Metagenomic analysis of the bacterial microbiome, resistome and virulome distinguishes Portuguese Serra da Estrela PDO cheeses from similar non-PDO cheeses: An exploratory approach.

This study aimed to evaluate the microbiome, resistome and virulome of two types of Portuguese cheese using high throughput sequencing (HTS). Culture-dependent chromogenic methods were also used for certain groups/microorganisms. Eight samples of raw ewe's milk cheese were obtained from four producers: two producers with cheeses with a PDO (Protected Designation of Origin) label and the other two producers with cheeses without a PDO label. Agar-based culture methods were used to quantify total mesophiles, Enterobacteriaceae, Escherichia coli, Staphylococcus, Enterococcus and lactic acid bacteria. The presence of Listeria monocytogenes and Salmonella was also investigated. The selected isolates were identified by 16S rRNA gene sequencing and evaluated to determine antibiotic resistance and the presence of virulence genes. The eight cheese samples analyzed broadly complied with EC regulations in terms of the microbiological safety criteria. The HTS results demonstrated that Leuconostoc mesenteroides, Lactococcus lactis, Lactobacillus plantarum, Lacticaseibacillus rhamnosus, Enterococcus durans and Lactobacillus coryniformis were the most prevalent bacterial species in cheeses. The composition of the bacterial community varied, not only between PDO and non-PDO cheeses, but also between producers, particularly between the two non-PDO cheeses. Alpha-diversity analyses showed that PDO cheeses had greater bacterial diversity than non-PDO cheeses, demonstrating that the diversity of spontaneously fermented foods is significantly higher in cheeses produced without the addition of food preservatives and dairy ferments. Despite complying with microbiological regulations, both PDO and non-PDO cheeses harbored potential virulence genes as well as antibiotic resistance genes. However, PDO cheeses exhibited fewer of these virulence and antibiotic resistance genes compared to non-PDO cheeses. Therefore, the combination of conventional microbiological methods and the metagenomic approach could contribute to improving the attribution of the PDO label to this type of cheese.

Multidrug Resistance in Enterococci Isolated from Cheese and Capable of Producing Benzalkonium Chloride-Resistant Biofilms.

This study aimed to investigate enterococci recovered from eight Portuguese cheeses made with raw ewe's milk, regarding antibiotic resistance, virulence genes, minimum inhibitory concentration (MIC) of benzalkonium chloride (BAC), biofilm formation capacity, and biofilm eradication (MBEC) by BAC. Antimicrobial resistance against seven antibiotics of five groups was evaluated using the disk diffusion method. The presence of the genes that encode resistance to the antibiotics penicillin (blaZ), erythromycin (ermA, ermB, and ermC), vancomycin (vanA and vanB), aminoglycoside (aac(6')-Ie-aph(2″)-Ia), and ß-lactam (pbp5) and the genes that encode virulence factors, frsB, cylA, gelE, esp, and agg, were investigated via multiplex PCR. The susceptibility of planktonic cells to BAC was evaluated by the MIC and MBC values of the isolates, using the broth microdilution method. To assess the biofilm-forming ability and resistance of biofilms to BAC, biofilms were produced on stainless steel coupons, followed by exposure to BAC. The results showed a high resistance to the antibiotics vancomycin (87.5%), erythromycin (75%), tetracycline (50%), and penicillin (37.5%). Multidrug resistance was observed in 68.8% of the isolates. Genes encoding the virulence factors FrsB (frsB) and gelatinase E (gelE) were detected in all isolates. The esp and cylA genes were found in 56.3% and 37.5% of the isolates, respectively. All isolates exhibited a biofilm-forming ability, regardless of incubation time and temperature tested. However, after 72 h at 37 °C, E. faecium and E. faecalis biofilms showed significant differences (p ≤ 0.05). Although most isolates (62.5%) were susceptible to BAC (MIC ≤ 10 mg/L), biofilms of the same isolates were, generally, resistant to the higher concentration of BAC (80 mg/mL) tested. This study using Enterococcus isolates from a ready-to-eat food, such as cheese, reveals the high percentages of vancomycin resistance and multidrug resistance, associated with the presence of virulence genes, in isolates also capable of producing biofilms resistant to BAC, an important active ingredient of many disinfectants. These results emphasize the need for effective control measures to ensure the safety and quality of dairy products.

