Evidence of microevolution of Salmonella Typhimurium during a series of egg-associated outbreaks linked to a single chicken farm

Abstract

BackgroundThe bacterium Salmonella enterica serovar Typhimurium (S. Typhimurium) is one of the most frequent causes of foodborne outbreaks of gastroenteritis. Between 2005–2008 a series of S. Typhimurium outbreaks occurred in Tasmania, Australia, that were all traced to eggs originating from a single chicken farm. We sequenced the genomes of 12 isolates linked to these outbreaks, in order to investigate the microevolution of a pathogenic S. Typhimurium clone in a natural, spatiotemporally restricted population.ResultsThe isolates, which shared a phage type similar to DT135 known locally as 135@ or 135a, formed a clade within the S. Typhimurium population with close similarity to the reference genome SL1334 (160 single nucleotide polymorphisms, or SNPs). Ten of the isolates belonged to a single clone (<23 SNPs between isolate pairs) which likely represents the population of S. Typhimurium circulating at the chicken farm; the other two were from sporadic cases and were genetically distinct from this clone. Divergence dating indicated that all 12 isolates diverged from a common ancestor in the mid 1990s, and the clone began to diversify in 2003–2004. This clone spilled out into the human population several times between 2005–2008, during which time it continued to accumulate SNPs at a constant rate of 3–5 SNPs per year or 1x10-6 substitutions site-1 year-1, faster than the longer-term (~50 year) rates estimated previously for S. Typhimurium. Our data suggest that roughly half of non-synonymous substitutions are rapidly removed from the S. Typhimurium population, after which purifying selection is no longer important and the remaining substitutions become fixed in the population. The S. Typhimurium 135@ isolates were nearly identical to SL1344 in terms of gene content and virulence plasmids. Their phage contents were close to SL1344, except that they carried a different variant of Gifsy-1, lacked the P2 remnant found in SL1344 and carried a novel P2 phage, P2-Hawk, in place SL1344’s P2 phage SopEϕ. DT135 lacks P2 prophage. Two additional plasmids were identified in the S. Typhimurium 135@ isolates, pSTM2 and pSTM7. Both plasmids were IncI1, but phylogenetic analysis of the plasmids and their bacterial hosts shows these plasmids are genetically distinct and result from independent plasmid acquisition events.ConclusionsThis study provides a high-resolution insight into short-term microevolution of the important human pathogen S. Typhimurium. It indicates that purifying selection occurs rapidly in this population (≤6 years) and then declines, and provides an estimate for the short-term substitution rate. The latter is likely to be more relevant for foodborne outbreak investigation than previous estimates based on longer time scales.

Defining the Core Genome of Salmonella enterica Serovar Typhimurium for Genomic Surveillance and Epidemiological Typing

“…The S. Typhimurium core genome contained 956 (33.45%) more genes than the Salmonella core genome, which contains only 2,882 genes as defined previously (16). We showed that the use of the STCG or the STCG plus core IGRs increases the resolution of the genome sequencing for epidemiological typing using published and epidemiologically investigated outbreaks (14,15,25,26).…”

“…To evaluate the performance of the S. Typhimurium core genome (STCG with and without core IGRs) typing, 21 strains from SARA sequenced in this study and 94 isolates from 13 different outbreaks were used (see Table S2 in the supplemental material) (14,15,25,26). The selection of outbreaks was either based on representative phage type or ST, including DT135, DT135a, U292, DT3, DT104, DT12, DT120, and ST313.…”

Genomic Variability of Serial Human Isolates of Salmonella enterica Serovar Typhimurium Associated with Prolonged Carriage

“…There are three different mutation rates reported for S. Typhimurium. The lowest rate is 1.9 ϫ 10 Ϫ7 substitution per site per year, estimated for ST313, causing invasive infections in Africa (26); the intermediate rate is 3.4 ϫ 10 Ϫ7 substitution per site per year, obtained from the epidemic DT104 infections (27); and the highest rate is 1.2 ϫ10 Ϫ6 substitution per site per year, obtained from a DT135a outbreak (28). Our estimate of the mutation rate is the highest but is only 12% higher than the previous highest estimate.…”

A Salmonella typhimurium Outbreak Associated With Food Served at a Wedding Reception

“…Bununla birlikte gıda kaynaklı Salmonella infeksiyonlarının irdelendiği birçok çalışmada S. enterica subsp. enterica serovar Typhimurium en sık etken olarak tespit edilmiş ve en çok tavuk eti ve çiğ yumurta tüketimiyle ilişkili bulunmuştur (6,16). Çalışmamızda her ne kadar gıda örneklerinde herhangi bir üreme tespit edilmemiş olsa da, tüm olguların aynı yemek şirketi tarafından hazırlanmış menüyü aynı öğünde yemiş olması, tavuk eti yiyenlerde yakınmaların gelişmesi, sadece pilav ve ayran tüketenlerde herhangi bir yakınmanın olmaması, salgının kaynağının tavuk eti olduğunu düşündürmüş-tür.…”

“…enterica serovar Typhimurium (S. typhimurium) ve S. enterica subsp. enterica serovar Enteritidis (S. enteritidis), gıda kaynaklı salgın dahil gastroenteritlerin en sık etkenidir (6). Salmonella infeksiyonlarına en çok neden olan gıdaların başında ise kontamine tavuk eti ve yumurta gelmektedir (7,8).…”