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We investigated bile salts' ability to induce phenotypic changes in biofilm production and protein expression of pathogenic Escherichia coli strains. For this purpose, 82 pathogenic E. coli strains isolated from humans (n = 70), and animals (n = 12), were examined for their ability to form biofilms in the presence or absence of bile salts. We also identified bacterial proteins expressed in response to bile salts using sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-electrophoresis) and liquid chromatography-mass spectrometry (LC-MS/MS). Lastly, we evaluated the ability of these strains to adhere to Caco-2 epithelial cells in the presence of bile salts. Regarding biofilm formation, two strains isolated from an outbreak in Republic of Georgia in 2009 were the only ones that showed a high and moderate capacity to form biofilm in the presence of bile salts. Further, we observed that those isolates, when in the presence of bile salts, expressed different proteins identified as outer membrane proteins (i.e. OmpC), and resistance to adverse growth conditions (i.e. F0F1, HN-S, and L7/L12). We also found that these isolates exhibited high adhesion to epithelial cells in the presence of bile salts. Together, these results contribute to the phenotypic characterization of E. coli O104: H4 strains.

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Fish is a nutritionally rich product; however, it is easily contaminated by pathogenic microorganisms, such as Salmonella spp. Therefore, this study aimed to identify the best concentration of sodium hypochlorite (NaClO), exposure time, and water temperature that allow the most effective antimicrobial effect on the viable population of Salmonella spp. Thus, Salmonella Enteritidis ATCC 13076 and Salmonella Schwarzengrund were exposed to different time frames, ranging from 5 min to 38.5 min, temperatures between 5 and 38.5 °C, and NaClO concentrations ranging from 0.36 to 6.36 ppm, through a central composite rotational design experiment (CCRD). The results demonstrated that the ATCC strain exhibited a quadratic response to sodium hypochlorite when combined with exposure time, indicating that initial contact would already be sufficient for the compound's action to inhibit the growth of the mentioned bacteria. However, for S. Schwarzengrund (isolated directly from fish cultivated in aquaculture), both NaClO concentration and exposure time significantly influenced inactivation, following a linear pattern. This suggests that increasing the exposure time of NaClO could be an alternative to enhance Salmonella elimination rates in fish slaughterhouses. Thus, the analysis indicates that the Salmonella spp. strains used in in vitro experiments were sensitive to concentrations equal to or greater than the recommended ones, requiring a longer exposure time combined with the recommended NaClO concentration in the case of isolates from aquaculture.

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Ultraviolet-C light-emitting diode (UVC-LED) and ultrasound (US) are two nonthermal technologies with the potential to destroy pathogens. However, little is known about their effectiveness in strains with a history of heat resistance. Thus, this study aimed to evaluate the phenotype and genotype of heat-resistant extraintestinal pathogenic Escherichia coli (ExPEC) with heat resistance genes after the application of US, UVC-LED, and UVC-LED+US. For this, two central composite rotatable designs were used to optimize the UVC-LED and US conditions in four ExPEC isolated from beef. From the genome of these isolates obtained in a previous study, possible genes for UVC resistance were analyzed. Results showed that US was ineffective in reducing >0.30 log colony-forming unit/mL, and that when used after UVC-LED, it showed a nonsynergic or antagonistic effect. Also, UVC-LED had the greatest effect at the maximum dose (4950 mJ/cm2 from 1.65 mW/cm2 for 50 min). However, the strains showed some recovery after that, which could be implicated in the expression of genes included in SOS system genes, some others present in the transmissible Locus of Stress Tolerance (trxBC and degP), and others (terC). Thus, ExPEC can overcome the conditions used in this study for US, UVC-LED, and UVC-LED+US, probably due to the history of resistance to other cellular damage. The result of this study will contribute to future studies that aim to find better treatment conditions for each food product.

