Comprehensive Review on the Biocontrol of Listeria monocytogenes in Food Products.

Listeria monocytogenes is a foodborne pathogen that causes listeriosis, a group of human illnesses that appear more frequently in countries with better-developed food supply systems. This review discusses the efficacy of actual biocontrol methods combined with the main types of food involved in illnesses. Comments on bacteriophages, lactic acid bacteria, bacteriocins, essential oils, and endolysins and derivatives, as main biological antilisterial agents, are made bearing in mind that, using them, food processors can intervene to protect consumers. Both commercially available antilisterial products and solutions presented in scientific papers for mitigating the risk of contamination are emphasized. Potential combinations between different types of antilisterial agents are highlighted for their synergic effects (bacteriocins and essential oils, phages and bacteriocins, lactic acid bacteria with natural or synthetic preservatives, etc.). The possibility to use various antilisterial biological agents in active packaging is also presented to reveal the diversity of means that food processors may adopt to assure the safety of their products. Integrating biocontrol solutions into food processing practices can proactively prevent outbreaks and reduce the occurrences of L. monocytogenes-related illnesses.

The impact of MYD88 and PIM1 in mature large B-cell non-Hodgkin lymphomas: Defining element of their evolution and prognosis.

Sequence studies of the entire exome and transcriptome of lymphoma tissues have identified MYD88 and PIM1 as involved in the development and oncogenic signaling. We aimed to determine the frequency of MYD88 and PIM1 mutations, as well as their expressions in conjunction with the clinicopathological parameters identified in mature large B-cell non-Hodgkin lymphomas. The ten-year retrospective study included 50 cases of mature large B-cell lymphoma, diagnosed at the Pathology Department of the Emergency County Hospital of Constanta and Sacele County Hospital of Brasov. They were statistically analyzed by demographic, clinicopathological, and morphogenetic characteristics. We used a real-time polymerase chain reaction technique to identify PIM1 and MYD88 mutations as well as an immunohistochemical technique to evaluate the expressions of the 2 genes. Patients with lymphoma in the small bowel, spleen, brain, and testis had a low-performance status Eastern Cooperative Oncology Group (P = .001). The Eastern Cooperative Oncology Group performance status represented an independent risk factor predicting mortality (HR = 9.372, P < .001). An increased lactate dehydrogenase value was associated with a low survival (P = .002). The international prognostic index score represents a negative risk factor in terms of patient survival (HR = 4.654, P < .001). In cases of diffuse large B-cell lymphoma (DLBCL), immunopositivity of MYD88 is associated with non-germinal center B-cell origin (P < .001). The multivariate analysis observed the association between high lactate dehydrogenase value and the immunohistochemical expression of PIM1 or with the mutant status of the PIM1 gene representing negative prognostic factors (HR = 2.066, P = .042, respectively HR = 3.100, P = .004). In conclusion, our preliminary data suggest that the oncogenic mutations of PIM1 and MYD88 in our DLBCL cohort may improve the diagnosis and prognosis of DLBCL patients in an advanced stage.

The Predictive Role of the Histopathological Scoring System in Adipose Tumors-Lipoma, Atypical Lipomatous Tumor, and Liposarcoma.

Lipomatous tumors are the most frequent soft tissue neoplasms. Sometimes their differential diagnosis is difficult to perform only by microscopic analysis. This study aims to create a histopathological scoring system and highlight the impact of intratumoral microvascular density. This study was conducted over 10 years. We analyzed the main pathogenic pathways (MDM2 and CDK4), as well as the tumor microvascularization (CD31 and CD34) by immunohistochemical tests. We also analyzed the status of the MDM2 gene by CISH. These data, together with the clinical and histopathological information, were statistically analyzed by appropriate tests. We identified 112 eligible cases, with most of the patients being in their sixth decade of life, with a slight predominance of the female sex. We found important associations like tumor location linked to nuclear pleomorphism severity and microvascularization density correlated with atypia severity. Also, we observed that a maximum diameter of a tumor of at least 69 mm is associated with the presence of tumor necrosis. The score designed in this study shows an increased sensitivity and specificity for the diagnosis of lipomas (100%, respectively, 97%), atypical lipomatous tumors (93.8%, respectively, 82.3%), and liposarcomas (100%, respectively, 90.5%). This present study enhances the present data by bringing to attention the histopathological score with a role in differential diagnosis, as well as in the prediction of immunohistochemical and genetic tests. Also, we highlighted the importance of microvascular density, especially in the diagnosis of liposarcomas.