The Socioeconomic Factors of Street Food Vending in Developing Countries and Its Implications for Public Health: A Systematic Review.

The sale of ready-to-eat (RTE) street food represents an important source of income in many developing countries. However, these foods are frequently implicated in outbreaks of gastrointestinal diseases. Street food vendors face several constraints that hamper improvement in the microbiological quality of their products. The aim of this review was to update knowledge about the main causes of foodborne illnesses in developing countries, including the growing concern with the microbial transmission of antibiotic resistance. Following PRISMA guidelines, this systematic review was conducted on original articles published from January 2010 to July 2023. The search was carried out using Scopus, PubMed, Web of Science, Food Science and Technology Abstracts (FSTA), the International Information System for Agricultural Sciences and Technology (AGRIS), as well as isolated searches of relevant articles from Google Scholar. The initial search identified 915 articles, 50 of which were included in this systematic review. The results indicate that, in the majority of the 15 countries examined, women constitute the predominant segment of street food vendors, representing more than 55% of the total number of these vendors. In 11 countries, street food vendors under the age of 18 were identified. Most vendors had a low level of education and, consequently, were unaware of good hygiene practices when handling food. The combination of factors such as poor hygiene practices on the part of food handlers and the lack of facilities, namely, the absence of available potable water, were frequently listed as the main causes of food contamination. Enterobacteriaceae such as Escherichia coli (61.9%), Salmonella (30.1%), and Shigella spp. (9.5%), as well as Staphylococcus aureus (30.1%) and Listeria monocytogenes (14.3%), were the most common pathogens found in RTE street foods. In 22 studies from 13 developing countries, 59% (13/22) reported high multidrug resistance in Enterobacteriaceae (40% to 86.4% in E. coli, 16.7 to 70% in Salmonella, and 31 to 76.4% in S. aureus). To address the challenges faced by street vendors and improve their economic activities, it is necessary for government entities, consumers, and vendors to work together collaboratively.

A rare cause of acute abdomen after a Good Friday.

Small bowel perforation caused by an ingested fish bone is rare but can involve the appendix or Meckel's diverticulum. We report the case of a 25-year-old man who presented to the emergency department with acute abdomen caused by perforation of a Meckel's diverticulum with a fish bone ingested in a Good Friday.

Storage Stability and In Vitro Bioaccessibility of Microencapsulated Tomato (Solanum Lycopersicum L.) Pomace Extract.

Tomato pomace is rich in carotenoids (mainly lycopene), which are related to important bioactive properties. In general, carotenoids are known to react easily under environmental conditions, which may create a barrier in producing stable functional components for food. This work intended to evaluate the storage stability and in vitro release of lycopene from encapsulated tomato pomace extract, and its bioaccessibility when encapsulates were incorporated in yogurt. Microencapsulation assays were carried out with tomato pomace extract as the core material and arabic gum or inulin (10 and 20 wt%) as wall materials by spray drying (160 and 200 °C). The storage stability results indicate that lycopene degradation was highly influenced by the presence of oxygen and light, even when encapsulated. In vitro release studies revealed that 63% of encapsulated lycopene was released from the arabic gum particles in simulated gastric fluid, whereas for the inulin particles, the release was only around 13%. The feed composition with 20% inulin showed the best protective ability and the one that enabled releasing the bioactives preferentially in the intestine. The bioaccessibility of the microencapsulated lycopene added to yogurt increased during simulated gastrointestinal digestion as compared to the microencapsulated lycopene alone. We anticipate a high potential for the inulin microparticles containing lycopene to be used in functional food formulations.

A high level of antibiotic resistance in Klebsiella and Aeromonas isolates from street water sold in Mozambique, associated with the prevalence of extended-spectrum and AmpC ß-lactamases.

