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The investigation of molecular characterization of presumptive Listeria monocytogenes isolates from a food-processing environment.

ABSTRACT: Background:Listeria is a Gram-positive, non-spore forming, facultative anaerobic intracellular bacterium. The most important pathogens in mammals include Listeria monocytogenes and Listeria ivanovii. The former generally causes disease and death in both humans and animals while the latter performs sporadically and primarily causes illness in ruminants. Aims:The aim of this project was to use conventional and molecular techniques to determine whether the provided samples were L. monocytogenes, and whether they were genetically similar or different. Methods:The provided presumptive Listeria cultures isolated from industrial processed food are conventionally assumed to be L. monocytogenes. All samples were cultured on brain heart infusion agar and broth first and then on blood agar. Later, hly gene amplification was applied. Results:The provided culture phenotypically resembled L. monocytogenes as it caused haemolysis on blood agar plates; however, the absence of the hly gene revealed that they were genotypically different. 16S rRNA confirmed three species of Listeria species including L. grayi, L. welshimeri and L. ivanovii. The results from 16S rRNA sequencing confirmed the results obtained from hly gene amplification. Conclusion:Enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC PCR) confirmed that all bacterial cultures were isolated from different sources depending on their ERIC PCR profile variation.

SUBMITTER: Ahmed MS

PROVIDER: S-EPMC6509916 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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The investigation of molecular characterization of presumptive <i>Listeria monocytogenes</i> isolates from a food-processing environment.

Ahmed M S MS

Iranian journal of veterinary research 20190101 1

<h4>Background</h4><i>Listeria</i> is a Gram-positive, non-spore forming, facultative anaerobic intracellular bacterium. The most important pathogens in mammals include <i>Listeria monocytogenes</i> and <i>Listeria ivanovii</i>. The former generally causes disease and death in both humans and animals while the latter performs sporadically and primarily causes illness in ruminants.<h4>Aims</h4>The aim of this project was to use conventional and molecular techniques to determine whether the provid ...[more]

PMID: 31191699

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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:BACKGROUND:Listeria monocytogenes is a widespread foodborne pathogen that can cause listeriosis, a potentially fatal infection. L. monocytogenes is subdivided into four phylogenetic lineages, with the highest incidence of listeriosis occurring within lineage I followed by lineage II. Strains of L. monocytogenes differ in their phenotypic characteristics, including virulence. However, the genetic bases for these observed differences are not well understood, and current efforts to monitor L. monocytogenes in food consider all strains to be equally virulent. We use a comparative genomics approach to identify genes and single nucleotide polymorphisms (SNPs) in 174 clinical and food isolates of L. monocytogenes that potentially contribute to virulence or the capacity to adapt to food environments. RESULTS:No SNPs are significantly associated with food or clinical isolates. No genes are significantly associated with food or clinical isolates from lineage I, but eight genes consisting of multiple homologues are associated with lineage II food isolates. These include three genes which encode hypothetical proteins, the cadmium resistance genes cadA and cadC, the multi-drug resistance gene ebrB, a quaternary ammonium compound resistance gene qac, and a regulatory gene. All eight genes are plasmid-borne, and most closed L. monocytogenes plasmids carry at least five of the genes (24/27). In addition, plasmids are more frequently associated with lineage II food isolates than with lineage II clinical isolates. CONCLUSIONS:We identify eight genes that are significantly associated with food isolates in lineage II. Interestingly, the eight genes are virtually absent in lineage II outbreak isolates, are composed of homologues which show a nonrandom distribution among lineage I serotypes, and the sequences are highly conserved across 27 closed Listeria plasmids. The functions of these genes should be explored further and will contribute to our understanding of how L. monocytogenes adapts to the host and food environments. Moreover, these genes may also be useful as markers for risk assessment models of either pathogenicity or the ability to proliferate in food and the food processing environment.

Dataset Information

The investigation of molecular characterization of presumptive Listeria monocytogenes isolates from a food-processing environment.

Publications

The investigation of molecular characterization of presumptive <i>Listeria monocytogenes</i> isolates from a food-processing environment.

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