Search Results (55)

Regarding the trout microbiota, most information is focused on lactic acid bacteria, which can show beneficial properties. However, in trout farming, mostly pathogenic Gram-positive species were reported, such as Staphylococcus aureus, Listeria monocytogenes, and/or Clostridium spp. In this study, free-living trout were analyzed for Gram-negative microbiota that can cause loss as disease-stimulating agents. Bacteriocin postbiotics should be one of the approaches used to eliminate these agents. In total, 21 strains of different species isolated from the intestinal tract of 50 trout in Slovakia (Salmo trutta and Salmo gairdnerii) were taxonomically allotted into 13 species and 9 genera. This method showed variability in microbiota identified using MALDI-TOF mass spectrometry with the following species: Acinetobacter calcoaceticus, Citrobacter gillenii, Citrobacter freundii, Escherichia coli, Hafnia alvei, Kluyvera cryocrescens, K. intermedia, Leclercia adecarboxylata, Raoultella ornithinolytica, Pseudomonas fragi, Ps. putida, Ps. lundensis, Ps. teatrolens, and Serratia fonticola. Most strains were susceptible to the antibiotics used, reaching inhibitory zones up to 29 mm. On the other hand, 3 out of 21 strains (14%) were susceptible to nine enterocins- postbiotics (Hafnia alvei Hal281, Pseudomonas putida Pp391, and Ps. fragi Pf 284), with inhibitory activity in the range of 100–6400 AU/mL. Full article

Antimicrobial food packaging is considered a promising technology to improve food safety by inhibiting or reducing the growth of food microorganisms and minimizing the need for preservatives. This study aimed to develop and evaluate carboxymethyl cellulose (CMC) films integrated with bacteriocins for antibacterial efficacy. Plantaricin W was assessed as a potential bacteriocin for activation of CMC to control the dangerous food-borne pathogen, Listeria monocytogenes. Minced beef samples were inoculated with L. monocytogenes ATCC BAA-679 and treated with plantaricin W-activated food packaging. The results showed a significant reduction of the target pathogen by approximately 1 log cycle compared to the control group. Enterocin F4-9 is a novel bacteriocin that acts on Gram-negative microbes that were not affected by plantaricin W. Therefore, a novel food packaging activated with plantaricin W and enterocin F4-9 was developed to broaden their antimicrobial activity. The effect of this film on meat-associated microbes was investigated. The results demonstrated that the film significantly reduced the counts of mesophilic and psychotropic bacteria by 86.67% and 96.67%, respectively. Additionally, the pH values of the treated meat samples were significantly lower than those of the untreated controls. The obtained findings indicated that bacteriocin-activated CMC films could potentially be utilized as antimicrobial packaging in modern food technology. Full article

Eurasian beaver (Castor fiber) populations have been reintroduced to European countries, though this bears the risk of novel wildlife pathogen reservoir establishment. The species nova E. thailandicus was described first in Thailand as a food-derived strain. Later, this species was detected in the feces of pigs, poultry, sewage, and humans. In those studies, the potential risk posed by this species was evaluated. Against that background, the aim of this study was to investigate the susceptibility to postbiotic active substances (enterocins) against fecal E. thailandicus strains from beavers caught in Poland. The strains were identified with the use of 16S rRNA gene similarity sequencing. These six E. thailandicus strains with low-grade biofilm-forming abilities and two strains with the presence of the gelE virulence factor gene were susceptible to seven enterocins produced by non-autochthonous strains, mostly of the species E. faecium and E. durans. The inhibitory activity against the tested strains reached 25,600 AU/mL. Although the studied E. thailandicus strains did not show a strictly pathogenic character, their growth inhibition by postbiotics was identified as a novel elimination strategy. Full article

