Search Results (20)

Background/Objectives: Listeria monocytogenes is a major foodborne pathogen responsible for listeriosis, a serious illness with high morbidity and mortality, particularly in vulnerable populations. Its persistence in food processing environments and resistance to conventional preservation methods pose significant food safety challenges. Lactic acid bacteria (LAB) offer a promising natural alternative due to their antimicrobial properties, especially through the production of bacteriocins. This study investigates the competitive interactions between Lactococcus lactis and L. monocytogenes under co-culture conditions, with a focus on changes in their secretomes to better understand how LAB-derived bacteriocins can help mitigate the Listeria burden. Methods: Proteomic approaches, including Tricine-SDS-PAGE, two-dimensional electrophoresis, and shotgun proteomics, were employed to analyze the molecular adaptations of both species in response to bacterial competition. Results: Our results reveal a significant increase in the secretion of enolase by L. monocytogenes when in competition with L. lactis, suggesting its role as a stress-responsive moonlighting protein involved in adhesion, immune evasion, and biofilm formation. Concurrently, L. lactis exhibited a shift in the production of its bacteriocin, nisin, favoring the expression of Nisin Z—a variant with improved solubility and diffusion properties. This differential regulation indicates that bacteriocin production is modulated by bacterial competition, likely as a defensive response to the presence of pathogens. Conclusions: These findings highlight the dynamic interplay between LAB and L. monocytogenes, underscoring the potential of LAB-derived bacteriocins as natural biopreservatives. Understanding the molecular mechanisms underlying microbial competition could enhance food safety strategies, particularly in dairy products, by reducing reliance on chemical preservatives and mitigating the risk of L. monocytogenes contamination. Full article

Background/Objectives: Diabetic foot infections (DFIs) are commonly associated with frequent hospitalizations, limb amputations, and premature death due to the profile of the bacteria infecting foot ulcers. DFIs are generally colonized by a polymicrobial net of bacteria that grows in biofilms, developing an increased antimicrobial resistance to multiple antibiotics. DFI treatment is a hurdle, and the need to develop new therapies that do not promote resistance is urgent. Therefore, the antibacterial efficacy of Nisin Z (antimicrobial peptide), a core–shell polycationic polyurea pharmadendrimer (PURE_G4OEI₄₈) (antimicrobial polymer), and amlodipine (antihypertensive drug) was evaluated against S. aureus and P. aeruginosa isolated from a DFI and previously characterized. Methods: The antibacterial activity was analyzed in vitro by determining the minimal inhibitory concentration (MIC) and in vivo in a Galleria mellonella model by assessing the larvae survival and health index. Results: The results indicate that Nisin Z exhibited antibacterial activity against S. aureus in vivo, allowing larvae full survival, and no antibacterial activity against P. aeruginosa. Nisin Z may have reduced the antibacterial effectiveness of both PURE_G4OEI₄₈ and amlodipine. PURE_G4OEI₄₈ significantly increased the survival of the larvae infected with P. aeruginosa, while amlodipine showed no activity against both bacteria in vivo. Conclusions: These findings suggest that both Nisin Z and PURE_G4OEI₄₈ could potentially be used individually as adjunct treatments for mild DFIs. However, further studies are needed to confirm these findings and assess the potential toxicity and efficacy of PURE_G4OEI₄₈ in more complex models. Full article

Reducing the use of antibiotics in animal husbandry is essential to limit the spread of resistance. A promising alternative to antibiotics resides in bacteriocins, which are antimicrobial peptides produced by bacteria showing a great diversity in terms of spectrum of activity, structure, and mechanism of action. In this study, the effects of diverse bacteriocins on the composition and metabolic activity of chicken cecal microbiota were examined in vitro, in comparison with antibiotics. Different impacts on microbiota composition were revealed by 16S metabarcoding, with colistin having the most dramatic impact on diversity. Bacteriocins produced by Gram-negative bacteria, microcins J25 and E492, did not significantly influence the microbiota composition. In contrast, bacteriocins from Gram-positive bacteria impacted the abundance of lactic acid bacteria, with nisin Z showing the most impact while pediocin PA-1 (M31L) exhibited a moderate effect at the highest concentration tested. This study emphasizes the potential of bacteriocins as alternatives to antibiotics in poultry to protect from pathogens such as Salmonella, Clostridium, and Enterococcus. Full article