This study aims to evaluate the resistance profile and the prevalence of antibiotic resistance genes in 30 isolates of Klebsiella spp. and Aeromonas spp. recovered from water sold in the streets of Maputo. Susceptibility profiles to 15 antibiotics were performed according to Clinical Laboratory Standard Institute guidelines with antibiotic disks on Mueller-Hinton agar plates. Multiplex PCRs were performed targeting 10 ß-lactamase genes, five ESBL (blaTEM-variants, blaOXA-variants, BlaSHV-variants, MCTX-M Group 1 and Group 9 variants) and five AmpC (ACC variants, FOX variants, MOX variants, CIT variants and DHA variants). The results showed a high prevalence of Klebsiella resistance to ß-lactam antibiotics, such as amoxicillin/clavulanic acid (62.5%), amoxicillin (56.3%), ampicillin (50%), cefoxitin (43.8%), and cefotaxime (43.8%). Aeromonas showed resistance to cefoxitin and ampicillin (71.4%), amoxicillin/clavulanic acid (57.1%) and imipenem (42.9%). ESBL blaOXA-variants, blaSVH-variants, MCTX-M Group 1 variants, and MCTX-M Group 9 variants were the most prevalent b-lactam genes, followed by the b-lactams AmpC, ACC variants and FOX variants. It is extremely important to improve waterborne disease control strategies, especially in terms of public awareness of the potential health implications of multidrug-resistant strains of Klebsiella and Aeromonas, which are often neglected.

Lactobacillus plantarum in Dual-Species Biofilms With Listeria monocytogenes Enhanced the Anti-Listeria Activity of a Commercial Disinfectant Based on Hydrogen Peroxide and Peracetic Acid.

The aim of this work was to investigate the effect of dual-species biofilms of Listeria monocytogenes with Lactobacillus plantarum on the anti-Listeria activity of a hydrogen peroxide/peracetic acid based commercial disinfectant (P3, Oxonia) when using conditions approaching the food industry environment. Nine strains of L. monocytogenes, including eight persistent strains collected from the meat industry and one laboratory control strain, were used in mono and in dual-species biofilms with a strain of L. plantarum. Biofilms were produced on stainless steel coupons (SSCs), at 11°C (low temperature) or at 25°C (control temperature), in TSB-YE (control rich medium) or in 1/10 diluted TSB-YE (mimicking the situation of biofilm formation after a deficient industrial cleaning procedure). The biofilm forming ability of the strains was evaluated by enumeration of viable cells, and the antibiofilm activity of P3 was assessed by the log reduction of viable cells on SSC. In both nutrient conditions and at low temperature, there was no significant difference (p > 0.05) between L. monocytogenes biofilm forming ability in mono- and in dual-species biofilms. In dual-species biofilms, L. monocytogenes was the dominant species. However, it was generally more susceptible to the lower concentration of P3 0.5% (v/v) than in pure culture biofilms. The presence of L. plantarum, although without significant interference in the number of viable cells of L. monocytogenes, enhanced the efficacy of the anti-Listeria activity of P3, since dual-species biofilms were easier to control. The results presented here reinforce the importance of the investigation into co-culture biofilms produced in food industry conditions, namely at low temperatures, when susceptibility to sanitizers is being assessed.

High Fecal Contamination and High Levels of Antibiotic-Resistant Enterobacteriaceae in Water Consumed in the City of Maputo, Mozambique.