The increasing consumer demand for natural and sustainable food preservation methods has highlighted the potential of lactic acid bacteria (LAB) and their bioactive metabolites, particularly bacteriocins, as effective antimicrobial agents. This study aimed to isolate and characterize Enterococcus faecium strains from Algerian traditional dried figs marinated in olive oil, a nutrient-dense and underexplored food matrix. Twelve isolates were identified as E. faecium using MALDI-TOF MS and 16S rRNA gene sequencing, ensuring precise taxonomic classification. Genotypic analyses (BOX-PCR, GTG-PCR, and ERIC-PCR) revealed substantial genetic diversity, with BOX-PCR demonstrating superior discriminatory power. Functional screening confirmed the presence of enterocin genes, including entA (100% of strains), entB (60%), and entL50A/B (20%), which correlated with inhibition zones against Enterococcus faecium VCY, Micrococcus luteus GPE 3001, Staphylococcus aureus ATCC 25923, Pseudomonas aeruginosa ATCC 27853, and Acinetobacter lwoffii GPE 3002. Genotype–phenotype correlation analysis identified strain HFM7 as the most potent antimicrobial strain, exhibiting the largest inhibition zone (20.0 ± 1.0 mm) and harboring three enterocin genes (entA, entL50A, and entL50B). Protease sensitivity confirmed the proteinaceous nature of the antimicrobial compounds. Importantly, no virulence factors (esp, gelE, and hyl) or antibiotic resistance genes (vanA, vanB, ermA, ermB, and aac(6′)-Ie-alph(2″)) were detected, underscoring the safety of these isolates for food applications. These findings suggest that E. faecium strains from traditional foods are promising candidates as natural biopreservatives and starter cultures in clean-label food systems. By bridging traditional food ecosystems and modern biotechnological advancements, this study provides a foundation for sustainable, minimally processed food preservation strategies with potential applications in enhancing food safety and shelf life. Full article

Background/Objectives: Antimicrobial-resistant (AMR) pathogens represent a serious threat to public health, particularly in food production systems where antibiotic use remains widespread. As a result, alternative antimicrobial treatments to antibiotics are essential for effectively managing bacterial infections. This study aimed to identify and characterize novel antimicrobial peptides produced by bacteria, known as bacteriocins, as well as to recognize safe bacteriocin-producing strains, sourced from poultry slaughterhouse effluents. Methods: A total of 864 bacterial isolates were collected across eight stages of a poultry slaughter line and screened for antimicrobial activity against Gram-positive and Gram-negative indicator strains. Whole-genome sequencing (WGS) was performed on 12 selected strains, including Enterococcus faecium (6 isolates), Lactococcus lactis (1 isolate), Lactococcus garvieae (1 isolate) and Escherichia coli (4 isolates). The presence of bacteriocin gene clusters (BGC), antibiotic resistance genes (ARG), and virulence factors (VF) was analyzed. The antimicrobial activity of a novel bacteriocin was further evaluated using in vitro cell-free protein synthesis (IV-CFPS). Results: WGS revealed multiple BGCs, including a novel class IId bacteriocin, lactococcin P1A (LcnP1A), in L. lactis SWD9. LcnP1A showed antimicrobial activity against various indicator strains, including Listeria monocytogenes. While most bacteriocin-encoding strains harbored ARGs and VFs, E. faecium SWG6 was notable for its absence of ARGs and minimal VFs, highlighting its potential as a probiotic. Conclusions: These findings underscore the importance of discovering novel bacteriocins and safer bacteriocin producing strains to address antimicrobial resistance in the food chain. Further research would validate the efficacy of both the novel lactococcin P1A bacteriocin and the E. faecium SWG6 isolate for application in processed food and animal production systems. Full article

Intrapartum antibiotic prophylaxis, considered able to prevent streptococcal transmission from mother to newborn and its severe negative consequences, leads to microbiota dysbiosis, described as having a negative impact on well-being in both elements of the dyad. Enterococcus faecium L3 is a probiotic strain capable of exerting strong antagonistic activity against most streptococci, including S. agalactiae, due to the production of bacteriocins (known as enterocins A and B). A proprietary probiotic mixture containing the strain L3 demonstrated, in 2016, a significant reduction in episodes of PROM in pregnant women, with a less-than-expected effect on the vaginal–rectal presence of the pathogen S. agalactiae. With the aim of confirming the role exerted by the probiotic mixture in PROM episodes and to better understand the value of its impact on the clinical detection of S. agalactiae, we have retrospectively analyzed the results obtained in 125 L3-treated (over 12 weeks) women versus 125 untreated controls. Despite some limitations, our analysis has confirmed the role exerted by the probiotic in significantly reducing the following: (1) episodes of PROM, (2) vaginal–rectal positivity for S. agalactiae, and (3) the need to administer intrapartum antibiotics for prophylaxis. It likely also suggests operating using a cultural method very specific to S. agalactiae when testing women who were administered an Enterococcus-based probiotic. Full article