Diabetic foot ulcers (DFU) are a major complication of diabetes mellitus and a public health concern worldwide. The ability of P. aeruginosa to form biofilms is a key factor responsible for the chronicity of diabetic foot infections (DFIs) and frequently associated with the presence of persister cells. These are a subpopulation of phenotypic variants highly tolerant to antibiotics for which new therapeutic alternatives are urgently needed, such as those based on antimicrobial peptides. This study aimed to evaluate the inhibitory effect of nisin Z on P. aeruginosa DFI persisters. To induce the development of a persister state in both planktonic suspensions and biofilms, P. aeruginosa DFI isolates were exposed to carbonyl cyanide m-chlorophenylhydrazone (CCCP) and ciprofloxacin, respectively. After RNA extraction from CCCP-induced persisters, transcriptome analysis was performed to evaluate the differential gene expression between the control, persisters, and persister cells exposed to nisin Z. Nisin Z presented a high inhibitory effect against P. aeruginosa persister cells but was unable to eradicate them when present in established biofilms. Transcriptome analysis revealed that persistence was associated with downregulation of genes related to metabolic processes, cell wall synthesis, and dysregulation of stress response and biofilm formation. After nisin Z treatment, some of the transcriptomic changes induced by persistence were reversed. In conclusion, nisin Z could be considered as a potential complementary therapy for treating P. aeruginosa DFI, but it should be applied as an early treatment or after wound debridement. Full article

Metabiotics are the structural components of probiotic bacteria, functional metabolites, and/or signaling molecules with numerous beneficial properties. A novel Lactococcus lactis strain, UTNCys6-1, was isolated from wild Amazonian camu-camu fruits (Myrciaria dubia), and various functional metabolites with antibacterial capacity were found. The genome size is 2,226,248 base pairs, and it contains 2248 genes, 2191 protein-coding genes (CDSs), 50 tRNAs, 6 rRNAs, 1 16S rRNA, 1 23S rRNA, and 1 tmRNA. The average GC content is 34.88%. In total, 2148 proteins have been mapped to the EggNOG database. The specific annotation consisted of four incomplete prophage regions, one CRISPR-Cas array, six genomic islands (GIs), four insertion sequences (ISs), and four regions of interest (AOI regions) spanning three classes of bacteriocins (enterolysin_A, nisin_Z, and sactipeptides). Based on pangenome analysis, there were 6932 gene clusters, of which 751 (core genes) were commonly observed within the 11 lactococcal strains. Among them, 3883 were sample-specific genes (cloud genes) and 2298 were shell genes, indicating high genetic diversity. A sucrose transporter of the SemiSWEET family (PTS system: phosphoenolpyruvate-dependent transport system) was detected in the genome of UTNCys6-1 but not the other 11 lactococcal strains. In addition, the metabolic profile, antimicrobial susceptibility, and inhibitory activity of both protein–peptide extract (PPE) and exopolysaccharides (EPSs) against several foodborne pathogens were assessed in vitro. Furthermore, UTNCys6-1 was predicted to be a non-human pathogen that was unable to tolerate all tested antibiotics except gentamicin; metabolized several substrates; and lacks virulence factors (VFs), genes related to the production of biogenic amines, and acquired antibiotic resistance genes (ARGs). Overall, this study highlighted the potential of this strain for producing bioactive metabolites (PPE and EPSs) for agri-food and pharmaceutical industry use. Full article

Lactococcus garvieae is a main ichthyopathogen in rainbow trout (Oncorhynchus mykiss, Walbaum) farming, although bacteriocinogenic L. garvieae with antimicrobial activity against virulent strains of this species have also been identified. Some of the bacteriocins characterized, such as garvicin A (GarA) and garvicin Q (GarQ), may show potential for the control of the virulent L. garvieae in food, feed and other biotechnological applications. In this study, we report on the design of Lactococcus lactis strains that produce the bacteriocins GarA and/or GarQ, either alone or together with nisin A (NisA) or nisin Z (NisZ). Synthetic genes encoding the signal peptide of the lactococcal protein Usp45 (SP_usp45), fused to mature GarA (lgnA) and/or mature GarQ (garQ) and their associated immunity genes (lgnI and garI, respectively), were cloned into the protein expression vectors pMG36c, which contains the P₃₂ constitutive promoter, and pNZ8048c, which contains the inducible P_nisA promoter. The transformation of recombinant vectors into lactococcal cells allowed for the production of GarA and/or GarQ by L. lactis subsp. cremoris NZ9000 and their co-production with NisA by Lactococcus lactis subsp. lactis DPC5598 and L. lactis subsp. lactis BB24. The strains L. lactis subsp. cremoris WA2-67 (pJFQI), a producer of GarQ and NisZ, and L. lactis subsp. cremoris WA2-67 (pJFQIAI), a producer of GarA, GarQ and NisZ, demonstrated the highest antimicrobial activity (5.1- to 10.7-fold and 17.3- to 68.2-fold, respectively) against virulent L. garvieae strains. Full article

