Comparison of fidaxomicin, thuricin CD, vancomycin and nisin highlights the narrow spectrum nature of thuricin CD.

Vancomycin and metronidazole are commonly used treatments for Clostridioides difficile infection (CDI). However, these antibiotics have been associated with high levels of relapse in patients. Fidaxomicin is a new treatment for CDI that is described as a narrow spectrum antibiotic that is minimally active on the commensal bacteria of the gut microbiome. The aim of this study was to compare the effect of fidaxomicin on the human gut microbiome with a number of narrow (thuricin CD) and broad spectrum (vancomycin and nisin) antimicrobials. The spectrum of activity of each antimicrobial was tested against 47 bacterial strains by well-diffusion assay. Minimum inhibitory concentrations (MICs) were calculated against a select number of these strains. Further, a pooled fecal slurry of 6 donors was prepared and incubated for 24 h with 100 µM of each antimicrobial in a mini-fermentation system together with a no-treatment control. Fidaxomicin, vancomycin, and nisin were active against most gram positive bacteria tested in vitro, although fidaxomicin and vancomycin produced larger zones of inhibition compared to nisin. In contrast, the antimicrobial activity of thuricin CD was specific to C. difficile and some Bacillus spp. The MICs showed similar results. Thuricin CD exhibited low MICs (<3.1 µg/mL) for C. difficile and Bacillus firmus, whereas fidaxomicin, vancomycin, and nisin demonstrated lower MICs for all other strains tested when compared to thuricin CD. The narrow spectrum of thuricin CD was also observed in the gut model system. We conclude that the spectrum of activity of fidaxomicin is comparable to that of the broad-spectrum antibiotic vancomycin in vitro and the broad spectrum bacteriocin nisin in a complex community.

Turbidimetric bioassays: A solution to antimicrobial activity detection in asymptomatic bacteriuria isolates against uropathogenic Escherichia coli.

Traditional bacteriocin screening methods often face limitations due to diffusion-related challenges in agar matrices, which can prevent the peptides from reaching their target organism. Turbidimetric techniques offer a solution to these issues, eliminating diffusion-related problems and providing an initial quantification of bacteriocin efficacy in producer organisms. This study involved screening the cell-free supernatant (CFS) from eight uncharacterized asymptomatic bacteriuria (ABU) isolates and Escherichia coli 83972 for antimicrobial activity against clinical uropathogenic E. coli (UPEC) strains using turbidimetric growth methods. ABU isolates exhibiting activity against five or more UPEC strains were further characterized (PUTS 37, PUTS 58, PUTS 59, S-07-4, and SK-106-1). The inhibition of the CFS by proteinase K suggested that the antimicrobial activity was proteinaceous in nature, potentially bacteriocins. The activity of E. coli PUTS 58 and SK-106-1 was enhanced in an artificial urine medium, with both inhibiting all eight UPECs. A putative microcin H47 operon was identified in E. coli SK-106-1, along with a previously identified microcin V and colicin E7 in E. coli PUTS 37 and PUTS 58, respectively. These findings indicate that ABU bacteriocin-producers could serve as viable prophylactics and therapeutics in the face of increasing antibiotic resistance among uropathogens.

In Vivo Mapping of Myocardial Injury Outside the Infarct Zone: Tissue at an Intermediate Pathological State.