This study aimed to investigate the probiogenomic features of artisanal bacteriocin-producing Enterococcus faecium BGPAS1-3 and the use of the improved pMALc5HisEk expression vector for overexpressing class II bacteriocins and the application of purified bacteriocin 31 in a milk model as a preservative against L. monocytogenes. The BGPAS1-3 strain was isolated from traditional fresh soft cheese manufactured in households on a small scale in rural locations surrounding Pale Mountain City in Bosnia and Herzegovina. The whole-genome sequencing approach and bioinformatics analyses revealed that the strain BGPAS1-3 was non-pathogenic to humans. The presence of bacteriocin operons suggested the ability of the isolate to suppress the growth of pathogens. Coding regions for three maturated bacteriocins (bacteriocin 31, bacteriocin 32, and enterocin P) produced by BGPAS1-3 were amplified and expressed in Escherichia coli ER2523 using the pMALc5HisEk system. All three bacteriocins were successfully overexpressed and purified after enterokinase cleavage but showed different antimicrobial activity. Bacteriocin 31 showed significantly stronger antimicrobial activity compared with bacteriocin 32. It was the only one that proved to be suitable for use as a food preservative against L. monocytogenes in a milk model. Full article

Nowadays, the use of probiotics, postbiotics, parabiotics and other beneficial natural substances dominate health status sustainability in animals. Mundticin-like substance EM 41/3 (MLS) is a thermo-stable substance of proteinaceous character—a bacteriocin (postbiotic) with a broad antimicrobial (inhibitory) spectrum produced by the horse-derived strain Enterococcus mundtii EM 41/3. The aim of this study was to find if this new one bacteriocin can have an effect and/or a similar effect in rabbits as was achieved previously by using enterocins detected and characterized in our laboratory. In addition, these tests have been conducted to secure the health of rabbits as food-derived animals. Moreover, the aim of this study was also to test the effect of MLS EM 41/3 in broiler rabbits as an animal model before its use in horses. The parameters tested were analyzed by validated methods. Administration of MLS EM 41/3 lead to a significant increase in the non-specific immunity parameter (phagocytic activity, p < 0.05), while microbiota in feces, caecum and appendix were not negatively influenced. However, the total enterococcal count was significantly increased in the EG compared to the CG (p < 0.001). Application of MLS EM 41/3 did not have a negative impact on blood biochemistry. No oxidative stress was noted. However, higher growth parameters were noted and also hydrolytic activity was increased after MLS EM 41/3 application. Finally, to check physico-chemical parameters, no negative influence on rabbit meat quality was noted. Postbiotic MLS EM 41/3 administration seems to be a contributing factor in the feeding strategy for animal health status sustainability. Full article

Lactic acid bacteria (LAB) can produce peptides known as bacteriocins with antagonistic activity against foodborne pathogens. The potential of LAB isolated from the surface of jalapeno peppers to produce bacteriocins with antagonistic activity against Listeria monocytogenes, Staphylococcus aureus, Escherichia coli O157:H7, and Salmonella Typhimurium was evaluated. Previously isolated LAB strains were reactivated, and their cell-free supernatants (CFSs) were evaluated. Out of 390 reactivated strains, 60 produced bacteriocin-like inhibitory substances (BLIS) since their antagonistic activity was lost after proteases addition. Subsequently, 16 BLIS showed heat resistance (HR-BLIS), retaining their bioactivity after heat treatment (121 °C for 15 min). By 16S rRNA gene sequencing and antibiotic susceptibility tests, LAB strains producing HR-BLIS were identified as Enterococcus lactis. Four HR-BLIS exhibited a minimum inhibitory concentration (MIC) of 80 mg/mL against L. monocytogenes. MIC and minimum bactericidal concentration (MBC) of HR-BLIS-67 for S. aureus (MIC = 80 mg/mL; MBC = 320 mg/mL), S. Typhimurium (MIC = 150 mg/mL; MBC = 250 mg/mL), and E. coli O157:H7 (MIC = 250 mg/mL; MBC = 400 mg/mL) were determined. LAB isolated from the surface of jalapeno pepper produced HR-BLIS (possibly enterocin) that exhibited broad-spectrum antagonistic activity against foodborne pathogens; therefore, they are a promising source of natural antimicrobials to ensure food safety. Full article

Beneficial/probiotic strains protect the host from pathogens by competitive displacement and production of antibacterial substances, i.e., bacteriocins. The antiparasitic potential of bacteriocins/enterocins and their producing strains in experimental murine trichinellosis were tested as a new therapeutic strategy. Enterocin M and Durancin-like and their producers Enterococcus faecium CCM8558 and Enterococcus durans ED26E/7 were administered daily to mice that were challenged with Trichinella spiralis. Our study confirmed the antiparasitic effect of enterocins/enterococci, which reduced the number of adults in the intestine (Enterocin M—43.8%, E. faecium CCM8558—54.5%, Durancin-like—16.4%, E. durans ED26E/7—35.7%), suppressed the Trichinella reproductive capacity ex vivo (Enterocin M—61%, E. faecium CCM8558—74%, Durancin-like—38%, E. durans ED26E/7—66%), and reduced the number of muscle larvae (Enterocin M—39.6%, E. faecium CCM8558—55.7%, Durancin-like—15%, E. durans ED26E/7—36.3%). The direct effect of enterocins on Trichinella fecundity was documented by an in vitro test in which Durancin-like showed a comparable reducing effect to Enterocin M (40–60%) in contrast to the ex vivo test. The reducing activity of T.spiralis infection induced by Enterocin M was comparable to its strain E. faecium CCM8558; Durancin-like showed lower antiparasitic activity than its producer E. durans ED26E/7. Full article

