Genomic Characterization of Listeria monocytogenes Isolates Associated with Clinical Listeriosis and the Food Production Environment in Ireland.

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.

Draft Genome Sequences of 25 Listeria monocytogenes Isolates Associated with Human Clinical Listeriosis in Ireland.

Listeria monocytogenes is a Gram-positive opportunistic pathogen that is the causative agent of listeriosis. Here, we report the draft genome sequences of 25 L. monocytogenes strains isolated from patients with clinical listeriosis in the Republic of Ireland between 2013 and 2015.

A 3-year multi-food study of the presence and persistence of Listeria monocytogenes in 54 small food businesses in Ireland.

The problem of assessing the occurrence of the food-borne pathogen Listeria monocytogenes in the food chain, and therefore the risk of exposure of the human population, is often challenging because of the limited scope of some studies. In this study the occurrence of L. monocytogenes in food from four major food groups, dairy products, meats, seafood and vegetables, and associated food processing environments in Ireland was studied over a three-year period. Fifty-four small food businesses participated in the study and sent both food and environmental samples every 2months between 2013 and 2015. L. monocytogenes was isolated using the ISO11290 standard method. Confirmation of L. monocytogenes and identification of serogroups were achieved using a multiplex PCR assay, and for some isolates serotype was determined using commercial antisera. Pulsed- field gel electrophoresis (PFGE) analysis was performed on all isolates allowing the relatedness of isolates from different food businesses to be compared nationwide. In total, 86 distinct pulsotypes were identified. The overall occurrence of L. monocytogenes in food samples was 4.2%, while in environmental samples it was 3.8%. In general, the occurrence of L. monocytogenes in food businesses decreased over the course of the study, presumably reflecting increased awareness and vigilance. The majority of the pulsotypes detected were unique to a particular food group (63/86), while only three pulsotypes were found in all four food groups investigated. The highest occurrence in food was found in the meat category (7.5%) while seafood had the lowest rate of occurrence (1.8%). Seventeen of the pulsotypes detected in the study were persistent, where persistence was defined as repeated isolation from a single facility with a minimum time interval of 6months. Using PFGE, 11 of the pulsotypes identified in this study were indistinguishable from those of 11 clinical isolates obtained from patients in Ireland over the last 4years, highlighting the fact that these pulsotypes are capable of causing disease. Overall, the study shows the diversity of L. monocytogenes strains in the Irish food chain and highlights the ability of many of these strains to persist in food processing environments. The finding that a significant proportion of these pulsotypes are also found in clinical settings highlights the need for continued vigilance by food producers, including frequent sampling and typing of isolates detected.

Draft Genome Sequences of Listeria monocytogenes Serotype 4b Strains 944 and 2993 and Serotype 1/2c Strains 198 and 2932.

Listeria monocytogenes is a foodborne pathogen and the causative agent of listeriosis among humans and animals. The draft genome sequences of L. monocytogenes serotype 4b strains 944 and 2993 and serotype 1/2c strains 198 and 2932 are reported here.

Occurrence, Persistence, and Virulence Potential of Listeria ivanovii in Foods and Food Processing Environments in the Republic of Ireland.

The aim of this study was to assess the occurrence of L. ivanovii in foods and food processing environments in Ireland, to track persistence, and to characterize the disease causing potential of the isolated strains. A total of 2,006 samples (432 food samples and 1,574 environmental swabs) were collected between March 2013 and March 2014 from 48 food business operators (FBOs) belonging to different production sectors (dairy, fish, meat, and fresh-cut vegetable). Six of the forty-eight FBOs had samples positive for L. ivanovii on at least one sampling occasion. L. ivanovii was present in fifteen samples (fourteen environmental samples and one food sample). All but one of those positive samples derived from the dairy sector, where L. ivanovii prevalence was 1.7%. Six distinguishable pulsotypes were obtained by PFGE analysis, with one pulsotype being persistent in the environment of a dairy food business. Sequence analysis of the sigB gene showed that fourteen isolates belonged to L. ivanovii subsp. londoniensis, while only one isolate was L. ivanovii subsp. ivanovii. Cell invasion assays demonstrated that the majority of L. ivanovii strains were comparable to L. monocytogenes EGDe in their ability to invade CACO-2 epithelial cells whilst four isolates had significantly higher invasion efficiencies.

Examination of Listeria monocytogenes in Seafood Processing Facilities and Smoked Salmon in the Republic of Ireland.

Listeria monocytogenes is a foodborne pathogen that causes listeriosis, a relatively rare but life-threatening disease primarily affecting immunocompromised individuals. The aim of this study was to determine the prevalence of L. monocytogenes in the seafood processing industry in the Republic of Ireland. The occurrence of L. monocytogenes was determined by regular sampling of both food samples and processing environment swabs at eight seafood processing facilities over two calendar years. All samples were analyzed by the International Organization for Standardization 11290-1 standard method, and the isolates were characterized by PCR, pulsed-field gel electrophoresis, serotyping, and the occurrence of some genes related to survival under stress (SSI-1, Tn6188, and bcrABC). A prevalence of 2.5% in 508 samples (433 environmental swabs and 75 food samples) was found. From the isolates obtained, eight different pulsed-field gel electrophoresis profiles were identified, two occurring in more than one facility and one occurring in food and the environment. Five of the eight pulsotypes identified contained at least one of the three stress survival-related genes tested. The tolerance of the isolates to benzalkonium chloride, a representative quaternary ammonium compound, was also examined and ranged from 5.5 ± 0.5 to 8.5 ± 0.5 ppm of benzalkonium chloride. To evaluate the ability of smoked salmon to support the growth of L. monocytogenes, including the T4 widespread pulsotype that was isolated, a challenge test was performed on cold-smoked salmon obtained from two separate producers. The results showed clearly that both types of smoked salmon supported the growth of L. monocytogenes. Although occurrence of L. monocytogenes on seafood was low, this study showed that the smoked salmon used in this study can support the growth of L. monocytogenes; therefore, vigilance is required in the processing facilities to reduce the associated risk.