BACKGROUND: The goal was to determine the feasibility of mapping the injured-but-not-infarcted myocardium using 99mTc-duramycin in the postischemic heart, with spatial information for its characterization as a pathophysiologically intermediate tissue, which is neither normal nor infarcted. METHODS AND RESULTS: Coronary occlusion was conducted in Sprague Dawley rats with preconditioning and 30-minute ligation. In vivo single-photon emission computed tomography was acquired after 3 hours (n=6) using 99mTc-duramycin, a phosphatidylethanolamine-specific radiopharmaceutical. The 99mTc-duramycin+ areas were compared with infarct and area-at-risk (n=8). Cardiomyocytes and endothelial cells were isolated for gene expression profiling. Cardiac function was measured with echocardiography (n=6) at 4 weeks. In vivo imaging with 99mTc-duramycin identified the infarct (3.9±2.4% of the left ventricle and an extensive area 23.7±2.2% of the left ventricle) with diffuse signal outside the infarct, which is pathologically between normal and infarcted (apoptosis 1.8±1.6, 8.9±4.2, 13.6±3.8%; VCAM-1 [vascular cell adhesion molecule 1] 3.2±0.8, 9.8±4.1, 15.9±4.2/mm2; tyrosine hydroxylase 14.9±2.8, 8.6±4.4, 5.6±2.2/mm2), with heterogeneous changes including scattered micronecrosis, wavy myofibrils, hydropic change, and glycogen accumulation. The 99mTc-duramycin+ tissue is quantitatively smaller than the area-at-risk (26.7% versus 34.4% of the left ventricle, P=0.008). Compared with infarct, gene expression in the 99mTc-duramycin+-noninfarct tissue indicated a greater prosurvival ratio (BCL2/BAX [B-cell lymphoma 2/BCL2-associated X] 7.8 versus 5.7 [cardiomyocytes], 3.7 versus 3.2 [endothelial]), and an upregulation of ion channels in electrophysiology. There was decreased contractility at 4 weeks (regional fractional shortening -8.6%, P<0.05; circumferential strain -52.9%, P<0.05). CONCLUSIONS: The injured-but-not-infarcted tissue, being an intermediate zone between normal and infarct, is mapped in vivo using phosphatidylethanolamine-based imaging. The intermediate zone contributes significantly to cardiac dysfunction.

The composition of Tibetan kefir grain TKG-Y and the antibacterial potential and milk fermentation ability of S. warneri KYS-164 screened from TKG-Y.

This study utilized high-throughput sequencing and SEM observation to elucidate the microbial composition of a Tibetan herder's homemade kefir grain named TKG-Y. Subsequently, S. warneri KYS-164 was isolated from TKG-Y, which can produce mixed protein substances with antibacterial activity, namely bacteriocin-like inhibitory substances (BLIS). BLIS can significantly reduce the growth rate of Escherichia coli 366-a, Staphylococcus aureus CICC 10384 and mixed strains at low concentrations (1 × MIC). The presence of the warnericin-centered gene cluster in KYS-164 may explain the antibacterial properties of the BLIS. Pepsin and an acidic environment can reduce the number of colonies of KYS-164 by 2.5 Log10 CFU mL-1 within 1 h, and reduce the antibacterial activity of BLIS by 21.48%. S. warneri KYS-164 showed no antibiotic resistance and biological toxicity after 80 subcultures, while BLIS produced by 40 generations of the strain retained their inhibitory efficacy against pathogenic bacteria. After 48-hour fermentation of milk with KYS-164, volatile compounds such as aldehydes, phenols, esters, and alcohols, giving it a floral, fruity, milky, oily, and nutty aroma, were released, enriching the sensory characteristics of dairy products. This study not only revealed the bacterial colony composition information of home-made kefir grain TKG-Y but also discovered and proved that S. warneri KYS-164 has the potential to inhibit bacteria and ferment dairy products. This will provide a basis for subsequent applied research on KYS-164.

Bacteriocins: potentials and prospects in health and agrifood systems.

Bacteriocins are highly diverse, abundant, and heterogeneous antimicrobial peptides that are ribosomally synthesized by bacteria and archaea. Since their discovery about a century ago, there has been a growing interest in bacteriocin research and applications. This is mainly due to their high antimicrobial properties, narrow or broad spectrum of activity, specificity, low cytotoxicity, and stability. Though initially used to improve food quality and safety, bacteriocins are now globally exploited for innovative applications in human, animal, and food systems as sustainable alternatives to antibiotics. Bacteriocins have the potential to beneficially modulate microbiota, providing viable microbiome-based solutions for the treatment, management, and non-invasive bio-diagnosis of infectious and non-infectious diseases. The use of bacteriocins holds great promise in the modulation of food microbiomes, antimicrobial food packaging, bio-sanitizers and antibiofilm, pre/post-harvest biocontrol, functional food, growth promotion, and sustainable aquaculture. This can undoubtedly improve food security, safety, and quality globally. This review highlights the current trends in bacteriocin research, especially the increasing research outputs and funding, which we believe may proportionate the soaring global interest in bacteriocins. The use of cutting-edge technologies, such as bioengineering, can further enhance the exploitation of bacteriocins for innovative applications in human, animal, and food systems.