The formation of biogenic amines (BAs) in artisan Galotyri PDO cheeses fermented with Sterptococcus thermophilus ST1 and the Greek indigenous nisin A-producing Lactococcus lactis spp. cremoris M78 (A1cheese), or with the A1 starter supplemented with either the enterocin A-B-P-producing Enterococcus faecium KE82 (A2cheese) or the multi-functional Lactiplantibacillus plantarum H25 (A4cheese) adjunct strains was evaluated. Three pilot-scale cheese trials, GL1, GL2, and GL3, made from boiled ewes’ milk, were analyzed for their BA contents before and after cold ripening at 4 °C for 30 days. Total BAs of the fresh GL1 and GL3 cheeses (pH 4.3–4.5) were below 50 mg/kg, except for the A1/GL1 and A1/GL3 cheeses, which contained ca. 300 mg/kg (81.2% histamine) and 1250 mg/kg (45.6% putrescine) BAs, respectively. Whereas due to an outgrowth (>7 log cfu/g) of post-thermal Gram-negative bacteria contaminants during fermentation, most fresh GL2 cheeses (pH 4.7–5.0) accumulated more than 1500 mg/kg of total BAs, which exceeded 3800 mg/kg in all GL2 cold-ripened cheeses due to major increases in cadaverine and putrescine. Tyramine and histamine exceeded 500 mg/kg in the fresh A1/GL2cheeses. Conversely, total BAs remained or declined below 50 mg/kg in all cold-ripened GL3 cheeses. None of the starter or adjunct cultures could be correlated with a specific BA increase, despite E. faecium KE82, which increased at 7.6–9.2 log cfu/g in the A2 cheeses is a strong tyramine producer in culture BA broth with 1% tyrosine in vitro. The adoption of strict hygienic measures during artisan Galotyri PDO cheese production (trial GL3) enabled the best performance of all starter LAB strain combinations and reduced BA formation, whereas the high presence of Gram-negative decarboxylating bacteria contaminants compromised cheese (trial GL2) safety. Full article

Enterocins are bacteriocins synthesized by Enterococcus strains that show an interesting antimicrobial effectiveness against foodborne pathogens such as Listeria monocytogenes. The objectives of this study were to identify and analyze the expression of enterocin genes of Enterococcus isolated from breast-fed infants and evaluate their ability to inhibit three human isolates of virulent L. monocytogenes, as well as some probiotic bacteria. The susceptibility of the strains of L. monocytogenes to fifteen antibiotics was tested, detecting their resistance to cefoxitin (constitutively resistant), oxacillin, and clindamycin. The production of enterocins A, B, and P was observed in Enterococcus faecium isolates, while enterocin AS-48 was detected in an Enterococcus faecalis isolate. AS-48 showed antilisterial activity by itself, while the joint action of enterocins A and B or B and P was necessary for inhibiting L. monocytogenes, demonstrating the synergistic effect of those combinations. The presence of multiple enterocin genes does not assure the inhibition of L. monocytogenes strains. However, the expression of multiple enterocin genes showed a good correlation with the inhibition capacity of these strains. Furthermore, the potential beneficial strains of lactobacilli and bifidobacteria examined were not inhibited by any of the enterocins produced individually or in combination, with the exception of Bifidobacterium longum BB536, which was inhibited by enterocin AS-48 and the joint production of enterocins A and B or B and P. The enterocins studied here could be candidates for developing alternative treatments against antibiotic-resistant bacterial infections. Moreover, these selected enterocin-producing E. faecium strains isolated from breast-fed infants could be used as probiotic strains due to their antilisterial effect, as well as the absence of virulence factors. Full article

