneumophila. The bacterial-protozoan interaction contributes to the amplification of Legionella population in water systems, represents a shelter against unfavourable environmental conditions, acts as a reservoir of infection and contributes to virulence by priming the pathogen to infect human cells. Legionella is able to survive as free organism for long periods within biofilms which are widespread in man-made water systems. Biofilm provides shelter and nutrients, exhibits a remarkable resistance to biocide compounds and chlorination, thus representing ecological niches for legionella persistence in such environments. Further knowledge on biofilm-associated legionellae may lead to effective control measures to prevent legionellosis. Lastly, new perspectives in controlling legionella contamination can arise from investigations on aquatic bacteria able to inhibit legionella growth in natural and artificial water systems.


Legionella contamination in hot water of italian hotels

Borella P, Montagna MT, Stampi S, Stancanelli G, Romano-Spica V, Triassi M, Marchesi I, Bargellini A, Tato D, Napoli C, Zanetti F, Leoni E, Moro M, Scaltriti S, Ribera D'Alcala G, Santarpia R, Boccia S.

Department of Hygiene and Microbiology, Via Campi, 287, I-41100 Modena, Italy.

Appl Environ Microbiol. 2005 Oct;71(10):5805-13.

ABSTRACT: A cross-sectional multicenter survey of Italian hotels was conducted to investigate Legionella spp. contamination of hot water. Chemical parameters (hardness, free chlorine concentration, and trace element concentrations), water systems, and building characteristics were evaluated to study risk factors for colonization. The hot water systems of Italian hotels were strongly colonized by Legionella; 75% of the buildings examined and 60% of the water samples were contaminated, mainly at levels of >/=10(3) CFU liter(-1), and Legionella pneumophila was the most frequently isolated species (87%). L. pneumophila serogroup 1 was isolated from 45.8% of the contaminated sites and from 32.5% of the hotels examined. When a multivariate logistic model was used, only hotel age was associated with contamination, but the risk factors differed depending on the contaminating species and serogroup. Soft water with higher chlorine levels and higher temperatures were associated with L. pneumophila serogroup 1 colonization, whereas the opposite was observed for serogroups 2 to 14. In conclusion, Italian hotels, particularly those located in old buildings, represent a major source of risk for Legionnaires' disease due to the high frequency of Legionella contamination, high germ concentration, and major L. pneumophila serogroup 1 colonization. The possible role of chlorine in favoring the survival of Legionella species is discussed.


Balamuthia mandrillaris, Free-Living Ameba and Opportunistic Agent of Encephalitis, Is a Potential Host for Legionella pneumophila Bacteria

Shadrach WS, Rydzewski K, Laube U, Holland G, Ozel M, Kiderlen AF, Flieger A.

Robert Koch-Institut, Cellular Immunology Unit, Nordufer 20, D-13353 Berlin, Germany.

Appl Environ Microbiol. 2005 May;71(5):2244-9.

ABSTRACT: Balamuthia mandrillaris is a free-living ameba and an opportunistic agent of granulomatous encephalitis in humans and other mammalian species. Other free-living amebas, such as Acanthamoeba and Hartmannella, can provide a niche for intracellular survival of bacteria, including the causative agent of Legionnaires' disease, Legionella pneumophila. Infection of amebas by L. pneumophila enhances the bacterial infectivity for mammalian cells and lung tissues. Likewise, the pathogenicity of amebas may be enhanced when they host bacteria. So far, the colonization of B. mandrillaris by bacteria has not been convincingly shown. In this study, we investigated whether this ameba could host L. pneumophila bacteria. Our experiments showed that L. pneumophila could initiate uptake by B. mandrillaris and could replicate within the ameba about 4 to 5 log cycles from 24 to 72 h after infection. On the other hand, a dotA mutant, known to be unable to propagate in Acanthamoeba castellanii, also did not replicate within B. mandrillaris. Approaching completion of the intracellular cycle, L. pneumophila wild-type bacteria were able to destroy their ameboid hosts. Observations by light microscopy paralleled our quantitative data and revealed the rounding, collapse, clumping, and complete destruction of the infected amebas. Electron microscopic studies unveiled the replication of the bacteria in a compartment surrounded by a structure resembling rough endoplasmic reticulum. The course of intracellular infection, the degree of bacterial multiplication, and the ultrastructural features of a L. pneumophila-infected B. mandrillaris ameba resembled those described for other amebas hosting Legionella bacteria. We hence speculate that B. mandrillaris might serve as a host for bacteria in its natural environment.



