Identification and Characterization of a Novel Genomic Island Harboring Cadmium and Arsenic Resistance Genes in Listeria welshimeri

Listeria monocytogenes, the bacterial foodborne pathogen responsible for the severe disease listeriosis, frequently exhibits heavy metal resistance. Concurrent resistance to cadmium and arsenic in L. monocytogenes is strongly associated with the 35-kb chromosomal island LGI2. LGI2 has been encountered repeatedly among L. monocytogenes serotype 4b hypervirulent clones but, surprisingly, not among non-pathogenic Listeria spp. Here we describe a novel LGI2 variant, LGI2-3, in two L. welshimeri strains from an urban aquatic environment. Whole genome sequence analysis revealed that the genomes were closely related except for one prophage region and confirmed a chromosomally integrated LGI2-3. It harbored a cystathionine beta-lyase gene previously only encountered in LGI2-1 of L. monocytogenes clonal complex 1 but was otherwise most closely related to LGI2. LGI2-3 harbored a novel cadAC cassette (cadA7C7) that, like LGI2′s cadA4C4, was associated with lower-level tolerance to cadmium (MIC 50 μg/mL) than other cadAC cassettes (MIC ≥ 140 μg/mL). CadA sequence analysis identified two amino acids that may be important for mediating different levels of cadmium tolerance. Our findings clearly demonstrated the potential for LGI2-like islands to be harbored by non-pathogenic Listeria spp. and generate intriguing hypotheses on the genetic diversity mediated by this island and its transfer among Listeria spp.

Virulence Characterization of Listeria monocytogenes, Listeria innocua, and Listeria welshimeri Isolated from Fish and Shrimp Using In Vivo Early Zebrafish Larvae Models and Molecular Study

Listeriosis is one of the most notable foodborne diseases and is characterized by high rates of mortality. L. monocytogenes is the main cause of human listeriosis outbreaks, however, there are isolated cases of disease caused by other species of the genus Listeria. The aim of this study was to evaluate strains of L. monocytogenes (n = 7), L. innocua (n = 6), and L. welshimeri (n = 2) isolated from fish and shrimps for their virulence based on the presence of virulence genes and the in vivo Danio rerio (zebrafish) larvae models. A total of 15 strains were analyzed. The zebrafish larvae model showed that the larvae injected with L. monocytogenes strains were characterized by the lowest survival rate (46.5%), followed by L. innocua strains (64.2%) and L. welshimeri (83.0%) strains. Multiplex PCRs were used for detection of selected virulence genes (luxS, actA2, prfA, inlB, rrn, iap, sigB, plcB, actA, hlyA), the majority of which were present in L. monocytogenes. Only a few virulence-related genes were found in L. welshimeri, however, no correlation between the occurrence of these genes and larval survival was confirmed. This research highlights the importance of the potential impact that Listeria spp. strains isolated from fish and shrimps may have on consumers.

Surveillance for Contamination of Salad with Listeria Monocytogenes in Some of Baghdad Restaurants

Background: Listeria monocytogenes, a member of the genus Listeria, is widely distributed in agricultural environments, such as soil, manure and water. The genus of Listeria bacteria is about 15-17 species. It is a pathogenic bacterium that can cause a rare but dangerous infection called listeriosis. Objectives: Studying the rate of salads contaminated with Listeria bacteria. and Listeria monocytogenes according to International, Arabic and Iraqi specifications and finding the correlation between commitments of restaurants to standard health conditions with contamination with these bacteria Methods: The study included 152 samples of salads taken from 39 restaurants chosen randomly and of different levels and places in Baghdad from the period between 1/9/2014 to 20/1/2015. The laboratory tests were carried out on samples based on internationally approved methods in addition to methods of the International Standards Organization. Results: The study revealed that 23 samples (15.13%) from the 152 samples taken from the restaurants were contaminated with Listeria species. of these, 3 (2%) were contaminated with Listeria monocytogenes and 20 (13.2%) were contaminated with other types of different and non-pathogenic Listeria as follows; (Listeria welshimeri, Listeria seeligeri, Listeria ivanovii, Listeria grayi, Listeria innocua) with the following prevalence (7(4.6%), 6(3.9%), 3(2%), 3(2%), 1 (0.7%) respectively). Conclusions: Contamination of salads taken from restaurants with Listeria bacteria is not uncommon. This indicates that routine examination is necessary and should be added to the Iraqi standard for salads.

