A secondary model for the effect of pH on the variability in growth fitness of Listeria innocua strains.

Variability in microbial growth is a keystone of modern Quantitative Microbiological Risk Assessment (QMRA). However, there are still significant knowledge gaps on how to model variability, with the most common assumption being that variability is constant. This is implemented by an error term (with constant variance) added on top of the secondary growth model (for the square root of the growth rate). However, this may go against microbial ecology principles, where differences in growth fitness among bacterial strains would be more prominent in the vicinity of the growth limits than at optimal growth conditions. This study coins the term "secondary models for variability", evaluating whether they should be considered in QMRA instead of the constant strain variability hypothesis. For this, 21 strains of Listeria innocua were used as case study, estimating their growth rate by the two-fold dilution method at pH between 5 and 10. Estimates of between-strain variability and experimental uncertainty were obtained for each pH using mixed-effects models, showing the lowest variability at optimal growth conditions, increasing towards the growth limits. Nonetheless, the experimental uncertainty also increased towards the extremes, evidencing the need to analyze both sources of variance independently. A secondary model was thus proposed, relating strain variability and pH conditions. Although the modelling approach certainly has some limitations that would need further experimental validation, it is an important step towards improving the description of variability in QMRA, being the first model of this type in the field.

Phenotypic and genotypic antimicrobial resistance of Listeria spp. in Spain.

Listeriosis is a zoonotic disease caused by Listeria monocytogenes and Listeria ivanovii. The genus Listeria currently includes 27 recognized species and is found throughout the environment. The number of systematic studies on antimicrobial resistance in L. monocytogenes isolates from domestic farms using antimicrobial substances is limited. Importantly, dairy ruminant farms are reservoir of hypervirulent lineage I L. monocytogenes isolates, previously associated with human clinical cases. Considering that the classes of antibiotics used in food-producing domestic animals are frequently the same or closely related to those used in human medicine, studies about the impact of antibiotic use on the acquisition of antibiotic resistance in Listeria spp. in domestic animal farms are, therefore, of high importance. Here, susceptibility to 25 antibiotics was determined. Eighty-one animal-related, 35 food and 21 human pathogenic Listeria spp. isolates and 114 animal-related non-pathogenic Listeria spp. isolates were tested. Whole genome sequencing data was used for molecular characterization. Regarding L. monocytogenes, 2 strains from the clinical-associated linage I showed resistance to erythromycin, both related to dairy ruminants. Acquired resistance to one antibiotic was exhibited in 1.5% of L. monocytogenes isolates compared with 14% of non-pathogenic Listeria spp. isolates. Resistance to tetracycline (7.9%), doxycycline (7.9%), penicillin (4.4%), and ampicillin (4.4%) were the most frequently observed in non-pathogenic Listeria spp. While resistance to two or more antibiotics (5.6%) was most common in Listeria spp., isolates, resistance to one antibiotic was also observed (1.6%). The present results show that non-pathogenic Listeria spp. harbour antimicrobial resistance genes.

Investigating processing practices and microbiological quality of minimally processed vegetables in Brazil.

Minimally processed vegetables (MPVs) are marketed as convenient and healthy choices for consumers. However, the absence of post-commercialization treatments raises concerns about their microbiological safety. This study investigated the processing practices of 28 Brazilian MPV plants and compared the microbiological quality of these products with fresh counterparts in the city of Sao Paulo, Brazil. Through cluster analysis, the processing plants were categorized into two groups: group 1 (nineteen plants) primarily uses chemical substances in the washing step, while group 2 (nine plants) avoids chemical use but employs similar rinsing practices. Microbiological analysis of 100 samples (49 unprocessed and 51 MPVs) revealed no significant differences in microbial group counts (Enterobacteriaceae, coliforms, and E. coli) between the in natura (unprocessed) and MPV products. However, the prevalence of E. coli was higher in natura vegetables than in MPVs. The results indicated the presence of Salmonella DNA (from either dead or live cells or residual DNA) in 4 samples (3 in natura and 1 MPV) using conventional PCR, suggesting the presence of the pathogen in these samples. Listeria monocytogenes was absent, but Listeria innocua was found in two unprocessed products. The study suggests that certain MPVs have microbial loads similar to unprocessed vegetables, potentially serving as carriers for pathogen transmission. These findings emphasize the importance of understanding practices in Brazilian MPV processing plants, informing the implementation of control measures to improve MPV safety and shelf-life, thus ensuring microbiological safety.