CMV and HIV Coinfection in Women from a Region in Eastern Europe.

(1) Background: Human cytomegalovirus (CMV) infection is one of the most frequent opportunistic infections in immunosuppressed patients. Romania has one of the highest incidences of patients living with human immunodeficiency virus (HIV) which determines an immunosuppressive state. The aim of this study was to establish the prevalence of CMV infection among women living with HIV in Southeastern Romania and also to evaluate and correlate antiretroviral therapy (ART) with CD4 level and CMV disease evolution. (2) Methods: Seventy women living with HIV from Southeastern Romania were screened for CMV infection using antigen quantification. Of these, 50 were included in the study. First, the patients filled out a questionnaire regarding social conditions and other associated diseases. Then, we explored the statistical correlations between the data and HIV status, CD4+ cell counts, viral load, and antiretroviral therapy (ART). (3) Results: Median age of the patients was 33 years. Twenty-nine cases were diagnosed with HIV after sexual life beginning and 21 before. Most of the patients had a CD4 level over 200 cells/µL. ART duration in the CD4 under 200 cells/µL group was a bit longer than that in the CD4 over 200 cells/µL group. Forty-one patients had undetectable viremia. CD4 average value in the lot of patients with undetectable viremia was 704.71 cells/µL and in the lot with detectable viremia was 452.44 cells/µL. Viremia values correlated negatively with CD4 level. A positive correlation between IgG CMV values and ART therapy length was identified. A negative significant correlation between values of IgG CMV and values of CD4 was identified. CD4 value correlated negatively with IgG CMV values and with CMV avidity. (4) Conclusions: IgG CMV values had a weak positive correlation with ART therapy length, and a negative statistically significant correlation with values of CD4. CMV avidity has a negative correlation with CD4 value.

Comparison of Ki67 Proliferation Index in Gastrointestinal Non-Hodgkin Large B-Cell Lymphomas: The Conventional Method of Evaluation or AI Evaluation?

The most common NHL subtype in SEEU is DLBCL (39%), and it manifests with a variety of cellular morphologies and a high proliferation index. Also, the GI tract is the most common site of extranodal NHLs, and most NHLs involving the GI tract are of B-cell lineage, of which diffuse large B-cell lymphoma is the most common subtype, irrespective of location. The last few years have seen digital pathology as a vital technology that has a positive impact on diagnostics, but studies on the use of DP for lymphoma identification, however, are still restricted to only determining whether a tumor is present or absent. Using the example of cases of malignant NHL, we aim to investigate the diagnostic utility of DP using QuPath software in evaluating the proliferation index and the prognostic significance and to show that improved visualization and analysis contribute to the convergence of these complementary diagnostic modalities for lymphomas. The average proliferation index (Ki67) was 58.33% with values between 10% and 85%. After the stratification of the cases, an increased proliferation index was observed in the majority of cases (53.33%), and this aspect was associated with the advanced age of the patients (p = 0.045). Visual assessment provides lower Ki67 values than automated digital image analysis. However, the agreement coefficient between the conventional method and the AI method indicates an excellent level of reliability (ICC1-0.970, ICC2-0.990). The multivariate analysis revealed that in the cases where the proliferation index Ki67 is high (˃70%), the IPI score represents an important risk factor predicting mortality (HR = 10.597, p = 0.033).

Hand hygiene practices during meal preparation-a ranking among ten European countries.

