Subinhibitory concentrations of nisin enhance virulence gene expression in Staphylococcus aureus and increase mortality in Galleria mellonella.

Alternative strategies for controlling Staphylococcus aureus and other pathogens have been continuously investigated, with nisin, a bacteriocin widely used in the food industry as a biopreservative, gaining increasing attention. In addition to its antimicrobial properties, bacteriocins have significant effects on genome functionality even at inhibitory concentrations. This study investigated the impact of subinhibitory concentrations of nisin on S. aureus. Culturing in the presence of 0.625 µmol l-1 nisin, led to the increased relative expression of hla, saeR, and sarA, genes associated with virulence while expression of the sea gene, encoding staphylococcal enterotoxin A (SEA), decreased. In an in vivo experiment, Galleria mellonella larvae inoculated with S. aureus cultured in the presence of nisin exhibited 97% mortality at 72 h post-infection, compared to over 40% of larvae mortality in larvae infected with S. aureus. A comprehensive understanding of the effect of nisin on the transcriptional response of virulence genes and the impact of these changes on the virulence of S. aureus can contribute to assessing the application of this bacteriocin in food and medical contexts.

Metal Uptake by Mitochondrial Carrier Family Proteins Using Lactococcus lactis.

Metal ion homeostasis in mitochondria is essential to maintaining proper cellular physiology. However, the ability of metals to bind off target or form complexes with multiple metabolites presents major challenges to understanding the mechanisms that govern this homeostasis. Adding further to the complexity, some of the major mitochondrial transporters have shown substrate promiscuity. In many cases, mitochondrial metals are found in the matrix compartment that is surrounded by the impermeable inner membrane. Four major classes of transporters facilitate the movement of solute across the inner membrane. These are mitochondrial carrier family, ATP-binding cassette transporters, mitochondrial pyruvate carriers, and sideroflexins. For iron, the matrix is the site of iron-sulfur clusters and heme synthesis and therefore transport must occur in a coordinated fashion with the cellular needs for these critical cofactors. Iron could be transported in numerous forms as it has been shown to form complexes with abundant metabolites such as citrate, nucleotides, or glutathione. Here, we describe assays to study iron (or any metal) transport by mitochondrial carrier family proteins expressed in Lactococcus lactis using a nisin-controlled expression system.

Thiolated chitosan nanoparticles encapsulated nisin and selenium: antimicrobial/antibiofilm/anti-attachment/immunomodulatory multi-functional agent.

