Pulsed-Field Gel Electrophoresis (PFGE) Pattern Analysis and Chlorine-Resistance of Legionella pneumophila Isolated from Hot Spring Water Samples

For a microbial ecological analysis, 20 strains of Legionella pneumophila isolated from both unchlorinated Noyu (unattended natural hot spring) samples and chlorinated hot spring bath water samples collected throughout Japan were subjected to a clustering analysis on the basis of a Pulsed-Field Gel Electrophoresis (PFGE) pattern analysis. The PFGE patterns obtained from 19 strains of L. pneumophila after digestion with Sfil were used to divide the strains into two groups (Groups A and B), although the similarity level was very low among the groups. Group A consisted of 8 strains, and all of these strains were isolated from hot spring bath water samples. Group B consisted of 11 strains, and all but two of these strains were isolated from Noyu samples. The chlorine resistance (99.9% CT values) of these isolates was experimentally confirmed, and we attempted to define the relationship between chlorine resistance and the geno-cluster. The average CT value of group A (8 strains from hot spring bath water) was 0.49 mg center dot min/1 and the average of group B (9 strains from Noyu samples) was 0.29 mg center dot min/1. No remarkable differences in the CT values for the groups were found. A chlorine-sensitive Noyu strain (0.14 mg center dot min/1) and a chlorine-resistant strain (0.62 mg center dot min/1) from hot spring bath water were then compared to identify any differences in their lipid composition. There was no notable difference in the ratio of saturated to unsaturated fatty acids between the chlorinesensitive and chlorine-resistant bacteria. However, the chlorine-sensitive and chlorine-resistant bacteria demonstrated differences in the relative percentages of cell wall and cell membrane fatty acids.