Presence and persistence of Legionella spp. in groundwater

Costa J, Tiago I, da Costa MS, Verissimo A.

Departamento de Zoologia and Centro de Neurociencias e Biologia Celular, Universidade de Coimbra, 3004-517 Coimbra, Portugal.

Appl Environ Microbiol. 2005 Feb;71(2):663-71.

ABSTRACT: Groundwater samples (111) from six different boreholes located in two geographical areas were examined for the presence of legionellae over a 7-year period. The number of Legionella isolates detected was generally low. The colonization of the aquifers was not uniform, and the persistence of Legionella was independent of the hydraulic pumps and the plumbing system present in the borehole. A total of 374 isolates identified by fatty acid methyl ester analysis belonged to Legionella pneumophila, L. oakridgensis, L. sainthelensi, and L. londiniensis. In area 1, L. oakridgensis constituted the major population detected, exhibiting only one random amplified polymorphic DNA (RAPD)-PCR profile. L. sainthelensi strains were less frequently isolated and also displayed a single RAPD profile, while L. pneumophila was only sporadically detected. In contrast, L. pneumophila comprised the vast majority of the isolates in area 2 and exhibited six distinct RAPD patterns, indicating the presence of different genetic groups; three L. londiniensis RAPD types were also detected. Two of the L. pneumophila and one of the L. londiniensis RAPD types were persistent in this environment for at least 12 years. The genetic structure of L. pneumophila groundwater populations, inferred from rpoB and dotA gene sequences, was peculiar, since the majority of the isolates were allied in a discrete group different from the lineages containing most of the type and reference strains of the three subspecies of L. pneumophila. Furthermore, gene exchange events related to the dotA allele could be envisioned.


Accumulation and fate of microorganisms and microspheres in biofilms formed in a pilot-scale water distribution system

Langmark J, Storey MV, Ashbolt NJ, Stenstrom TA.

Department of Parasitology, Mycology and Water Microbiology, Swedish Institute for Infectious Disease Control, SE-171 82 Solna, Sweden.

Appl Environ Microbiol. 2005 Feb;71(2):706-12.

ABSTRACT: The accumulation and fate of model microbial "pathogens" within a drinking-water distribution system was investigated in naturally grown biofilms formed in a novel pilot-scale water distribution system provided with chlorinated and UV-treated water. Biofilms were exposed to 1-mum hydrophilic and hydrophobic microspheres, Salmonella bacteriophages 28B, and Legionella pneumophila bacteria, and their fate was monitored over a 38-day period. The accumulation of model pathogens was generally independent of the biofilm cell density and was shown to be dependent on particle surface properties, where hydrophilic spheres accumulated to a larger extent than hydrophobic ones. A higher accumulation of culturable legionellae was measured in the chlorinated system compared to the UV-treated system with increasing residence time. The fate of spheres and fluorescence in situ hybridization-positive legionellae was similar and independent of the primary disinfectant applied and water residence time. The more rapid loss of culturable legionellae compared to the fluorescence in situ hybridization-positive legionellae was attributed to a loss in culturability rather than physical desorption. Loss of bacteriophage 28B plaque-forming ability together with erosion may have affected their fate within biofilms in the pilot-scale distribution system. The current study has demonstrated that desorption was one of the primary mechanisms affecting the loss of microspheres, legionellae, and bacteriophage from biofilms within a pilot-scale distribution system as well as disinfection and biological grazing. In general, two primary disinfection regimens (chlorination and UV treatment) were not shown to have a measurable impact on the accumulation and fate of model microbial pathogens within a water distribution system.


Legionella waterline colonization: detection of Legionella species in domestic, hotel and hospital hot water systems

Leoni E, De Luca G, Legnani PP, Sacchetti R, Stampi S, Zanetti F.

Department of Medicine and Public Health, University of Bologna , Bologna , Italy .

J Appl Microbiol. 2005 Feb;98(2):373-379.