BACKGROUND: The increase in the resistance of bacterial strains to antibiotics has led to research into the bactericidal potential of non-antibiotic compounds. This study aimed to evaluate in vitro antibacterial/ antibiofilm properties of nisin and selenium encapsulated in thiolated chitosan nanoparticles (N/Se@TCsNPs) against prevalent enteric pathogens including standard isolates of Vibrio (V.) cholerae O1 El Tor ATCC 14,035, Campylobacter (C.) jejuni ATCC 29,428, Salmonella (S.) enterica subsp. enterica ATCC 19,430, Shigella (S.) dysenteriae PTCC 1188, Escherichia (E.) coli O157:H7 ATCC 25,922, Listeria (L.) monocytogenes ATCC 19,115, and Staphylococcus (S.) aureus ATCC 29,733. METHODS: The synthesis and comprehensive analysis of N/Se@TCsNPs have been completed. Antibacterial and antibiofilm capabilities of N/Se@TCsNPs were evaluated through broth microdilution and crystal violet assays. Furthermore, the study included examining the cytotoxic effects on Caco-2 cells and exploring the immunomodulatory effects of N/Se@TCsNPs. This included assessing the levels of both pro-inflammatory (IL-6 and TNFα) and anti-inflammatory (IL-10 and TGFß) cytokines and determining the gene expression of TLR2 and TLR4. RESULTS: The N/Se@TCsNPs showed an average diameter of 136.26 ± 43.17 nm and a zeta potential of 0.27 ± 0.07 mV. FTIR spectroscopy validated the structural features of N/Se@TCsNPs. Scanning electron microscopy (SEM) images confirmed their spherical shape and uniform distribution. Thermogravimetric Analysis (TGA)/Differential Scanning Calorimetry (DSC) tests demonstrated the thermal stability of N/Se@TCsNPs, showing minimal weight loss of 0.03%±0.06 up to 80 °C. The prepared N/Se@TCsNPs showed a thiol content of 512.66 ± 7.33 µmol/g (p < 0.05), an encapsulation efficiency (EE) of 69.83%±0.04 (p ≤ 0.001), and a drug release rate of 74.32%±3.45 at pH = 7.2 (p ≤ 0.004). The synthesized nanostructure demonstrated potent antibacterial activity against various isolates, with effective concentrations ranging from 1.5 ± 0.08 to 25 ± 4.04 mg/mL. The ability of N/Se@TCsNPs to reduce bacterial adhesion and internalization in Caco-2 cells underscored their antibiofilm properties (p ≤ 0.0001). Immunological studies indicated that treatment with N/Se@TCsNPs led to decreased levels of inflammatory cytokines IL-6 (14.33 ± 2.33 pg/mL) and TNFα (25 ± 0.5 pg/mL) (p ≤ 0.0001), alongside increased levels of anti-inflammatory cytokines IL-10 (46.00 ± 0.57 pg/mL) and TGFß (42.58 ± 2.10 pg/mL) in infected Caco-2 cells (p ≤ 0.0001). Moreover, N/Se@TCsNPs significantly reduced the expression of TLR2 (0.22 ± 0.09) and TLR4 (0.16 ± 0.05) (p < 0.0001). CONCLUSION: In conclusion, N/Se@TCsNPs exhibited significant antibacterial/antibiofilm/anti-attachment/immunomodulatory effectiveness against selected Gram-positive and Gram-negative enteric pathogens. However, additional ex-vivo and in-vivo investigations are needed to fully assess the performance of nanostructured N/Se@TCsNPs.

Facile Halogenation of Antimicrobial Peptides As Demonstrated by Producing Bromotryptophan-Labeled Nisin Variants with Enhanced Antimicrobial Activity.

Antimicrobial peptides (AMPs) have raised significant interest, forming a potential new class of antibiotics in the fight against multi-drug-resistant bacteria. Various AMPs are ribosomally synthesized and post-translationally modified peptides (RiPPs). One post-translational modification found in AMPs is the halogenation of Trp residues. This modification has, for example, been shown to be critical for the activity of the potent AMP NAI-107 from Actinoallomurus. Due to the importance of organohalogens, establishing methods for facile and selective halogen atom installation into AMPs is highly desirable. In this study, we introduce an expression system utilizing the food-grade strain Lactococcus lactis, facilitating the efficient incorporation of bromo-Trp (BrTrp) into (modified) peptides, exemplified by the lantibiotic nisin with a single Trp residue or analogue incorporated at position 1. This provides an alternative to the challenges posed by halogenase enzymes, such as poor substrate selectivity. Our method yields expression levels comparable to that of wild-type nisin, while BrTrp incorporation does not interfere with the post-translational modifications of nisin (dehydration and cyclization). One brominated nisin variant exhibits a 2-fold improvement in antimicrobial activity against two tested pathogens, including a WHO priority pathogen, while maintaining the same lipid II binding and bactericidal activity as wild-type nisin. The work presented here demonstrates the potential of this methodology for peptide halogenation, offering a new avenue for the development of diverse antimicrobial products labeled with BrTrp.

Label-Free Monitoring of Coculture System Dynamics: Probing Probiotic and Cancer Cell Interactions via Infrared Spectroscopic Imaging.

Evaluating the dynamic interaction of microorganisms and mammalian cells is challenging due to the lack of suitable platforms for examining interspecies interactions in biologically relevant coculture conditions. In this work, we demonstrate the interaction between probiotic bacteria (Lactococcus lactis and Escherichia coli) and A498 human cancer cells in vitro, utilizing a hydrogel-based platform in a label-free manner by infrared spectroscopy. The L. lactis strain recapitulated in the compartment system secretes polypeptide molecules such as nisin, which has been reported to trigger cell apoptosis. We propose a mid-infrared (IR) spectroscopic imaging approach to monitor the variation of biological components utilizing kidney cells (A498) as a model system cocultured with bacteria. We characterized the biochemical composition (i.e., nucleic acids, protein secondary structures, and lipid conformations) label-free using an unbiased measurement. Several IR spectral features, including unsaturated fatty acids, ß-turns in protein, and nucleic acids, were utilized to predict cellular response. These features were then applied to establish a quantitative relationship through a multivariate regression model to predict cellular dynamics in the coculture system to assess the effect of nisin on A498 kidney cancer cells cocultured with bacteria. Overall, our study sheds light on the potential of using IR spectroscopic imaging as a label-free tool to monitor complex microbe-host cell interactions in biological systems. This integration will enable mechanistic studies of interspecies interactions with insights into their underlying physiological processes.

