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Whole-Genome Sequencing-Based Characterization of Listeria monocytogenes from Fish and Fish Production Environments in Poland.

ABSTRACT: Listeria monocytogenes, an important foodborne pathogen, may be present in different kinds of food and in food processing environments where it can persist for a long time. In this study, 28 L. monocytogenes isolates from fish and fish manufactures were characterized by whole genome sequencing (WGS). Core genome multilocus sequence typing (cgMLST) analysis was applied to compare the present isolates with publicly available genomes of L. monocytogenes strains recovered worldwide from food and from humans with listeriosis. All but one (96.4%) of the examined isolates belonged to molecular serogroup IIa, and one isolate (3.6%) was classified to serogroup IVb. The isolates of group IIa were mainly of MLST sequence types ST121 (13 strains) and ST8 (four strains) whereas the isolate of serogroup IVb was classified to ST1. Strains of serogroup IIa were further subtyped into eight different sublineages with the most numerous being SL121 (13; 48.1% strains) which belonged to six cgMLST types. The majority of strains, irrespective of the genotypic subtype, had the same antimicrobial resistance profile. The cluster analysis identified several molecular clones typical for L. monocytogenes isolated from similar sources in other countries; however, novel molecular cgMLST types not present in the Listeria database were also identified.

SUBMITTER: Wieczorek K

PROVIDER: S-EPMC7764581 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Whole-Genome Sequencing-Based Characterization of Listeria monocytogenes from Fish and Fish Production Environments in Poland.

Wieczorek Kinga K Bomba Arkadiusz A Osek Jacek J

International journal of molecular sciences 20201210 24

Listeria monocytogenes, an important foodborne pathogen, may be present in different kinds of food and in food processing environments where it can persist for a long time. In this study, 28 L. monocytogenes isolates from fish and fish manufactures were characterized by whole genome sequencing (WGS). Core genome multilocus sequence typing (cgMLST) analysis was applied to compare the present isolates with publicly available genomes of L. monocytogenes strains recovered worldw ...[more]

PMID: 33321935

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Project description:Listeria monocytogenes is frequently found in foods and processing facilities, where it can persist, creating concerns for the food industry. Its ability to survive under a wide range of environmental conditions enhances the potential for cross-contamination of the final food products, leading to possible outbreaks of listeriosis. In this study, whole-genome sequencing (WGS) was applied as a tool to characterize and track 100 L. monocytogenes isolates collected from three food processing environments. These WGS data from environmental and food isolates were analyzed to (i) assess the genomic diversity of L. monocytogenes, (ii) identify possible source(s) of contamination, cross-contamination routes, and persistence, (iii) detect absence/presence of antimicrobial resistance-encoding genes, (iv) assess virulence genotypes, and (v) explore in vivo pathogenicity of selected L. monocytogenes isolates carrying different virulence genotypes. The predominant L. monocytogenes sublineages (SLs) identified were SL101 (21%), SL9 (17%), SL121 (12%), and SL5 (12%). Benzalkonium chloride (BC) tolerance-encoding genes were found in 62% of these isolates, a value that increased to 73% among putative persistent subgroups. The most prevalent gene was emrC followed by bcrABC, qacH-Tn6188, and qacC. The L. monocytogenes major virulence factor inlA was truncated in 31% of the isolates, and only one environmental isolate (L. monocytogenes CFS086) harbored all major virulence factors, including Listeria pathogenicity island 4 (LIPI-4), which has been shown to confer hypervirulence. A zebrafish embryo infection model showed a low (3%) embryo survival rate for all putatively hypervirulent L. monocytogenes isolates assayed. Higher embryo survival rates were observed following infection with unknown virulence potential (20%) and putatively hypovirulent (53 to 83%) L. monocytogenes isolates showing predicted pathogenic phenotypes inferred from virulence genotypes.IMPORTANCE This study extends current understanding of the genetic diversity among L. monocytogenes from various food products and food processing environments. Application of WGS-based strategies facilitated tracking of this pathogen of importance to human health along the production chain while providing insights into the pathogenic potential for some of the L. monocytogenes isolates recovered. These analyses enabled the grouping of selected isolates into three putative virulence categories according to their genotypes along with informing selection for phenotypic assessment of their pathogenicity using the zebrafish embryo infection model. It has also facilitated the identification of those isolates with genes conferring tolerance to commercially used biocides. Findings from this study highlight the potential for the application of WGS as a proactive tool to support food safety controls as applied to L. monocytogenes.