BACKGROUND: The aim of this paper was to map consumers' food hygiene practices from 10 European countries and evaluate which demographic groups are more likely to be exposed to foodborne pathogens and establish a ranking of adherence to food hygiene practices in 10 European countries. METHODS: The research design consisted of a cross-national quantitative consumer survey regarding food safety and hygiene practices during meal preparation (SafeConsume project) and was conducted in ten European countries (France, Denmark, Germany, Greece, Hungary, Norway, Portugal, Romania, Spain and UK). The survey questions were based on recommended hand hygiene practices and on observed practices from a field study performed in 90 European households from six of the countries covered by the survey (France, Hungary, Norway, Portugal, Romania, and UK). SPSS Statistics 26 (IBM Software Group, Chicago, IL) was used for the descriptive and regression analyses of the data. Regression analyses were used to check the relation between demographic characteristics, country of origin and self-reported hand hygiene practices. RESULTS: According to the regression models, families with elderly members aged over 65 showed a higher tendency to follow proper hand washing practices compared to families without elderly members. Meanwhile, families with children under the age of 6 reported being up to twice as likely to wash their hands at critical moments compared to families without children. Overall, taking into consideration the likelihood of washing hands after touching raw chicken and the percentages scores for proper hand cleaning methods and key moments for hand washing, the rank of the countries regarding proper hand hygiene practices was the following: Denmark, Greece, Norway, Romania, Hungary, Germany, UK, Portugal, France, and Spain. CONCLUSIONS: Information and education should point both at the key moments as suggested by the Royal Society for Public Health (RSPH) and the International Scientific Forum on Home Hygiene (IFH) and safe practices. Public health burden generated by improper hand washing may be significantly reduced if education is targeted on consumers' behaviour and practices.

Deterministic Approach and Monte Carlo Simulation to Predict Listeria monocytogenes Time to Grow on Refrigerated Ham: A Study Supporting Risk-based Decisions for Consumers' Health.

This study assessed the growth of Listeria monocytogenes in ready-to-eat (RTE) ham during storage under conditions simulating domestic practices with the intention to offer support in the elaboration of food safety policies that should better protect consumers against food poisoning at home. RTE ham, artificially contaminated at either medium (102-103 CFU/g) or high (104-105 CFU/g) concentration, was stored at both isothermal (4℃ in a refrigerator able to maintain a relatively constant temperature and 5℃ and 7℃ in a refrigerator with fluctuating temperature) and dynamic (5℃ and 7℃ with intermittent exposure to ambient temperature, e.g. 25℃) conditions. Under isothermal conditions, the increasing storage temperature determined a significantly increased (p < 0.05) capacity of L. monocytogenes to grow. The kinetic growth parameters were derived by fitting the Baranyi and Roberts model to the experimental data and, based on the maximum specific growth rates, it was estimated the temperature dependence of L. monocytogenes growth in RTE ham. At medium contamination level, sanitary risk time calculation revealed that, unlike storage at 5℃ and 7℃, storage at 4℃ of the RTE ham extends the time period during which the product is safe for consumption by ∼40 and 52%, respectively. However, the real temperature fluctuations included in the Monte Carlo simulations at low L. monocytogenes counts (1, 5 and 10 CFU/g) have shortened the safety margins. Stochastic models also proved to be useful tools for describing the pathogen's behavior when refrigeration of the RTE ham alternates with periods of ham being kept at room temperature, considered dynamic conditions of growth.

Kitchen layouts and consumers' food hygiene practices: Ergonomics versus safety.