Antimicrobial peptides and proteins as alternative antibiotics for porcine semen preservation.

BACKGROUND: Antimicrobial resistance (AMR) is nowadays a major emerging challenge for public health worldwide. The over- and misuse of antibiotics, including those for cell culture, are promoting AMR while also encouraging the research and employment of alternative drugs. The addition of antibiotics to the cell media is strongly recommended in sperm preservation, being gentamicin the most used for boar semen. Because of its continued use, several bacterial strains present in boar semen have developed resistance to this antibiotic. Antimicrobial peptides and proteins (AMPPs) are promising candidates as alternative antibiotics because their mechanism of action is less likely to promote AMR. In the present study, we tested two AMPPs (lysozyme and nisin; 50 and 500 µg/mL) as possible substitutes of gentamicin for boar semen preservation up to 48 h of storage. RESULTS: We found that both AMPPs improved sperm plasma membrane and acrosome integrity during semen storage. The highest concentration tested for lysozyme also kept the remaining sperm parameters unaltered, at 48 h of semen storage, and reduced the bacterial load at comparable levels of the samples supplemented with gentamicin (p > 0.05). On the other hand, while nisin (500 µg/mL) reduced the total Enterobacteriaceae counts, it also decreased the rapid and progressive sperm population and the seminal oxidation-reduction potential (p < 0.05). CONCLUSIONS: The protective effect of lysozyme on sperm function together with its antimicrobial activity and inborn presence in body fluids, including semen and cervical mucus, makes this enzyme a promising antimicrobial agent for boar semen preservation.

Microfluidic preparation and optimization of (Kollicoat ® IR-b-PCL) polymersome for co-delivery of Nisin-Curcumin in breast cancer application.

This work aimed to develop amphiphilic nanocarriers such as polymersome based diblock copolymer of Kollicoat ® IR -block-poly(ε-caprolactone) (Kollicoat ® IR-b-PCL) for potential co-delivery of Nisin (Ni) and Curcumin (CUR) for treatment of breast cancer. To generate multi-layered nanocarriers of uniform size and morphology, microfluidics was used as a new technology. In order to characterise and optimize polymersome, design of experiments (Design-Expert) software with three levels full factorial design (3-FFD) method was used. Finally, the optimized polymersome was produced with a spherical morphology, small particle size (dH < 200 nm), uniform size distribution (PDI < 0.2), and high drug loading efficiency (Ni 78 % and CUR 93 %). Furthermore, the maximum release of Ni and CUR was found to be roughly 60 % and 80 % in PBS, respectively. Cytotoxicity assays showed a slight cytotoxicity of Ni and CUR -loaded polymersome (N- Ni /CUR) towards normal cells while demonstrating inhibitory activity against cancer cells compared to the free drugs. Also, the apoptosis assays and cellular uptake confirmed the obtained results from cytotoxic analysis. In general, this study demonstrated a microfluidic approach for preparation and optimization of polymersome for co-delivery of two drugs into cancer cells.

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.

Sensitivity of Salmonella Typhimurium to nisin in vitro and in orange juice under refrigeration.

The influence of environmental factors on Salmonella sensitivity to nisin in vitro and in refrigerated orange juice were investigated. Nisin activity was observed in the different conditions, but the highest efficiency was achieved at lower pH (4.0) and with higher bacteriocin concentration (174 µM). Moreover, the bactericidal action was directly proportional to the incubation period. When tested in orange juice, nisin caused a reduction of up to 4.05 logarithm cycles in the Salmonella population. So, environmental factors such as low pH and low temperature favored the sensitization of Salmonella cells to the bactericidal action of nisin. Therefore, this may represent an alternative to control Salmonella in refrigerated foods.