ABSTRACT: Aims: An evaluation was made of the prevalence of Legionella species in hot water distribution systems in the city of Bologna ( Italy ) and their possible association with bacterial contamination (total counts and Pseudomonadaceae) and the chemical characteristics of the water (pH, Ca, Mg, Fe, Mn, Cu, Zn and Total Organic Carbon, TOC). Methods and Results: A total of 137 hot water samples were analysed: 59 from the same number of private apartments, 46 from 11 hotels and 32 from five hospitals, all using the same water supply. Legionella species were detected in 40.0% of the distribution systems, L. pneumophila in 33.3%. The highest colonization was found in the hot water systems of hospitals (93.7% of samples positive for L. pneumophila, geometric mean: 2.4 x 10(3) CFU l(-1)), followed by the hotels (60.9%, geometric mean: 127.3 CFU l(-1)) and the apartments with centralized heating (41.9%, geometric mean: 30.5 CFU l(-1)). The apartments with independent heating systems showed a lower level of colonization (3.6% for Legionella species), with no evidence of L. pneumophila. Correlation analysis suggests that copper exerts an inhibiting action, while the TOC tends to favour the development of L. pneumophila. No statistically significant association was seen with Pseudomonadaceae, which were found at lower water temperatures than legionellae and in individual distribution points rather than in the whole network. Conclusions: The water recirculation system used by centralized boilers enhances the spreading of legionellae throughout the whole network, both in terms of the number of colonized sites and in terms of CFU count. Significance and Impact of the Study: Differences in Legionella colonization between types of buildings are not due to a variation in water supply but to other factors. Besides the importance of water recirculation, the study demonstrates the inhibiting action of copper and the favourable action of TOC on the development of L. pneumophila.


Legionella species diversity in an acidic biofilm community in yellowstone national park

Sheehan KB, Henson JM, Ferris MJ.

Division of Health Sciences, WWAMI Medical Program, 308 Leon Johnson Hall, Montana State University, P.O. Box 173080, Bozeman, MT 59717-3080.

Appl Environ Microbiol. 2005 Jan;71(1):507-11.

ABSTRACT: Legionella species are frequently detected in aquatic environments, but their occurrence in extreme, acidic, geothermal habitats has not been explored with cultivation-independent methods. We investigated a predominately eukaryotic algal mat community in a pH 2.7 geothermal stream in Yellowstone National Park for the presence of Legionella and potential host amoebae. Our analyses, using PCR amplification with Legionella-specific primers targeting 16S rRNA genes, detected four known Legionella species, as well as Legionella sequences from species that are not represented in sequence databases, in mat samples and cultivated isolates. The nonrandom occurrence of sequences detected at lower (30 degrees C) and higher (35 to 38 degrees C) temperatures suggests that natural thermal gradients in the stream influence Legionella species distributions in this mat community. We detected only one sequence, Legionella micdadei, from cultivated isolates. We cultured and sequenced partial 18S rRNA gene regions from two potential hosts, Acanthamoeba and Euglena species.


Intracellular proliferation of Legionella pneumophila in Hartmannella vermiformis in aquatic biofilms grown on plasticized polyvinyl chloride

Kuiper MW, Wullings BA, Akkermans AD, Beumer RR, van der Kooij D.

Laboratories of Food Microbiology, Wageningen University and Research Center, Nieuwegein, The Netherlands.

Appl Environ Microbiol. 2004 Nov;70(11):6826-33.

ABSTRACT: The need for protozoa for the proliferation of Legionella pneumophila in aquatic habitats is still not fully understood and is even questioned by some investigators. This study shows the in vivo growth of L. pneumophila in protozoa in aquatic biofilms developing at high concentrations on plasticized polyvinyl chloride in a batch system with autoclaved tap water. The inoculum, a mixed microbial community including indigenous L. pneumophila originating from a tap water system, was added in an unfiltered as well as filtered (cellulose nitrate, 3.0-microm pore size) state. Both the attached and suspended biomasses were examined for their total amounts of ATP, for culturable L. pneumophila, and for their concentrations of protozoa. L. pneumophila grew to high numbers (6.3 log CFU/cm2) only in flasks with an unfiltered inoculum. Filtration obviously removed the growth-supporting factor, but it did not affect biofilm formation, as determined by measuring ATP. Cultivation, direct counting, and 18S ribosomal DNA-targeted PCR with subsequent sequencing revealed the presence of Hartmannella vermiformis in all flasks in which L. pneumophila multiplied and also when cycloheximide had been added. Fluorescent in situ hybridization clearly demonstrated the intracellular growth of L. pneumophila in trophozoites of H. vermiformis, with 25.9% +/- 10.5% of the trophozoites containing L. pneumophila on day 10 and >90% containing L. pneumophila on day 14. Calculations confirmed that intracellular growth was most likely the only way for L. pneumophila to proliferate within the biofilm. Higher biofilm concentrations, measured as amounts of ATP, gave higher L. pneumophila concentrations, and therefore the growth of L. pneumophila within engineered water systems can be limited by controlling biofilm formation.


