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Genomic characterization of a large outbreak of Legionella pneumophila serogroup 1 strains in Quebec City, 2012.

Lévesque S, Plante PL, Mendis N, Cantin P, Marchand G, Charest H, Raymond F, Huot C, Goupil-Sormany I, Desbiens F, Faucher SP, Corbeil J, Tremblay C - PLoS ONE (2014)

Bottom Line: Two new Legionellaceae plasmids were found only in the epidemic strain.The LVH type IV secretion system was found in the 2012 outbreak isolates but not in the ones from the 1996 outbreak and only in half of the contemporary human isolates.At least four Icm/Dot effectors in the epidemic strains were absent in the environmental strains suggesting that some effectors could impact the intracellular replication in human macrophages.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Santé Publique du Québec (LSPQ)/Institut National de Santé Publique du Québec, Québec, Canada.

ABSTRACT
During the summer of 2012, a major Legionella pneumophila serogroup 1 outbreak occurred in Quebec City, Canada, which caused 182 declared cases of Legionnaire's disease and included 13 fatalities. Legionella pneumophila serogroup 1 isolates from 23 patients as well as from 32 cooling towers located in the vicinity of the outbreak were recovered for analysis. In addition, 6 isolates from the 1996 Quebec City outbreak and 4 isolates from patients unrelated to both outbreaks were added to allow comparison. We characterized the isolates using pulsed-field gel electrophoresis, sequence-based typing, and whole genome sequencing. The comparison of patients-isolated strains to cooling tower isolates allowed the identification of the tower that was the source of the outbreak. Legionella pneumophila strain Quebec 2012 was identified as a ST-62 by sequence-based typing methodology. Two new Legionellaceae plasmids were found only in the epidemic strain. The LVH type IV secretion system was found in the 2012 outbreak isolates but not in the ones from the 1996 outbreak and only in half of the contemporary human isolates. The epidemic strains replicated more efficiently and were more cytotoxic to human macrophages than the environmental strains tested. At least four Icm/Dot effectors in the epidemic strains were absent in the environmental strains suggesting that some effectors could impact the intracellular replication in human macrophages. Sequence-based typing and pulsed-field gel electrophoresis combined with whole genome sequencing allowed the identification and the analysis of the causative strain including its likely environmental source.

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Production of biofilms by L. pneumophila serogroup 1 strains.One strain from patient (ID119442), one strain from the outbreak source's cooling tower (ID120292), and three from environment (other that the outbreak source) (ID120145, ID120086 and ID 120329) were tested. Star represent difference statistically significant (p = 0.0065) with control strain JR32. Error bars represent standard deviations derived from four independent experiments.
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pone-0103852-g003: Production of biofilms by L. pneumophila serogroup 1 strains.One strain from patient (ID119442), one strain from the outbreak source's cooling tower (ID120292), and three from environment (other that the outbreak source) (ID120145, ID120086 and ID 120329) were tested. Star represent difference statistically significant (p = 0.0065) with control strain JR32. Error bars represent standard deviations derived from four independent experiments.

Mentions: WGS was performed to characterize the strains for putative virulence markers that could have contributed to the major Quebec City Legionella outbreak. The epidemic strain harbored a 50.5 kb plasmid comprising 55 genes and a 144 kb plasmid harboring 158 genes. The smaller one had two sections similar to two plasmids from Fluoribater dumofii plasmids: pLDNY1 and pLD-TEX-KL. It mainly contained genes for pili formation and conjugation with 25% hypothetical genes. The bigger one was similar to pLELO plasmid from Legionella pneumophila. It contained genes associated with biological and metabolic processes. The longest plasmid also harboured genes associated with drug resistance: an erythromycin ABC transporter ATP-binding protein (similar to gene lpg1616) and a beta-lactamase AmpS (similar to gene LPC_1045). However no clinical resistance to these antibiotics was detected in the patient strains (data not shown). The larger plasmid had nearly 50% hypothetical genes. Type II and Dot/Icm Type IV secretion system were present in all strains. LVH Type IV secretion system was found in all strains except for those unrelated to the outbreak patient strains and 1996 strains. We have identified at least 4 Icm/Dot effectors in the epidemic strain that were absent in environmental strains (Table 2), from a list of 140 effectors found in Lp1 strains. We also found that gene lpg1057 was missing in the outbreak strain's genome but was present in environmental strains (Table 1). Recently, it was shown that lpg1057 is linked with biofilm formation [21]. We performed a biofilm formation assay measured by crystal violet staining to confirm that strains lacking lpg1057 were producing less biofilm than other strains harboring the gene. Only the strain ID120292 from the environmental source had a significantly lower level of biofilm production (p = 0.0065) (Figure 3).