Our paper emphasizes the importance of the kitchen layout in facilitating consumers' food hygiene practices. A significant correlation was found between the sink placement (inside or outside the kitchen) and hygienic practices during food handling based on a survey performed on consumers from ten European countries, indicating that those who had the sink in the kitchen were more likely to perform proper hygiene practices than those who have not. The self-reported practices were supported by observed practices in 64 households from five European countries. The observational study combined with the examination of kitchen layouts revealed that the kitchen work triangle with its apexes represented by the kitchen sink, cooking stove and refrigerator, which is recommended for ergonomic reasons by architects and designers, did not necessarily support food hygiene practices in kitchens. Cross-contamination events were associated with the sink - countertop distances longer than 1 m. Based on this, a new kitchen triangle with its apexes represented by the kitchen sink, working place (usually countertop) and cooking stove, with the distance between the sink and the working place less than 1 m is proposed to be used as norm in kitchen designs for combining ergonomics with safety. This triangle is proposedly named the food safety triangle and is aimed to mitigate the risks of foodborne illnesses by creating an arrangement that facilitates hygiene practices. This study is the first to highlight the importance of implementing the concept of food safety in the kitchen design based on significant correlations between kitchen equipment placement and consumers' food safety practices.

Data on European kitchen layouts belonging to vulnerable consumers (elderly people and young families with children or pregnant women) and risk-takers (young single men).

The data presented here capture the structure of kitchen layouts belonging to consumers vulnerable to foodborne diseases and food risk-takers. Data were collected in the frame of the SafeConsume project by multidisciplinary research teams that visited consumers during preparing a meal and had the possibility to examine their cooking routines. Distances between sink and stove, sink and refrigerator, stove and refrigerator, sink and working place (countertop or table), stove and working place were analyzed to correlate food safety practices applied during cooking with kitchen arrangements. The results arising from analyzing the ergonomics of kitchens versus potential cross-contamination events are presented in Mihalache et al., [1]. These data contribute to a better understanding of real kitchen layouts and can be used as a starting point for future research regarding food safety-oriented arrangements instead of ergonomics-focused designs, for food safety risk assessments, as study cases for explaining specific measures that can be established to improve food handling and hygiene practices in homes and for sociological research pointing consumers' behavior during cooking.

Efficacy of Removing Bacteria and Organic Dirt from Hands-A Study Based on Bioluminescence Measurements for Evaluation of Hand Hygiene When Cooking.

The objective of this study was to evaluate the efficacy of dirt removal (bacteria and organic matter) of several hand-cleaning procedures. The results from the hand hygiene experiment indicated that washing hands with warm water and soap for 20 s is the most effective method investigated when hands are either dirty or greasy. Even if not proper washing, rinsing under running water for 5 s is a cleaning procedure that may significantly reduce the probability of cross-contamination, as it removes 90% of the hands' dirt. Although less effective than water and soap, the usage of antibacterial wipes was significantly more effective than wet wipes, indicating that they are a better choice when water and soap are not available. The results of this study enable us to inform consumers about the effectiveness of hand-cleaning procedures applied in their homes when cooking. Moreover, it can make consumers understand why, during the COVID-19 pandemic, authorities recommended washing hands as a preventive measure of infection and using an anti-bacterial hand gel or wiping hands with an antimicrobial wipe if water and soap are not available.

Cross-contamination of lettuce with Campylobacter spp. via cooking salt during handling raw poultry.

Campylobacter spp. are the most common bacterial pathogens associated with human gastroenteritis in industrialized countries. Contaminated chicken is the food vehicle associated with the majority of reported cases of campylobacteriosis, either by the consumption of undercooked meat or via cross- contamination of ready-to-eat (RTE) foods during the handling of contaminated raw chicken parts and carcasses. Our results indicate that cooking salt (used for seasoning) is a potential vehicle for Campylobacter spp. cross-contamination from raw chicken to lettuce, through unwashed hands after handling contaminated chicken. Cross-contamination events were observed even when the chicken skin was contaminated with low levels of Campylobacter spp. (ca. 1.48 Log CFU/g). The pathogen was recovered from seasoned lettuce samples when raw chicken was contaminated with levels ≥ 2.34 Log CFU/g. We also demonstrated that, once introduced into cooking salt, Campylobacter spp. are able to survive in a culturable state up to 4 hours. After six hours, although not detected following an enrichment period in culture medium, intact cells were observed by transmission electron microscopy. These findings reveal a "novel" indirect cross-contamination route of Campylobacter in domestic settings, and a putative contamination source to RTE foods that are seasoned with salt, that might occur if basic food hygiene practices are not adopted by consumers when preparing and cooking poultry dishes.