Microbiological evaluation of dental unit water systems in general dental practice in Europe

Walker JT, Bradshaw DJ, Finney M, Fulford MR, Frandsen E, OStergaard E, Ten Cate JM, Moorer WR, Schel AJ, Mavridou A, Kamma JJ, Mandilara G, Stosser L, Kneist S, Araujo R, Contreras N, Goroncy-Bermes P, O'Mullane D, Burke F, Forde A, O'Sullivan M, Marsh PD.

Health Protection Agency, Porton Down, Salisbury, UK.

Eur J Oral Sci. 2004 Oct;112(5):412-8.

ABSTRACT: A range of opportunistic pathogens have been associated with dental unit water systems (DUWS), particularly in the biofilms that can line the tubing. This study therefore aimed to assess the microbiology of DUWS and biofilms in general dental practices across seven European countries, including the United Kingdom (UK), Ireland (IRL), Greece (GR), Spain (ES), Germany (D), Denmark (DK) and the Netherlands (NL). Water supplied by 51% of 237 dental unit water lines exceeded current American Dental Association recommendations of </= 200 colony-forming units (CFU) ml(-1). Microbiological loading of the source waters was between 0 (Denmark, the Netherlands and Spain) and 4.67 (IRL) log CFU ml(-1); water line samples from the DUWS ranged from 1.52 (ES) to 2.79 (GR) log CFU ml(-1); and biofilm counts ranged from 1.49 (GR) to 3.22 (DK) log Opportunistic pathogens such as legionellae (DK and ES), including Legionella pneumophila SG1 (DK and GR), and Mycobacterium spp. (DK, NL, GR, D and ES) were recovered occasionally. Presumptive oral streptococci (ES and NL), oral anaerobes (GR), Candida spp. (UK, NL and ES) and blood (GR and IRL) were detected at relatively low frequencies, but their presence indicated a failure of the 3-in-1 antiretraction valve, leading to back siphonage of oral fluids into the water and biofilm phase. These findings confirm that a substantial proportion of DUWS have high levels of microbial contamination, irrespective of country, type of equipment and source water. The study emphasizes the need for effective mechanisms to reduce the microbial burden within DUWS, and highlights the risk of occupational exposure and cross-infection in general dental practice.

Detection and identification of legionella species from groundwaters

Brooks T, Osicki R, Springthorpe V, Sattar S, Filion L, Abrial D, Riffard S.

Centre for Research on Environmental Microbiology, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada.

J Toxicol Environ Health A. 2004;67(20-22):1845-1859.

ABSTRACT: Legionellae are opportunistic bacterial pathogens causing Legionnaires' disease and Pontiac fever and are ubiquitous in surface waters and in infrastructure to contain or distribute water, including pipes, cooling towers, and whirlpool spas. Infection in community-acquired and nosocomial outbreaks is by exposure to contaminated aerosols. Little is known about the presence of legionellae in groundwater. This study used samples from various locations in the United States and Canada to determine if legionellae could be isolated from water and biofilms derived from groundwaters not known to be under the direct influence of surface water. Of the 114 total samples of water and biofilm tested, 29.1% and 28.2% were positive for Legionella by cultivation and polymerase chain reaction (PCR), respectively. Legionellae were found in both warm and colder groundwaters, with more isolates from samples incubated at 30 degrees C than the 35 degrees C conventional temperature for Legionella isolation. The concentration of Legionella found in the water samples ranged from 102 to 105 CFU/L and up to 1.2 x 102 CFU/cm2 in the biofilm. The species of Legionella identified included both known pathogenic species and species that have not yet been identified as human pathogens. Millions of people in Canada, and around the world, rely on groundwater as their source for drinking. This study shows that legionellae are widespread in groundwater and have the potential to seed derived water supplies and biofilms in public distribution systems. This further widens the known sphere of Legionella colonization and the implications of its presence for public health.