Impairment of Listeria monocytogenes biofilm developed on industrial surfaces by Latilactobacillus curvatus CRL1579 bacteriocin.

The effect of lactocin AL705, bacteriocin produced by Latilactobacillus (Lat.) curvatus CRL1579 against Listeria biofilms on stainless steel (SS) and polytetrafluoroethylene (PTFE) coupons at 10 °C was investigated. L. monocytogenes FBUNT showed the greatest adhesion on both surfaces associated to the hydrophobicity of cell surface. Partially purified bacteriocin (800 UA/mL) effectively inhibited L. monocytogenes preformed biofilm through displacement strategy, reducing the pathogen by 5.54 ± 0.26 and 4.74 ± 0.05 log cycles at 3 and 6 days, respectively. The bacteriocin-producer decreased the pathogen biofilm by ∼2.84 log cycles. Control and Bac- treated samples reached cell counts of 7.05 ± 0.18 and 6.79 ± 0.06 log CFU/cm2 after 6 days of incubation. Confocal scanning laser microscopy (CLSM) allowed visualizing the inhibitory effect of lactocin AL705 on L. monocytogenes preformed biofilms under static and hydrodynamic flow conditions. A greater effect of the bacteriocin was found at 3 days independently of the surface matrix and pathogen growth conditions at 10 °C. As a more realistic approach, biofilm displacement strategy under continuous flow conditions showed a significant loss of biomass, mean thickness and substratum coverage of pathogen biofilm. These findings highlight the anti-biofilm capacity of lactocin AL705 and their potential application in food industries.

Evaluation of the Effect of Lactobacillus acidophilus ATCC 4356 Bacteriocin against Staphylococcus aureus.

Background: Lactobacillus acidophilus is lactic acid bacteria that produce bacteriocins. Bacteriocins are antimicrobial peptides or proteins that exhibit activity against closely related bacteria. The aim of this study was to determine the effect of L. acidophilus ATCC 4356 bacteriocin against Staphylococcus aureus. Material and Methods. We used four different phenotypic methods for antimicrobial activities against two standard strains: methicillin-resistant S. aureus (MRSA) ATCC 33591 and methicillin-susceptible S. aureus (MSSA) ATCC 25923. The methods were (1) agar well diffusion, (2) overlay soft agar, (3) paper disk, and (4) modification of punch hole. The ammonium sulfate method was used to concentrate crude bacteriocin, and ultrafiltration and dialysis tubes were used to remove ammonium sulfate from the bacteriocins. Each method was repeated in triplicate. Result: L. acidophilus ATCC 4356 showed antimicrobial activity against both MRSA and MSSA standard strains only by the overlay soft agar method and not by the agar well diffusion, punch hole modification, and paper disk methods. No antimicrobial effects were observed in crude bacteriocins concentrated. Conclusion: The growth inhibition of S. aureus in overlay soft agar method may be due to the production of bacteriocin-like substances. The overlay soft agar method is a qualitative test, so there is a need for further study to optimize the conditions for the production of bacteriocin-like substances in the culture supernatant and precise comparison between the inhibitory activity and pheromone secretion of different strains.

An update on the taxonomy and functional properties of the probiotic Enterococcus faecium SF68.

Enterococcus faecium SF68 (SF68) is a well-known probiotic with a long history of safe use. Recent changes in the taxonomy of enterococci have shown that a novel species, Enterococcus lactis, is closely related with E. faecium and occurs together with other enterococci in a phylogenetically well-defined E. faecium species group. The close phylogenetic relationship between the species E. faecium and E. lactis prompted a closer investigation into the taxonomic status of E. faecium SF68. Using phylogenomics and ANI, the taxonomic analysis in this study showed that probiotic E. faecium SF68, when compared to other E. faecium and E. lactis type and reference strains, could be re-classified as belonging to the species E. lactis. Further investigations into the functional properties of SF68 showed that it is potentially capable of bacteriocin production, as a bacteriocin gene cluster encoding the leaderless bacteriocin EntK1 together with putative Lactococcus lactis bacteriocins LsbA, and LsbB-like putative immunity peptide (LmrB) were found located in an operon on plasmid pF9. However, bacteriocin expression was not studied. Competitive exclusion experiments in co-culture over 7 days at 37 °C showed that the probiotic SF68 could inhibit the growth of specific E. faecium and Listeria monocytogenes strains, while showing little or no inhibitory activity towards an entero-invasive Escherichia coli and a Salmonella Typhimurium strain, respectively. In cell culture experiments with colon carcinoma HT29 cells, the probiotic SF68 was also able to strain-specifically inhibit adhesion and/or invasion of enterococcal and L. monocytogenes strains, while such adhesion and invasion inhibition effects were less pronounced for E. coli and Salmonella strains. This study therefore provides novel data on the taxonomy and functional properties of SF68, which can be reclassified as Enterococcus lactis SF68, thereby enhancing the understanding of its probiotic nature.