Salmonella in eggs: From shopping to consumption-A review providing an evidence-based analysis of risk factors.

Nontyphoidal salmonellae are among the most prevalent foodborne pathogens causing gastrointestinal infections worldwide. A high number of cases and outbreaks of salmonellosis are associated with the consumption of eggs and egg products, and several of these occur at the household level. The aim of the current study is to critically evaluate the current status of knowledge on Salmonella in eggs from a consumer's perspective, analyzing the hazard occurrence and the good practices that should be applied to reduce salmonellosis risk. Following a HACCP (Hazard Analysis and Critical Control Point) based approach, some steps along the food journey were identified as Critical Consumer Handling (CCH)-steps in which consumers, through their behavior or choice, can significantly reduce the level of Salmonella in eggs and egg products. From shopping/collecting to consumption, each of these steps is discussed in this review to provide an evidence-based overview of risk factors of human salmonellosis related to egg consumption. The main message to consumers is to choose Salmonella-free eggs (those that some official entity or producer guarantees that does not contain Salmonella), when available, especially for dishes that are not fully heat treated. Second, as guaranteed Salmonella-free eggs are only available in a few countries, refrigerated storage from the point of collection and proper cooking will significantly reduce the risk of salmonellosis. This will require a revision of the actual recommendations/regulations, as not all ensure that eggs are maintained at temperatures that prevent growth of Salmonella from collection until the time of purchasing.

Analysis of temporal gene regulation of Listeria monocytogenes revealed distinct regulatory response modes after exposure to high pressure processing.

BACKGROUND: The pathogen Listeria (L.) monocytogenes is known to survive heat, cold, high pressure, and other extreme conditions. Although the response of this pathogen to pH, osmotic, temperature, and oxidative stress has been studied extensively, its reaction to the stress produced by high pressure processing HPP (which is a preservation method in the food industry), and the activated gene regulatory network (GRN) in response to this stress is still largely unknown. RESULTS: We used RNA sequencing transcriptome data of L. monocytogenes (ScottA) treated at 400 MPa and 8∘C, for 8 min and combined it with current information in the literature to create a transcriptional regulation database, depicting the relationship between transcription factors (TFs) and their target genes (TGs) in L. monocytogenes. We then applied network component analysis (NCA), a matrix decomposition method, to reconstruct the activities of the TFs over time. According to our findings, L. monocytogenes responded to the stress applied during HPP by three statistically different gene regulation modes: survival mode during the first 10 min post-treatment, repair mode during 1 h post-treatment, and re-growth mode beyond 6 h after HPP. We identified the TFs and their TGs that were responsible for each of the modes. We developed a plausible model that could explain the regulatory mechanism that L. monocytogenes activated through the well-studied CIRCE operon via the regulator HrcA during the survival mode. CONCLUSIONS: Our findings suggest that the timely activation of TFs associated with an immediate stress response, followed by the expression of genes for repair purposes, and then re-growth and metabolism, could be a strategy of L. monocytogenes to survive and recover extreme HPP conditions. We believe that our results give a better understanding of L. monocytogenes behavior after exposure to high pressure that may lead to the design of a specific knock-out process to target the genes or mechanisms. The results can help the food industry select appropriate HPP conditions to prevent L. monocytogenes recovery during food storage.

The complete genome sequence of Listeria monocytogenes strain S2542 and expression of selected genes under high-pressure processing.

OBJECTIVES: The study aims to generate the whole genome sequence of L. monocytogenes strain S2542 and to compare it to the genomes of strains RO15 and ScottA. In addition, we aimed to compare gene expression profiles of L. monocytogenes strains S2542, ScottA and RO15 after high-pressure processing (HPP) using ddPCR. RESULTS: The whole genome sequence of L. monocytogenes S2542 indicates that this strain belongs to serotype 4b, in contrast to the previously reported serotype 1/2a. Strain S2542 appears to be more susceptible to the treatment at 400 MPa compared to RO15 and ScottA strains. In contrast to RO15 and ScottA strains, viable cell counts of strain S2542 were below the limit of detection after HPP (400 MPa/8 min) when stored at 8 °C for 24 and 48 h. The transcriptional response of all three strains to HPP was not significantly different.

