Genomic Characterization of Listeria monocytogenes Isolates Associated with Clinical Listeriosis and the Food Production Environment in Ireland - PubMed (original) (raw)
Genomic Characterization of Listeria monocytogenes Isolates Associated with Clinical Listeriosis and the Food Production Environment in Ireland
Amber Hilliard et al. Genes (Basel). 2018.
Abstract
Listeria monocytogenes is a major human foodborne pathogen that is prevalent in the natural environment and has a high case fatality rate. Whole genome sequencing (WGS) analysis has emerged as a valuable methodology for the classification of L. monocytogenes isolates and the identification of virulence islands that may influence infectivity. In this study, WGS was used to provide an insight into 25 L. monocytogenes isolates from cases of clinical infection in Ireland between 2013 and 2015. Clinical strains were either lineage I (14 isolates) or lineage II (11 isolates), with 12 clonal complexes (CC) represented, of which CC1 (6) and CC101 (4) were the most common. Single nucleotide polymorphism (SNP) analysis demonstrated that clinical isolates from mother-infant pairs (one isolate from the mother and one from the infant) were highly related (3 SNP differences in each) and also identified close similarities between isolates from otherwise distinct cases (1 SNP difference). Clinical strains were positive for common virulence-associated loci and 13 isolates harbour the LIPI-3 locus. Pulsed-field gel electrophoresis (PFGE) was used to compare strains to a database of 1300 Irish food and food processing environment isolates and determined that 64% of clinical pulsotypes were previously encountered in the food or food processing environment. Five of the matching food and food processing environment isolates were sequenced and results demonstrated a correlation between pulsotype and genotype. Overall, the work provides insights into the nature of L. monocytogenes strains currently causing clinical disease in Ireland and indicates that similar isolates can be found in the food or food processing environment.
Keywords: Listeria monocytogenes; SNP; clinical; genome; genomics; sequence; single nucleotide polymorphism.
Conflict of interest statement
The authors declare no conflict of interest.
Figures
Figure 1
(A) Clustering of isolates based upon multi-locus sequence typing (MLST). The evolutionary history was inferred by using the Maximum Likelihood method based on the Tamura 3-parameter model [30] as outlined in detail in Materials and Methods. The tree with the highest log likelihood (−5971.3729) is shown. The percentage of trees in which the associated taxa clustered together is shown next to the branches. The tree is drawn to scale, with branch lengths measured in the number of substitutions per site. Evolutionary analyses were conducted in MEGA7 [31]; (B) Clustering of isolates based upon pulsed field gel electrophoresis (PFGE). Dendograms were generated with BioNumerics v7.0 software (Applied Maths) using UPGMA (unweighted pair group method with averages) and the Pearson coefficient with 1% tolerance. Using either method (MLST or PFGE) strains cluster into two distinct groups dependent upon lineage. ST: sequence type; CC: clonal complex.
Figure 2
Phylogeny of the isolates as determined by single nucleotide polymorphism (SNP) analysis. (A) All serotype 4b isolates were compared with strain F2365 as the reference genome. The evolutionary history was inferred by using the Maximum Likelihood method based on the General Time Reversible model [34]. The tree with the highest log likelihood (−67933.7374) is shown. The percentage of trees in which the associated taxa clustered together is shown next to the branches. Initial tree(s) for the heuristic search were obtained automatically by applying Neighbor-Join and BioNJ algorithms to a matrix of pairwise distances estimated using the Maximum Composite Likelihood (MCL) approach, and then selecting the topology with superior log likelihood value. The tree is drawn to scale, with branch lengths measured in the number of substitutions per site. The analysis involved 18 nucleotide sequences. Codon positions included were 1st + 2nd + 3rd + Noncoding. All positions with less than 95% site coverage were eliminated. That is, fewer than 5% alignment gaps, missing data, and ambiguous bases were allowed at any position. There were a total of 12,069 positions in the final dataset. Evolutionary analyses were conducted in MEGA7 [31]. (B) All serotype 1/2a isolates were compared with strain EGDe as the reference genome. The evolutionary history was inferred by using the Maximum Likelihood method based on the General Time Reversible model [34]. The tree with the highest log likelihood (−214301.5165) is shown. The tree is drawn to scale, with branch lengths measured in the number of substitutions per site. The analysis involved 15 nucleotide sequences. Codon positions included were 1st + 2nd + 3rd + Noncoding. All positions with less than 95% site coverage were eliminated. That is, fewer than 5% alignment gaps, missing data, and ambiguous bases were allowed at any position. There were a total of 30,102 positions in the final dataset. Evolutionary analyses were conducted in MEGA7 [31]. * Indicates strains from a food production source.
Figure 3
Pan-genome, constructed using GView [35], of 34 genomes including 25 clinical isolates, six food-associated isolates and three reference genomes. The pangenome is constructed by iteratively appending unique regions onto the initial seed genome in this case F2365. Gaps indicate that the region is missing in a particular gene but is found in others. Strains are grouped according to CC and colour coded as indicated.
Figure 4
The presence and absence of genes can be used to cluster the isolates into serotypes and sequence types.
References
- Health Protection Surveillance Centre (HPSC) HPSC Annual Epidemiological Report 2015. Volume 75 Health Protection Surveillance Centre; Dublin, Ireland: 2016.
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