A two-step regulatory circuit involving Spo0A-AbrB activates mersacidin biosynthesis in Bacillus subtilis.

Due to intramolecular ring structures, the ribosomally produced and post-translationally modified peptide mersacidin shows antimicrobial properties comparable to those of vancomycin without exhibiting cross-resistance. Although the principles of mersacidin biosynthesis are known, there is no information on the molecular control processes for the initial stimulation of mersacidin bioproduction. By using Bacillus subtilis for heterologous biosynthesis, a considerable amount of mersacidin could be produced without the mersacidin-specific immune system and the mersacidin-activating secretory protease. By using the established laboratory strain Bacillus subtilis 168 and strain 3NA, which is used for high cell density fermentation processes, in combination with the construction of reporter strains to determine the promoter strengths within the mersacidin core gene cluster, the molecular regulatory circuit of Spo0A, a master regulator of cell differentiation including sporulation initiation, and the global transcriptional regulator AbrB, which is involved in cell adaptation processes in the transient growth phase, was identified to control the initial stimulation of the mersacidin core gene cluster. In a second downstream regulatory step, the activator MrsR1, encoded in the core gene cluster, acts as a stimulatory element for mersacidin biosynthesis. These findings are important to understand the mechanisms linking environmental conditions and microbial responses with respect to the bioproduction of bioactive metabolites including antimicrobials such as mersacidin. This information will also support the construction of production strains for bioactive metabolites with antimicrobial properties.

Exploring the feasibility of bacteriocins EntK1 and EntEJ97s in treatment of systemic vancomycin resistant enterococci infections in mice.

AIMS: Enterocins K1 and EJ97 have specific antimicrobial activity against Enterococcus faecium and Enterococcus faecalis, respectively. The aim of this study was to investigate the utility of these enterocins for in vivo treatment of systemic enterococcal infections. METHODS AND RESULTS: The antimicrobial effect in blood was analysed and compared against the effect in saline. Colony forming unit counts revealed that the enterocins killed all the bacteria within 1 hour. Additionally, the bactericidal effect against E. faecalis was more rapid in blood, indicating a possible synergy between EntEJ97 and blood. Importantly, no enterocin resistant mutants emerged in these experiments. Injecting the enterocins intraperitoneally in an in vivo mouse model and using fluorescence and minimum inhibitory concentration determination to estimate concentrations of the peptides in plasma, indicate that the enterocins exist in circulation in therapeutic concentrations. Alanine aminotransferase detection, and haemolysis analysis indicates that there is no detectable liver damage or haemolytic effect after injection. CONCLUSIONS: The study revealed that EntK1 and EntEJ97 are able to kill all bacteria ex vivo in the presence of blood. In vivo experiments determine that the enterocins exist in circulation in therapeutic concentrations without causing liver damage or haemolysis. Future experiments should test these peptides for treatment of infection in a relevant in vivo model.

Suppression of planktonic and biofilm of Escherichia coli by the synergistic lantibiotics-polymyxins combinations.

Escherichia coli are generally resistant to the lantibiotic's action (nisin and warnerin), but we have shown increased sensitivity of E. coli to lantibiotics in the presence of subinhibitory concentrations of polymyxins. Synergistic lantibiotic-polymyxin combinations were found for polymyxins B and M. The killing of cells at the planktonic and biofilm levels was observed for two collection and four clinical multidrug-resistant E. coli strains after treatment with lantibiotic-polymyxin B combinations. Thus, 24-h treatment of E. coli mature biofilms with warnerin-polymyxin B or nisin-polymyxin B leads to five to tenfold decrease in the number of viable cells, depending on the strain. AFM revealed that the warnerin and polymyxin B combination caused the loss of the structural integrity of biofilm and the destruction of cells within the biofilm. It has been shown that pretreatment of cells with polymyxin B leads to an increase of Ca2+ and Mg2+ ions in the culture medium, as detected by atomic absorption spectroscopy. The subsequent exposure to warnerin caused cell death with the loss of K+ ions and cell destruction with DNA and protein release. Thus, polymyxins display synergy with lantibiotics against planktonic and biofilm cells of E. coli, and can be used to overcome the resistance of Gram-negative bacteria to lantibiotics.