Project description:Listeria monocytogenes (L. monocytogenes) is a ubiquitous organism that can easily enter the food chain. Infection with L. monocytogenes can cause invasive listeriosis. Since 2014, in Austria, L. monocytogenes isolates from human and food/food-associated samples have been provided on a mandatory basis by food producers and laboratories to the National Reference Laboratory. Since 2017, isolates undergo routinely whole genome sequencing (WGS) and core genome Multilocus Sequence Typing (cgMLST) for cluster analyses. Aims of this study were to characterize isolates and clusters of 2017 by using WGS data and to assess the usefulness of this isolate-based surveillance for generating hypotheses on sources of invasive listeriosis in real-time. WGS data from 31 human and 1744 non-human isolates originating from 2017, were eligible for the study. A cgMLST-cluster was defined as two or more isolates differing by ?10 alleles. We extracted the sequence types (STs) from the WGS data and analyzed the food subcategories meat, fish, vegetable and diary for associations with the ten most prevalent STs among food, through calculating prevalence ratios (PR) with 95% confidence intervals (CI). The three most frequent STs among the human isolates were ST1 (7/31; 22.6%), ST155 (4/31; 12.9%) and ST451 (3/31; 9.7%) and among the non-human isolates ST451 (614/1744; 35.2%), ST8 (173/1744, 10.0%) and ST9 (117/1744; 6.7%). We found ST21 associated with vegetables (PR: 11.39, 95% CI: 8.32-15.59), ST121 and ST155 with fish (PR: 7.05, 95% CI: 4.88-10.17, PR: 3.29, 95% CI: 1.86-5.82), and ST511, ST7 and ST451 with dairy products (PR: 8.55, 95% CI: 6.65-10.99; PR: 5.05, 95% CI: 3.83-6.66, PR: 3.03, 95% CI: 2.02-4.55). We identified 132 cgMLST-clusters. Six clusters contained human isolates (ST155, ST1, ST101, ST177, ST37 and ST7) and for five of those cgMLST-based cluster analyses solely was able to hypothesize the source: an Austrian meat processing company, two Austrian cheese manufacturers and two vegetable processing companies, one based in Austria and the other in Belgium. Determining routinely STs in food isolates by WGS allows to associate STs with food products. Real-time WGS of L. monocytogenes isolates provided mandatorily, proved to be useful in promptly generating hypotheses on sources of invasive listeriosis.

Project description:Listeria monocytogenes is a foodborne pathogen that can cause invasive disease with high mortality in immunocompromised individuals and can survive in a variety of food-associated environments for a long time. L. monocytogenes clonal complex (CC) 87 is composed of ST87 and three other STs and has been identified as the most common subgroup associated with both foods and human clinical infections in China. Therefore, the persistence of CC87 L. monocytogenes in food-associated environments poses a significant concern for food safety. In this study, 83 draft genomes of CC87 L. monocytogenes, including 60 newly sequenced genomes, were analyzed with all isolates from our previous surveillance in Zigong, Sichuang, China. Sixty-eight of the studied isolates were isolated from one retail market (M1 market), while the others were from seven other markets (M2-M8 markets) in the same city. Whole-genome multilocus sequence typing (wg-MLST) and the whole-genome single nucleotide polymorphism (wg-SNP) analysis were performed. Three persistent contamination routes were identified in the M1 market, caused by 2 clusters (A and B) and a wgST31 type. Cluster A isolates were associated with the persistent contamination in a raw meat stall (M1-S77), while Cluster B isolates caused a persistent contamination in aquatic foods stalls. Five wgST31 isolates caused persistent contamination in a single aquatic stall (M1-S65). A pLM1686-like plasmid was found in all Cluster A isolates. A novel plasmid, pLM1692, a truncated pLM1686 plasmid without the cadmium, and other heavy metal resistance genes were conserved in all wgST31 isolates. By comparing persistent and putative non-persistent isolates, four genes that were all located in the prophage comK might be associated with persistence. These findings enhanced our understanding of the underlying mechanisms of contamination and assist in formulating targeted strategies for the prevention and control of L. monocytogenes transmission from the food processing chain to humans. IMPORTANCE Contamination of food by Listeria monocytogenes at retail level leads to potential consumption of contaminated food with high risk of human infection. Our previous study found persistent contamination of CC87 L. monocytogenes from a retail market in China through pulsed-field gel electrophoresis and multilocus sequence typing. In this study, whole-genome sequencing was used to obtain the highest resolution inference of the source and reasons for persistent contamination; meat grinders and minced meat were the major reservoir of persistent contamination in meat stalls, whereas fishponds were the major reservoir in seafood stalls, with different L. monocytogenes isolates involved. These isolates carried different properties such as plasmids and prophages, which may have contributed to their ability to survive or adapt to the different environments. Our findings suggest that whole-genome sequencing will be an effective surveillance tool to detect persistent L. monocytogenes contamination in retail food markets and to design new control strategies to improve food safety.

Dataset Information

Whole-Genome Sequencing-Based Characterization of Listeria monocytogenes from Fish and Fish Production Environments in Poland.

Publications

Whole-Genome Sequencing-Based Characterization of <i>Listeria monocytogenes</i> from Fish and Fish Production Environments in Poland.

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