Listeria monocytogenes is a foodborne pathogen that contaminates food-processing environments and persists within biofilms on equipment, utensils, floors, and drains, ultimately reaching final products by cross-contamination. This pathogen grows even under high salt conditions or refrigeration temperatures, remaining viable in various food products until the end of their shelf life. While the estimated incidence of listeriosis is lower than other enteric illnesses, infections caused by L. monocytogenes are more likely to lead to hospitalizations and fatalities. Despite the description of L. monocytogenes occurrence in Brazilian food-processing facilities and foods, there is a lack of consistent data regarding listeriosis cases and outbreaks directly associated with food consumption. Listeriosis requires rapid treatment with antibiotics and most drugs suitable for Gram-positive bacteria are effective against L. monocytogenes. Only a minority of clinical antibiotic-resistant L. monocytogenes strains have been described so far; whereas many strains recovered from food-processing facilities and foods exhibited resistance to antimicrobials not suitable against listeriosis. L. monocytogenes control in food industries is a challenge, demanding proper cleaning and application of sanitization procedures to eliminate this foodborne pathogen from the food-processing environment and ensure food safety. This review focuses on presenting the L. monocytogenes distribution in food-processing environment, food contamination, and control in the food industry, as well as the consequences of listeriosis to human health, providing a comparison of the current Brazilian situation with the international scenario.

Bacteriocinogenic Enterococcus hirae ST57ACC and Pediococcus pentosaceus ST65ACC strains, previously isolated from artisanal cheese, were evaluated for their safety with the aim to determine whether they could be used as beneficial strains, especially in the control of Listeria monocytogenes. Both isolates survived simulated gastrointestinal conditions and showed high levels of auto- and co-aggregation with L. monocytogenes, although the hydrophobicity of cells varied. Using the agar-spot test with 33 commercial drugs from different groups, only anti-inflammatory drugs and drugs containing loratadine and propranolol hydrochloride were able to affect the growth of the tested strains. Both strains were resistant to 3 out of 11 antibiotics tested by the disc diffusion method, and low frequencies of antibiotic resistance-encoding genes were observed by PCR analysis. Tested strains neither presented biogenic amine-related genes nor produced these substances. Aside from some antibiotic resistance characteristics, the tested strains were considered safe as they lack other virulence-related genes. E. hirae ST57ACC and P. pentosaceus ST65ACC both presented beneficial properties, particularly their ability to survive gastrointestinal conditions and to aggregate with L. monocytogenes, which can facilitate the elimination of this pathogen. Further studies should be conducted to better understand these interactions.

Pseudomonas aeruginosa is an opportunistic pathogen that can, like other bacterial species, exist in antimicrobial resistant sessile biofilms and as free-swimming, planktonic cells. Specific virulence factors are typically associated with each lifestyle and several two-component response regulators have been shown to reciprocally regulate transition between biofilm-associated chronic, and free-swimming acute infections. Quorum sensing (QS) signal molecules belonging to the las and rhl systems are known to regulate virulence gene expression by P. aeruginosa. However the impact of a recently described family of novel quorum sensing signals produced by the Pseudomonas Quinolone Signal (PQS) biosynthetic pathway, on the transition between these modes of infection is less clear. Using clonal isolates from a patient developing ventilator-associated pneumonia, we demonstrated that clinical observations were mirrored by an in vitro temporal shift in isolate phenotype from a non-secreting, to a Type III cytotoxin secreting (TTSS) phenotype and further, that this phenotypic change was PQS-dependent. While intracellular type III cytotoxin levels were unaffected by PQS concentration, cytotoxin secretion was dependent on this signal molecule. Elevated PQS concentrations were associated with inhibition of cytotoxin secretion coincident with expression of virulence factors such as elastase and pyoverdin. In contrast, low concentrations or the inability to biosynthesize PQS resulted in a reversal of this phenotype. These data suggest that expression of specific P. aeruginosa virulence factors appears to be reciprocally regulated and that an additional level of PQS-dependent posttranslational control, specifically governing type III cytotoxin secretion, exists in this species.

Listeria monocytogenes is an important foodborne pathogen commonly isolated from food processing environments and food products. This organism can multiply at refrigeration temperatures, form biofilms on different materials and under various conditions, resist a range of environmental stresses, and contaminate food products by cross-contamination. L. monocytogenes is recognized as the causative agent of listeriosis, a serious disease that affects mainly individuals from high-risk groups, such as pregnant women, newborns, the elderly, and immunocompromised individuals. Listeriosis can be considered a disease that has emerged along with changing eating habits and large-scale industrial food processing. This disease causes losses of billions of dollars every year with recalls of contaminated foods and patient medical treatment expenses. In addition to the immune status of the host and the infecting dose, the virulence potential of each strain is crucial for the development of disease symptoms. While many isolates are naturally virulent, other isolates are avirulent and unable to cause disease; this may vary according to the presence of molecular determinants associated with virulence. In the last decade, the characterization of genetic profiles through the use of molecular methods has helped track and demonstrate the genetic diversity among L. monocytogenes isolates obtained from various sources. The purposes of this review were to summarize the main methods used for isolation, identification, and typing of L. monocytogenes and also describe its most relevant virulence characteristics.