Microbiological water quality in a large in-building, water recycling facility

Birks R, Colbourne J, Hills S, Hobson R.

Thames Water Innovation and Development, Manor Farm Road, Reading, RG2 0JN, UK.

Water Sci Technol. 2004;50(2):165-72.

ABSTRACT: The Thames Water recycling plant at the Millennium Dome, London, reclaimed three sources of water: greywater from the washbasins, rainwater from the Dome roof and groundwater from a borehole on site. These were pre-treated separately, and the mixed stream filtered using ultrafiltration and reverse osmosis membranes. Monitoring for indicator microorganisms was undertaken throughout the plant and in the reclaimed water distribution system, as well as ad-hoc monitoring for the presence of pathogens in the raw waters. Treatment to the level of ultrafiltration was more than adequate to produce a water quality meeting existing worldwide reclaimed water guidelines for toilet flushing. Owing to the excellent quality of the water leaving the plant, no significant microbiological growth was observed in the reclaimed water distribution system during the year. The raw greywater exhibited a higher faecal bacterial load than the rainwater and groundwater, as predicted from more human contact (i.e. hand washing). Environmental strains of Legionella were observed in the three raw greywater samples analysed for pathogens, as was Cryptosporidium, Giardia and faecal enterococci. The rainwater had relatively high levels of faecal bacteria, probably of avian origin. Giardia was detected in one rainwater sample confirming the potential for this water source to contain pathogens.

Isolation and characterization of a nonfluorescent strain of Legionella parisiensis

Igel L, Helbig JH, Luck PC.

Institut fur Medizinische Mikrobiologie und Hygiene, TU Dresden, Fiedlerstr. 42, D-01307 Dresden, Germany.

J Clin Microbiol. 2004 Jun;42(6):2877-8.



Legionella drancourtii sp. nov., a strictly intracellular amoebal pathogen

Scola BL, Birtles RJ, Greub G, Harrison TJ, Ratcliff RM, Raoult D.

Unite des Rickettsies, CNRS UPRESA 6020, Faculte de Medecine, Universite de la Mediterrannee, 27 Boulevard Jean Moulin, 13385 Marseille Cedex 05, France.

Int J Syst Evol Microbiol. 2004 May;54(Pt 3):699-703.

ABSTRACT: A Legionella-like amoebal pathogen (LLAP), formerly named LLAP12(T), was characterized on the basis of microscopic appearance, staining characteristics, growth in Acanthamoeba polyphaga at different temperatures, DNA G+C content, serological cross-reactivity and 16S rRNA and macrophage infectivity potentiator (mip) gene sequence analysis. LLAP12(T) was found to be a motile, Gram-negative bacterium that grew within cytoplasmic vacuoles in infected amoebae. The infecting bacteria induced lysis of their amoebal hosts and time taken to do so was dependent on incubation temperature. Recovery of LLAP12(T) from amoebae onto axenic media could not be achieved. Phylogenetic analysis of LLAP12(T), based on 16S rRNA and mip gene sequence analysis, indicated that it lay within the radiation of the Legionellaceae and that it clustered specifically with Legionella lytica and Legionella rowbothamii. The divergence observed between LLAP12(T) and these two species was of a degree equal to, or greater than, that observed between other members of the family. In support of this delineation, LLAP12(T) was found not to cross-react serologically with any other Legionella species. The mip and 16S rRNA gene sequence-based analyses also indicated that LLAP12(T) was related very closely to two other previously identified LLAP isolates, LLAP4 and LLAP11. Taken together, these results support the proposal of LLAP12(T) as the type strain of Legionella drancourtii sp. nov.


Rare occurrence of heterotrophic bacteria with pathogenic potential in potable water

Stelma GN Jr, Lye DJ, Smith BG, Messer JW, Payment P.

National Exposure Research Laboratory, U.S. Environmental Protection Agency, Cincinnati, OH 45268, USA.

Int J Food Microbiol. 2004 May 1;92(3):249-54.

