Listeria monocytogenes, a bacterium responsible for listeriosis, is an environmental and food-borne pathogen that poses a particular risk to pregnant women and the elderly. While traditionally associated with animal products, ready-to-eat salads are increasingly recognised as a source of Listeria outbreaks. However, little is known about the potential virulence of Listeria isolates from the fresh produce environment. This study assessed the virulence potential of nine L. monocytogenes strains from the fresh produce chain using the Galleria mellonella invertebrate infection model. Larvae were infected with 106 CFU of each strain via their circulatory system and compared to a reference strain L. monocytogenes (EGD-e) and Listeria ivanovii. Virulence was evaluated by measuring mortality rates, health index score of larvae, viable bacterial counts in the larvae, and the larvae’s immune. Significant differences in larval mortality were observed among strains. Strains NLmo4 and NLmo5 caused the highest mortality rates (98.8% and 96.7%, respectively at 7 days post-infection), while strain NLmo20 had a significantly lower mortality rate of 65% at the same time point (p<0.05). Six isolates that caused varied mortality rates were then selected and tested for their ability to replicate both in vitro and in vivo and their impact on larval haemocyte density. In vitro growth rates were not significantly different among L. monocytogenes strains or compared to Listeria ivanovii. However, L. monocytogenes strains persisted and replicated in larvae up to 7d days post-infection, whereas Listeria ivanovii was reduced by 5 logs CFU by day 7. The presence of these L. monocytogenes strains caused organ damage in larvae, indicated by increased melanisation and subsequent larval death. Haemocyte density showed insignificant fluctuations following infection. In conclusion, the results of this study suggest L. monocytogenes strains from fresh produce food chain have varying pathogenicity levels and can pose potential risk to human health.

Listeria monocytogenes is sporadically detected on a range of ready to eat fresh produce lines, such as spinach and rocket, and is a threat to public health. However, little is known about the diversity of L. monocytogenes present on fresh produce and their potential pathogenicity. In this work, fifteen Listeria monocytogenes isolates from the UK fresh produce supply chain were characterised using whole genome sequencing (WGS). Additionally, isolates were characterised based on their ability to form biofilm. Whole genome sequencing data was used to determine the sequence type of isolates based on multi-locus sequence typing (MLST), construct a core single nucleotide polymorphism (SNP) phylogeny and determine the presence of virulence and resistance associated genes. MLST revealed 9 distinct sequence types (STs) spanning 2 lineages (I & II) with one isolate belonging to the ST6 subtype, strains from which have been recently implicated in two large, food-associated L. monocytogenes outbreaks in South Africa and across Europe. Although most of the 15 isolates were different, comparison of core genome SNPs showed 4 pairs of 'indistinguishable' strains (<5 SNPs difference). Virulence profiling revealed that some isolates completely lacked the Listeria pathogenicity island-3 (LIPI-3) amongst other virulence factors. Investigation of the inlA gene showed that no strains in this study contained a premature stop codon (PMSC), an indicator of attenuated virulence. Assessment of biofilm production showed that isolates found in the fresh produce supply chain differ in their ability to form biofilm. This trait is considered important for L. monocytogenes to persist in environments associated with food production and processing. Overall the work indicates that a genetically diverse range of L. monocytogenes strains is present in the UK fresh produce supply chain and the virulence profiles found suggests that at least some of the strains are capable of causing human illness. Interestingly, the presence of some genetically indistinguishable isolates within the 15 isolates examined suggests that cross-contamination in the fresh produce environment does occur. These findings have useful implications in terms of food safety and for informing microbial surveillance programmes in the UK fresh produce supply chain.

Listeria monocytogenes is an intracellular human pathogen which enters the body through contaminated food stuffs and is known to contaminate fresh leafy produce such as spinach, lettuce and rocket. Routinely, fresh leafy produce is grown and processed on a large scale before reaching the consumer through various products such as sandwiches and prepared salads. From farm to fork, the fresh leafy produce supply chain (FLPSC) is complex and contains a diverse range of environments where L. monocytogenes is sporadically detected during routine sampling of produce and processing areas. This review describes sources of the bacteria in the FLPSC and outlines the physiological and molecular mechanisms behind its survival in the different environments associated with growing and processing fresh produce. Finally, current methods of source tracking the bacteria in the context of the food supply chain are discussed with emphasis on how these methods can provide additional, valuable information on the risk that L. monocytogenes isolates pose to the consumer.