Antimicrobial susceptibility, multilocus sequence typing, and virulence of listeria isolated from a slaughterhouse in Jiangsu, China.

BACKGROUND: Listeria monocytogenes is one of the deadliest foodborne pathogens. The bacterium can tolerate severe environments through biofilm formation and antimicrobial resistance. This study aimed to investigate the antimicrobial susceptibility, resistance genes, virulence, and molecular epidemiology about Listeria from meat processing environments. METHODS: This study evaluated the antibiotic resistance and virulence of Listeria isolates from slaughtering and processing plants. All isolates were subjected to antimicrobial susceptibility testing using a standard microbroth dilution method. The harboring of resistant genes was identified by polymerase chain reaction. The multilocus sequence typing was used to determine the subtyping of the isolates and characterize possible routes of contamination from meat processing environments. The virulence of different STs of L. monocytogenes isolates was evaluated using a Caco-2 cell invasion assay. RESULTS: A total of 59 Listeria isolates were identified from 320 samples, including 37 L. monocytogenes isolates (62.71%). This study evaluated the virulence of L. monocytogenes and the antibiotic resistance of Listeria isolates from slaughtering and processing plants. The susceptibility of these 59 isolates against 8 antibiotics was analyzed, and the resistance levels to ceftazidime, ciprofloxacin, and lincomycin were as high as 98.31% (L. m 37; L. innocua 7; L. welshimeri 14), 96.61% (L. m 36; L. innocua 7; L. welshimeri 14), and 93.22% (L. m 35; L. innocua 7; L. welshimeri 13), respectively. More than 90% of the isolates were resistant to three to six antibiotics, indicating that Listeria isolated from meat processing environments had high antimicrobial resistance. Up to 60% of the isolates harbored the tetracycline-resistance genes tetA and tetM. The frequency of ermA, ermB, ermC, and aac(6')-Ib was 16.95, 13.56, 15.25, and 6.78%, respectively. Notably, the resistant phenotype and genotype did not match exactly, suggesting that the mechanisms of antibiotic resistance of these isolates were likely related to the processing environment. Multilocus sequence typing (MLST) revealed that 59 Listeria isolates were grouped into 10 sequence types (STs). The dominant L. monocytogenes STs were ST5, ST9, and ST121 in the slaughtering and processing plant of Jiangsu province. Moreover, ST5 subtypes exhibited high invasion in Caco-2 cells compared with ST9 and ST121 cells. CONCLUSION: The dominant L. monocytogenes ST5 persisted in the slaughtering and processing plant and had high antimicrobial resistance and invasion characteristics, illustrating a potential risk in food safety and human health.

Proteomic Characterization of Antibiotic Resistance in Listeria and Production of Antimicrobial and Virulence Factors.

Some Listeria species are important human and animal pathogens that can be found in contaminated food and produce a variety of virulence factors involved in their pathogenicity. Listeria strains exhibiting multidrug resistance are known to be progressively increasing and that is why continuous monitoring is needed. Effective therapy against pathogenic Listeria requires identification of the bacterial strain involved, as well as determining its virulence factors, such as antibiotic resistance and sensitivity. The present study describes the use of liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) to do a global shotgun proteomics characterization for pathogenic Listeria species. This method allowed the identification of a total of 2990 non-redundant peptides, representing 2727 proteins. Furthermore, 395 of the peptides correspond to proteins that play a direct role in Listeria pathogenicity; they were identified as virulence factors, toxins and anti-toxins, or associated with either antibiotics (involved in antibiotic-related compounds production or resistance) or resistance to toxic substances. The proteomic repository obtained here can be the base for further research into pathogenic Listeria species and facilitate the development of novel therapeutics for these pathogens.

Nationwide genomic atlas of soil-dwelling Listeria reveals effects of selection and population ecology on pangenome evolution.

