Hydrogen peroxide, sodium dichloro-s-triazinetriones and quaternary alcohols significantly inactivate the dry-surface biofilms of Staphylococcus aureus and Pseudomonas aeruginosa more than quaternary ammoniums.

Globally, healthcare-associated infections (HAI) are the most frequent adverse outcome in healthcare delivery. Although bacterial biofilms contribute significantly to the incidence of HAI, few studies have investigated the efficacy of common disinfectants against dry-surface biofilms (DSB). The objective of this study was to evaluate the bactericidal efficacy of seven Environmental Protection Agency (EPA)-registered liquid disinfectants against DSB of Staphylococcus aureus and Pseudomonas aeruginosa. We hypothesized that overall, there will be significant differences among the bactericidal efficacies of tested disinfectants by product type and active ingredient class. We also hypothesized that depending on the species, higher bactericidal efficacies against DSB will be exhibited after 24 h of dehydration compared to 72 h. Wet-surface biofilms of S. aureus and P. aeruginosa were grown following EPA-MLB-SOP-MB-19 and dehydrated for 24 and 72 h to establish DSB. Seven EPA-registered disinfectants were tested against dehydrated DSB following EPA-MLB-SOP-MB-20. Overall, quaternary ammonium plus alcohol, sodium dichloro-s-triazinetrione and hydrogen peroxide products were more efficacious against DSB than quaternary ammoniums for both tested species. While there was no significant difference in the log10 reductions between 24 and 72 h S. aureus biofilms, significantly higher log10 reductions were observed when products were challenged with 24 h P. aeruginosa DSB compared to 72 h P. aeruginosa DSB. Species type, active ingredient class and dry time significantly impact disinfectant efficacy against DSB of S. aureus or P. aeruginosa.

Integrative Assessment of Reduced Listeria monocytogenes Susceptibility to Benzalkonium Chloride in Produce Processing Environments.

For decades, quaternary ammonium compounds (QAC)-based sanitizers have been broadly used in food processing environments to control foodborne pathogens such as Listeria monocytogenes. Still, there is a lack of consensus on the likelihood and implication of reduced Listeria susceptibility to benzalkonium chloride (BC) that may emerge due to sublethal exposure to the sanitizers in food processing environments. With a focus on fresh produce processing, we attempted to fill multiple data and evidence gaps surrounding the debate. We determined a strong correlation between tolerance phenotypes and known genetic determinants of BC tolerance with an extensive set of fresh produce isolates. We assessed BC selection on L. monocytogenes through a large-scale and source-structured genomic survey of 25,083 publicly available L. monocytogenes genomes from diverse sources in the United States. With the consideration of processing environment constraints, we monitored the temporal onset and duration of adaptive BC tolerance in both tolerant and sensitive isolates. Finally, we examined residual BC concentrations throughout a fresh produce processing facility at different time points during daily operation. While genomic evidence supports elevated BC selection and the recommendation for sanitizer rotation in the general context of food processing environments, it also suggests a marked variation in the occurrence and potential impact of the selection among different commodities and sectors. For the processing of fresh fruits and vegetables, we conclude that properly sanitized and cleaned facilities are less affected by BC selection and unlikely to provide conditions that are conducive for the emergence of adaptive BC tolerance in L. monocytogenes. IMPORTANCE Our study demonstrates an integrative approach to improve food safety assessment and control strategies in food processing environments through the collective leveraging of genomic surveys, laboratory assays, and processing facility sampling. In the example of assessing reduced Listeria susceptibility to a widely used sanitizer, this approach yielded multifaceted evidence that incorporates population genetic signals, experimental findings, and real-world constraints to help address a lasting debate of policy and practical importance.

Genomic and Transcriptomic Analysis of Biofilm Formation in Persistent and Transient Listeria monocytogenes Isolates from the Retail Deli Environment Does Not Yield Insight into Persistence Mechanisms.