Consumer practices and prevalence of Campylobacter, Salmonella and norovirus in kitchens from six European countries.

About 40% of foodborne infections are acquired in the home. The aim of the present study was to track contamination of pathogens during domestic food preparation and link the contamination to preparation practices. Research participants from 87 households in six European countries were observed and interviewed during shopping and preparation of a chicken and vegetable meal. The presence of Salmonella spp., Campylobacter spp. and norovirus on raw chicken, kitchen surfaces, cloths and sponges was determined. The prevalence of Campylobacter on raw chicken varied from 8.3% in Norway (NO) to 80% in France (FR) and Portugal (PT), with a mean prevalence of 57%. Campylobacter was found on half of the products that had been frozen and appeared to be less prevalent on chicken from supermarkets than other sources. Salmonella was found in 8.6% of raw chicken samples, exclusively from Hungary (HU). A relationship between observed practices and spread of pathogens to kitchen surfaces was found only for the use of cutting boards for chicken and/or vegetables. After food preparation, Campylobacter and Salmonella were isolated from 23% (samples derived from HU, RO, UK) and 8.7% (HU), respectively of cutting boards. Research participants in France and Portugal were more likely to buy products that fitted their recipe, with less need for using cutting boards. Using the same board and knife for vegetables after using it for chicken and without washing with detergent was common in Portugal and Romania, but not in the other countries. Contamination with Campylobacter to other kitchen surfaces or washing utensils were found in five households (UK, RO, PT). Rinsing chicken in sinks was common in three countries (PT, HU, RO), and washing vegetables in the same sink was also usual. Prevalence of Norovirus was low, with detection in one out of 451 samples. The participants' awareness of the risk posed by pathogens from raw chicken differed among the six countries, with higher awareness in Norway and the UK than the other countries studied. In conclusion, practices intended to avoid cross-contamination from chicken to kitchen surfaces and washing utensils are not established among consumers in all European countries. Nevertheless, cross-contamination events that disseminate infectious doses of pathogens seems to be rare, probably due to the relatively low levels of pathogens in food combined with food preferences. Food safety interventions must consider the national food culture, preferences, practices and the prevalence and levels of pathogens in food. Emphasis should be on providing and promoting chicken products with lower risk (prevalence of pathogens, ready-to-cook) and safe use of cutting boards.

High-pressure processing-induced transcriptome response during recovery of Listeria monocytogenes.

BACKGROUND: High-pressure processing (HPP) is a commonly used technique in the food industry to inactivate pathogens, including L. monocytogenes. It has been shown that L. monocytogenes is able to recover from HPP injuries and can start to grow again during long-term cold storage. To date, the gene expression profiling of L. monocytogenes during HPP damage recovery at cooling temperature has not been studied. In order identify key genes that play a role in recovery of the damage caused by HPP treatment, we performed RNA-sequencing (RNA-seq) for two L. monocytogenes strains (barotolerant RO15 and barosensitive ScottA) at nine selected time points (up to 48 h) after treatment with two pressure levels (200 and 400 MPa). RESULTS: The results showed that a general stress response was activated by SigB after HPP treatment. In addition, the phosphotransferase system (PTS; mostly fructose-, mannose-, galactitol-, cellobiose-, and ascorbate-specific PTS systems), protein folding, and cobalamin biosynthesis were the most upregulated genes during HPP damage recovery. We observed that cell-division-related genes (divIC, dicIVA, ftsE, and ftsX) were downregulated. By contrast, peptidoglycan-synthesis genes (murG, murC, and pbp2A) were upregulated. This indicates that cell-wall repair occurs as a part of HPP damage recovery. We also observed that prophage genes, including anti-CRISPR genes, were induced by HPP. Interestingly, a large amount of RNA-seq data (up to 85%) was mapped to Rli47, which is a non-coding RNA that is upregulated after HPP. Thus, we predicted that Rli47 plays a role in HPP damage recovery in L. monocytogenes. Moreover, gene-deletion experiments showed that amongst peptidoglycan biosynthesis genes, pbp2A mutants are more sensitive to HPP. CONCLUSIONS: We identified several genes and mechanisms that may play a role in recovery from HPP damage of L. monocytogenes. Our study contributes to new information on pathogen inactivation by HPP.