Natural bacterial populations can display enormous genomic diversity, primarily in the form of gene content variation caused by the frequent exchange of DNA with the local environment. However, the ecological drivers of genomic variability and the role of selection remain controversial. Here, we address this gap by developing a nationwide atlas of 1,854 Listeria isolates, collected systematically from soils across the contiguous United States. We found that Listeria was present across a wide range of environmental parameters, being mainly controlled by soil moisture, molybdenum and salinity concentrations. Whole-genome data from 594 representative strains allowed us to decompose Listeria diversity into 12 phylogroups, each with large differences in habitat breadth and endemism. 'Cosmopolitan' phylogroups, prevalent across many different habitats, had more open pangenomes and displayed weaker linkage disequilibrium, reflecting higher rates of gene gain and loss, and allele exchange than phylogroups with narrow habitat ranges. Cosmopolitan phylogroups also had a large fraction of genes affected by positive selection. The effect of positive selection was more pronounced in the phylogroup-specific core genome, suggesting that lineage-specific core genes are important drivers of adaptation. These results indicate that genome flexibility and recombination are the consequence of selection to survive in variable environments.

Listeria cossartiae sp. nov., Listeria farberi sp. nov., Listeria immobilis sp. nov., Listeria portnoyi sp. nov. and Listeria rustica sp. nov., isolated from agricultural water and natural environments.

A total of 27 Listeria isolates that could not be classified to the species level were obtained from soil samples from different locations in the contiguous United States and an agricultural water sample from New York. Whole-genome sequence-based average nucleotide identity blast (ANIb) showed that the 27 isolates form five distinct clusters; for each cluster, all draft genomes showed ANI values of <95 % similarity to each other and any currently described Listeria species, indicating that each cluster represents a novel species. Of the five novel species, three cluster with the Listeria sensu stricto clade and two cluster with sensu lato. One of the novel sensu stricto species, designated L. cossartiae sp. nov., contains two subclusters with an average ANI similarity of 94.9%, which were designated as subspecies. The proposed three novel sensu stricto species (including two subspecies) are Listeria farberi sp. nov. (type strain FSL L7-0091T=CCUG 74668T=LMG 31917T; maximum ANI 91.9 % to L. innocua), Listeria immobilis sp. nov. (type strain FSL L7-1519T=CCUG 74666T=LMG 31920T; maximum ANI 87.4 % to L. ivanovii subsp. londoniensis) and Listeria cossartiae sp. nov. [subsp. cossartiae (type strain FSL L7-1447T=CCUG 74667T=LMG 31919T; maximum ANI 93.4 % to L. marthii) and subsp. cayugensis (type strain FSL L7-0993T=CCUG 74670T=LMG 31918T; maximum ANI 94.7 % to L. marthii). The two proposed novel sensu lato species are Listeria portnoyi sp. nov. (type strain FSL L7-1582T=CCUG 74671T=LMG 31921T; maximum ANI value of 88.9 % to L. cornellensis and 89.2 % to L. newyorkensis) and Listeria rustica sp. nov. (type strain FSL W9-0585T=CCUG 74665T=LMG 31922T; maximum ANI value of 88.7 % to L. cornellensis and 88.9 % to L. newyorkensis). L. immobilis is the first sensu stricto species isolated to date that is non-motile. All five of the novel species are non-haemolytic and negative for phosphatidylinositol-specific phospholipase C activity; the draft genomes lack the virulence genes found in Listeria pathogenicity island 1 (LIPI-1), and the internalin genes inlA and inlB, indicating that they are non-pathogenic.

Characterization of a Novel Group of Listeria Phages That Target Serotype 4b Listeria monocytogenes.