Summary Advances in whole‐genome sequencing (WGS) technologies have documented genetic diversity and epidemiology of the major foodborne pathogen Listeria monocytogenes (Lm) in Europe and North America, but data concerning South America are scarce. Here, we examined the population structure and genetic diversity of this major foodborne pathogen collected in Brazil. Based on core genome multilocus sequence typing (cgMLST), isolates from lineages I (n = 22; 63%) and II (n = 13; 37%) were distributed into 10 different sublineages (SLs) and represented 31 new cgMLST types (CTs). The most prevalent SLs were SL9 (n = 9; 26%), SL3 (n = 6; 17%) and SL2 and SL218 (n = 5; 14%). Isolates belonging to CTs L2‐SL9‐ST9‐CT4420 and L1‐SL315‐ST520‐CT4429 were collected 3 and 9 years apart, respectively, revealing long‐term persistence of Lm in Brazil. Genetic elements associated with stress survival were present in 60% of isolates (57% SSI‐1 and 3% SSI‐2). Pathogenic islands were present in 100% (LIPI‐1), 43% (LIPI‐3) and 6% (LIPI‐4) of the isolates. Mutations leading to premature stop codons were detected in the prfA and inlA virulence genes. This study is an important contribution to understanding the genomic diversity and epidemiology of Lm in South America. In addition, the results highlight the importance of using WGS to reveal Lm long‐term persistence.

The present study aimed to assess the activity of cell-free supernatant (CFS) containing bacteriocins on the formation and maintenance of biofilms developed by Listeria monocytogenes, and the associated effect of bacteriocins and ethylene-diamine-tetra-acetic acid (EDTA) on the formed biofilm. CFS from 9 lactic acid bacteria (LAB) strains was tested for inhibitory activity against 85 L. monocytogenes isolates and 21 LAB strains. Then, 12 L. monocytogenes strains were selected based on genetic profiles and sensitivity to CFS and were subjected to an in vitro assay to assess biofilm formation in microtiter plates, considering different culture media and incubation conditions. Based on these results, 6 L. monocytogenes strains were subjected to the same in vitro procedure to assess biofilm formation, being co-inoculated with CFS. In addition, these strains were subjected to the same in vitro procedure, modified by adding the CFS after biofilm formation. Relevant decrease in biofilm formation was observed in the first experiment, but CFS added after biofilm formation did not eliminate them. CFS from Lactobacillus curvatus ET31 were selected due to its anti-biofilm activity, being associated to EDTA at different concentrations and tested for biofilm control of three strains of L. monocytogenes, using the same in vitro procedure described previously. Concentrated bacteriocin presented poor performance in eliminating formed biofilms, and EDTA concentration presented no evident interference on biofilm elimination. Twelve selected L. monocytogenes strains were positive for investigated virulence makers and negative for luxS gene, recognized as being involved in biofilm formation. Selected L. monocytogenes strains were able to produce biofilms under different conditions. CFSs have the potential to prevent biofilm formation, but they were not able to destroy already formed biofilms. Nevertheless, low concentrations of CFS combined with EDTA caused a relevant reduction in already formed biofilms, but this association was not able to eliminate them. The activity of selected CFS was demonstrated against L. monocytogenes-formed biofilms, being more effective when associated to EDTA at different concentrations.

We isolated and characterized bacteriocin producers Enterococcus hirae ST57ACC and Pediococcus pentosaceus ST65ACC from raw milk artisanal cheeses. Their bacteriocins were tolerant to temperatures from 4°C to 100°C and under sterilization conditions (121°C for 15 min). Additionally, the tested bacteriocins remained active after being exposed to pH 2.0 to 10.0 for 2 h. The activity of the bacteriocins was affected by proteolytic enzymes but remained stable after treatment with EDTA, sodium dodecyl sulfate, NaCl, skim milk, and Tween 80. Cell-free supernatants were capable of inhibiting Listeria innocua and several strains of Listeria monocytogenes obtained from different sources and belonging to different serotypes. When L. monocytogenes 211 and L. monocytogenes 422 were treated with bacteriocins, growth was completely inhibited over 12 h. Cocultures of bacteriocinogenic strains and L. monocytogenes 422 in skim milk showed that E. hirae ST57ACC could control the growth of the pathogen in the matrix after 48 h. None of the selected isolates presented positive results on a screening panel for 25 bacteriocin-related genes, however, indicating that both strains might express novel bacteriocins.

The Listeria monocytogenes strains selected in the present study exhibited similar behavior in biofilm formation, independently of the tested conditions (bacteriocin from L. plantarum ST8SH, vancomycin, propolis (a natural antimicrobial product) and EDTA (chelating agent)), individual or in associations. The individual application of vancomycin had better inhibitory activity than that of propolis and EDTA; however, the association of the previously mentioned antimicrobial agents with bacteriocins resulted in better performance. However, when we compared the effects of vancomycin, propolis and EDTA, we could clearly observe that the combined application of bacteriocin and vancomycin was more effective than the combination of bacteriocin and propolis, and bacteriocin and EDTA. Considering the current need to reduce the use of antimicrobials and chemical substances in food processing, propolis can represent an alternative to improve the inhibitory effect of bacteriocins against L. monocytogenes biofilm formation, based on the obtained results. In general, high concentrations of bacteriocin produced by L. plantarum ST8SH were more effective in biofilm inhibition, and similar results were observed for vancomycin and propolis; however, all tested EDTA concentrations had similar effect on biofilm formation.