Genome mining of Bacillus licheniformis MCC2514 for the identification of lasso peptide biosynthetic gene cluster and its characterization.

The probiotic strain Bacillus licheniformis MCC2514 has been shown to produce a strong antibacterial peptide and the whole genome sequence of this strain is also reported in our previous study. The present study is focused on the genome level investigation of this peptide antibiotic and its characterization. Genome mining of the culture revealed the presence of three putative bacteriocin clusters, viz. lichenicidin, sonorensin and lasso peptide. Hence, the mode of action of the peptide was investigated by reporter assay, scanning electron microscopy, and Fourier Transform Infrared spectroscopy. Additionally, the peptide treated groups of Kocuria rhizophila showed a reduction in the fold expression for transcription-related genes. The gene expression studies, quantitative ß-galactosidase induction assay using the RNA stress reporter strain, yvgS along with the homology studies concluded that lasso peptide is responsible for the antibacterial activity of the peptide which acts as an inhibitor of RNA biosynthesis. Gene expression analysis showed a considerable increase in fold expression of lasso peptide genes at various fermentation hours. Also, the peptide was isolated, and its time-kill kinetics and minimum inhibitory concentration against the indicator pathogen K. rhizophila were examined. The peptide was also purified and the molecular weight was determined to be ~ 2 kDa. Our study suggests that this bacteriocin can function as an effective antibacterial agent in food products as well as in therapeutics as it contains lasso peptide, which inhibits the RNA biosynthesis.

Bad to the bone? - Genomic analysis of Enterococcus isolates from diverse environments reveals that most are safe and display potential as food fermentation microorganisms.

Enterococci comprise a group of lactic acid bacteria (LAB) with considerable potential to serve as food fermentation microorganisms. Unfortunately, enterococci have received a lot of negative attention, due to the occurrence of pathogenic and multidrug resistant strains. In this study, we used genomics to select safe candidates among the forty-four studied enterococcal isolates. The genomes of the forty-four strains were fully sequenced and assessed for presence of virulence and antibiotic resistance genes. Nineteen isolates belonging to the species Enterococcus lactis, Enterococcus faecium, Enterococcus durans, and Enterococcus thailandicus, were deemed safe from the genome analysis. The presence of secondary metabolite gene clusters for bacteriocins was assessed, and twelve candidates were found to secrete antimicrobial compounds effective against Listeria monocytogenes isolated from cheese and Staphylococcus aureus. Physiological characterization revealed nineteen industrial potentials; all strains grew well at 42 °C and acidified 1.5 hours faster than their mesophilic counterpart Lactococcus lactis, with which they share metabolism and flavor forming ability. We conclude that a large fraction of the examined enterococci were safe and could serve as excellent food fermentation microorganisms with inherent bioprotective abilities.

Lactobacillus-derived components for inhibiting biofilm formation in the food industry.

Biofilm, a microbial community formed by especially pathogenic and spoilage bacterial species, is a critical problem in the food industries. It is an important cause of continued contamination by foodborne pathogenic bacteria. Therefore, removing biofilm is the key to solving the high pollution caused by foodborne pathogenic bacteria in the food industry. Lactobacillus, a commonly recognized probiotic that is healthy for consumer, have been proven useful for isolating the potential biofilm inhibitors. However, the addition of surface components and metabolites of Lactobacillus is not a current widely adopted biofilm control strategy at present. This review focuses on the effects and preliminary mechanism of action on biofilm inhibition of Lactobacillus-derived components including lipoteichoic acid, exopolysaccharides, bacteriocins, secreted protein, organic acids and some new identified molecules. Further, the review discusses several modern biofilm identification techniques and particularly interesting new technology of biofilm inhibition molecules. These molecules exhibit stronger inhibition of biofilm formation, playing a pivotal role in food preservation and storage. Overall, this review article discusses the application of biofilm inhibitors produced by Lactobacillus, which would greatly aid efforts to eradicate undesirable bacteria from environment in the food industries.