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AIMS: Characterize Escherichia coli and E. coli -producing (STEC) isolates from Brazilian beef to determine heat resistance and the presence of the transmissible locus of stress tolerance (tLST). METHODS AND RESULTS: Twenty-two STEC previously isolated from beef and characterized as STEC by PCR were subjected to different heat survival challenges (60°C and 71°C). Furthermore, the three tLST-positive isolates and one tLST-negative isolate by PCR were selected for WGS analysis. Phenotypic results indicated that 3/22 (13.64%) were heat resistant, 12/22 (54.54%) were moderately resistant, and 7/22 (31.82%) were sensitive to heat treatments. WGS analyses showed that three isolates with heat resistance showed tLST with up to 80% and 42% of similarity by BLAST analysis, with the major tLST genes being responsible for the homeostasis module. However, WGS showed the absence of stx genes associated with tLST-positive isolates, albeit with virulence and resistance genes found in extraintestinal pathogenic E. coli (ExPEC). CONCLUSION: Our findings demonstrate the presence of heat-resistant E. coli as well as confirm some tLST genes in E. coli isolated from Brazilian beef.

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Escherichia coli harboring a transmissible locus of stress tolerance (tLST) and the ability to form biofilms represent a serious risk in dairy production. Thus, we aimed to evaluate the microbiological quality of pasteurized milk from two dairy producers in Mato Grosso, Brazil, with a focus on determining the possible presence of E. coli with heat resistance (60 °C/6 min), biofilm-forming potential phenotypes and genotypes, and antimicrobial susceptibility. For this, fifty pasteurized milk samples from producers named A and B were obtained for 5 weeks to investigate the presence of Enterobacteriaceae members, coliforms, and E. coli. For heat resistance, E. coli isolates were exposed to a water bath at 60 °C for 0 and 6 min. In antibiogram analysis, eight antibiotics belonging to six antimicrobial classes were analyzed. The potential to form biofilms was quantified at 570 nm, and curli expression by Congo Red was analyzed. To determine the genotypic profile, we performed PCR for the tLST and rpoS genes, and pulsed-field gel electrophoresis (PFGE) was used to investigate the clonal profile of the isolates. Thus, producer A presented unsatisfactory microbiological conditions regarding Enterobacteriaceae and coliforms for weeks 4 and 5, while all samples analyzed for producer B were contaminated at above-the-limit levels established by national and international legislation. These unsatisfactory conditions enabled us to isolate 31 E. coli from both producers (7 isolates from producer A and 24 isolates from producer B). In this way, 6 E. coli isolates (5 from producer A and 1 from producer B) were highly heat resistant. However, although only 6 E. coli showed a highly heat-resistant profile, 97% (30/31) of all E. coli were tLST-positive. In contrast, all isolates were sensitive to all antimicrobials tested. In addition, moderate or weak biofilm potential was verified in 51.6% (16/31), and the expression of curli and presence of rpoS was not always related to this biofilm potential. Therefore, the results emphasize the spreading of heat-resistant E. coli with tLST in both producers and indicate the biofilm as a possible source of contamination during milk pasteurization. However, the possibility of E. coli producing biofilm and surviving pasteurization temperatures cannot be ruled out, and this should be investigated.

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The species Mycobacterium tuberculosis variant bovis (M. tuberculosis var. bovis) is associated with tuberculosis, mainly in cattle and buffaloes. This pathogen has the potential to infect other mammals, including humans. Tuberculosis caused by M. tuberculosis var. bovis is a zoonosis clinically identical to tuberculosis caused by Mycobacterium tuberculosis, and the recommended treatment in humans results in the use of antibiotics. In this study, we used the whole genome sequencing (WGS) methodology Illumina NovaSeq 6000 System platform to characterize the genome of M. tuberculosis var. bovis in cattle circulating in Mato Grosso, identify mutations related to drug resistance genes, compare with other strains of M. tuberculosis var. bovis brazilian and assess potential drug resistance. Four isolates of M. tuberculosis var. bovis of cattle origin representing the main livestock circuits, which had been more prevalent in previous studies in the state of Mato Grosso, were selected for the genomic study. The genome sizes of the sequenced strains ranged from 4,306,423 to 4,332,964 bp, and the GC content was 65.6%. The four strains from Mato Grosso presented resistance genes to pncA (pyrazinamide), characterized as drug-resistant strains. In addition to verifying several point mutations in the pncA, rpsA, rpsL, gid, rpoB, katG, gyrB, gyrA, tlyA, embA, embB, embC, fgd, fbiB, and fbiC genes, these genes were similar to antibiotic resistance in more than 92% of the Brazilian strains. Therefore, our results indicated a high genetic diversity between our isolates and other M. tuberculosis var. bovis isolated in Brazil. Thus, multiple transmission routes of this pathogen may be present in the production chain. So, to achieve a bovine tuberculosis-free health status, the use of the WGS as a control and monitoring tool will be crucial to determine these transmission routes.