Listeria monocytogenes serotype 4b strains are the most prevalent clinical isolates and are widely found in food processing environments. Bacteriophages are natural viral predators of bacteria and are a promising biocontrol agent for L. monocytogenes. The aims of this study were to characterize phages that specifically infect serotype 4b strains and to assess their ability to inhibit the growth of serotype 4b strains. Out of 120 wild Listeria phages, nine phages were selected based on their strong lytic activity against the model serotype 4b strain F2365. These nine phages can be divided into two groups based on their morphological characteristics and host range. Comparison to previously characterized phage genomes revealed one of these groups qualifies to be defined as a novel species. Phages LP-020, LP-027, and LP-094 were selected as representatives of these two groups of phages for further characterization through one-step growth curve and inhibition of serotype 4b L. monocytogenes experiments. Listeria phages that target serotype 4b showed an inhibitory effect on the growth of F2365 and other serotype 4 strains and may be useful for biocontrol of L.monocytogenes in food processing environments.

Prevalence and transmission characteristics of Listeria species from ruminants in farm and slaughtering environments in China.

Listeria monocytogenes is an important foodborne pathogen, and is ubiquitously distributed in the natural environment. Cattle and sheep, as natural hosts, can transmit L. monocytogenes to related meat and dairy products. In this study, the prevalence, distribution, and transmission characteristics of Listeria were analysed by investigating 5214 samples of cattle and sheep in farm and slaughtering environments in China. A low contamination incidence of L. monocytogenes (0.5%, 20/4430) was observed in farm environment, but there was a high contamination incidence in slaughtering environment (9.4%, 74/784). The incidence of L. innocua in cattle and sheep farm and slaughtering environments is more common and significantly higher (9.7%, 508/5214) than that of L. monocytogenes (1.8%, 94/5214). The distinct molecular and genetic characteristics of Listeria by PFGE and MLST indicated that L. monocytogenes and L. innocua were gradually transmitted from the farm and slaughtering environments to end products, such as beef and mutton along the slaughtering chain. The ST7, ST9, ST91, and ST155 found in our study were associated with the human listeriosis cases in China. In addition, the findings of virulence markers (inlC, inlJ, LIPI-3, LIPI-4, and ECIII) concerned with the pathogenesis of human listeriosis and antibiotics resistance of L. monocytogenes in this study implies a potential public health risk. This study fills the gap in the epidemiology of beef cattle and sheep that carry Listeria in farm and slaughtering environments in major cattle and sheep producing areas in China.

Genetic Carriers and Genomic Distribution of cadA6-A Novel Variant of a Cadmium Resistance Determinant Identified in Listeria spp.

Listeria monocytogenes is a pathogen responsible for severe cases of food poisoning. Listeria spp. strains occurring in soil and water environments may serve as a reservoir of resistance determinants for pathogenic L. monocytogenes strains. A large collection of Listeria spp. strains (155) isolated from natural, agricultural, and urban areas was screened for resistance to heavy metals and metalloids, and the presence of resistance determinants and extrachromosomal replicons. Of the tested strains, 35% were resistant to cadmium and 17% to arsenic. Sequence analysis of resistance plasmids isolated from strains of Listeria seeligeri and Listeria ivanovii, and the chromosome of L. seeligeri strain Sr73, identified a novel variant of the cadAC cadmium resistance efflux system, cadA6, that was functional in L. monocytogenes cells. The cadA6 cassette was detected in four Listeria species, including strains of L. monocytogenes, isolated from various countries and sources-environmental, food-associated, and clinical samples. This resistance cassette is harbored by four novel composite or non-composite transposons, which increases its potential for horizontal transmission. Since some cadAC cassettes may influence virulence and biofilm formation, it is important to monitor their presence in Listeria spp. strains inhabiting different environments.

Listeria valentina sp. nov., isolated from a water trough and the faeces of healthy sheep.

In the context of a study on the occurrence of Listeria species in an animal farm environment in Valencia, Spain, six Listeria-like isolates could not be assigned to any known species. Phylogenetic analysis based on the 16S rRNA gene and on 231 Listeria core genes grouped these isolates in a monophyletic clade within the genus Listeria, with highest similarity to Listeria thailandensis. Whole-genome sequence analyses based on in silico DNA-DNA hybridization, the average nucleotide blast and the pairwise amino acid identities against all currently known Listeria species confirmed that these isolates constituted a new taxon within the genus Listeria. Phenotypically, these isolates differed from other Listeria species mainly by the production of acid from inositol, the absence of acidification in presence of methyl α-d-glucoside, and the absence of α-mannosidase and nitrate reductase activities. The name Listeria valentina sp. nov. is proposed for this novel species, and the type strain is CLIP 2019/00642T (=CIP 111799T=DSM 110544T).