Natural approach of using nisin and its nanoform as food bio-preservatives against methicillin resistant Staphylococcus aureus and E.coli O157:H7 in yoghurt.

BACKGROUND: Natural antimicrobial agents such as nisin were used to control the growth of foodborne pathogens in dairy products. The current study aimed to examine the inhibitory effect of pure nisin and nisin nanoparticles (nisin NPs) against methicillin resistant Staphylococcus aureus (MRSA) and E.coli O157:H7 during the manufacturing and storage of yoghurt. Nisin NPs were prepared using new, natural, and safe nano-precipitation method by acetic acid. The prepared NPs were characterized using zeta-sizer and transmission electron microscopy (TEM). In addition, the cytotoxicity of nisin NPs on vero cells was assessed using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The minimum inhibitory concentrations (MICs) of nisin and its nanoparticles were determined using agar well-diffusion method. Further, fresh buffalo's milk was inoculated with MRSA or E.coli O157:H7 (1 × 106 CFU/ml) with the addition of either nisin or nisin NPs, and then the inoculated milk was used for yoghurt making. The organoleptic properties, pH and bacterial load of the obtained yoghurt were evaluated during storage in comparison to control group. RESULTS: The obtained results showed a strong antibacterial activity of nisin NPs (0.125 mg/mL) against MRSA and E.coli O157:H7 in comparison with control and pure nisin groups. Notably, complete eradication of MRSA and E.coli O157:H7 was observed in yoghurt formulated with nisin NPs after 24 h and 5th day of storage, respectively. The shelf life of yoghurt inoculated with nisin nanoparticles was extended than those manufactured without addition of such nanoparticles. CONCLUSIONS: Overall, the present study indicated that the addition of nisin NPs during processing of yoghurt could be a useful tool for food preservation against MRSA and E.coli O157:H7 in dairy industry.

Milk-derived antimicrobial peptides incorporated whey protein film as active coating to improve microbial stability of refrigerated soft cheese.

Nisin is the first FDA-approved antimicrobial peptide and shows significant antimicrobial activity against Gram-positive bacteria, but only a weakly inhibitory effect on Gram-negative bacteria. The aim of this study was to prepare whey protein-based edible films with the incorporation of milk-derived antimicrobial peptides (αs2-casein151-181 and αs2-casein182-207) and compare their mechanical properties and potential application in cheese packaging with films containing nisin. These two antimicrobial peptides showed similar activity against B. subtilis and much higher activity against E. coli than bacteriocin nisin, representing that these milk-derived peptides had great potential to be applied as food preservatives. Antimicrobial peptides in whey protein films caused an increase in film opaqueness and water vapor barrier properties but decreased the tensile strength and elongation at break. Compared to other films, the whey protein film containing αs2-casein151-181 had good stability in salt or acidic solution, as evidenced by the results from scanning electron microscope and Fourier transform infrared spectroscopy. Whey protein film incorporated with αs2-casein151-181 could inhibit the growth of yeasts and molds, and control the growth of psychrotrophic bacteria present originally in the soft cheese at refrigerated temperature. It also exhibited significant inhibitory activity against the development of mixed culture (E. coli and B. subtilis) in the cheese due to superficial contamination during storage. Antimicrobial peptides immobilized in whey protein films showed a higher effectiveness than their direct application in solution. In addition, films containing αs2-casein151-181 could act as a hurdle inhibiting the development of postprocessing contamination on the cheese surface during the 28 days of storage. The films in this study exhibited the characteristics desired for active packaging materials.

Development of transcriptional factor-based whole-cell biosensors to monitor and degrade antibiotics using mutant cells obtained via adaptive laboratory evolution.