Diabetes mellitus (DM) patients frequently develop diabetic foot ulcers (DFU) which are generally infected by a community of microorganisms, mainly Staphylococcus aureus and Pseudomonas aeruginosa. These bacteria exhibit a multi-drug resistance profile and biofilm-forming ability which represent a hurdle in the treatment of diabetic foot infections (DFI). We aimed to evaluate the potential of Nisin Z, an antimicrobial peptide (AMP), as an alternative treatment for severe DFI. Nisin Z shows antibacterial activity against Gram-positive and Gram-negative bacteria and an increased antibacterial effect against Gram-negatives when added to EDTA. As such, Minimum Inhibitory Concentration (MIC), Minimum Bactericidal Concentration (MBC), Minimum Biofilm Inhibitory Concentration (MBIC), and Minimum Biofilm Eradication Concentration (MBEC) were determined for Nisin Z, Nisin Z + EDTA (0.4%), and Nisin Z + EDTA incorporated into guar gum, in order to test its efficacy against S. aureus and P. aeruginosa isolated from the same DFU. Results showed that Nisin Z added to the chelation agent EDTA displayed higher antibacterial and bacteriostatic efficacy against mono and dual co-cultures of S. aureus and P. aeruginosa, and higher antibiofilm efficiency against monocultures. Nisin Z was moderately cytotoxic at 200 µg/mL. Prospect in vivo studies are needed to confirm the potential of Nisin Z supplemented with EDTA to be used as a complement to conventional antibiotic therapy for severe DFI. Full article

Burn wounds can lead to numerous severe complications including bacterial infections causing patient morbidity and mortality, mostly in low- and middle-income countries. The considerable increase in microbial resistance against traditional antibiotics is leading towards alternative strategies to treat bacterial infections. Nisin Z is an antimicrobial peptide which exhibits a significant antibacterial activity against Gram-positive bacteria. Its efficacy against Gram-negative bacteria is limited, nonetheless it can be improved with the addition of surfactants, such as ethylenediaminetetraacetic acid (EDTA). The incorporation of peptide and other biomolecules within a biopolymer matrix provides protection maintaining their antimicrobial potential. Bacterial nanocellulose (BNC) has been widely used as wound dressings. Its impressive water retention capacity (>99%) and porosity are beneficial to manage wounds due to its potential to absorb exudates, providing a breathable and humid environment. In this work, the functionalization of BNC with Nisin Z (BNC-NZ) via vacuum filtration is reported. The entrapment of the peptide inside the BNC films was confirmed through morphological characterization using attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectrometry. Typical absorbance peaks of Nisin Z are easily identifiable at 1647 cm⁻¹ (amide group) and 1520 cm⁻¹ (bending of primary amines). Thermal gravimetric analysis (TGA) suggested that Nisin Z did not interfere with the BNC matrix. The antimicrobial activity of Nisin Z against five of the most common bacteria found in burn wounds was verified by minimum bactericidal concentration (MBC) ranging 8.0–256.0 µg/mL. Agar diffusion and shake flask methods revealed the potential of BNC-NZ for prospective applications in burn wound dressings. Full article

The growing environmental concern with the inappropriate disposal of conventional plastics has driven the development of eco-friendly food packaging. However, the intrinsic characteristics of polymers of a renewable origin, e.g., poor mechanical properties, continue to render their practical application difficult. For this, the present work studied the influence of ionic strength (IS) from 0 to 500 mM to modulate the physicochemical properties of methylcellulose (MC). Moreover, for protection against biological risks, Nisin-Z was incorporated into MC’s polymeric matrices, providing an active function. The incorporation of salts (LiCl and MgCl₂) promoted an increase in the equilibrium moisture content in the polymer matrix, which in turn acted as a plasticizing agent. In this way, films with a hydrophobic surface (98°), high true strain (85%), and low stiffness (1.6 mPa) can be manufactured by addition of salts, modulating the IS to 500 mM. Furthermore, films with an IS of 500 mM, established with LiCl, catalyzed antibacterial activity against E. coli, conferring synergism and extending protection against biological hazards. Therefore, we demonstrated that the IS control of MC dispersion presents a new alternative to achieve films with the synergism of antibacterial activity against Gram-negative bacteria in addition to flexibility, elasticity, and hydrophobicity required in various applications in food packaging. Full article