Occurrence of Listeria and Escherichia coli in frozen fruit and vegetables collected from retail and catering premises in England 2018-2019.

Frozen vegetables have previously been associated with outbreaks of listeriosis in both the USA and Europe. An outbreak of Listeria monocytogenes serogroup 4 caused 53 cases in five European countries between 2015 and 2018. Whole genome sequencing (WGS) indicated that frozen sweet corn from a producer in Hungary was the source of illness. However, limited data is available on the prevalence of Listeria in frozen produce. A study of frozen fruit and vegetables from catering and retail premises in England was therefore carried out to assess their microbiological quality with respect to Listeria and Escherichia coli. Between December 2018 and April 2019, 1050 frozen fruit and vegetable samples were collected. Of these, 99% were of a satisfactory or borderline microbiological quality. Eleven samples (1%) contained ≥100 cfu/g of Escherichia coli (considered unsatisfactory in products labelled as ready-to-eat). Listeria monocytogenes or other Listeria species were detected in six samples (2%) of fruit compared to 167 samples (24%) of vegetables and six samples (26%) of fruit and vegetable mixes, but none at a level of ≥100 cfu/g. Characterisation by WGS of 74 L. monocytogenes isolates identified ten genetic clusters indicating a common source. For 8 of the 10 clusters, the isolates came from homogenous food types: four were sweet corn, and there was one cluster each for beans, peas, peppers and broccoli. There were five genetic associations between isolates from frozen vegetables and from clinical cases of listeriosis, including two cultures from frozen beans that were indistinguishable from the 2015-2018 sweet corn outbreak strain. This study indicates that L. monocytogenes was present in 10% of frozen vegetables and even though products are generally not ready-to-eat and are intended to be cooked prior to consumption, these have the potential to cause illness. Clear cooking and handling instructions are therefore required on these products to ensure that the health of consumers is not put at risk, and appropriate Good Manufacturing Practice measures should be followed by all fruit and vegetable freezing plants in order to reduce contamination with Listeria during processing.

Prevalence of Listeria species and L. monocytogenes in ready-to-eat foods in the West Pomeranian region of Poland: Correlations between the contamination level, serogroups, ingredients, and producers.

Poland is one of the largest food producers in Europe, and the West Pomeranian region of Poland is a large producer of RTE food. Thus, the objective of this study was to determine the prevalence of Listeria sp. and L. monocytogenes (LM) in RTE foods manufactured by 13 selected Polish food producers whose processing plants are located in this region. In total, 650 samples of RTE foods, and 263 ingredients of salads and desserts were analyzed. Almost 18% of the RTE foods failed to meet the zero tolerance limit for Listeria, which means they should not be allowed for retail. LM was isolated from 13.5% of the samples, with counts of 10-100 CFU/g noted in half of them. Products with meat and dairy ingredients, and fish products, sandwiches, sprouts and sushi, were at the highest statistically significant risk of LM contamination. Four serogroups were identified among the LM isolated from RTE foods, of which the 4b-4d-4e serogroup was predominating. The samples most heavily contaminated with LM contained even 2 serogroups. Results were subjected to the cluster analysis and principal component analysis to determine correlations between food groups, food ingredients, producers, contamination level, and serogroups of LM.

Prevalence of Listeria spp. in produce handling and processing facilities in the Pacific Northwest.