Staphylococcus epidermidis bacteriocin A37 kills natural competitors with a unique mechanism of action.

Many bacteria produce antimicrobial compounds such as lantibiotics to gain advantage in the competitive natural environments of microbiomes. Epilancins constitute an until now underexplored family of lantibiotics with an unknown ecological role and unresolved mode of action. We discovered production of an epilancin in the nasal isolate Staphylococcus epidermidis A37. Using bioinformatic tools, we found that epilancins are frequently encoded within staphylococcal genomes, highlighting their ecological relevance. We demonstrate that production of epilancin A37 contributes to Staphylococcus epidermidis competition specifically against natural corynebacterial competitors. Combining microbiological approaches with quantitative in vivo and in vitro fluorescence microscopy and cryo-electron tomography, we show that A37 enters the corynebacterial cytoplasm through a partially transmembrane-potential-driven uptake without impairing the cell membrane function. Upon intracellular aggregation, A37 induces the formation of intracellular membrane vesicles, which are heavily loaded with the compound and are essential for the antibacterial activity of the epilancin. Our work sheds light on the ecological role of epilancins for staphylococci mediated by a mode of action previously unknown for lantibiotics.

Automated workflow for characterization of bacteriocin production in natural producers Lactococcus lactis and Latilactobacillus sakei.

BACKGROUND: Lactic acid bacteria are commonly used as protective starter cultures in food products. Among their beneficial effects is the production of ribosomally synthesized peptides termed bacteriocins that kill or inhibit food-spoiling bacteria and pathogens, e.g., members of the Listeria species. As new bacteriocins and producer strains are being discovered rapidly, modern automated methods for strain evaluation and bioprocess development are required to accelerate screening and development processes. RESULTS: In this study, we developed an automated workflow for screening and bioprocess optimization for bacteriocin producing lactic acid bacteria, consisting of microcultivation, sample processing and automated antimicrobial activity assay. We implemented sample processing workflows to minimize bacteriocin adsorption to producer cells via addition of Tween 80 and divalent cations to the cultivation media as well as acidification of culture broth prior to cell separation. Moreover, we demonstrated the applicability of the automated workflow to analyze influence of media components such as MES buffer or yeast extract for bacteriocin producers Lactococcus lactis B1629 and Latilactobacillus sakei A1608. CONCLUSIONS: Our automated workflow provides advanced possibilities to accelerate screening and bioprocess optimization for natural bacteriocin producers. Based on its modular concept, adaptations for other strains, bacteriocin products and applications are easily carried out and a unique tool to support bacteriocin research and bioprocess development is provided.

Complete nucleotide sequence and comparative genomic analysis of microcin B17 plasmid pMccB17.

We present a comprehensive sequence and bioinformatic analysis of the prototypical microcin plasmid, pMccb17, which includes a definitive sequence for the microcin operon, mcb. Microcin B17 (MccB17) is a ribosomally synthesized and posttranslationally modified peptide produced by Escherichia coli. It inhibits bacterial DNA gyrase similarly to quinolone antibiotics. The mcb operon, which consists of seven genes encoding biosynthetic and immunity/export functions, was originally located on the low copy number IncFII plasmid pMccB17 in the Escherichia coli strain LP17. It was later transferred to E. coli K-12 through conjugation. In this study, the plasmid was extracted from the E. coli K-12 strain RYC1000 [pMccB17] and sequenced twice using an Illumina short-read method. The first sequencing was conducted with the host bacterial chromosome, and the plasmid DNA was then purified and sequenced separately. After assembly into a single contig, polymerase chain reaction primers were designed to close the single remaining gap via Sanger sequencing. The resulting complete circular DNA sequence is 69,190 bp long and includes 81 predicted genes. These genes were initially identified by Prokka and subsequently manually reannotated using BLAST. The plasmid was assigned to the F2:A-:B- replicon type with a MOBF12 group conjugation system. A comparison with other IncFII plasmids revealed a large proportion of shared genes, particularly in the conjugative plasmid backbone. However, unlike many contemporary IncFII plasmids, pMccB17 lacks transposable elements and antibiotic resistance genes. In addition to the mcb operon, this plasmid carries 25 genes of unknown function.