Persistence of Listeria monocytogenes in retail deli environments is a serious food safety issue, potentially leading to cross-contamination of ready-to-eat foods such as deli meats, salads, and cheeses. We previously discovered strong evidence of L. monocytogenes persistence in delis across multiple states. We hypothesized that this was correlated with isolates' innate characteristics, such as biofilm-forming capacity or gene differences. To test this hypothesis, we sequenced the genomes of 21 L. monocytogenes isolates previously collected longitudinally from the retail deli environment. Isolates were chosen to represent varying attachment capacity and sanitizer tolerance as well as persistence or transience. We used single-nucleotide polymorphism analysis to characterize the isolates' genetic relationships and used BLAST to search the isolates' genomes for antibiotic resistance elements, quaternary ammonium tolerance genes, and stress survival islets. We further chose four isolates for RNA-sequencing analysis and compared their global biofilm transcriptome with their global planktonic transcriptome. We did not find genetic content that explained persistence. The presence of stress survival islet-1 correlated to increased attachment capacity (p < 0.05), but not persistence. Further, the presence of sanitizer tolerance elements was not significantly correlated with phenotypic sanitizer tolerance. Analysis of biofilm versus planktonic gene expression did not show the expected differences in gene expression patterns. Overall, L. monocytogenes persistence in the deli environment is likely a matter of poor sanitation and/or facility design, rather than isolates' biofilm-forming capacity, sanitizer tolerance, or genomic content.

Lytic Capacity Survey of Commercial Listeria Phage Against Listeria spp. with Varied Genotypic and Phenotypic Characteristics.

Listeria monocytogenes is regularly isolated from food processing environments and is endemic in some facilities. Bacteriophage have potential as biocontrol strategies for L. monocytogenes. In this study, the lytic capacity of a commercial Listeria phage cocktail was evaluated against a library of 475 Listeria spp. isolates (426 L. monocytogenes and 49 other Listeria spp.) with varied genotypic and phenotypic characteristics. The lytic capacity of the Listeria phages was measured by spot assays where lysis was scored on a scale of 0-3 (0 = no lysis; 1 = slight lysis; 2 = moderate lysis; 3 = confluent lysis). Only 5% of all tested Listeria spp. isolates, including L. monocytogenes, were either moderately or highly susceptible (score 2 or 3) to lysis by Listeria phage when scores were averaged across temperature and phage concentration; 155 of 5700 treatment (multiplicity of infection [MOI] and temperature) and characteristic (genotype, sanitizer tolerance, and attachment capacity) combinations resulted in confluent lysis (score = 3). Odds ratios for susceptibility to lysis were calculated using multinomial logistic regression. The odds of susceptibility to lysis by phage decreased (p < 0.05) if the L. monocytogenes isolate was previously found to persist or if the phage-bacteria culture was incubated at 30°C; neither isolate persistence or temperature was significant (p ≥ 0.05) when all factors were considered. In addition, lytic efficacy varied (p < 0.05) among pulse field gel electrophoresis (PFGE) pulsotypes and may be affected by host MOI (p < 0.05). There was no effect (p > 0.05) of attachment capacity or sanitizer tolerance on phage susceptibility. This study underscores the complexity of using Listeria phage as a biocontrol for Listeria spp. in food processing facilities and highlights that phage susceptibility is most greatly impacted by genotype. Further studies are needed to evaluate these findings within a processing environment.

Food safety in Peru: A review of fresh produce production and challenges in the public health system.

Peru has a commodities-based economy where agriculture plays an essential role in the nation's development. Among agricultural products, fruits and vegetables are foundational to Peruvian culture and a healthy and nutritious diet. Produce is also the primary income source for thousands of small-scale farmers and producers throughout the country. Peru has significant potential to export agricultural and value-added products. Nevertheless, the Peruvian food chain has weak food safety and quality standards, limiting access to international markets. The inherent lack of food safety surveillance and management systems negatively affects public health. In the past decade, fresh and raw produce has been associated with several foodborne outbreaks worldwide, resulting in significant health and economic losses. This alarming situation for public health officials and regulators has called for the strengthening of produce safety standards and food safety risk management for safer food and to reduce the incidence of foodborne illnesses. This review summarizes the current status of produce safety in Peru and explores opportunities (e.g., policy, university capacity development) toward a safer food system.