Multidrug-resistant Clostridium perfringens infections are a major threat to the poultry industry. Effective alternatives to antibiotics are urgently needed to prevent these infections and limit the spread of multidrug-resistant bacteria. The aim of the study was to produce by chemical synthesis a set of enterocins of different subgroups of class II bacteriocins and to compare their spectrum of inhibitory activity, either alone or in combination, against a panel of twenty C. perfringens isolates. Enterocins A, P, SEK4 (class IIa bacteriocins), B (unsubgrouped class II bacteriocin), and L50 (class IId leaderless bacteriocin) were produced by microwave-assisted solid-phase peptide synthesis. Their antimicrobial activity was determined by agar well diffusion and microtitration methods against twenty C. perfringens isolates and against other pathogens. The FIC_INDEX of different combinations of the selected enterocins was calculated in order to identify combinations with synergistic effects. The results showed that synthetic analogs of L50A and L50B were the most active against C. perfringens. These peptides also showed the broadest spectrum of activity when tested against other non-clostridial indicator strains, including Listeria monocytogenes, methicillin-resistant Staphylococcus aureus, Streptococcus suis, Streptococcus pyogenes, Enterococcus cecorum, Enterococcus faecalis, as well as Gram-negative bacteria (Campylobacter coli and Pseudomonas aeruginosa), among others. The selected synthetic enterocins were combined on the basis of their different mechanisms of action, and all combinations tested showed synergy or partial synergy against C. perfringens. In conclusion, because of their high activity against C. perfringens and other pathogens, the use of synthetic enterocins alone or as a consortium can be a good alternative to the use of antibiotics in the poultry sector. Full article

Bacteriocins (enterocins) represent a new therapeutic strategy in various intestinal and non-intestinal infections. In antiparasitic defence, an oxidative inflammation of phagocytes is effective in destroying new-born Trichinella spiralis larvae. The strains Enterococcus faecium CCM8558 and E. durans ED26E/7 and their enterocins, enterocin M and a durancin-like enterocin, respectively, were administered daily, and mice were then infected with T. spiralis larvae on the seventh day of treatment. Phagotest and Bursttest kits were used to detect the phagocytosis and respiratory burst in blood leukocytes. T. spiralis infection inhibited phagocytosis from day 11 post-infection (dpi) during the migration of new-born larvae into the muscles. E. faecium CCM8558, E. durans ED26E/7, and the durancin-like enterocin increased phagocytic activity from day 11 dpi. Both strains and their enterocins (enterocin M and durancin-like) stimulated the ingestion capability of phagocytes from 18 to 32 dpi. Enterococci/enterocins therapy prevented a reduction in cells with respiratory burst caused by T. spiralis infection from 11 dpi. The enzymatic activity of phagocytes was stimulated on 18 and 25 dpi, particularly by E. faecium CCM8558 and enterocin M. Enterocin M and the durancin-like enterocin were as effective in stimulating phagocytosis as the bacterial strains that produce them. The stimulation of phagocytosis could contribute to decreased larval migration and reduced parasite burden in the host. Full article

This study sought to evaluate the probiotic properties and the food preservation ability of lactic acid bacteria isolates collected from the intestines of wild marine fishes (gilthead seabream (Sparus aurata) (n = 60) and whiting fish (Merlangius merlangus) (n = 40)) from the Mediterranean sea in the area of Mostaganem city, Algeria. Forty-two isolates were identified as: Enterococcus durans (n = 19), Enterococcus faecium (n = 15), Enterococcus faecalis (n = 4), Lactococcus lactis subp. lactis (n = 3), and Lactobacillus plantarum (n = 1). All isolates showed inhibition to at least one indicator strain, especially against Listeria monocytogenes, Staphylococcus aureus, Paenibacillus larvae, Vibrio alginolyticus, Enterococcus faecalis, Bacillus cereus, and Bacillus subtilis. In all collected isolates, PCR analysis of enterocin-encoding genes showed the following genes: entP (n = 21), ent1071A/B (n = 11), entB (n = 8), entL50A/B (n = 7), entAS48 (n = 5), and entX (n = 1). Interestingly, 15 isolates harbored more than one ent gene. Antimicrobial susceptibility, phenotypic virulence, and genes encoding virulence factors were investigated by PCR. Resistance to tetracycline (n = 8: tetL + tetK), erythromycin (n = 7: 5 ermA, 2 msrA, and 1 mef(A/E)), ciprofloxacin (n = 1), gentamicin (n = 1: aac(6′)-aph(2″)), and linezolid (n = 1) were observed. Three isolates were gelatinase producers and eight were α-hemolytic. Three E. durans and one E. faecium harbored the hyl gene. Eight isolates showing safety properties (susceptible to clinically relevant antibiotics, free of genes encoding virulence factors) were tested to select probiotic candidates. They showed high tolerance to low pH and bile salt, hydrophobicity power, and co-culture ability. The eight isolates showed important phenotypic and genotypic traits enabling them to be promising probiotic candidates or food bio-conservers and starter cultures. Full article