Occurrence and antibiogram of Listeria species in raw pork, beef, and chicken meats marketed in Enugu State, Southeast Nigeria

Aim: This study was undertaken to isolate Listeria (L.) species from raw meats sold in markets in Enugu State, Southeast Nigeria, and to determine the antibacterial resistance profile. Materials and Methods: Twenty-five grams of beef (n=144), chicken meat (n=144), and pork (n=144) were collected randomly from supermarkets and general markets in Enugu State. Isolation of Listeria was done using half and full Fraser broths, and polymyxin acriflavine lithium chloride ceftazidime aesculin mannitol agar. Identification of isolates was done using an analytical profile index kit specific for Listeria. Confirmation of the genus Listeria was done by a polymerase chain reaction. The resistance of the isolates was determined using the disk diffusion method. Results: Listeria was isolated from 39/144 (27.1%) chicken meat, 19/144 (13.2%) pork, and 66/144 (45.8%) beef samples cultured. Listeria innocua was the predominant species in chicken meat (52.6%) and beef (81.8%) samples. Listeria grayi, Listeria welshimeri, and Listeria ivanovii were also isolated from the beef and chicken meat samples. More than 65% of the isolates were resistant to penicillin, rifampicin, ciprofloxacin, sulfamethoxazole-trimethoprim, and cephalothin. All the isolates from beef and pork samples and 23 (92%) from chicken meat samples, were resistant to ≥3 classes of antibacterial agents. Mean multiple antibiotic resistance index (MARI) was 0.77 (range=0.42-1.00), 0.58 (range=0.25-0.83), and 0.79 (range=0.58-0.92) for the isolates from beef, chicken meat, and pork samples, respectively. All the isolates had MARI >0.2. Conclusion: Multidrug-resistant Listeria strains contaminate raw beef, pork, and chicken meats marketed in Enugu State, Southeast Nigeria.

Salmonella, Listeria monocytogenes, and Indicator Microorganisms on Hass Avocados Sold at Retail Markets in Guadalajara, Mexico

ABSTRACT Hass avocados may become contaminated with Salmonella and Listeria monocytogenes at the farm and the packing facility or later during transportation and at retail. In Mexico, avocados are frequently sold in bulk at retail markets, where they are stored at room temperature for several hours or days and exposed to potential sources of microorganisms. These conditions may favor the entry, adhesion, survival, and biofilm formation of Salmonella and L. monocytogenes. The aim of this study was to determine the occurrence of Salmonella, L. monocytogenes, and other Listeria species and the levels of indicator microorganisms on the surface of avocados sold at retail markets. A total of 450 samples (Persea americana var. Hass) were acquired from retail markets located in Guadalajara, Mexico. One group of 225 samples was evaluated for the presence of Salmonella and for enumeration of aerobic plate counts, yeasts and molds, Enterobacteriaceae, coliforms, and Escherichia coli. The other 225 samples were processed for isolation of L. monocytogenes and other Listeria species. Microbial counts (log CFU per avocado) were 4.3 to 9.0 for aerobic plate counts, 3.3 to 7.1 for yeasts and molds, 3.3 to 8.2 for Enterobacteriaceae, 3.3 to 8.4 for coliforms, and 3.3 to 6.2 for E. coli. Eight samples (3.5%) were positive for Salmonella. Listeria spp. and L. monocytogenes were detected in 31 (13.8%) and 18 (8.0%) of 225 samples, respectively. Listeria innocua, Listeria welshimeri, and Listeria grayi were isolated from 7.6, 1.3, and 0.9% of samples. These results indicate that avocados may carry countable levels of microorganisms and could be a vehicle for transmission of Salmonella and L. monocytogenes. HIGHLIGHTS

Distribution of Listeria spp. on Carcasses of Regularly Slaughtered Swine for Italian Dry Cured Ham