Salmonella enterica subsp. enterica Serovar Heidelberg Food Isolates Associated with a Salmonellosis Outbreak Have Enhanced Stress Tolerance Capabilities.

Salmonella enterica serovar Heidelberg is currently the 12th most common serovar of Salmonella enterica causing salmonellosis in the United States and results in twice the average incidence of blood infections caused by nontyphoidal salmonellae. Multiple outbreaks of salmonellosis caused by Salmonella Heidelberg resulted from the same poultry processor, which infected 634 people during 2013 and 2014. The hospitalization and invasive illness rates were 38% and 15%, respectively. We hypothesized that the outbreak strains of Salmonella Heidelberg had enhanced stress tolerance and virulence capabilities. We sourced nine food isolates collected during the outbreak investigation and three reference isolates to assess their tolerance to heat and sanitizers, ability to attach to abiotic surfaces, and invasiveness in vitro We performed RNA sequencing on three isolates (two outbreak-associated isolates and a reference Salmonella Heidelberg strain) with various levels of heat tolerance to gain insight into the mechanism behind the isolates' enhanced heat tolerance. We also performed genomic analyses to determine the genetic relationships among the outbreak isolates. Ultimately, we determined that (i) six Salmonella Heidelberg isolates associated with the foodborne outbreak had enhanced heat tolerance, (ii) one outbreak isolate with enhanced heat tolerance also had an enhanced biofilm-forming ability under stressful conditions, (iii) exposure to heat stress increased the expression of Salmonella Heidelberg multidrug efflux and virulence genes, and (iv) outbreak-associated isolates were likely transcriptionally primed to better survive processing stresses and, potentially, to cause illness.IMPORTANCE This study provides a deep analysis of the intrinsic stress tolerance and virulence capabilities of Salmonella Heidelberg that may have contributed to the length and severity of a recent salmonellosis outbreak. Additionally, this study provides a comprehensive analysis of the transcriptomic response of S. enterica strains to heat stress conditions and compares baseline stationary-phase gene expression among outbreak- and non-outbreak-associated Salmonella Heidelberg isolates. These data can be used in assay development to screen isolates for stress tolerance and subsequent survival. This study adds to our understanding of the strains associated with the outbreak and informs ongoing regulatory discussions on Salmonella in poultry.

Prevalence, Persistence, and Antimicrobial Resistance of Campylobacter spp. from Eggs and Laying Hens Housed in Five Commercial Housing Systems.

Husbandry practices for laying hens in commercial egg production is a topic of interest from a social, economic, and regulatory standpoint. Animal welfare concerns regarding the use of conventional cages have arisen and consumer perceptions of hen welfare have led to a higher demand for cage-free eggs. The aim of this study was to assess the impact of housing systems on prevalence, persistence, and antimicrobial resistance (AMR) of Campylobacter from laying hens and shell eggs. A total of 425 samples were collected over a 10-month period from the North Carolina Layer Performance and Management Test and Campylobacter isolates were identified by serological, biochemical, and molecular tests. Genetic variability was evaluated using pulsed-field gel electrophoresis (PFGE) and AMR testing was performed. Prevalence of Campylobacter spp. ranged from 11.1% in the enrichable cages to 19.7% in the conventional systems. A greater prevalence of Campylobacter was found in the fecal swab samples from free-range birds compared with those of birds housed in the more intensive housing systems (p > 0.05). Overall, 72 isolates were confirmed as Campylobacter spp. by PCR. More than 90% of the isolates (n = 66) were identified as Campylobacter jejuni, followed by Campylobacter coli (n = 6). C. jejuni isolates displayed high levels of resistance to tetracycline (67%). Genetic variability of Campylobacter was high, with more than 20 PFGE patterns identified. Pattern "a" comprised 42% of isolates from all housing systems and was also the most persistent. This study suggests that housing systems of laying hens used for commercial shell egg production may impact the rate of Campylobacter shedding by layers. Isolation rates and tetracycline resistance levels of this pathogen are still of concern, emphasizing the need for well-implemented biosecurity measures on the farm.