Listeria monocytogenes is a significant concern for the produce industry; however, there is limited information to support the practical decision-making to mitigate this risk. This study investigated the prevalence of Listeria spp. and L. monocytogenes in seven produce handling and processing (PHP) facilities in the Pacific Northwest. PHP facilities were defined as facilities that receive raw agricultural commodities and further handle, pack, wash, or process prior to distribution into the retail sector. Environmental swabs (n = 50/facility) were collected in high-risk areas (e.g., near raw product entry points) from seven PHP facilities over two visits. Listeria spp. were isolated using modified ISO 11290-1 method and speciated with Microgen® Listeria-ID. Listeria spp., including L. monocytogenes, were found in 5/7 PHP. Prevalence of Listeria spp. ranged from 2% to 26% in these five facilities. Drains, entry areas, and portable equipment consistently tested positive for Listeria spp. during active production. Two additional sampling rounds (n = 50/round) were conducted in the highest prevalence facility (Facility #1). Overall, Listeria spp. were detected in 44/150 (29.3%) swabs collected from Facility #1. This study demonstrated the high prevalence of Listeria spp. near raw product entry points across PHP facilities.

Prevalence and antimicrobial resistance of Listeria species and molecular characterization of Listeria monocytogenes isolated from retail ready-to-eat foods.

A wide variety of foods can be contaminated with Listeria species, especially L. monocytogenes. Ready-to-eat (RTE) foods are predominantly associated with human listeriosis caused by L. monocytogenes. Therefore, this study aimed to assess the presence of Listeria species in RTE foods and to characterize L. monocytogenes isolates by means of detection of virulence markers, serotypes and genetic relatedness. Of the 300 RTE food samples, 59 (19.7%) were positive for Listeria species: L. innocua (13.3%), L. monocytogenes (5%), L. welshimerii (2.3%), L. grayi subsp. murrayi (1.3%), L. grayi (1%), L. ivanovii (1%) and L. ivanovi subsp. londoniensis (0.3%). All L. monocytogenes isolates identified were positive for the actA, iap, inlA, inlB, inlC, inlJ, plcA and prfA virulence genes and biofilm. The isolates were serotyped as 1/2c (33.3%), 4b (26.7%), 1/2a (26.7%), 1/2b (6.7%) and 3c (6.7%) by the multiplex-PCR and agglutination methods. PCR-restriction fragment length polymorphism with AluI and MluCI resulted in three and two profiles, respectively. Pulsed-field gel electrophoresis differentiated the L. monocytogenes isolates into 15 ApaI and 12 AscI patterns. Antimicrobial resistance of all Listeria isolates was determined by the disk diffusion method. Most L. monocytogenes isolates were sensitive to antimicrobials used in the treatment of listeriosis. This study shows the presence of potential pathogenic and antimicrobial-resistant L. monocytogenes in RTE foods that may lead to consumer health risks.

Sentinel Listeriosis Surveillance in Selected Hospitals, China, 2013-2017.

During 2013-2017, a total of 211 cases of listeriosis were reported by 64 sentinel hospitals in China to a national foodborne disease surveillance network. The average case-fatality rate was 31.2% for perinatal cases and 16.4% for nonperinatal cases. Sequence types 87 and 8 were the most prevalent types.

Immuno-detection and differentiation of Listeria monocytogenes and Listeria ivanovii in stone fruits.

AIMS: The aim of this study was to develop a rapid detection and differentiation method for pathogenic Listeria species in stone fruits. METHODS AND RESULTS: We utilized activated charcoal enrichment media (ACM) to induce overexpression and hypersecretion of pathogenic Listeria virulence proteins which can subsequently be detected via immunoblot analysis. Plum and nectarine slices spiked with either L. monocytogenes or L. ivanovii were incubated in pre-enrichment broth followed by enrichment in ACM. Secreted proteins were precipitated and subjected to SDS-PAGE and immunoblot analysis using a combination of L. monocytogenes-specific antibody (α-listeriolysin O) and antibody specific for both L. monocytogenes and L. ivanovii (α-Internalin C). As low as 1 CFU per gram of L. monocytogenes in plum and nectarine was detected, whereas a detection limit of 10 CFU per gram was achieved for L. ivanovii in each food tested following a 20-h enrichment period. Nonpathogenic Listeria species and non-Listeria bacterial pathogens tested were negative. CONCLUSIONS: These results demonstrate the highly sensitive and specific nature of the detection method for pathogenic Listeria in stone fruits using activated charcoal enrichment as well as the capability to discriminate between L. monocytogenes and L. ivanovii. SIGNIFICANCE AND IMPACT OF THE STUDY: This method is the first to identify and differentiate L. monocytogenes and L. ivanovii in select stone fruit enrichments within 24 h using immunological techniques. The rapidity and sensitivity of the method could aid in the reduction of exposure to the public in the event of an outbreak and expedite the administration of appropriate antibiotics to infected individuals.