With the widespread use of antibiotics and increasing environmental concerns regarding antibiotic abuse, the detection and degradation of antibiotic residues in various samples has become a pressing issue. Transcriptional factor (TF)-based whole-cell biosensors are low-cost, easy-to-use, and flexible tools for detecting chemicals and controlling bioprocesses. However, because of cytotoxicity caused by antibiotics, the application of such biosensors is limited in the presence of antibiotics. In this study, we used antibiotic-tolerant mutants obtained via adaptive laboratory evolution (ALE) to develop TF-based whole-cell biosensors for antibiotic monitoring and degradation. The biosensors had high performance and stability in detecting relatively high concentrations of tetracycline (Tc) and nisin. The ALE mutant-based Tc biosensor exhibited a 10-fold larger linear detection range than the wild-type strain-based biosensor. Then, the Tc biosensor was employed to detect residual amounts of Tc in mouse stool, serum, and urine samples and facilitate Tc biodegradation in mouse stool, demonstrating its high utility. Considering that ALE has been demonstrated to enhance cell tolerance to various toxic chemicals, our strategy might facilitate the development of whole-cell biosensors for most antibiotics and other toxic ligands.

In Bacterial Membranes Lipid II Changes the Stability of Pores Formed by the Antimicrobial Peptide Nisin.

Resistance to available antibiotics poses a growing challenge to modern medicine, as this often disallows infections to be controlled. This problem can only be alleviated by the development of new drugs. Nisin, a natural lantibiotic with broad antimicrobial activity, has shown promise as a potential candidate for combating antibiotic-resistant bacteria. However, nisin is poorly soluble and barely stable at physiological pH, which despite attempts to address these issues through mutant design has restricted its use as an antibacterial drug. Therefore, gaining a deeper understanding of the antimicrobial effectiveness, which relies in part on its ability to form pores, is crucial for finding innovative ways to manage infections caused by resistant bacteria. Using large-scale molecular dynamics simulations, we find that the bacterial membrane-specific lipid II increases the stability of pores formed by nisin and that the interplay of nisin and lipid II reduces the overall integrity of bacterial membranes by changing the local thickness and viscosity.

Assessing the synergistic potential of bacteriophage endolysins and antimicrobial peptides for eradicating bacterial biofilms.

Phage-encoded endolysins have emerged as a potential substitute to conventional antibiotics due to their exceptional benefits including host specificity, rapid host killing, least risk of resistance. In addition to their antibacterial potency and biofilm eradication properties, endolysins are reported to exhibit synergism with other antimicrobial agents. In this study, the synergistic potency of endolysins was dissected with antimicrobial peptides to enhance their therapeutic effectiveness. Recombinantly expressed and purified bacteriophage endolysin [T7 endolysin (T7L); and T4 endolysin (T4L)] proteins have been used to evaluate the broad-spectrum antibacterial efficacy using different bacterial strains. Antibacterial/biofilm eradication studies were performed in combination with different antimicrobial peptides (AMPs) such as colistin, nisin, and polymyxin B (PMB) to assess the endolysin's antimicrobial efficacy and their synergy with AMPs. In combination with T7L, polymyxin B and colistin effectively eradicated the biofilm of Pseudomonas aeruginosa and exhibited a synergistic effect. Further, a combination of T4L and nisin displayed a synergistic effect against Staphylococcus aureus biofilms. In summary, the obtained results endorse the theme of combinational therapy consisting of endolysins and AMPs as an effective remedy against the drug-resistant bacterial biofilms that are a serious concern in healthcare settings.

Preparation of modified chitosan-based nano-TiO2-nisin composite packaging film and preservation mechanism applied to chilled pork.

Here, we developed a nano-TiO2-nisin-modified chitosan composite packaging film and investigated its properties and antibacterial activity, as well as its effect on chilled pork preservation time. The results indicated that the preservation time of chilled pork coated with a nano-TiO2-nisin-modified chitosan film (including 0.7 g/L nano-TiO2, irradiated with ultraviolet light for 40 min, and dried for 6 h) followed by modified atmosphere packaging (50% CO2 + 50% N2) increased from 7 to 20 days at 4 °C. Both nano-TiO2 and nisin enhanced the mechanical strength of the chitosan film, and nisin promoted nano-TiO2 dispersion and compatibility in chitosan. Treatment with 0.4 g/L nano-TiO2 for 60 min considerably inhibited spoilage bacteria, particularly Acinetobacter johnnii XBB1 (A. johnnii XBB1). As nano-TiO2 concentration and photocatalytic time increased, K+, Ca2+, and Mg2+ leakage in A. johnnii XBB1 increased but Na+/K+-ATPase and Ca2+/Mg2+-ATPase activities decreased. In A. johnnii XBB1, TiO2 significantly downregulated the expression of putrefaction-related genes such as cysM and inhibited cell self-regulation and membrane wall system repair. Therefore, our nano-TiO2-nisin-modified chitosan film could extend the shelf life without the addition of any chemical preservatives, demonstrating great potential for application in food preservation.