Whole-Genome Sequencing Allows for Improved Identification of Persistent Listeria monocytogenes in Food-Associated Environments.

While the food-borne pathogen Listeria monocytogenes can persist in food associated environments, there are no whole-genome sequence (WGS) based methods to differentiate persistent from sporadic strains. Whole-genome sequencing of 188 isolates from a longitudinal study of L. monocytogenes in retail delis was used to (i) apply single-nucleotide polymorphism (SNP)-based phylogenetics for subtyping of L. monocytogenes, (ii) use SNP counts to differentiate persistent from repeatedly reintroduced strains, and (iii) identify genetic determinants of L. monocytogenes persistence. WGS analysis revealed three prophage regions that explained differences between three pairs of phylogenetically similar populations with pulsed-field gel electrophoresis types that differed by ≤3 bands. WGS-SNP-based phylogenetics found that putatively persistent L. monocytogenes represent SNP patterns (i) unique to a single retail deli, supporting persistence within the deli (11 clades), (ii) unique to a single state, supporting clonal spread within a state (7 clades), or (iii) spanning multiple states (5 clades). Isolates that formed one of 11 deli-specific clades differed by a median of 10 SNPs or fewer. Isolates from 12 putative persistence events had significantly fewer SNPs (median, 2 to 22 SNPs) than between isolates of the same subtype from other delis (median up to 77 SNPs), supporting persistence of the strain. In 13 events, nearly indistinguishable isolates (0 to 1 SNP) were found across multiple delis. No individual genes were enriched among persistent isolates compared to sporadic isolates. Our data show that WGS analysis improves food-borne pathogen subtyping and identification of persistent bacterial pathogens in food associated environments.

Persistent and transient Listeria monocytogenes strains from retail deli environments vary in their ability to adhere and form biofilms and rarely have inlA premature stop codons.

Based on recent risk assessments, up to 83% of listeriosis cases from deli meat in the United States are predicted to be from ready-to-eat deli meats contaminated during processing at retail grocery stores. Listeria monocytogenes is known to use sanitizer tolerance and biofilm formation to survive, but interplay of these mechanisms along with virulence potential and persistence mechanisms specific to deli environments had yet to be elucidated. In this study, 442 isolates from food and nonfood contact surfaces in 30 retail delis over 9 months were tested for inlA premature stop codons (PMSCs); inlA encodes InlA, which is necessary to cause listeriosis. A total of 96 isolates, composed of 23 persistent and 73 transient strains, were tested for adhesion and biofilm-forming ability and sanitizer tolerance. Only 10/442 isolates had inlA PMSCs (p<0.001). Strains with PMSCs were not persistent, even in delis with other persistent strains. Most (7/10) PMSC-containing isolates were collected from food contact surfaces (p<0.001); 6/10 PMSC-containing isolates were found in moderate prevalence delis (p<0.05). Persistent strains had enhanced adhesion on day 1 of a 5-day adhesion-biofilm formation assay. However, there was no significant difference in sanitizer tolerance between persistent and transient strains. Results suggest that foods contaminated with persistent L. monocytogenes strains from the retail environment are (1) likely to have wild-type virulence potential and (2) may persist due to increased adhesion and biofilm formation capacity rather than sanitizer tolerance, thus posing a significant public health risk.

Transcriptional and phenotypic responses of Listeria monocytogenes to chlorine dioxide.

