A community outbreak of Legionnaires' disease: evidence of a cooling tower as the source

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.

mentioning

confidence: 99%

High Diversity and Abundance of Legionella spp. in a Pristine River and Impact of Seasonal and Anthropogenic Effects

Parthuisot

West

Lebaron

et al. 2010

“…L. pneumophila is more frequently isolated from man-made water systems with high water temperature (9,17) than from relatively cold freshwater environments (4,16). This fact suggests that thermal conditions affect the relationship between L. pneumophila and protozoa, a notion supported by some reports (1,3,7,11).…”

mentioning

confidence: 99%

Temperature-Dependent Parasitic Relationship between Legionella pneumophila and a Free-Living Amoeba ( Acanthamoeba castellanii )

Ohno¹,

Kato

Sakamoto

et al. 2008

We analyzed the effects of temperature on the interaction of Legionella pneumophila with Acanthamoeba castellanii. At <20°C, overexpression of type 1 metacaspase, a stimulator of A. castellanii encystation, was associated with a reduced number of bacteria within amoeba. At low temperatures, A. castellanii seems to eliminate L. pneumophila by encystation and digestion.The intracellular pathogen Legionella pneumophila causes Legionnaires' disease and exploits aquatic protozoa for replication. L. pneumophila is more frequently isolated from man-made water systems with high water temperature (9, 17) than from relatively cold freshwater environments (4,16). This fact suggests that thermal conditions affect the relationship between L. pneumophila and protozoa, a notion supported by some reports (1, 3, 7, 11). The trophozoite of protozoa transforms into a cyst under harmful environments such as starvation, cold, and certain chemicals used in medical treatment. The effect of protozoal encystation on Legionella infection is not well understood although it seems that encystation is enhanced in a freshwater environment at low temperature. With these considerations in mind, we investigated whether the host-parasite relationship between L. pneumophila and protozoa is temperature dependent.Two strains of L. pneumophila serogroup 1, Suzuki and Lp01, and Acanthamoeba castellanii ATCC 30234 were used in the present study. Intracellular growth kinetics assays using A. castellanii were performed as described previously (15 Total RNA was purified using an RNeasy minikit (Qiagen, Valencia, CA) according to the instructions provided by the manufacturer. Total RNA was reverse transcribed into cDNA using an RNA PCR kit (Takara Bio Inc., Shiga, Japan). Real-time PCR with Sybr greenER (Invitrogen Life Sciences) was performed using the ABI Prism 7000 system (Applied Biosystems, Foster City, CA). Table 1 lists the primer pairs for target and internal control genes. Data were analyzed by Student's t test, and a two-tailed P value of Ͻ0

“…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.The involvement of cooling towers in some community outbreaks of legionellosis has been demonstrated (3,6,8,9,11), and it has been suggested that there may be an association between high Legionella counts in cooling towers and the occurrence of an outbreak of legionellosis (12). Although not all environmental isolates are associated with human disease, the present guidelines aimed at preventing legionellosis have proposed ranges of risk involving different environmental insults based on Legionella counts (4).…”

mentioning

confidence: 99%

Legionella pneumophila in Cooling Towers: Fluctuations in Counts, Determination of Genetic Variability by Pulsed-Field Gel Electrophoresis (PFGE), and Persistence of PFGE Patterns

Ragull¹,

García-Núñez²,

Pedro‐Botet³

et al. 2007

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.The involvement of cooling towers in some community outbreaks of legionellosis has been demonstrated (3,6,8,9,11), and it has been suggested that there may be an association between high Legionella counts in cooling towers and the occurrence of an outbreak of legionellosis (12). Although not all environmental isolates are associated with human disease, the present guidelines aimed at preventing legionellosis have proposed ranges of risk involving different environmental insults based on Legionella counts (4). However, the fluctuations in Legionella counts over short periods of time have led some authors to question these risk ranges (1).Since the recovery of a high inoculum of Legionella in a cooling tower is not necessarily related to an outbreak or sporadic cases of legionellosis, genetic studies are necessary to establish the link between environmental and clinical isolates. The "gold standard" technique for genotyping Legionella is the analysis of chromosomal restriction patterns by pulsed-field gel electrophoresis (PFGE). Isolation of Legionella strains from a cooling tower which are genetically indistinguishable from clinical isolates is considered to be clear evidence of the implication of the tower in an outbreak (1, 9). However, little is known regarding the genetic variability of Legionella within the same tower and in cooling towers from areas in the vicinity. Likewise, to our knowledge, no study has determined the persistence of molecular patterns in isolates in cooling towers. All these circumstances have epidemiological and legal consequences when assigning responsibility for an outbreak of legionellosis to a particular installation.The main objectives of this study were to establish the fluctuations in Legionella counts over time and to determine the genetic variability and persistence of PFGE patterns in cooling towers.Fifteen cooling towers (A to O) were selected within a radius of 3 km. Four of them (J, K, M, and N) shared the same main line for water distribution. The other cooling towers had a different domestic distribution system but were supplied by the same municipal water network. All the cooling towers followed the controls and hygiene measures required by Spanish regulations (RD 865/2003), and we were able to verify that the levels of disinfectant (sodium hypochlorite [0.2 to 0.5 ppm residual] in conjunction with the discontinuous dosage of nonoxidant biocide [dibromonitrilopropionamide]) were in accordance with legislation at the time of sampling. Cooling water samples were cultured for Legionella fortnightly over a 1-year period. The samples were concentrated by filtration, and four plates of GVPC agar (Oxoid, Wesel, Germany) we...