Bacterial nanocellulose loaded with bromelain and nisin as a promising bioactive material for wound debridement.

The bacterial nanocellulose (BnC) membranes were produced extracellularly by a novel aerobic acetic acid bacterium Komagataeibacter melomenusus. The BnC was modified in situ by adding carboxymethyl cellulose (CMC) into the culture media, obtaining a BnC-CMC product with denser fibril arrangement, improved rehydration ratio and elasticity in comparison to BnC. The proteolytic enzyme bromelain (Br) and antimicrobial peptide nisin (N) were immobilized to BnC matrix by ex situ covalent binding and/or adsorption. The optimal Br immobilization conditions towards the maximized specific proteolytic activity were investigated by response surface methodology as factor variables. At optimal conditions, i.e., 8.8 mg/mL CMC and 10 mg/mL Br, hyperactivation of the enzyme was achieved, leading to the specific proteolytic activity of 2.3 U/mg and immobilization efficiency of 39.1 %. The antimicrobial activity was observed against Gram-positive bacteria (S. epidermidis, S. aureus and E. faecalis) for membranes with immobilized N and was superior when in situ modified BnC membranes were used. N immobilized on the BnC or BnC-CMC membranes was cytocompatible and did not cause changes in normal human dermal fibroblast cell morphology. BnC membranes perform as an efficient carrier for Br or N immobilization, holding promise in wound debridement and providing antimicrobial action against Gram-positive bacteria, respectively.

Comparison of controlling capabilities of lactobionic acid, nisin, and K-Sorbate against dairy-borne Staphylococcus aureus, Escherichia coli, and mold in soft cheese.

Background: The utilization of chemical preservatives holds the promise of effectively controlling microbial growth in soft cheese. Aim: The first trial aimed to compare the effectiveness of lactobionic acid (LBA) and K-Sorbate in controlling the proliferation of Staphylococcus aureus, Escherichia coli, and mold in white soft cheese. The subsequent part of the study explored the inhibitory effects of K-Sorbate, nisin, and LBA on mold populations in cheese whey. Methods: Two sets of soft cheese were produced. One set was contaminated with S. aureus, while the other was with E. coli, each at concentrations of 1 log CFU/ml and 1 log CFU/100 ml. Different concentrations of LBA were incorporated into these sets of cheese. Similar cheese samples were treated with K-Sorbate. For the subsequent part of the study, it was manufactured and divided into groups that inoculated with LBA with different concentrations, K-Sorbate, and nisin. Results: With higher S. aureus inoculation, by day 18, the positive control exhibited growth exceeding 5 log CFU/g. In contrast, the LBA treatment dropped below limit of detection (LOD) and K-Sorbate yielded 4.8 log CFU/g. While with lower S. aureus inoculation, the positive control reached log CFU/g, while LBA treatment fell below LOD by day 14, and K-Sorbate reached 2.9 log CFU/g. For E. coli inoculation, with higher concentrations, by day 18, the positive control exceeded 5 log CFU/g. Conversely, LBA treatment greatly decreased and K-Sorbate treatment measured 5.1 log CFU/g. With lower E. coli concentrations, the positive control surpassed 3 log CFU/g, yet LBA treatment dropped below LOD by day 3. Mold counts indicated some inhibition with the K-Sorbate treatment, while control groups showed growth. LBA treatments exhibit noticeable growth inhibition. About the other part of the study, the outcomes demonstrated that while growth of mold occurred in the control group, inhibitory effects were apparent in the treatment groups, and significant distinctions existed between K-Sorbate, nisin, LBA treatments, and the control group. Conclusion: Our findings suggest that LBA has the potential to effectively control the growth of E. coli, S. aureus, and mold in soft cheese. Moreover, LBA displays greater preservative efficacy compared to K-Sorbate and nisin.