Significant food-borne disease outbreaks have occurred from consumption of ready-to-eat foods, including produce, contaminated with Listeria monocytogenes. Challenging food matrices (e.g., cantaloupe, sprouts) with limited processing steps postharvest to reduce pathogen loads have underscored a need for new mitigation strategies. Chlorine dioxide (ClO2) is increasingly being used in produce and other food systems to reduce food-borne pathogen levels. The goal of this study was to characterize the transcriptional response and survival of L. monocytogenes 10403S exposed to ClO2. The transcriptional profile of log-phase cells exposed to 300 mg/liter ClO2 for 15 min was defined by whole-genome microarray. A total of 340 genes were significantly differentially expressed. Among the differentially expressed genes, 223 were upregulated (fold change ≥ 1.5; adjusted P value < 0.05) in role categories responsible for protein fate, cellular processes, and energy metabolism. There were 113 and 16 genes differentially expressed belonging to regulatory networks of σ(B) and CtsR, respectively. We assessed L. monocytogenes 10403S survival after exposure to 100, 300, and 500 mg/liter aqueous ClO2 in brain heart infusion (BHI) broth; there was a significant difference between cells exposed to 500 mg/liter ClO2 and those exposed to all other conditions over time (P value < 0.05). Isogenic ΔsigB and ΔctsR mutants exposed to 300 mg/liter ClO2 were more sensitive to ClO2 than the wild type under the same conditions. These results provide an initial insight into the mechanisms that L. monocytogenes employs to survive sublethal ClO2 and further our understanding of the inactivation mechanisms of this increasingly used sanitizer.

Correction: Wiedmann, M., et al. Exploration of the Role of the Non-Coding RNA SbrE in L. monocytogenes Stress Response. Int. J. Mol. Sci. 2013, 14, 378-393.

The original version of the paper in Section 3.8 reports that "The peptide mass tolerance and fragment mass tolerance values were 10 ppm and 30 mDa, respectively" [1] (p. 387). To help others who may want to use the same methods in the future, the authors would like to correct the wording to: "The peptide mass tolerance and fragment mass tolerance values were 30 ppm and 0.15 Da, respectively. In order to decrease the probability of false peptide identification, only peptides with significance scores above the identity threshold (at the 95% confidence interval), defined by Mascot probability analysis (www.matrixscience.com/help/scoring_help.html#PBM), were considered to be confidently identified and used for protein identification.  Furthermore, only proteins identified in all four iTRAQ samples through at least two peptides with a p-value of <0.05 (expectation value) were further analyzed". The authors would like to apologize for any inconvenience this may have caused to the readers of this journal.

Evaluation of automated floor cleaning, disinfection, and application methods against Staphylococcus aureus.

BACKGROUND: Optimization of automated floor disinfection practices using different application methods and product types is important to ensure that pathogens such as Staphylococcus aureus do not transfer from contaminated floors to other high contact areas resulting in healthcare-associated infections (HAIs). We hypothesized that there would be significant differences among the disinfectants and a cleaner under different application methods. Also, performance of application methods would be dependent upon type of product used. METHODS: We tested and compared efficacies of 5 EPA registered disinfectants and one cleaner using an automated Taski 455B floor cleaner against S aureus ATCC 6538 on 2 meters of contaminated vinyl flooring using 3 application methods. RESULTS: Hydrogen peroxide and quaternary ammonium compounds were more efficacious against S aureus than the neutral cleaner. There were no significant differences among the sampling areas tested and application methods regardless of product type. Mean log10 densities recovered from different machine parts and wastewater collected were statistically higher for the cleaner than disinfectants. CONCLUSIONS: All disinfectants had more bactericidal efficacy than the cleaner for all sampling zones on the tested floor. Overall, performance of the floor machine is dependent upon the type of product used.

Contact time has limited impact on the efficacy of disinfectant towelettes when tested under conditions reflective of realistic use.