In the city of Maputo, Mozambique, food and water are often sold on the streets. Street water is packaged, distributed, and sold not paying attention to good hygienic practices, and its consumption is often associated with the occurrence of diarrheal diseases. Coincidentally, the increase of diarrheal diseases promotes the inappropriate use of antibiotics that might cause the emergence of antibiotic-resistant bacterial strains. In this context, the present study aimed to assess the microbiological quality of water sold on the streets of Maputo, as well as the antibiotic resistance profile of selected Enterobacteriaceae isolates. The 118 water samples analyzed were from street home-bottled water (n = 81), municipal water distribution systems (tap water) (n = 25), and selected supply wells in several neighborhoods (n = 12). The samples were analyzed for total mesophilic microorganisms, fecal enterococci, fecal coliforms, Escherichia coli, and Vibrio spp. The results showed a high level of fecal contamination in all types of water samples. In home-bottled water, fecal coliforms were found in 88% of the samples, and E. coli in 66% of the samples. In tap water, fecal coliforms were found in 64%, and E. coli in 28% of the samples. In water from supply wells, fecal coliforms and E. coli were found in 83% of the samples. From 33 presumptive Vibrio spp. colonies, only three were identified as V. fluvialis. The remaining isolates belonged to Aeromonas spp. (n = 14) and Klebsiella spp. (n = 16). Of 44 selected Enterobacteriaceae isolates from water samples (28 isolates of E. coli and 16 isolates of Klebsiella spp.), 45.5% were not susceptible to the beta-lactams ampicillin and imipenem, 43.2% to amoxicillin, and 31.8% to amoxicillin/clavulanic acid. Regarding non-beta-lactam antibiotics, there was a high percentage of isolates with tolerance to tetracycline (52.3%) and azithromycin (31.8%). In conclusion, water in Maputo represents a risk for human health due to its high fecal contamination. This situation is made more serious by the fact that a relatively high percentage of isolates with multidrug resistance (40%) were found among Enterobacteriaceae. The dissemination of these results can raise awareness of the urgent need to reduce water contamination in Maputo and other cities in Mozambique.

Pineapple (Ananas comosus L.) By-Products Valorization: Novel Bio Ingredients for Functional Foods.

Pineapple is consumed on a large scale around the world due to its appreciated sensorial characteristics. The industry of minimally processed pineapple produces enormous quantities of by-products (30-50%) which are generally undervalued. The end-of-life of pineapple by-products (PBP) can be replaced by reuse and renewal flows in an integrated process to promote economic growth by reducing consumption of natural resources and diminishing food waste. In our study, pineapple shell (PS) and pineapple core (PC), vacuum-packed separately, were subjected to moderate hydrostatic pressure (225 MPa, 8.5 min) (MHP) as abiotic stress to increase bromelain activity and antioxidant capacity. Pressurized and raw PBP were lyophilized to produce a stable powder. The dehydrated samples were characterized by the following methodologies: chemical and physical characterization, total phenolic compounds (TPC), antioxidant capacity, bromelain activity, microbiology, and mycotoxins. Results demonstrated that PBP are naturally rich in carbohydrates (66-88%), insoluble (16-28%) and soluble (2-4%) fiber, and minerals (4-5%). MHP was demonstrated to be beneficial in improving TPC (2-4%), antioxidant activity (2-6%), and bromelain activity (6-32%) without affecting the nutritional value. Furthermore, microbial and mycotoxical analysis demonstrated that powdered PC is a safe by-product. PS application is possible but requires previous decontamination to reduce the microbiological load.

High Fecal Contamination and High Levels of Antibiotic-Resistant Enterobacteriaceae in Water Consumed in the City of Maputo, Mozambique

In the city of Maputo, Mozambique, food and water are often sold on the streets. Street water is packaged, distributed, and sold not paying attention to good hygienic practices, and its consumption is often associated with the occurrence of diarrheal diseases. Coincidentally, the increase of diarrheal diseases promotes the inappropriate use of antibiotics that might cause the emergence of antibiotic-resistant bacterial strains. In this context, the present study aimed to assess the microbiological quality of water sold on the streets of Maputo, as well as the antibiotic resistance profile of selected Enterobacteriaceae isolates. The 118 water samples analyzed were from street home-bottled water (n = 81), municipal water distribution systems (tap water) (n = 25), and selected supply wells in several neighborhoods (n = 12). The samples were analyzed for total mesophilic microorganisms, fecal enterococci, fecal coliforms, Escherichia coli, and Vibrio spp. The results showed a high level of fecal contamination in all types of water samples. In home-bottled water, fecal coliforms were found in 88% of the samples, and E. coli in 66% of the samples. In tap water, fecal coliforms were found in 64%, and E. coli in 28% of the samples. In water from supply wells, fecal coliforms and E. coli were found in 83% of the samples. From 33 presumptive Vibrio spp. colonies, only three were identified as V. fluvialis. The remaining isolates belonged to Aeromonas spp. (n = 14) and Klebsiella spp. (n = 16). Of 44 selected Enterobacteriaceae isolates from water samples (28 isolates of E. coli and 16 isolates of Klebsiella spp.), 45.5% were not susceptible to the beta-lactams ampicillin and imipenem, 43.2% to amoxicillin, and 31.8% to amoxicillin/clavulanic acid. Regarding non-beta-lactam antibiotics, there was a high percentage of isolates with tolerance to tetracycline (52.3%) and azithromycin (31.8%). In conclusion, water in Maputo represents a risk for human health due to its high fecal contamination. This situation is made more serious by the fact that a relatively high percentage of isolates with multidrug resistance (40%) were found among Enterobacteriaceae. The dissemination of these results can raise awareness of the urgent need to reduce water contamination in Maputo and other cities in Mozambique.