Genomic characterization of a large outbreak of Legionella pneumophila serogroup 1 strains in Quebec City, 2012.

Lévesque S, Plante PL, Mendis N, Cantin P, Marchand G, Charest H, Raymond F, Huot C, Goupil-Sormany I, Desbiens F, Faucher SP, Corbeil J, Tremblay C - PLoS ONE (2014)

Production of biofilms by L. pneumophila serogroup 1 strains.One strain from patient (ID119442), one strain from the outbreak source's cooling tower (ID120292), and three from environment (other that the outbreak source) (ID120145, ID120086 and ID 120329) were tested. Star represent difference statistically significant (p = 0.0065) with control strain JR32. Error bars represent standard deviations derived from four independent experiments.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4126679&req=5

pone-0103852-g003: Production of biofilms by L. pneumophila serogroup 1 strains.One strain from patient (ID119442), one strain from the outbreak source's cooling tower (ID120292), and three from environment (other that the outbreak source) (ID120145, ID120086 and ID 120329) were tested. Star represent difference statistically significant (p = 0.0065) with control strain JR32. Error bars represent standard deviations derived from four independent experiments.
Mentions: WGS was performed to characterize the strains for putative virulence markers that could have contributed to the major Quebec City Legionella outbreak. The epidemic strain harbored a 50.5 kb plasmid comprising 55 genes and a 144 kb plasmid harboring 158 genes. The smaller one had two sections similar to two plasmids from Fluoribater dumofii plasmids: pLDNY1 and pLD-TEX-KL. It mainly contained genes for pili formation and conjugation with 25% hypothetical genes. The bigger one was similar to pLELO plasmid from Legionella pneumophila. It contained genes associated with biological and metabolic processes. The longest plasmid also harboured genes associated with drug resistance: an erythromycin ABC transporter ATP-binding protein (similar to gene lpg1616) and a beta-lactamase AmpS (similar to gene LPC_1045). However no clinical resistance to these antibiotics was detected in the patient strains (data not shown). The larger plasmid had nearly 50% hypothetical genes. Type II and Dot/Icm Type IV secretion system were present in all strains. LVH Type IV secretion system was found in all strains except for those unrelated to the outbreak patient strains and 1996 strains. We have identified at least 4 Icm/Dot effectors in the epidemic strain that were absent in environmental strains (Table 2), from a list of 140 effectors found in Lp1 strains. We also found that gene lpg1057 was missing in the outbreak strain's genome but was present in environmental strains (Table 1). Recently, it was shown that lpg1057 is linked with biofilm formation [21]. We performed a biofilm formation assay measured by crystal violet staining to confirm that strains lacking lpg1057 were producing less biofilm than other strains harboring the gene. Only the strain ID120292 from the environmental source had a significantly lower level of biofilm production (p = 0.0065) (Figure 3).

Bottom Line: Two new Legionellaceae plasmids were found only in the epidemic strain.The LVH type IV secretion system was found in the 2012 outbreak isolates but not in the ones from the 1996 outbreak and only in half of the contemporary human isolates.At least four Icm/Dot effectors in the epidemic strains were absent in the environmental strains suggesting that some effectors could impact the intracellular replication in human macrophages.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Santé Publique du Québec (LSPQ)/Institut National de Santé Publique du Québec, Québec, Canada.

ABSTRACT
During the summer of 2012, a major Legionella pneumophila serogroup 1 outbreak occurred in Quebec City, Canada, which caused 182 declared cases of Legionnaire's disease and included 13 fatalities. Legionella pneumophila serogroup 1 isolates from 23 patients as well as from 32 cooling towers located in the vicinity of the outbreak were recovered for analysis. In addition, 6 isolates from the 1996 Quebec City outbreak and 4 isolates from patients unrelated to both outbreaks were added to allow comparison. We characterized the isolates using pulsed-field gel electrophoresis, sequence-based typing, and whole genome sequencing. The comparison of patients-isolated strains to cooling tower isolates allowed the identification of the tower that was the source of the outbreak. Legionella pneumophila strain Quebec 2012 was identified as a ST-62 by sequence-based typing methodology. Two new Legionellaceae plasmids were found only in the epidemic strain. The LVH type IV secretion system was found in the 2012 outbreak isolates but not in the ones from the 1996 outbreak and only in half of the contemporary human isolates. The epidemic strains replicated more efficiently and were more cytotoxic to human macrophages than the environmental strains tested. At least four Icm/Dot effectors in the epidemic strains were absent in the environmental strains suggesting that some effectors could impact the intracellular replication in human macrophages. Sequence-based typing and pulsed-field gel electrophoresis combined with whole genome sequencing allowed the identification and the analysis of the causative strain including its likely environmental source.

Show MeSH
Related in: MedlinePlus