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Background    Legionella  is the causative agent of Legionnaires' disease, and hot springs are a major source of outbreaks of this disease. It is important from a public health perspective to survey hot spring environments for the presence of  Legionella .    Methods  Prospective surveillance of the extent of  Legionella  pollution was conducted at three hot spring recreational areas in Beijing, China in 2011. Pulsed-field gel electrophoresis (PFGE) and sequence-based typing (SBT) were used to describe the genetic polymorphism of isolates. The intracellular growth ability of the isolates was determined by interacting with J774 cells and plating the dilutions onto BCYE agar plates.    Results  Overall, 51.9% of spring water samples showed  Legionella- positive, and their concentrations ranged from 1 CFU/liter to 2,218 CFU/liter. The positive rates of  Legionella  were significantly associated with a free chlorine concentration of ≥0.2 mg/L, urea concentration of ≥0.05 mg/L, total microbial counts of ≥400 CFU/ml and total coliform of ≥3 MPN/L (p<0.01). The  Legionella  concentrations were significantly associated with sample temperature, pH, total microbial counts and total coliform (p<0.01).  Legionella pneumophila  was the most frequently isolated species (98.9%), and the isolated serogroups included serogroups 3 (25.3%), 6 (23.4%), 5 (19.2%), 1 (18.5%), 2 (10.2%), 8 (0.4%), 10 (0.8%), 9 (1.9%) and 12 (0.4%). Two hundred and twenty-eight isolates were analyzed by PFGE and 62 different patterns were obtained. Fifty-seven  L. pneumophila  isolates were selected for SBT analysis and divided into 35 different sequence types with 5 main clonal groups. All the 57 isolates had high intracellular growth ability.    Conclusions  Our results demonstrated high prevalence and genetic polymorphism of  Legionella  in springs in Beijing, China, and the SBT and intracellular growth assay results suggested that the  Legionella  isolates of hot spring environments were pathogenic. Improved control and prevention strategies are urgently needed.

High Prevalence, Genetic Diversity and Intracellular Growth Ability of Legionella in Hot Spring Environments