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Sous-vide is a cooking technique which is considered safe for food preparation, however, the recommended temperatures and times for preparing seafood and fish may not be sufficient to ensure safety, and additionally, thermal stress may cause changes in virulence and resistance. The objective of the present investigation was to determine the reduction of Listeria monocytogenes isolated from fish after sous-vide preparation. For this purpose flow cytometry was used. The impact of stress during the preparation of the fish-based dish using the sous-vide technique on the virulence potential and resistance profile was determined geno- and phenotypically. The study was conducted at four different temperatures, that are typical for fish preparation (42, 52, 57 and 60 °C) showing that none of them could be effective against L. monocytogenes. In addition, the investigation of changes in the virulence, phenotypic antibiotic resistance and gene expression showed that the stress during the preparation of sous-vide decrease virulence potential of the L. monocytogenes strains. It was observed that the greatest changes in antibiotic resistance profile were noted for tetracycline and notable changes were also observed for some strains in the case of fosfomycin and ciprofloxacin. Therefore it is important not to consider the sous-vide method as ensuring the microbiological quality of products, especially those with high contamination levels, and choose products of the best quality for processing.

High-pressure processing (HPP) is currently one of the leading methods of non-thermal food preservation as an alternative to traditional methods based on thermal processing. The application of HPP involves the simultaneous action of a combination of several factors—pressure values (100–600 MPa), time of operation (a few–several minutes), and temperature of operation (room temperature or lower)—using a liquid medium responsible for pressure transfer. The combination of these three factors results in the inactivation of microorganisms, thus extending food shelf life and improving the food’s microbiological safety. HPP can provide high value for the sensory and quality characteristics of products and reduce the population of pathogenic microorganisms such as L. monocytogenes to the required safety level. Nevertheless, the technology is not without impact on the cellular response of pathogens. L. monocytogenes cells surviving the HPP treatment may have multiple damages, which may impact the activation of mechanisms involved in the repair of cellular damage, increased virulence, or antibiotic resistance, as well as an increased expression of genes encoding pathogenicity and antibiotic resistance. This review has demonstrated that HPP is a technology that can reduce L. monocytogenes cells to below detection levels, thus indicating the potential to provide the desired level of safety. However, problems have been noted related to the possibilities of cell recovery during storage and changes in virulence and antibiotic resistance due to the activation of gene expression mechanisms, and the lack of a sufficient number of studies explaining these changes has been reported.

High-pressure processing (HPP) is one of the non-thermal methods of food preservation considered to be safe but may cause an increase/decrease in virulence potential and antibiotic resistance. The aim of the present study was to evaluate the survival of L. monocytogenes isolates after high-pressure processing (200 and 400 MPa for 5 min) and to determine changes in phenotypic and genotypic antibiotic resistance and virulence after this treatment. The 400 MPa treatment was shown to be effective in reducing pathogens to safe levels; however, the potential for cell recovery during storage was observed. In addition, studies on changes in virulence indicated possibilities related to a decrease in actA gene expression, overexpression of the hly and osfX gene, and an increase in biofilm-forming ability. The studies on changes in antibiotic resistance of isolates showed that all isolates showing initial susceptibility to lincomycin, fosfomycin, trimethoprim/sulfamethoxazole, and tetracycline became resistant to these antibiotics, which was associated with an increase in the values of minimum inhibitory concentrations. An increase in the expression of antibiotic resistance genes (mainly tetA_1, tetA_3, tetC) was also observed (mainly after the application of 200 MPa pressure), which was isolate dependent. However, it is noteworthy that the induced changes were permanent, i.e., they persisted even after the restoration of optimal environmental conditions. The results presented in our work indicate that the stress occurring during HPP can affect both phenotypic and genotypic changes in the virulence and antibiotic resistance potential of pathogens isolated from food and food processing environments. The potential associated with cell recovery and persistence of changes may influence the spread of virulent isolates of pathogens with increased antibiotic resistance in the food and food processing environment.

Listeria monocytogenes is a particularly foodborne pathogen associated with listeriosis, which can be disseminated in food and food processing environments. This study aimed to determine the serotypes and characteristics of virulence factors and antibiotic resistance among 40 L. monocytogenes strains isolated from food (n = 27) purchased in Olsztyn (Warmia and Mazury region, Poland) and food processing environments in Poland (n = 13). Isolates were assigned to serotypes 1/2a, 1/2c, 3a, and 3c using polymerase chain reaction (PCR). The results showed that serotype 1/2a (66.7%) was the most prevalent among strains from food, and serotype 1/2c (53.8%) among strains from the food processing environments. Five different virulence factors (hlyA, prfA, inlB, luxS, sigB) were detected in all isolates from the food processing environments using PCR. The hlyA (100.0%), prfA (100.0%), and inlB (96.3%) were the most prevalent in food strains. Seven (25.9%) of the strains of food and ten (76.9%) strains from the food processing environments showed the ability to form biofilm. The tested isolates were subjected to antibiotic susceptibility testing against 12 antibiotics used in the mitigation of listeriosis, using the disk diffusion method. The most frequent were intermediate resistance and resistance to clindamycin. Twelve (92.3%) strains from the food processing environments, and twenty-three (85.2%) from food were non-susceptible to clindamycin. Generally, antibacterial resistance determinants (Lde, aadB, aac(3)-IIa(aacC2)a, penA, mefA, lnuA, lnuB, sulI, sulII) were detected in sixteen (59.0%) strains from food and four (30.8%) from the food processing environments, by PCR. The most frequent were the mefA-lnuA (n = 7; 20.0%) and lnuA (n = 6; 17.1%) genotypes. From this research, we can conclude that virulent and antimicrobial-resistant strains of L. monocytogenes are present in food and the food processing environment in Poland, which may pose a potential health risk to consumers. Monitoring for the control of virulent and antimicrobial-resistant L. monocytogenes strains in the food system can contribute to effective planning and prevention of their spread.

Ceviche is a marinated raw fish dish ready for consumption; it is a part of the cuisine of various countries on the Pacific coast and its preparation may differ among them. Although the process uses the traditional method of food preservation by lowering the pH, the exposure time is very limited, so the aim of the study was to determine the viability of bacteria often isolated from fish after the process of preparing traditional ceviche. For this purpose, the traditional plate method and flow cytometry were used, and for pathogenic L. monocytogenes strains, the influence of stress during the preparation of the dish on the pathogenic potential was determined. The study showed that the highest percentage of viable cells was observed in the case of L. monocytogenes and remained at the level of 98.54%, slightly less for L. innocua, 96.93%. For the remaining species the reduction did not exceed 10%, for E. faecalis it was 92.76%, for S. liqefaciens 91.44%, H. alvei 93.68%. In addition, the study of the antibacterial properties of individual ingredients showed that habanero and coriander did not show any bactericidal effect, while for onions the amount of live cells was 99.11%, and for lime juice 97.26%, Additionally, the study of changes in virulence, antibiotic resistance and gene expression showed that the stress during the preparation of ceviche has different effects depending on the strain and may cause virulence potential increase, levofloxacin and daptomycin minimum inhibiotory concentration increase and some crucial virulence gene expression induction; therefore, it is important to take care of the quality of the products used to prepare the ceviche and accurate pretreatment.

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