New endemic Legionella pneumophila serogroup I clones, Ontario, Canada

Abstract

The water-borne pathogen Legionella pneumophila serogroup 1 (Lp1) is the most commonly reported etiologic agent of legionellosis. To examine the genetic diversity, the long-term epidemiology, and the molecular evolution of Lp1 clinical isolates, we conducted sequence-based typing on a collection of clinical isolates representing 3 decades of culture-confirmed legionellosis in Ontario, Canada. Analysis showed that the population of Lp1 in Ontario is highly diverse and combines lineages identified worldwide with local strains. Identical types were identified in sporadic and outbreak-associated strains. In the past 15 years, the incidence of some lineages distributed worldwide has tended to decrease, and local endemic clones and lineages have emerged. Comparative geographic distribution analysis suggests that some lineages are specific to eastern North America. These findings have general clinical implications for the study of Lp1 molecular evolution and for the identification of Lp1 circulating strains in North America.

Figure 1

Phylogenetic analysis of flaA, pilE, asd, mip, mompS, proA, and neuA concatenated sequences from the 62 Legionella pneumophila serogroup 1 sequence types (STs) identified in Ontario. The tree was constructed with ClustalW2 (www.ebi.ac.uk/Tools/clustalw2/index.html) and the neighbor-joining method with 1,000 bootstrap replicates. Scale bar indicates genetic distances between sequences. STs in boldface were detected in outbreaks.

Figure 2

Dendrogram created by the unweighted pair group method with arithmetic mean method based on the 62 allelic profiles of 194 Legionella pneumophila serogroup 1 isolates. Clonal groups (CGs) identified by eBURST (http://eBURST.mlst.net) are indicated with solid lines, and STs included in CGs are in boldface. Ontario STs included in clonal complexes (CC) identified by comparative eBURST analysis with the European Working Group for Legionella Infections database are indicated with dashed lines. The 3 major clusters are indicated on the right of the figure with bold lines. The number of strains isolated in Ontario is indicated below CG. Scale bar indicates linkage distances.

Figure 3

Representation of Legionella pneumophila serogroup 1 clonal complexes (CC) A, B, C, D obtained by comparative eBURST (http://eBURST.mlst.net) analysis between the Ontario collection and the European Working Group for Legionella Infections database. Each circle represents a single sequence type (ST). Size of the circle is proportional to the number of isolates. Dark circles represent predicted founder of each CC. Labels in boldface indicate STs found in both datasets, regular black characters indicate STs absent from the Ontario collection, light gray characters indicate STs exclusively found in Ontario. Solid lines represent single-locus variants.

Figure 4

Prevalence of Legionella pneumophila serogroup 1 sequence type 1 (ST1) (A) and ST211 (B) endemic strains in Ontario. Black bar sections indicate proportion of strains from isolated cases and white bar sections indicate proportion of isolates from outbreaks.

Figure 5

Incidence of Ontario Legionella pneumophila serogroup 1 isolates from clonal complexes (CC) A and C. CC-A (A) and CC-C (B) were identified by eBURST (http://eBURST.mlst.net) comparative analysis using the Ontario and the European Working Group for Legionella Infections international databases. Black bar sections indicate proportion of strains isolated during sporadic cases. White bar sections indicate proportion of outbreak isolates.

Figure 6

Geographic distribution of phylogenetic clusters II and I/III from 1990 through 2007. Rates are cases of infection with Legionella pneumophilia serogroup 1 clones per 100,000 persons per year. The province of Ontario was divided into 7 health regions (OHRs) with populations ranging from ≈0.5 to 2 million persons: Toronto, South West (SW), Central South (CS), Central West (CW), Central East (CE), East, and North.