Odontogenic cutaneous sinus tracts due to infection with nontuberculous mycobacteria: a report of three cases.

BACKGROUND: Soft tissue or skin infections due to nontuberculous mycobacteria (NTM) have been reported frequently and are mostly associated with trauma or cosmetic interventions like plastic surgery. However, infection with NTM as a result of a dental procedure have rarely been described and the lack of clinical suspicion and a clear clinical manifestation makes diagnosis challenging. CASE PRESENTATION: We report on three patients with a facial cutaneous sinus tract of dental origin, due to an infection with respectively Mycobacterium fortuitum, M. abscessus and M. peregrinum. The infection source was the dental unit waterlines (DUWLs), which were colonized with NTM. CONCLUSIONS: Water of the DUWL can pose a health risk. This report emphasizes the need for quality control and certification of water flowing through DUWLs, including the absence of NTM. Our report also shows the need for a rapid recognition of NTM infections and accurate laboratory diagnosis in order to avoid long-term ineffective antibiotic treatment.

Listeria monocytogenes cells under nutrient deprivation showed reduced ability to infect the human intestinal cell line HT-29.

PURPOSE: This study aimed to evaluate the effect of two types of stress, cold and nutritional, on the viability and the in vitro virulence of the foodborne pathogenic bacteria Listeria monocytogenes. METHODOLOGY: Ten diverse isolates were kept in phosphate-buffered saline (PBS) at optimal (37 °C) or at refrigeration temperature (7 °C), for 1 and 7 days. The viability of the cells [log colony-forming units (c.f.u.)/ml] and their in vitro virulence, before and after storage in these conditions, were investigated. In vitro virulence (log PFA) was evaluated using the human intestinal epithelial cell line HT-29 in plaque-forming assays (PFAs).Results/Key findings. In general, when compared with the conditions at 37 °C, the exposure at 7 °C for 7 days seemed to increase the resistance of the isolates to nutritional stress. Nutritional stress per se acted significantly to decrease the in vitro virulence of the isolates. After 7 days of nutrient deprivation, whether at optimal or at refrigeration temperature, the majority of the isolates assumed a low-virulence phenotype. CONCLUSION: Our results suggest that when L. monocytogenes are in refrigerated post-processing environments that are unable to support their growth they may increase their resistance to nutritional stress and may decrease their virulence. This should be considered when performing risk assessments for refrigerated ready-to-eat (RTE) foods.

Inmunopatogenia de la psoriasis: nuevos avances / Inmunopatogenia of psoriasis. New progress

La psoriasis es una enfermedad inflamatoria crónica de la piel. Su etiología es multifactorial e incluye susceptibilidad genética, factores inmunológicos y múltiples elementos ambientales, que pueden desencadenar y/o exacerbar la enfermedad. En las últimas décadas se han realizado investigaciones minuciosas sobre la patogénesis de la psoriasis, han sido reconocidos varios subtipos de células T que tienen un papel fundamental en el establecimiento de la inflamación en lesiones cutáneas. Los estudios genéticos brindan las bases para la construcción del modelo de la enfermedad, demostrando que las células dendríticas, los linfocitos T y los queratinocitos desempeñan un rol clave en la patología de esta entidad, así como también un conjunto de citoquinas que impulsan la inflamación psoriásica, dentro de las que se incluyen TNFα, IL-22, IL-23 e IL-17, las cuales promueven la respuesta inflamatoria de queratinocitos, y la producción de péptidos antimicrobianos, citoquinas y quimiocinas, perpetuando así la respuesta inflamatoria. En la actualidad, el desarrollo de varios fármacos biológicos altamente eficaces ha revolucionado el tratamiento de la psoriasis en placas de moderada a severa. Estos medicamentos son un reflejo de una mayor comprensión de la patogénesis de la psoriasis, incluyendo la importancia central de IL-23 e IL17 y las diferentes vías de señalización. El objetivo de este trabajo es realizar una revisión crítica de la literatura sobre la psoriasis y los mecanismos implicados en su imnunopatogenia(AU)

