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. 2013 Jan 28:13:17.
doi: 10.1186/1471-2180-13-17.

Molecular diversity and high virulence of Legionella pneumophila strains isolated from biofilms developed within a warm spring of a thermal spa

Zineddine Chaabna  1 , Françoise Forey, Monique Reyrolle, Sophie Jarraud, Danièle Atlan, Dominique Fontvieille, Christophe Gilbert
Affiliations

Molecular diversity and high virulence of Legionella pneumophila strains isolated from biofilms developed within a warm spring of a thermal spa

Zineddine Chaabna et al. BMC Microbiol. .

Abstract

Background: Several cases of legionellosis have been diagnosed in the same French thermal spa in 1986, 1994 and 1997. L. pneumophila serogroup 1 (Lp1) strains have been isolated from several patients, but the source of contamination was not identified despite the presence of different Lp1 in water samples of the three natural springs feeding the spa at this period.

Results: Our strategy was to investigate L. pneumophila (Lp) strains from natural biofilms developed in a sulphur-rich warm spring of this contaminated site. Biofilm analysis revealed the presence of three Lp serogroups (Lp1, Lp10 and Lp12). Surprisingly, Lp10 and Lp12 were not reported in the previous described studies from water samples. Besides, the new seven Lp1 we isolated exhibit a high molecular diversity and have been differentiated in five classes according to their DNA genome patterns obtained by PFGE and mip sequences. It must be noted that these DNA patterns are original and unknown in databases. Interestingly, the 27 Lp environmental strains we isolated display a higher cytotoxicity and virulence towards the amoeba Acanthamoeba castellanii than those of known Lp1 epidemic strains.

Conclusion: The characteristics of Legionella pneumophila Lp1 strains isolated from the warm spring are in agreement with their presence in biofilms and their probable long-term persistence in this ecosystem.

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Figures

Figure 1
Figure 1
Examples of PCR Amplification of several Legionella pneumophila genes: lpg0774, lpg1905,wzm and mip. The ladder was the GeneRuler 1kb DNA ladder (Fermentas SM0311).
Figure 2
Figure 2
Legionella pneumophila typing. The dendogramm represents the relationships of environmental and clinical strains of Legionella pneumophila. Patterns were generated by pulse field gel electrophoresis (PFGE) of total bacterial DNA and then clustered by unweighted pair group method with arithmetic averages algorithm.
Figure 3
Figure 3
Phylogenetic tree (Neighbor-joining) of mip sequences from L. pneumophila sg 1 clinical and environmental (mip1, mip2 and mip3) strains and L. non-pneumophila strains.
Figure 4
Figure 4
Quantification of cytotoxicity and virulence of environmental L. pneumophila strains towards the amoeba Acanthamoeba castellanii. Lp1 dotA : dotA mutant of Lp1 Lens; Lp1 clin: means of cytotoxicities (a) and virulences (c) of three clinical Lp1 strains (Lens, Paris and Lorraine). These means of cytotoxicities (b) and virulences (d) of three clinical Lp1 strains were compared to those of five independent pulsotypes (PST1 to PST5) of environmental Lp1 strains.
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