Whole-Genome Sequencing-Based Characterization of Listeria Isolates from Produce Packinghouses and Fresh-Cut Facilities Suggests Both Persistence and Reintroduction of Fully Virulent L. monocytogenes

Abstract

Listeria monocytogenes is of particular concern to the produce industry due to its frequent presence in natural environments as well as its ability to survive in packinghouses and fresh-cut processing facilities over time. The use of whole-genome sequencing, which provides high discriminatory power for the characterization of Listeria isolates, along with detailed source data (isolation date and sample location) shows that the presence of Listeria in produce operations appears to be due to random and continued reintroduction as well as to the persistence of highly related strains in both packinghouses and fresh-cut facilities.

Characterization and Antibiotic Resistance of Listeria monocytogenes Strains Isolated from Greek Myzithra Soft Whey Cheese and Related Food Processing Surfaces over Two-and-a-Half Years of Safety Monitoring in a Cheese Processing Facility

“…The present study provides insights into the pathogenic potential and persistence of the L. monocytogenes isolates recovered from a cheese processing facility. The findings of this work should be useful as they could be utilized for epidemiological investigations of L. monocytogenes in the food processing environment, revealing possible contamination scenarios, acquired antimicrobial resistance along the food production chain and providing critical information for post-outbreak management in the facility [ 103 , 104 , 105 ]. These findings also suggest that further research is needed for definitive genome mapping of the isolates.…”

Intensive environmental sampling and whole genome sequence-based characterization ofListeriain small and medium sized dairy plants reveal opportunities for simplified and size-appropriate environmental monitoring strategies

“…In this study, the 303 putatively persistent Listeria isolates characterized through WGS were assigned into initial hqSNP clusters based on a relatively high initial hqSNP distance cut-off (i.e., ≤50 pairwise hqSNP differences between one or more isolates within a given hqSNP cluster). While some studies have reported using a similar strategy (Pettengill et al, 2022; Sullivan et al, 2022; Wang et al, 2018), others with a similar number of isolates have reported using lower SNP distance cut-offs, such as ≤20 or ≤10 SNPs, to assign clusters for further analysis (Andrews et al, 2023; Castro et al, 2021; Lucchini et al, 2023), as they may be more likely to indicate that two or more isolates originated from the same source, e.g., the same plant (Pightling et al, 2018). Importantly, in this study we identified key trends among hqSNP clusters that contained isolates with >21 hqSNP differences that might not have been captured if a lower initial SNP distance cut-off was employed.…”

“…Isolates <100 kSNPs apart identified based on kSNP3 analysis were then analyzed using the Center for Food Safety and Applied Nutrition (CFSAN) SNP pipeline v. 2.2.1 to identify any preserved hqSNPs existing between groups of isolates (Davis et al, 2015). The reference assembly for the CFSAN SNP pipeline was selected as previously described (Sullivan et al, 2022). After hqSNP analysis was performed for groups of isolates with <100 kSNP differences, isolates that showed >50 pairwise hqSNP differences from all other isolates in the group were removed, and the group was re-examined to obtain “hqSNP clusters” which only included isolates that showed ≤50 pairwise hqSNP differences from at least one other isolate in the cluster.…”

“…Molecular subtyping tools can allow food plants to identify re-occurring subtypes that may be persisting in their food production environments (Beno et al, 2016; Vongkamjan et al, 2013). Whole genome sequencing (WGS) is now typically the preferred molecular subtyping tool for Listeria characterization (Jackson et al, 2016), as it can provide valuable insights about Listeria transmission and persistence in food processing environments through high-resolution subtyping approaches such as core genome multi-locus sequencing typing (cgMLST) or high quality single nucleotide polymorphism (hqSNP) analysis (Jagadeesan et al, 2019; Sullivan et al, 2022). Additionally, WGS data can be used to screen for high priority genes that may contribute to Listeria virulence, as well as tolerance to sanitizers and other environmental stressors (Y.…”

Combining Whole Genome Sequencing Data from Human and Non-Human Sources: Tackling Listeria monocytogenes Outbreaks

“…The initial positive food samples from the monitoring were not enough to pinpoint the exact source of contamination in five of the nine clusters, emphasizing that the point in the production chain at which the food product tests positive does not have to be the origin of the contamination. Nevertheless, this information can still indicate that hygienic measures have to be taken or that shelf-life studies should be reviewed at different levels of the production chain due to previously underestimated levels of cross-or post-contamination [24].…”