Enhancing Antibiotic Efficacy in Regenerative Endodontics by Improving Biofilm Susceptibility.

INTRODUCTION: Various strategies have been researched to enhance the susceptibility of biofilms, given their tolerance to antibiotics. This study evaluated the effect of the anti-microbial peptide nisin in association with antibiotics used in regenerative endodontics, exploring different treatment times and biofilm growth conditions. METHODS: A mixture of 10 bacterial species was cultivated on dentin specimens anaerobically for 21 days. Biofilms were treated with 1 mL of high-purity nisin Z (nisin ZP, 200 µg/mL) and a triple antibiotic mixture (TAP: ciprofloxacin + metronidazole + minocycline, 5 mg/mL), alone or in combination. The effectiveness of antimicrobial agents was assessed after 1 and 7 days. During the 7-day period, biofilms were treated under 2 conditions: a single dose in a nutrient-depleted setting (ie, no replenishment of growth medium) and multiple doses in a nutrient-rich environment (ie, renewal of medium and antimicrobial agents every 48 h). After treatments, biofilm cells were dispersed, and total colony-forming units were counted. RESULTS: After 1 d-treatment, nisin ZP + TAP resulted in 2-log cell reduction compared to TAP alone (P < .05). After 7 d-treatment with a single dose, nisin ZP + TAP and TAP reduced bacteria to nonculturable levels (P < .05), whereas repeated antimicrobial doses did not eliminate bacteria in a nutrient-rich environment. No bacterial reduction was observed with nisin ZP alone in any treatment time. CONCLUSIONS: The additional use of nisin improved the TAP activity only after a short exposure time. Longer exposure to TAP or nisin + TAP in a nutrient-deprived environment effectively eliminated biofilms.

Nisin variants: What makes them different and unique?

Nisin A is a lantibiotic bacteriocin typically produced by strains of Lactococcus lactis. This bacteriocin has been approved as a natural food preservative since the late 1980 s and shows antimicrobial activity against a range of food-borne spoilage and pathogenic microorganisms. The therapeutic potential of nisin A has also been explored increasingly both in human and veterinary medicine. Nisin has been shown to be effective in treating bovine mastitis, dental caries, cancer, and skin infections. Recently, it was demonstrated that nisin has an affinity for the same receptor used by SARS-CoV-2 to enter human cells and was proposed as a blocker of the viral infection. Several nisin variants produced by distinct bacterial strains or modified by bioengineering have been described since the discovery of nisin A. These variants present modifications in the peptide structure, biosynthesis, mode of action, and spectrum of activity. Given the importance of nisin for industrial and therapeutic applications, the objective of this study was to describe the characteristics of the nisin variants, highlighting the main differences between these molecules and their potential applications. This review will be useful to researchers interested in studying the specifics of nisin A and its variants.

Potential diagnostic and drug target markers in glioblastoma.

Glioblastoma multiforme (GBM) IDH-wildtype is the most prevalent brain malignancy in adults. However, molecular mechanisms, which leads to GBM have not been completely elucidated. Granulocyte colony-stimulating factor (GCSF), Granulocyte colony-stimulating factor receptor GCSFR, and Signal transducers and activators of transcription 3 (STAT3) have been involved in the occurrence and development of various cancers, but their role in GBM is little known. Herein, we have investigated the gene and protein expression of GCSF, GCSFR, and STAT3 in 21 tissue biopsy samples and also in tumor associated normal tissue (TANT) samples derived from glioblastoma patients, which revealed significantly differential expression of these genes. To validate our findings, we performed a comprehensive integrated analysis of transcriptomic and proteomic profiling of respective genes by retrieving GBM RNA-sequence data from Genome Atlas Databases. GO and KEGG analysis revealed enrichment in disease-related pathways, such as JAK/STAT pathway activation, which were associated with GBM progression. We further performed computational docking analysis of potential drug candidate Nisin against GCSF, and the results were validated in vitro through cytotoxic activity assay using a human glioblastoma cell line SF-767 in a dose-dependent manner. Our comprehensive analysis reveals that GCSF augments glioma progression, and its blockade with anticancer bacteriocin peptide Nisin can potentially inhibit the growth and metastasis of GBM.