Psoriasis is a chronic inflammatory disease of the skin. Its etiology involves several agents such as genetic susceptibility, immunological factors and multiple environmental elements, which can trigger and / or exacerbate the disease. In recent decades thorough research has been conducted on the pathogenesis of psoriasis, several T-cell subtypes that play a key role in the establishment of inflammation in skin lesions have been recognized. Genetic studies provide the basis for the construction of the disease model, demonstrating that dendritic cells, T lymphocytes and keratinocytes play a key role in the pathology of this entity, as well as a set of cytokines that drive psoriatic inflammation , such as include TNF , IL-22, IL-23 and IL-17, which promote the inflammatory response of keratinocytes, and the production of antimicrobial peptides, cytokines and chemokines, thus perpetuating the inflammatory response. At present, the development of several highly effective biological drugs has revolutionized the treatment of moderate to severe plaque psoriasis. These drugs are a reflection of a greater understanding of the pathogenesis of psoriasis, including the central importance of IL-23 and IL-17 and different signaling pathways. The objective of this work is to perform a critical review of the literature on psoriasis and the mechanisms involved in its imnunopathogenesi(AU)

Biofilm Formation and Disinfectant Susceptibility of Persistent and Nonpersistent Listeria monocytogenes Isolates from Gorgonzola Cheese Processing Plants.

The aim of this study was to investigate whether the biofilm-forming ability and/or the disinfectant susceptibility accounted for the persistence of Listeria monocytogenes in Gorgonzola cheese processing plants. For this purpose, a set of 16 L. monocytogenes isolates collected in the 2004-2007 period was analyzed, including 11 persistent isolates collected in different years, within the collection period, and displaying identical or highly correlated pulsotypes. The evaluation of biofilm-forming ability was assessed using crystal violet (CV) staining and the enumeration of viable cells on stainless steel coupons (SSC). Absorbance values obtained with CV staining for persistent and nonpersistent isolates were not significantly different (rm-ANOVA p > 0.05) and the cell counts from nonpersistent isolates showed to be higher compared with persistent isolates (rm-ANOVA p < 0.05). A simulation of disinfectant treatments was performed on spot inoculated coupons in clean and dirty conditions, according to EN 13697, and on biofilms on SSC, grown in nutrient-rich (dirty) and limiting (clean) conditions using acid acetic-hydrogen peroxide (P3) and acid citric-hydrogen peroxide (MS) commercial disinfectants. The treatment was considered effective when a 4 Log reduction in viable cell count was observed. The Log reductions of persistent and nonpersistent isolates, obtained with both the assays in clean and dirty conditions, were compared and no significant differences were detected (rm-ANOVA p > 0.05). A greater influence of organic matter on MS could explain why P3 was efficient in reducing to effective levels the majority of the isolates at the lowest concentration suggested by the manufacturer (0.2% [v/v]), while the same purpose required a higher concentration (1% [v/v]) of MS. In conclusion, our results demonstrate that the persistence of these isolates in Gorgonzola cheese processing plants was linked neither to the biofilm-forming ability nor to their susceptibility to hydrogen peroxide-based disinfectants; therefore, other factors should contribute to the persistent colonization of the dairies.

Lactobacillus plantarum LB95 impairs the virulence potential of Gram-positive and Gram-negative food-borne pathogens in HT-29 and Vero cell cultures.