ABSTRACT: Since the discovery of Legionella pneumophila, an opportunistic pathogen that is indigenous to water, microbiologists have speculated that there may be other opportunistic pathogens among the numerous heterotrophic bacteria found in potable water. The US Environmental Protection Agency (USEPA) developed a series of rapid in vitro assays to assess the virulence potential of large numbers of bacteria from potable water to possibly identify currently unknown pathogens. Results of surveys of potable water from several distribution systems using these tests showed that only 50 of the approximately 10,000 bacterial colonies expressed one or more virulence characteristics. In another study, 45 potable water isolates that expressed multiple virulence factors were tested for pathogenicity in immunocompromised mice. None of the isolates infected mice that were compromised either by treatment with carrageenan (CG), to induce susceptibility to facultative intracellular pathogens, or by cyclophosphamide (CY), to induce susceptibility to extracellular pathogens. These results indicate that there are very few potential pathogens in potable water and that the currently developed in vitro virulence screening tests give an overestimation of the numbers of heterotrophic bacteria that may be pathogens. Current efforts are focused on using the animal models to screen concentrated samples of waters known to contain large numbers of heterotrophic bacteria and newly discovered Legionella-like organisms that parasitize amoebae.


Microorganisms resistant to free-living amoebae

Greub G, Raoult D.

Unite des Rickettsies, Faculte de Medecine, Universite de la Mediterranee, Marseille, France.

Clin Microbiol Rev. 2004 Apr;17(2):413-33.

ABSTRACT: Free-living amoebae feed on bacteria, fungi, and algae. However, some microorganisms have evolved to become resistant to these protists. These amoeba-resistant microorganisms include established pathogens, such as Cryptococcus neoformans, Legionella spp., Chlamydophila pneumoniae, Mycobacterium avium, Listeria monocytogenes, Pseudomonas aeruginosa, and Francisella tularensis, and emerging pathogens, such as Bosea spp., Simkania negevensis, Parachlamydia acanthamoebae, and Legionella-like amoebal pathogens. Some of these amoeba-resistant bacteria (ARB) are lytic for their amoebal host, while others are considered endosymbionts, since a stable host-parasite ratio is maintained. Free-living amoebae represent an important reservoir of ARB and may, while encysted, protect the internalized bacteria from chlorine and other biocides. Free-living amoebae may act as a Trojan horse, bringing hidden ARB within the human "Troy," and may produce vesicles filled with ARB, increasing their transmission potential. Free-living amoebae may also play a role in the selection of virulence traits and in adaptation to survival in macrophages. Thus, intra-amoebal growth was found to enhance virulence, and similar mechanisms seem to be implicated in the survival of ARB in response to both amoebae and macrophages. Moreover, free-living amoebae represent a useful tool for the culture of some intracellular bacteria and new bacterial species that might be potential emerging pathogens.


Microbial Characterization of Free Floating Condensate aboard the Mir Space Station

Ott CM, Bruce RJ, Pierson DL.

EASI/Wyle Laboratories, Microbiology Laboratory, Johnson Space Center, 1290 Hercules Drive, 77058, Houston, TX, USA.

Microb Ecol. 2004 Feb;47(2):133-6.

ABSTRACT: Three samples of humidity condensate that had accumulated behind panels aboard the Russian space station Mir were collected and returned to earth for analysis. As these floating masses of liquid come into contact with the astronauts and the engineering systems, they have the potential to affect both crew health and systems performance. Using a combination of culturing techniques, a wide variety of organisms were isolated included Escherichia coli, Serratia marcescens, and a presumed Legionella species. In addition, microscopic analysis indicated the presence of protozoa, dust mites, and spirochetes. These findings suggest the need for more comprehensive microbial analysis of the environment through the use of new methodologies to allow a more thorough risk assessment of spacecraft


Investigation of natural biofilms formed during the production of drinking water from surface water embankment filtration

Emtiazi F, Schwartz T, Marten SM, Krolla-Sidenstein P, Obst U.

Department of Environmental Microbiology, Forschungszentrum Karlsruhe GmbH, Institute for Technical Chemistry-Water Technology and Geotechnology Division, P.O. Box 3640 , Karlsruhe D-76021, Germany .

Water Res. 2004 Mar;38(5):1197-206.