The challenge of challenge testing to monitor Listeria monocytogenes growth on ready-to-eat foods in Europe by following the European Commission (2014) Technical Guidance document.

European Regulation (EC) No. 2073/2005 lays down the microbiological criteria for certain microorganisms in foods and the implementing rules to be complied with by food business operators (FBOs) in Europe when implementing general and specific hygiene measures. In relation to Listeria monocytogenes, this regulation covers primarily ready-to-eat (RTE) food products, and requires different microbiological criteria depending on the ability of the food product to support growth of L. monocytogenes. In addition, this regulation establishes that food safety is the responsibility of the FBO. The FBO can conduct studies to evaluate the growth of L. monocytogenes that may be present in the product during the shelf-life under reasonably foreseeable storage conditions of distribution, storage and use in order to investigate compliance with the criteria throughout the shelf-life of the product. The European Union Community Reference Laboratory for L. monocytogenes published a revised technical guidance document in June 2014 for conducting shelf-life studies on L. monocytogenes in RTE foods. This review article describes the recently published European guidance document, with special focus on the design of challenge studies to determine the growth potential of L. monocytogenes on foods. Information is given particularly on what a challenge test is and when one is advisable. The factors to be considered and the laboratory methodology to be applied when performing a challenge test to determine the growth potential of L. monocytogenes in a defined food matrix are also described. Results of recent research articles applying challenge tests to determine the growth of L. monocytogenes in a range of foodstuffs are summarized and discussed. Finally, recommendations for obtaining data that can contribute to any further revision of the guidance document and for addressing the main challenges of challenge testing for FBOs are presented.

A novel erythromycin resistance plasmid from Bacillus sp. strain HS24, isolated from the marine sponge Haliclona simulans.

A better understanding of the origin and natural reservoirs of resistance determinants is fundamental to efficiently tackle antibiotic resistance. This paper reports the identification of a novel 5.8 kb erythromycin resistance plasmid, from Bacillus sp. HS24 isolated from the marine sponge Haliclona simulans. pBHS24B has a mosaic structure and carries the erythromycin resistance gene erm(T). This is the first report of an erythromycin resistance plasmid from a sponge associated bacteria and of the Erm(T) determinant in the genus Bacillus.

Monitoring occurrence and persistence of Listeria monocytogenes in foods and food processing environments in the Republic of Ireland.

Although rates of listeriosis are low in comparison to other foodborne pathogenic illness, listeriosis poses a significant risk to human health as the invasive form can have a mortality rate as high as 30%. Food processors, especially those who produce ready-to-eat (RTE) products, need to be vigilant against Listeria monocytogenes, the causative pathogen of listeriosis, and as such, the occurrence of L. monocytogenes in food and in the food processing environment needs to be carefully monitored. To examine the prevalence and patterns of contamination in food processing facilities in Ireland, 48 food processors submitted 8 samples every 2 months from March 2013 to March 2014 to be analyzed for L. monocytogenes. No positive samples were detected at 38% of the processing facilities tested. Isolates found at the remaining 62% of facilities were characterized by serotyping and Pulsed Field Gel Electrophoresis (PFGE). A general L. monocytogenes prevalence of 4.6% was seen in all samples analyzed with similar rates seen in food and environmental samples. Differences in prevalence were seen across different food processors, food sectors, sampling months etc. and PFGE analysis allowed for the examination of contamination patterns and for the identification of several persistent strains. Seven of the food processing facilities tested showed contamination with persistent strains and evidence of bacterial transfer from the processing environment to food (the same pulsotype found in both) was seen in four of the food processing facilities tested.

Determination of Listeria monocytogenes Growth during Mushroom Production and Distribution.

In the EU, food is considered safe with regard to Listeria monocytogenes if its numbers do not exceed 100 CFU/g throughout the shelf-life of the food. Therefore, it is important to determine if a food supports growth of L. monocytogenes. Challenge studies to determine the ability of a food to support growth of L. monocytogenes are essential as predictive modelling often overestimates the growth ability of L. monocytogenes. The aim of this study was to determine if growth of L. monocytogenes was supported during the production and distribution of mushrooms. A three-strain mixture of L. monocytogenes was inoculated onto three independent batches of whole mushrooms, sliced mushrooms, mushroom casing and mushroom substrate at a concentration of about 100-1000 CFU/g. The batches were incubated at potential abuse temperatures, as a worst case scenario, and at intervals during storage L. monocytogenes numbers, % moisture and pH were determined. The results showed that the sliced and whole mushrooms had the ability to support growth, while mushroom casing allowed survival but did not support growth. Mushroom substrate showed a rich background microflora that grew on Listeria selective media and this hindered enumeration of L. monocytogenes. In the case of this study, Combase predictions were not always accurate, indicating that challenge studies may be a necessary part of growth determination of L.monocytogenes.