Listeria monocytogenes, Salmonella enterica and verocytotoxigenic Escherichia coli (VTEC) are amongst the most important agents responsible for food outbreaks occurring worldwide. In this work, two Lactobacillus spp. strains (LABs), Lactobacillus plantarum (LB95) and Lactobacillus paraplantarum (LB13), previously isolated from spontaneously fermenting olive brines, and two reference probiotic strains, Lactobacillus casei Shirota and Lactobacillus rhamnosus GG, were investigated for their ability to attenuate the virulence of the aforementioned pathogens using animal cell culture assays. In competitive exclusion assays, the relative percentages of adhesion and invasion of S. enterica subsp. enterica serovar Enteritidis were significantly reduced when the human HT-29 cell line was previously exposed to LB95. The relative percentage of invasion by Listeria monocytogenes was significantly reduced when HT-29 cells were previously exposed to LB95. In the cytotoxicity assays, the cell-free supernatant of the co-culture (CFSC)of VTEC with LB95 accounted for the lowest value obtained amongst the co-cultures of VTEC with LABs, and was significantly lower than the value obtained with the co-culture of VTEC with the two probiotic reference strains. The cytotoxicity of CFSC of VTEC with both LB95 and LB13 exhibited values not significantly different from the cell-free supernatant of the nonpathogenic E. coli B strain. Our results suggested that LB95 may be able to attenuate the virulence of Gram-positive and Gram-negative food-borne pathogens; together with other reported features of these strains, our data reveal their possible use in probiotic foods due to their interesting potential in preventing enteric infections in humans.

Differences in the Expression of Cold Stress-Related Genes and in the Swarming Motility Among Persistent and Sporadic Strains of Listeria monocytogenes.

The persistence of certain Listeria monocytogenes strains in food-related environments suggests niche adaptation of these strains and therefore constitutes a major risk to consumer health and results in economic losses for the food producer. In this study, a set of 23 L. monocytogenes isolates, including a group of persistent and a group of sporadic strains, was evaluated regarding their swarming motility at 11°C. In each group, significant (p<0.05) differences in motility were observed. The transcript levels of nine cold stress-related genes were analyzed by real-time quantitative PCR in two representatives of persistent (CBISA3077) and sporadic (CBISA3049) strains isolated from the dairy environment, and significant (p<0.05) differences between the two strains were observed. The persistent strain showed significantly higher transcript levels of dtpT and sigB genes, and significantly lower levels of flaA, oppA, lmo1722, and lmo0866 genes. In the persistent strain, the upregulation of sigB, involved in the tolerance to low temperature and to osmotic stress, could account for the persistence of this strain in its original dairy environment. In a similar way, the downregulation of two helicase-encoding genes lmo1722 and lmo0866, in this strain, may be an evolutionary trait that could facilitate cold stress adaptation. Even though this analysis should be extended to more sporadic and more persistent strains, the results presented here strongly suggest gene expression networks differently adjusted, in the two strains, to the low-temperature environment from where they were collected. Moreover, our findings suggest that bacterial motility per se should not be considered a key feature for the persistence of L. monocytogenes in the food environment.

Bisphenol A disrupts transcription and decreases viability in aging vascular endothelial cells.

Bisphenol A (BPA) is a widely utilized endocrine disruptor capable of mimicking endogenous hormones, employed in the manufacture of numerous consumer products, thereby interfering with physiological cellular functions. Recent research has shown that BPA alters epigenetic cellular mechanisms in mammals and may be correlated to enhanced cellular senescence. Here, the effects of BPA at 10 ng/mL and 1 µg/mL, concentrations found in human samples, were analyzed on HT29 human colon adenocarcinona cell line and Human Umbilical Vein Endothelial Cells (HUVEC). Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) transcriptional analysis of the Long Interspersed Element-1 (LINE-1) retroelement showed that BPA induces global transcription deregulation in both cell lines, although with more pronounced effects in HUVEC cells. Whereas there was an increase in global transcription in HT29 exclusively after 24 h of exposure, this chemical had prolonged effects on HUVEC. Immunoblotting revealed that this was not accompanied by alterations in the overall content of H3K9me2 and H3K4me3 epigenetic marks. Importantly, cell viability assays and transcriptional analysis indicated that prolonged BPA exposure affects aging processes in senescent HUVEC. To our knowledge this is the first report that BPA interferes with senescence in primary vascular endothelial cells, therefore, suggesting its association to the etiology of age-related human pathologies, such as atherosclerosis.