Multistate Outbreak of Listeriosis Associated with Packaged Leafy Green Salads, United States and Canada, 2015-2016.

We investigated an outbreak of listeriosis detected by whole-genome multilocus sequence typing and associated with packaged leafy green salads. Nineteen cases were identified in the United States during July 5, 2015-January 31, 2016; isolates from case-patients were closely related (median difference 3 alleles, range 0-16 alleles). Of 16 case-patients interviewed, all reported salad consumption. Of 9 case-patients who recalled brand information, all reported brands processed at a common US facility. The Public Health Agency of Canada simultaneously investigated 14 cases of listeriosis associated with this outbreak. Isolates from the processing facility, packaged leafy green salads, and 9 case-patients from Canada were closely related to US clinical isolates (median difference 3 alleles, range 0-16 alleles). This investigation led to a recall of packaged leafy green salads made at the processing facility. Additional research is needed to identify best practices and effective policies to reduce the likelihood of Listeria monocytogenes contamination of fresh produce.

[Fluorescence immunosorbent assay for quantitative detection of Listeria monocytogenes].

OBJECTIVE: Establishment of fuorescence immunosorbent assay for quantitative detection of Listeria monocytogenes. METHODS: The coupled mAbs named 10 E7 H6 and 10 A11 were screened from seven strains of anti-L. monocytogenes mAbs using a sandwich enzyme-linked immunosorbent assay(ELISA). The fluorescent immunoassay was established for L. monocytogenes detection by using biotinylated 10 A11 mAbs as detection antibody, 10 E7 H6 mAbs as capture antibody, and streptavidin-labeled fluorescent microspheres as detection probes, respectively. RESULTS: The optimum concentrations of capture and detection antibodies were 10 µg/mL and 5 µg/mL, respectively, and the optimum reaction pH was 7. 4. Under these conditions, the limit of detection of the proposed method for L. monocytogenes detection was 10~5 CFU/mL, which improved by two orders of magnitude compared to conventional ELISA; and it has a certain cross-reaction with several other Listeria but no significant cross-reactivity with other pathogenic bacteria. CONCLUSION: The method can be used for the detection of L. monocytogenes in pure culture solution, and can also be used for rapid immunological screening test of several other Listeria species in the genus Listeria.

Characterization of Listeria prophages in lysogenic isolates from foods and food processing environments.

Prophages are commonly found in Listeria genomes, potentially enhancing survival or fitness of Listeria spp. Currently, there is still limited information on the distribution of prophages among Listeria isolates of different allelic types and from various sources. In this study, by using mitomycin C induction, prophages were found in 23/144 isolates (16.0%), including 13 L. monocytogenes and 10 Listeria spp. isolates, resulting in 28 and 11 induced phages, respectively. These prophage-carrying isolates (lysogens) were obtained from foods and food-related environments presenting 3 common allelic types (ATs) of L. monocytogenes (lineage I, II and IV), 4 ATs of L. innocua and 1 AT of L. welshimeri. The likelihood of prophage-carrying isolates of L. monocytogenes was 14.4 (95% CI: 4.9-35.4), and 18.5 (95% CI: 4.8-50.2) for Listeria spp. The 39 induced phages were classified into 3 lysis groups by the host range test against 9 major serotypes of L. monocytogenes and 5 species of Listeria. Most phages were host-specific with higher ability to lyse L. monocytogenes serotype 4 than other serotypes. The genome size of phages ranged from 35±2 kb to 50±2 kb and belonged to two common phage families, Myoviridae and Siphoviridae. Restriction analysis classified 19 selected phages into 16 restriction profiles, suggesting highly diverse prophages with at least 16 types. This may contribute to the variation in the genomes of Listeria. Information obtained here provides basic knowledge for further study to understand the overall role of prophages in Listeria, including roles in survival or fitness in foods and food processing environments.