ABSTRACT: Populations of bacteria in biofilms from different sites of a drinking water production system were analysed. Polymerase chain reaction (PCR) and denaturing gradient gel electrophoresis (DGGE) analyses revealed changing DNA band patterns, suggesting a population shift during bank filtration and processing at the waterworks. In addition, common DNA bands that were attributed to ubiquitous bacteria were found. Biofilms even developed directly after UV disinfection (1-2m distance). Their DNA band patterns only partly agreed with those of the biofilms from the downstream distribution system. Opportunistic pathogenic bacteria in biofilms were analysed using PCR and Southern blot hybridisation (SBH). Surface water appeared to have a direct influence on the composition of biofilms in the drinking water distribution system. In spite of preceding filtration and UV disinfection, opportunistic pathogens such as atypical mycobacteria and Legionella spp. were found in biofilms of drinking water, and Pseudomonas aeruginosa was detected sporadically. Enterococci were not found in any biofilm. Bacterial cell counts in the biofilms from surface water to drinking water dropped significantly, and esterase and alanine-aminopeptidase activity decreased. [Formula: see text] -glucosidase activity was not found in the biofilms. Contrary to the results for planktonic bacteria, inhibitory effects were not observed in biofilms. This suggested an increased tolerance of biofilm bacteria against toxic compounds.


Health effects of Acanthamoeba spp. and its potential for waterborne transmission

Nwachuku N, Gerba CP.

Office of Science and Technology, Office of Water, U.S. Environmental Protection Agency, 1200 Pennsylvania Ave. N.W., Mc 4304T, Washington, DC 20460, USA.

Rev Environ Contam Toxicol. 2004;180:93-131

ABSTRACT: Risk from Acanthamoeba keratitis is complex, depending upon the virulence of the particular strain, exposure, trauma, or other stress to the eye, and host immune response. Bacterial endosymbionts may also play a factor in the pathogenicity of Acanthamoeba. Which factor(s) may be the most important is not clear. The ability of the host to produce IgA antibodies in tears may be a significant factor. The immune response of the host is a significant risk factor for GAE infection. If so, then a certain subpopulation with an inability to produce IgA in the tears may be at greatest risk. There was no sufficient data on the occurrence or types of Acanthamoeba in tapwater in the U.S. Published work on amoebal presence in tapwater does not provide information on the type of treatment the water received or the level of residual chlorine. Assessment of the pathogenicity by cell culture and molecular methods of Acanthamoeba in tapwater would also be useful in the risk assessment process for drinking water. The possibility that Acanthamoeba spp. might serve as vectors for bacterial infections from water sources also should be explored. The bacterial endosymbionts include an interesting array of pathogens such as Vibrio cholerae and Legionella pneumophila, both of which are well recognized waterborne/water-based pathogens. Work is needed to determine if control of Acanthamoeba spp. is needed to control water-based pathogens in water supplies.


Clinical and environmental distributions of Legionella strains in France are different

Doleans A, Aurell H, Reyrolle M, Lina G, Freney J, Vandenesch F, Etienne J, Jarraud S.

Laboratoire de Microbiologie, EA 3090, ISPB, Universite Claude Bernard Lyon 1, 69373 Lyon, France.

J Clin Microbiol. 2004 Jan;42(1):458-60.

ABSTRACT: In France, the clinical distribution of Legionella species and serogroups does not correspond to their environmental distribution. Legionella pneumophila serogroup 1 is more prevalent among clinical isolates (95.4%) than in the environment (28.2%), whereas L. anisa is more frequent in the environment (13.8%) than in the clinical setting (0.8%).

Relationship between mineral content of domestic hot water and microbial contamination.

Borella P, Montagna MT, Romano-Spica V, Stampi S, Stancanelli G, Triassi M, Marchesi I, Bargellini A, Neglia R, Paglionico N, Spilotros G, Moscato U, Casati G, Legnani PP, Sacchetti R, Ossi C, Moro M, Ribera G.

Department of Hygiene, Microbiology and Biostatistics, University of Modena, Modena, Italy.

J Trace Elem Med Biol, 2003; 17 (suppl.1): 37-43.