Molecular profiling of the colon cancer in South-Eastern Romania: Results from the MERCUR study.

ABSTRACT: Colorectal cancer is a heterogeneous disease with multiple epigenetic alterations and different molecular features. The molecular classification is based on 2 major distinct pathways: microsatellite stable pathway and the microsatellite instability pathway. Molecular profiling of colorectal cancer provides important information regarding treatment and prognosis. Aim of the study was to assess the frequency of microsatellite instability in colon cancer and the clinicopathological characteristics of the tumors with high level of microsatellite instability (MSI-H) in our region. The secondary outcome was to assess the frequency of v-raf murine sarcoma viral oncogene homolog B1 (BRAF) mutations in colon cancer.The study included 129 patients with colon cancer fit for surgery. Demographic data, clinical and pathological data, immunohistochemistry staining pattern (4 mismatch repair proteins were investigated), and BRAF gene mutations were assessed. According to microsatellite instability status by polymerase chain reaction, patients were divided into 3 groups: microsatellite stable (MSS) = 108 patients, high level of microsatellite instability (MSI-H) = 15 patients and low level of microsatellite instability (MSI-L) = 6 patients. Different clinicopathological comparisons between MSS and MSI-H patients, and between MSS and MSI-L patients were performed.Microsatellite instability was found in 16.3% patients: 11.6% had MSI-H and 4.7% had MSI-L. Significantly more patients in the MSI-H group than in the MSS group were female (P = .01) and had a family history of colon cancer (P < .001). MSI-H and MSI-L groups were associated with the ascending colon location of the tumors, were mostly type G3, T2, and stage I whereas MSS tumors were mostly G2, pT3, and stage III. Overall, BRAF mutations were identified in 18/129 patients (13.9%). BRAF mutant tumors were predominantly associated with MSI-H and MSI-L tumors. Immunohistochemistry had a sensitivity of 76% and a specificity of 89% in detecting MSI tumors and an accuracy of 87.6%.The frequency of microsatellite instability in our study was 16.3%. MSI-H is a distinct molecular phenotype of colon cancer with particular features: female gender, family history of colorectal cancer, a predilection for the ascending colon, poorly differentiated, predominantly T2, and stage I. The frequency of BRAF mutations was 13.9% and mutations were more often present in the MSI tumors.

Genomic characterization of the most barotolerant Listeria monocytogenes RO15 strain compared to reference strains used to evaluate food high pressure processing.

BACKGROUND: High pressure processing (HPP; i.e. 100-600 MPa pressure depending on product) is a non-thermal preservation technique adopted by the food industry to decrease significantly foodborne pathogens, including Listeria monocytogenes, from food. However, susceptibility towards pressure differs among diverse strains of L. monocytogenes and it is unclear if this is due to their intrinsic characteristics related to genomic content. Here, we tested the barotolerance of 10 different L. monocytogenes strains, from food and food processing environments and widely used reference strains including clinical isolate, to pressure treatments with 400 and 600 MPa. Genome sequencing and genome comparison of the tested L. monocytogenes strains were performed to investigate the relation between genomic profile and pressure tolerance. RESULTS: None of the tested strains were tolerant to 600 MPa. A reduction of more than 5 log10 was observed for all strains after 1 min 600 MPa pressure treatment. L. monocytogenes strain RO15 showed no significant reduction in viable cell counts after 400 MPa for 1 min and was therefore defined as barotolerant. Genome analysis of so far unsequenced L. monocytogenes strain RO15, 2HF33, MB5, AB199, AB120, C7, and RO4 allowed us to compare the gene content of all strains tested. This revealed that the three most pressure tolerant strains had more than one CRISPR system with self-targeting spacers. Furthermore, several anti-CRISPR genes were detected in these strains. Pan-genome analysis showed that 10 prophage genes were significantly associated with the three most barotolerant strains. CONCLUSIONS: L. monocytogenes strain RO15 was the most pressure tolerant among the selected strains. Genome comparison suggests that there might be a relationship between prophages and pressure tolerance in L. monocytogenes.