BACKGROUND: Disinfectant towelettes are increasingly being used as a means to prevent transmission of clinically important pathogens which could lead to healthcare-associated infections (HAIs). However, the efficacy of disinfectant towelette products when tested under realistic use conditions is understudied. A test model was designed to replicate realistic wiping conditions. The objective of this study was to determine the impact of varied contact time on disinfectant towelette efficacy under these conditions. METHODS: Five product types were tested against Staphylococcus aureus (ATCC 6538) and Pseudomonas aeruginosa (ATCC 15,442) at five contact times (30 s, one min, two min, three min, and 10 min) on hard, non-porous laminate templates to determine the impact of contact time on disinfectant towelette efficacy when tested under realistic use. RESULTS: Product type had a significant impact on the efficacy of disinfectant towelettes when tested under conditions reflective of realistic use. The effect of contact time was limited and no differences in efficacy were seen at a contact time of one min compared with the other contact times tested. Only one disinfectant towelette product achieved a mean 5-log reduction under the tested conditions. CONCLUSION: Efficacy of disinfectant towelettes was primarily impacted by product type when applied in a model designed to replicate realistic use in which only a limited effect of contact time was observed. There is a need for further investigation into which factors have the greatest impact on disinfectant towelette efficacy when applied in clinical settings.

Contact time and disinfectant formulation significantly impact the efficacies of disinfectant towelettes against Candida auris on hard, non-porous surfaces.

There has been an increase in Candida auris healthcare-associated infections, which result from cross-contamination from surfaces and equipment. In this study, we tested the efficacies of EPA-registered disinfectant towelettes products that are increasingly used for infection control against C. auris at a range of contact times following modifications to standard EPA protocol MB-33-00. Hydrogen peroxide (HP)-based disinfectant towelettes were more efficacious against C. auris than the quaternary ammonium chloride (QAC)-alcohol-based disinfectant towelettes irrespective of tested contact times. Thirty s contact time was significantly less effective in reducing C. auris compared to 1-, 2-, 3-, and 10-min contact times. However, there were no statistically significant differences in the level of disinfection among 1-min and longer contact times regardless of product chemistry. None of the products achieved a standard six-log10 reduction at any tested contact times. Overall, the HP-based disinfectant towelette was significantly more fungicidal than the QAC-alcohol-based disinfectant towelette. For all product types, 30 s contact time did not achieve the same level of disinfection as 1-min or longer contact times. Overall, disinfectant towelette efficacy is dependent upon product formulation and contact time.

Structured expert elicitation about Listeria monocytogenes cross-contamination in the environment of retail deli operations in the United States.

Listeria monocytogenes is among the foodborne pathogens with the highest death toll in the United States. Ready-to-eat foods contaminated at retail are an important source of infection. Environmental sites in retail deli operations can be contaminated. However, commonly contaminated sites are unlikely to come into direct contact with food and the public health relevance of environmental contamination has remained unclear. To identify environmental sites that may pose a considerable cross-contamination risk, to elucidate potential transmission pathways, and to identify knowledge gaps, we performed a structured expert elicitation of 41 experts from state regulatory agencies and the food retail industry with practical experience in retail deli operations. Following the "Delphi" method, the elicitation was performed in three consecutive steps: questionnaire, review and discussion of results, second questionnaire. Hands and gloves were identified as important potential contamination sources. However, bacterial transfers to and from hands or gloves represented a major data gap. Experts agreed about transfer probabilities from cutting boards, scales, deli cases, and deli preparation sinks to product, and about transfer probabilities from floor drains, walk-in cooler floors, and knife racks to food contact surfaces. Comparison of experts' opinions to observational data revealed a tendency among experts with certain demographic characteristics and professional opinions to overestimate prevalence. Experts' votes clearly clustered into separate groups not defined by place of employment, even though industry experts may have been somewhat overrepresented in one cluster. Overall, our study demonstrates the value and caveats of expert elicitation to identify data gaps and prioritize research efforts.