Is the exoproteome important for bacterial pathogenesis? Lessons learned from interstrain exoprotein diversity in Listeria monocytogenes grown at different temperatures.

Bacterial exoproteomes vary in composition and quantity among species and within each species, depending on the environmental conditions to which the cells are exposed. This article critically reviews the literature available on exoproteins synthesized by the foodborne pathogenic bacterium Listeria monocytogenes grown at different temperatures. The main challenges posed for exoproteome analyses and the strategies that are being used to overcome these constraints are discussed. Over thirty exoproteins from L. monocytogenes are considered, and the multifunctionality of some of them is discussed. Thus, at the host temperature of 37°C, good examples are provided by Lmo0443, a potential marker for low virulence, and by the virulence factors internalin C (InlC) and listeriolysin O (LLO). Based on the reported LLO-induced mucin exocytosis, a model is proposed for the involvement of extracellular LLO in optimizing the conditions for InlC intervention in the invasion of intestinal epithelial cells. At lower growth temperatures, exoproteins such as flagellin (FlaA) and oligopeptide permease (OppA) may explain the persistence of particular strains in the food industry environment, eventually allowing the development of new tools to eradicate L. monocytogenes, a major concern for public health.

Comparison of Listeria monocytogenes Exoproteomes from biofilm and planktonic state: Lmo2504, a protein associated with biofilms.

The food-borne pathogen Listeria monocytogenes is the causative agent of the severe human and animal disease listeriosis. The persistence of this bacterium in food processing environments is mainly attributed to its ability to form biofilms. The search for proteins associated with biofilm formation is an issue of great interest, with most studies targeting the whole bacterial proteome. Nevertheless, exoproteins constitute an important class of molecules participating in various physiological processes, such as cell signaling, pathogenesis, and matrix remodeling. The aim of this work was to quantify differences in protein abundance between exoproteomes from a biofilm and from the planktonic state. For this, two field strains previously evaluated to be good biofilm producers (3119 and J311) were used, and a procedure for the recovery of biofilm exoproteins was optimized. Proteins were resolved by two-dimensional difference gel electrophoresis and identified by electrospray ionization-tandem mass spectrometry. One of the proteins identified in higher abundance in the biofilm exoproteomes of both strains was the putative cell wall binding protein Lmo2504. A mutant strain with deletion of the gene for Lmo2504 was produced (3119Δlmo2504), and its biofilm-forming ability was compared to that of the wild type using the crystal violet and the ruthenium red assays as well as scanning electron microscopy. The results confirmed the involvement of Lmo2504 in biofilm formation, as strain 3119Δlmo2504 showed a significantly (P < 0.05) lower biofilm-forming ability than the wild type. The identification of additional exoproteins associated with biofilm formation may lead to new strategies for controlling this pathogen in food processing facilities.

The Tat pathway is prevalent in Listeria monocytogenes lineage II and is not required for infection and spread in host cells.

Listeria monocytogenes, a foodborne pathogenic bacterium, remains a serious public health concern due to its frequent occurrence in food products coupled with a high mortality rate. Bacterial pathogenicity depends greatly on the ability to secrete virulence factors to or beyond the bacterial cell surface. The Tat pathway, one of the secretion systems present in L. monocytogenes, was until now only investigated in silico. In L. monocytogenes strain EGDe two genes constitute this pathway, tatC(lmo0361) and tatA(lmo0362). Here we show that tatC and tatA are cotranscribed in a bicistronic- and growth-phase-dependent manner, being downregulated in the stationary phase. An EGDe tatAC mutant strain (EGDe ΔtatAC) was constructed, confirming that the Tat pathway is not essential for L.monocytogenes survival or biofilm-forming ability. When compared to the wild-type EGDe, deletion of tatAC did not decrease the virulence potential of EGDe ΔtatAC in HT-29 human epithelial cell line and even increased (p < 0.05) the virulence potential for mice. Moreover, we show that tat genes are prevalent in L. monocytogenes strains belonging to genetic lineage II and are generally absent from lineage I, which is more associated with human cases, thus excluding the possibility of using the Tat system as a target for novel antimicrobial compounds targeting L.monocytogenes.