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The aim of the present study was to investigate Salmonella behavior in meat stored in cool conditions (between 0 °C and 7.5 °C), by employing a systematic review and meta-analysis. The data were obtained from research articles published in SciELO, PubMed, the Web of Science, and Scopus databases. The results of the retrieved studies were obtained from meat (beef, chicken, pork, poultry, and turkey), fish, shellfish, and broth media samples The data were extracted as sample size (n), initial concentration (Xi), final concentration (Xf), standard deviation (SD), standard error (SE), and microbial behavior effects (reduction or growth). A meta-analysis was carried out using the metaphor package from R software. A total of 654 articles were initially retrieved. After applying the exclusion criteria, 83 articles were selected for the systematic review, and 61 of these were used for the meta-analysis. Most studies were conducted at 0 °C to 4.4 °C storage temperatures under normal atmosphere package conditions. Salmonella Typhimurium, S. Enteritidis, and a cocktail (strain mixture) were inoculated at 5.0 and 6.0 log CFU mL−1. Articles both with and without the addition of antimicrobial compounds were found. Salmonella concentration decreases were observed in most studies, estimated for all study combinations as −0.8429 ± 0.0931 log CFU g−1 (95% CI; −1.0254, −0.6604) (p < 0.001), varying for each subgroup analysis. According to this survey, Salmonella concentration decreases are frequent during cool storage, although concentration increases and no bacterial inactivation were observed in some studies.

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First-line drugs for the treatment of listeriosis are the same around the world, but particular conditions might reduce their efficacy, including antimicrobial resistance. Therefore, this study aimed to verify, based on a systematic review and meta-analysis, whether the prevalence of antimicrobial resistance in Listeria monocytogenes from animal foods is higher for first- or second-line antimicrobials. From the total of 302 identified studies, 16 met all the eligibility criteria from 2008 to 2021 and were included in this meta-analysis. They comprised a dataset of 1152 L. monocytogenes isolates, obtained from different animal food products, food processing environment, and live animals. The included studies were developed in South America (n = 5), Europe (n = 4), Asia (n = 3), Africa (n = 2), and North America (n = 2), testing a total of 35 different antimicrobials, 11 of them classified as first-line drugs. Complete lack of antimicrobial resistance across the studies (all L. monocytogenes isolates tested as susceptible) was only observed for linezolid, while widespread antimicrobial resistance (all L. monocytogenes isolates tested resistant) was described for amoxicillin, benzylpenicillin, cefoxitin, fusidic acid, imipenem, sulfamethoxazole, and vancomycin. Overall, the meta-analysis results indicated no evidence that antimicrobial resistance in L. monocytogenes isolated from animal-based food is higher for first-line antimicrobials compared to second-line compounds (p=0.37). A greater volume of publication, together with better characterization of the isolates, is still needed for a more precise estimate of the real prevalence of antimicrobial resistance in L. monocytogenes.

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Four Escherichia coli isolates with moderate or high heat resistance were sequenced. Through sequencing, truncated transmissible locus of stress tolerance (tLST) variants tLST1 and tLSTa were identified in the three isolates. The most identified tLST genes (clpK and hsp) are responsible for the homeostasis module.