Genome Sequencing Links Persistent Outbreak of Legionellosis in Sydney (New South Wales, Australia) to an Emerging Clone of Legionella pneumophila Sequence Type 211

Abstract

The city of Sydney, Australia, experienced a persistent outbreak of serogroup 1 (Lp1) pneumonia in 2016. To elucidate the source and guide public health actions, the genomes of clinical and environmental Lp1 isolates recovered over 7 weeks were examined. A total of 48 isolates from human cases and cooling towers were sequenced and compared using single-nucleotide polymorphism (SNP)-based core-genome multilocus sequencing typing (MLST) and pangenome approaches. All three methods confirmed phylogenetic relatedness between isolates associated with outbreaks in the Central Business District (CBD) in March and May and those in suburb 1. These isolates were designated the "main cluster" and consisted of isolates from two patients from the CBD March outbreak, one patient and one tower isolate from suburb 1, and isolates from two cooling towers and three patients from the CBD May outbreak. All main cluster isolates were sequence type 211 (ST211), which previously has only been reported in Canada. Significantly, pangenome analysis identified mobile genetic elements containing a unique type IV A F-type secretion system (T4ASS), which was specific to the main cluster, and cocirculating clinical strains, suggesting a potential mechanism for increased fitness and persistence of the outbreak clone. Genome sequencing enabled linking of the geographically dispersed environmental sources of infection among the spatially and temporally coinciding cases of legionellosis in a highly populated urban setting. The discovery of a unique T4ASS emphasizes the role of genome recombination in the emergence of successful Lp1 clones. A new emerging clone has been responsible for a prolonged legionellosis outbreak in Sydney, Australia. The use of whole-genome sequencing linked two outbreaks thought to be unrelated and confirmed the outliers. These findings led to the resampling and subsequent identification of the source, guiding public health actions and bringing the outbreak to a close. Significantly, the outbreak clone was identified as sequence type 211 (ST211). Our study reports this ST in the Southern Hemisphere and presents a description of ST211 genomes from both clinical and environmental isolates. A unique mobile genetic element containing a type IV secretion system was identified in Lp1 ST211 isolates linked to the main cluster and Lp1 ST42 isolates that were cocirculating at the time of the outbreak.

Contaminant DNA in bacterial sequencing experiments is a major source of false genetic variability

“…The datasets comprised 8 MTB studies and 12 studies of other 13 relevant pathogenic species. Nineteen of these datasets were publicly available beforehand [34][35][36][37][38][39][40][41][42][43][44][45][46][47][48][49][50][51][52]. To include a dataset generated in our laboratory, we sequenced 138 MTB samples from Mozambique in the Illumina MiSeq platform.…”

Contaminant DNA in bacterial sequencing experiments is a major source of false genetic variability

“…The datasets comprised 8 MTB studies and 12 studies of other 13 relevant pathogenic species. Nineteen of these datasets were publicly available beforehand (22)(23)(24)(25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39)(40). To include a dataset generated in our laboratory, we sequenced 138 MTB samples from Mozambique in the Illumina MiSeq platform (Supplementary Methods 1).…”

Complete microbial genomes for public health in Australia and the Southwest Pacific

“…However, pathogenic bacteria are not generally composed of uniform panmictic populations. Instead, they represent numerous diverse clades, with many being endemic to particular regions or jurisdictions [17–23]. We define the latter as clones that are repeatedly observed in a given region with evidence of ongoing local transmission, but are not commonly observed in other parts of the world; a more practical definition is given by clones observed in local outbreaks for which no suitable reference genome is available in the public domain.…”