ABSTRACT: The relationship between Legionella and Pseudomonas spp contamination and mineral content of domestic hot waters was investigated in a cross-sectional Italian survey. Pseudomonas spp contamination was associated with elevated Ca, but lower Fe and free chlorine content in the water. Waters with Cu concentrations > 50 microg/L prevented Legionella spp colonisation (OR = 0.14, 95% CI = 0.02-1.13), whereas the risk of legionellae occurrence increased in waters with Mn levels > 3 microg/L (OR = 2.37, 95% CI = 1.06-5.30). Furthermore, Mn was positively associated with the risk of colonisation by eterotrophic bacteria growing at either 22 and 36 degrees C. Legionella species and serogroups were differently distributed according to Cu and Mn concentrations, suggesting that Legionella strains may have a different sensibility/resistance to trace elements. A specific action of Cu as decontamination factor is suggested and the consideration of Mn as a risk indicator for bacterial colonisation and biofilm presence is proposed.


Morphology of Legionella pneumophila according to their location within Hartmanella vermiformis

Greub G, Raoult D.

Unite des Rickettsies, Faculte de Medecine, Universite de la Mediterranee, 27, Boulevard Jean Moulin, 13385, Marseille, France.

Res Microbiol. 2003 Nov;154(9):619-21

ABSTRACT: The morphology of Legionella pneumophila grown within Hartmanella vermiformis was studied by electron microscopy. Different morphologies were observed, including a replicative form and a mature intracellular form, that had previously been reported in Hela cells. L. pneumophila was also detected within the cyst wall of the amoeba.


Isolation of Legionella anisa from multiple sites of a hospital water system: the eradication of Legionella contamination

Yamamoto N, Kubota T, Tateyama M, Koide M, Nakasone C, Tohyama M, Shinzato T, Higa F, Kusano N, Kawakami K, Saito A.

First Department of Internal Medicine, University of the Ryukyus, Okinawa, Japan.

J Infect Chemother. 2003 Jun;9(2):122-5.

ABSTRACT: For the prevention of nosocomial cases of legionellosis in the Ryukyu University Hospital neonatal wards, we examined nine shower units and a sink tap water unit for the presence of Legionella, over a 6-year period. We isolated Legionella-like organisms (LLO) from showerheads by culturing sediments from the water samples on buffered charcoal yeast extracts (BCYE). We used DNA-DNA hybridization to determine that the organisms were L. anisa. A fingerprinting technique called random amplified polymorphism DNA (RAPD) also showed that all the organisms were identical at the genome level. Replacement of the shower heads harboring colonies of L. anisa prevented further contamination. Nosocomial cases of legionellosis have not been reported from the wards during the period of this survey. This is the first description of the isolation of L. anisa from multiple sites within a hospital, and RAPD analysis suggests that these may be the spread of a single clone.


Factors influencing survival of Legionella pneumophila serotype 1 in hot spring water and tap water

Ohno A, Kato N, Yamada K, Yamaguchi K.

Departments of Microbiology. Chemistry, Toho University School of Medicine, Tokyo.

Department of Chemistry, Faculty of Science, Toho University , Chiba , Japan.

Appl Environ Microbiol. 2003 May; 69(5): 2540-7.

ABSTRACT: The factors involved in the survival of Legionella pneumophila in the microcosms of both hot spring water and tap water were studied by examining cultivability and metabolic activity. L. pneumophila could survive by maintaining metabolic activity but was noncultivable in all microcosms at 42 degrees C, except for one microcosm with a pH of <2.0. Lower temperatures supported survival without loss of cultivability. The cultivability declined with increasing temperature, although metabolic activity was observed at temperatures of up to 45 degrees C. The optimal range of pH for survival was between 6.0 and 8. The metabolic activity could be maintained for long periods even in microcosms with high concentrations of salt. The cultivability of organisms in the post-exponential phase in a tap water microcosm with a low inoculum size was more rapidly reduced than that of organisms in the exponential phase. In contrast, the loss of cultivability in microcosms of a high inoculum size was significant in the exponential phase. Random(ly) amplified polymorphic DNA analysis of microcosms where cultivability was lost but metabolic activity was retained showed no change compared to cells grown freshly, although an effect on the amplified DNA band pattern by production of stress proteins was expected. Resuscitation by the addition of Acanthamoeba castellanii to the microcosm in which cultivability was completely lost but metabolic activity was maintained was observed only in part of the cell population. Our results suggest that L. pneumophila cell populations can potentially survive as free organisms for long periods by maintaining metabolic activity but temporarily losing cultivability under strict environments and requiring resuscitation by ingestion by amoebas.