Characterization of the biofilm phenotype of a Listeria monocytogenes mutant deficient in agr peptide sensing.

Listeria monocytogenes is a food-borne human pathogen and a serious concern in food production and preservation. Previous studies have shown that biofilm formation of L. monocytogenes and presence of extracellular DNA (eDNA) in the biofilm matrix varies with environmental conditions and may involve agr peptide sensing. Experiments in normal and diluted (hypoosmotic) complex media at different temperatures revealed reduced biofilm formation of L. monocytogenes EGD-e ΔagrD, a mutant deficient in agr peptide sensing, specifically in diluted Brain Heart Infusion at 25°C. This defect was not related to reduced sensitivity to DNase treatment suggesting sufficient levels of eDNA. Re-analysis of a previously published transcriptional profiling indicated that a total of 132 stress-related genes, that is 78.6% of the SigB-dependent stress regulon, are differentially expressed in the ΔagrD mutant. Additionally, a number of genes involved in flagellar motility and a large number of other surface proteins including internalins, peptidoglycan binding and cell wall modifying proteins showed agr-dependent gene expression. However, survival of the ΔagrD mutant in hypoosmotic conditions or following exposure to high hydrostatic pressure was comparable to the wild type. Also, flagellar motility and surface hydrophobicity were not affected. However, the ΔagrD mutant displayed a significantly reduced viability upon challenge with lysozyme. These results suggest that the biofilm phenotype of the ΔagrD mutant is not a consequence of reduced resistance to hypoosmotic or high pressure stress, motility or surface hydrophobicity. Instead, agr peptide sensing seems to be required for proper regulation of biosynthesis, structure and function of the cell envelope, adhesion to the substratum, and/or interaction of bacteria within a biofilm.

Characterization of Biofilms Formed by Foodborne Methicillin-Resistant Staphylococcus aureus.

The objective of this study was to evaluate the capacity of 49 methicillin resistant Staphylococcus aureus (MRSA) from foods of animal origin (42 from dairy products and 7 from meat and meat products) to form biofilms. Overall, a higher biofilm biomass was observed for those MRSA strains harboring SCCmec type IV, while 8 MRSA strains (5 from dairy products and 3 from meat and meat products) were classified as strong biofilm formers in standard Tryptic Soy Broth medium. When a prolonged incubation period (48 h) was applied for those 8 MRSA strains, an increased biofilm biomass accumulation was observed during the time course, whereas the number of viable cells within the biofilms decreased as the biomass increased. The capacity of biofilm production correlated pretty well between the experiments using polystyrene microtiter plates and stainless steel micro-well plates, and significant higher values were observed in stainless steel when glucose was added to TSB during the enrichment. Biofilms were further characterized by confocal laser scanning microscope (CLSM), confirming that proteins and α-polysaccharides were the predominant components inside the extracellular polymeric matrix of biofilms formed by MRSA strains. In conclusion, our results confirm that MRSA isolates from foods of animal origin have significant capacity for forming biofilms with a high protein content, which can play a key role for the successful dissemination of MRSA lineages via food. Knowledge of the capacity of MRSA strains to produce biofilms, as well as characterization of the main MRSA biofilms matrix components, can help both to counteract the mechanisms involved in biofilm formation and resistance and to define more rational control strategies by using tailor-made cleaning agents.