Characterization of the broad-spectrum antibacterial activity of bacteriocin-like inhibitory substance-producing probiotics isolated from fermented foods.

Antimicrobial peptides, such as bacteriocin, produced by probiotics have become a promising novel class of therapeutic agents for treating infectious diseases. Selected lactic acid bacteria (LAB) isolated from fermented foods with probiotic potential were evaluated for various tests, including exopolysaccharide production, antibiotic susceptibility, acid and bile tolerance, antibacterial activity, and cell adhesion and cytotoxicity to gastric cell lines. Six selected LAB strains maintained their high viability under gastrointestinal conditions, produced high exopolysaccharides, showed no or less cytotoxicity, and adhered successfully to gastric cells. Furthermore, three strains, Weissella confusa CYLB30, Lactiplantibacillus plantarum CYLB47, and Limosilactobacillus fermentum CYLB55, demonstrated a strong antibacterial effect against drug-resistant Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Salmonella enterica serovar Choleraesuis, Enterococcus faecium, and Staphylococcus aureus. Whole genome sequencing was performed on these three strains using the Nanopore platform; then, the results showed that all three strains did not harbor genes related to toxins, superantigens, and acquired antimicrobial resistance, in their genome. The bacteriocin gene cluster was found in CYLB47 genome, but not in CYLB30 and CYLB55 genomes. In SDS-PAGE, the extract of CYLB30 and CYLB47 bacteriocin-like inhibitory substance (BLIS) yielded a single band with a size of less than 10 kDa. These BLIS inhibited the growth and biofilm formation of drug-resistant P. aeruginosa and methicillin-resistant S. aureus (MRSA), causing membrane disruption and inhibiting adhesion ability to human skin HaCaT cells. Moreover, CYLB30 and CYLB47 BLIS rescued the larvae after being infected with P. aeruginosa and MRSA infections. In conclusion, CYLB30 and CYLB47 BLIS may be potential alternative treatment for multidrug-resistant bacteria infections.

In Silico Screening of Bacteriocin Gene Clusters within a Set of Marine Bacillota Genomes.

Due to their potential application as an alternative to antibiotics, bacteriocins, which are ribosomally synthesized antimicrobial peptides produced by bacteria, have received much attention in recent years. To identify bacteriocins within marine bacteria, most of the studies employed a culture-based method, which is more time-consuming than the in silico approach. For that, the aim of this study was to identify potential bacteriocin gene clusters and their potential producers in 51 marine Bacillota (formerly Firmicutes) genomes, using BAGEL4, a bacteriocin genome mining tool. As a result, we found out that a majority of selected Bacillota (60.78%) are potential bacteriocin producers, and we identified 77 bacteriocin gene clusters, most of which belong to class I bacteriocins known as RiPPs (ribosomally synthesized and post-translationally modified peptides). The identified putative bacteriocin gene clusters are an attractive target for further in vitro research, such as the production of bacteriocins using a heterologous expression system.

Controlling soft rot of green pepper by bacteriocin paracin wx3 and its effect on storage quality of green pepper.

A bacteriocin paracin wx3 was investigated as a candidate of natural preservative to control green pepper soft rot. Firstly, paracin wx3 was heterologously expressed in Pichia pastoris X33 with an improved yield of 0.537 g/L. Its size and amino acid sequence were confirmed by Tricine-SDS-PAGE and LC-MS/MS. Then, result of antibacterial activity showed that its MIC value against Pectobacterium carotovorum was 16 µg/mL. In vitro, paracin wx3 completely killed the pathogen at high concentrations ≥8 × MIC. In vivo, disease incidence of green pepper soft rot was decreased from 90% (control) to <2% (8 × MIC). Subsequently, results of action mode showed that paracin wx3 inhibited the growth of pathogen by pore-formation on cell membrane. Last, paracin wx3 treatment reduced losses of weight, firmness, total soluble solid, Vc of green pepper during storage. It also inhibited the production of soft rot volatile p-xylene, 1-butanol, 2-methyl-2-propanol, 3-hydroxybutan-2-one-D, 2-pentyl furan, butanal, etc.