ABSTRACT In recent years, the role of Listeria monocytogenes as a foodborne pathogen in public health has increased. Its presence poses a risk for humans, especially in ready-to-eat foods, such as ham. Understanding the presence and distribution of Listeria spp. on swine carcasses meant for Italian dry-cured hams can be a useful tool to improve food safety. This study assessed the distribution of Listeria spp. (as a marker of contamination with L. monocytogenes) on slaughtered pig carcasses intended for the production of high-quality, Italian, dry-cured ham and examined the roles of the site sampled on the carcass, farming cycle (open versus closed), farm-to-slaughterhouse distance, and time spent in lairage. Samples were collected from swine carcasses (n = 150) before refrigeration, from three different carcass locations (head, shoulder, and thigh), and assessed for the presence of listeriae. A total of 115 carcasses were contaminated with Listeria spp. in at least one location. In all, 178 listeriae were isolated and identified: 130 Listeria innocua, 28 Listeria welshimeri, 17 Listeria ivanovii, and 3 L. monocytogenes. Listeriae were detected on 62.7% of heads, on 25.3% of shoulders, and on 30.7% of thighs, with significant differences between heads versus shoulders and thighs. Animals reared in closed-cycle farms were more contaminated (P < 0.05) than were animals from open-cycle farms (90 versus 71.8%). The distance between farms and slaughterhouse was not related to the contamination rate. Carcasses of swine that stayed in lairage before slaughtering for more than 10 h showed a higher degree of contamination (90%) and were positive for Listeria spp. in more sample sites (55%) compared with those held for less than 2 h (73% of carcasses and 33.3% of samples). Our results show that heads should be detached from carcasses immediately after slaughter for meat-safety purposes and the amount of time animals stay in lairage should be limited. These results will be useful for a more-valid implementation of good manufacturing processes for slaughtering. HIGHLIGHTS

Prevalence and Characterization of Listeria Species from Raw Milk and Dairy Products from Çanakkale Province

The objective of this study was to determine the prevalence of Listeria species, specifically Listeria monocytogenes, in raw milk, pasteurized milk, white cheese, and homemade cheese. A total of 200 food samples were collected and analyzed to examine the presence of Listeria spp. The EN ISO 11290-1 method was used for isolation of Listeria. API Listeria test kit was used for biochemically characterization. Listeria spp. were isolated in 25 of the 200 samples (12.5%). The largest number of Listeria spp. was detected in homemade cheese (24%), followed by raw milk (18%), and white cheese (8%). Listeria spp. were not isolated from the pasteurized milk. The most common species isolated were Listeria innocua (5.5%); the remaining Listeria isolates were Listeria ivanovi (3.5%), Listeria welshimeri (3%), and Listeria monocytogenes (0.5%). Listeria monocytogenes was detected in only raw milk.

Listeria monocytogenes and Listeria spp. Contamination Patterns in Retail Delicatessen Establishments in Three U.S. States

Postprocessing contamination in processing plants has historically been a significant source of Listeria monocytogenes in ready-to-eat delicatessen meats, and therefore a major cause of human listeriosis cases and outbreaks. Recent risk assessments suggest that a majority of human listeriosis cases linked to consumption of contaminated deli meats may be due to L. monocytogenes contamination that occurs at the retail level. To better understand the ecology and transmission of Listeria spp. in retail delicatessens, food and nonfood contact surfaces were tested for L. monocytogenes and other Listeria spp. in a longitudinal study conducted in 30 retail delis in three U.S. states. In phase I of the study, seven sponge samples were collected monthly for 3 months in 15 delis (5 delis per state) prior to start of daily operation; in phase II, 28 food contact and nonfood contact sites were sampled in each of 30 delis during daily operation for 6 months. Among the 314 samples collected during phase I, 6.8% were positive for L. monocytogenes. Among 4,503 samples collected during phase II, 9.5% were positive for L. monocytogenes; 9 of 30 delis showed low L. monocytogenes prevalence (<1%) for all surfaces. A total of 245 Listeria spp. isolates, including 184 Listeria innocua, 48 Listeria seeligeri, and 13 Listeria welshimeri were characterized. Pulsed-field gel electrophoresis (PFGE) was used to characterize 446 L. monocytogenes isolates. PFGE showed that for 12 of 30 delis, one or more PFGE types were isolated on at least three separate occasions, providing evidence for persistence of a given L. monocytogenes subtype in the delis. For some delis, PFGE patterns for isolates from nonfood contact surfaces were distinct from patterns for occasional food contact surface isolates, suggesting limited cross-contamination between these sites in some delis. This study provides longitudinal data on L. monocytogenes contamination patterns in retail delis, which should facilitate further development of control strategies in retail delis.