Exploration of the role of the non-coding RNA SbrE in L. monocytogenes stress response.

SbrE is a ncRNA in Listeria monocytogenes, reported to be up-regulated by the alternative sigma factor σB. Initial quantitative RT-PCR (qRT-PCR) experiments on parent strains and isogenic ΔsigB strains demonstrated σB-dependent expression of SbrE across the four L. monocytogenes lineages and in L. innocua. Microarray and proteomics (MDLC/MS/MS with iTRAQ labeling) experiments with the L. monocytogenes parent strain and an isogenic ΔsbrE strain identified a single gene (lmo0636) and two proteins (Lmo0637 and Lmo2094) that showed lower expression levels in the ΔsbrE strain. qRT-PCR demonstrated an increase in SbrE transcript levels in stationary phase L. monocytogenes and in bacteria exposed to oxidative stress (mean log2 transcript levels 7.68 ± 0.57 and 1.70 ± 0.71 greater than in mid-log phase cells, respectively). However, no significant differences in growth or survival between the parent strain and ΔsbrE strain were confirmed under a variety of environmental stress conditions tested. Our data suggest that σB-dependent transcription of SbrE represents a conserved mechanism that contributes, across Listeria species, to fine-tuning of gene expression under specific environmental conditions that remain to be defined.

Listeria monocytogenes 10403S Alternative Sigma-54 Factor σL Has a Negative Role on Survival Ability Under Bile Exposure.

Listeria monocytogenes is a Gram-positive bacterium causing listeriosis in animals and humans. To initiate a foodborne infection, L. monocytogenes has to pass through the host gastrointestinal tract (GIT). In this study, we evaluated survival abilities of L. monocytogenes 10403S wild type (WT) and its isogenic mutants in alternative sigma (σ) factor genes (i.e., sigB, sigC, sigH, and sigL) under simulated gastric, duodenal, and bile fluids. Within 10min of exposures, only bile fluid was able to significantly reduce survival ability of L. monocytogenes WT by 2 logs CFU/ml. Loss of sigL showed the greatest bile resistance among 16 strains tested, p<0.0001, (i.e., WT, four single alternative σ factor mutants, six double mutants, four triple mutants, and one quadruple mutant). To further investigate the role of σL in bile response, RNA-seq was conducted to compare the transcriptional profiles among L. monocytogenes 10403S ΔBCH triple mutant (lacking sigB, sigC, and sigH genes; expressing housekeeping σA and σL) and ΔBCHL quadruple mutant (lacking all alternative sigma factor genes; expressing only σA) strains under BHI and 1% bile conditions. A total of 216 and 176 differentially expressed genes (DEGs) were identified in BHI and bile, respectively. We confirmed that mpt operon was shown to be strongly activated by σL. Interestingly, more than 80% of DEGs were found to be negatively regulated in the presence of σL. This includes PrfA regulon and its mediated genes (i.e., hly, hpt, inlB, clpP, clpE, groL, and inlC) which were downregulated in response to bile in the presence of σL. This result suggests the potential negative role of σL on bile survival, and the roles of σL and σB might be in a seesaw model prior to host cell invasion.

Enterobacteriaceae, coliform, yeast, and mold contamination patterns in peanuts compared to production, storage, use practices, and knowledge of food safety among growers in Senegal.

Peanuts and peanut products are significant revenue sources for smallholder farmers in the Senegalese peanut basin. However, microbial contamination during production and storage can greatly affect market access for producers. Peanut products have emerged as possible sources of foodborne illness, encouraging discussions on international standards for peanuts. In this study, we interviewed 198 households throughout the Senegalese peanut basin to assess current production practices, storage methods, and producers' prior knowledge of microbial contamination using a 162-question survey. A member of each household orally completed the survey with a trained enumerator and the results were compared to microbiological results obtained from peanut samples collected at the time of the interview using linear regression and an analysis of variance model. Samples were collected from stored peanuts at each household; peanuts were shelled and total Enterobacteriaceae, coliform, and yeast and mold populations were enumerated. Of the 198 samples analyzed, 13.0% and 13.6% were greater than the upper detection limits for Enterobacteriaceae and coliforms, respectively. A total of 21.2% of samples were above the detection limit for yeast and mold populations. Only 22.7% and 18.7% of producers were aware of pathogenic bacteria or aflatoxins, respectively; there were no significant differences in observed microbial populations between household who took preventative measures against microbial contamination and those who did not. Additionally, four households reported washing their kitchen utensils before using them to eat and 60.1% reported always washing their hands before eating. Enumerators were asked to report peanut storage container type and if the containers were stored off the ground at the time of collection. While the interaction between storage container type and if the container was stored off the ground was significant for Enterobacteriaceae and coliforms, it was not significant for yeast and mold. Additionally, when storage container type and if peanuts were stored off the ground were included in the regression model, these methods were predictive of contamination levels for Enterobacteriaceae and coliforms. To our knowledge, this is the first study to analyze the relationship among Enterobacteriaceae, coliforms, and yeast and mold contamination and producer knowledge of Senegalese peanuts. These results provide preliminary data to inform future studies to determine pathogen prevalence and impactful preventative measures to minimize microbial contamination of peanuts produced in Senegal.

Alternative σ Factors Regulate Overlapping as Well as Distinct Stress Response and Metabolic Functions in Listeria monocytogenes under Stationary Phase Stress Condition.

Listeria monocytogenes can regulate and fine-tune gene expression, to adapt to diverse stress conditions encountered during foodborne transmission. To further understand the contributions of alternative sigma (σ) factors to the regulation of L. monocytogenes gene expression, RNA-Seq was performed on L. monocytogenes strain 10403S and five isogenic mutants (four strains bearing in-frame null mutations in three out of four alternative σ factor genes, ΔCHL, ΔBHL, ΔBCL, and ΔBCH, and one strain bearing null mutations in all four genes, ΔBCHL), grown to stationary phase. Our data showed that 184, 35, 34, and 20 genes were positively regulated by σB, σL, σH, and σC (posterior probability > 0.9 and Fold Change (FC) > 5.0), respectively. Moreover, σB-dependent genes showed the highest FC (based on comparisons between the ΔCHL and the ΔBCHL strain), with 44 genes showing an FC > 100; only four σL-dependent, and no σH- or σC-dependent genes showed FC >100. While σB-regulated genes identified in this study are involved in stress-associated functions and metabolic pathways, σL appears to largely regulate genes involved in a few specific metabolic pathways, including positive regulation of operons encoding phosphoenolpyruvate (PEP)-dependent phosphotransferase systems (PTSs). Overall, our data show that (i) σB and σL directly and indirectly regulate genes involved in several energy metabolism-related functions; (ii) alternative σ factors are involved in complex regulatory networks and appear to have epistatic effects in stationary phase cells; and (iii) σB regulates multiple stress response pathways, while σL and σH positively regulate a smaller number of specific pathways.

Occurrence of Chemical Contaminants in Peruvian Produce: A Food-Safety Perspective.

The presence of chemical contaminants in agricultural products is a continued food-safety challenge in Peru. This country has robust agriculture potential, but its output of fruits and vegetables is severely impacted by massive mining activities, as well as poor farming practices, including the use of polluted irrigation water, misuse of pesticides, and inadequate postharvest conditions. This review examines the current scientific knowledge on the levels of pesticide residues, heavy metals, and mycotoxins on crops produced in Peru. The available data shows that several crop varieties are contaminated with these classes of chemical contaminants, and at levels that exceed the national and international permissible limits. The abundance of chemical contaminants in produce indicates a relevant food-safety issue, which increases the risks of chronic human diseases, like cancer-a leading cause of death in Peru. Finally, this review presents recommendations to address these contamination problems in produce grown in the Andean country.