At present, antibiotic resistance is considered a real problem. Therefore, for decades scientists have been looking for novel strategies to treat bacterial infections. Nisin Z, an antimicrobial peptide (AMP), can be considered an option, but its usage is mainly limited by the poor stability and short duration of its antimicrobial activity. In this context, cyclodextrin (CD)-based nanosponges (NSs), synthesized using carbonyldiimidazole (CDI) and pyromellitic dianhydride (PMDA), were chosen for nisin Z loading. To determine the minimum inhibitory of nisin Z loaded on CD-NS formulations, agar well diffusion plates were used. Then, the bactericide concentrations of nisin Z loaded on CD-NS formulations were determined against Gram-positive (Staphylococcus aureus) and -negative (Escherichia coli) bacteria, using microdilution brain heart infusion (BHI) and tetrazolium salt 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT). The minimum and bactericide inhibitory values of the nisin complex with NSs were potentially decreased against both bacteria, compared with the nisin-free sample, while the nisin complex with β-CD showed lower antibacterial activity. The antimicrobial effect was also demonstrated by free NSs. Furthermore, the total viable counts (TVCs) antibacterial experiment indicated that the combination of nisin Z in both PMDA and CDI β-CD-based NSs, especially CDI, can provide a better conservative effect on cooked chicken meat. Generally, the present study outcomes suggest that the cross-linked β-CD-based NSs can present their own antimicrobial potency or serve as promising carriers to deliver and enhance the antibacterial action of nisin Z. Full article

Probiotics are a viable alternative to traditional chemotherapy agents to control infectious diseases in aquaculture. In this regard, Lactococcus lactis subsp. cremoris WA2-67 has previously demonstrated several probiotic features, such as a strong antimicrobial activity against ichthyopathogens, survival in freshwater, resistance to fish bile and low pH, and hydrophobicity. The aim of this manuscript is an in silico analysis of the whole-genome sequence (WGS) of this strain to gain deeper insights into its probiotic properties and their genetic basis. Genomic DNA was purified, and libraries prepared for Illumina sequencing. After trimming and assembly, resulting contigs were subjected to bioinformatic analyses. The draft genome of L. cremoris WA2-67 consists of 30 contigs (2,573,139 bp), and a total number of 2493 coding DNA sequences (CDSs). Via in silico analysis, the bacteriocinogenic genetic clusters encoding the lantibiotic nisin Z (NisZ) and two new bacteriocins were identified, in addition to several probiotic traits, such as the production of vitamins, amino acids, adhesion/aggregation, and stress resistance factors, as well as the absence of transferable antibiotic resistance determinants and genes encoding detrimental enzymatic activities and virulence factors. These results unveil diverse beneficial properties that support the use of L. cremoris WA2-67 as a probiotic for aquaculture. Full article

The great variability of cancer types demands novel drugs with broad spectrum, this is the case of Nisin, a polycyclic antibacterial peptide that recently has been considered for prevention of cancer cells growth. As an accepted food additive, this drug would be very useful for intestinal cancers, but the peptide nature would make easier its degradation by digestion procedures. For that reason, the aim of present study to investigate the protective effect of two different β-cyclodextrin-based nanosponges (carbonyl diimidazole and pyromellitic dianhydride) and their anti-cancer enhancement effect of Nisin-Z encapsulated with against colon cancer cells (HT-29). To extend its possible use, a comparison with breast (MCF-7) cancer cell was carried out. The physicochemical properties, loading efficiency, and release kinetics of Nisin complex with nanosponges were studied. Then, tricin-SDS-PAGE electrophoresis was used to understand the effect of NSs on stability of Nisin-Z in the presence of gastric peptidase pepsin. In addition, the cytotoxicity and cell membrane damage of Nisin Z were evaluated by using the MTT and LDH assay, which was complemented via Annexin-V/ Propidium Iodide (PI) by using flowcytometry. CD-NS are able to complex Nisin-Z with an encapsulation efficiency around 90%. A protective effect of Nisin-Z complexed with CD-NSs was observed in presence of pepsin. An increase in the percentage of apoptotic cells was observed when the cancer cells were exposed to Nisin Z complexed with nanosponges. Interestingly, Nisin Z free and loaded on PMDA/CDI-NSs is more selectively toxic towards HT-29 cells than MCF-7 cancer cells. These results indicated that nanosponges might be good candidates to protect peptides and deliver drugs against intestinal cancers. Full article

Lactococcus lactis is a commonly used fermenting bacteria in cheese, beverages and meat products. Due to the lack of simplified chassis strains, it has not been widely used in the fields of synthetic biology. Thus, the construction of lactic acid bacteria chassis strains becomes more and more important. In this study, we performed whole genome sequencing, annotation and analysis of L. lactis N8. Based on the genome analysis, we found that L. lactis N8 contains two large plasmids, and the function prediction of the plasmids shows that some regions are related to carbohydrate transport/metabolism, multi-stress resistance and amino acid uptake. L. lactis N8 contains a total of seven prophage-related fragments and twelve genomic islands. A gene cluster encoding a hybrid NRPS–PKS system that was found in L. lactis N8 reveals that the strain has the potential to synthesize novel secondary metabolites. Furthermore, we have constructed a simplified genome chassis of L. lactis N8 and achieved the largest amount of deletion of L. lactis so far. Taken together, the present study offers further insights into the function and potential role of L. lactis N8 as a model strain of lactic acid bacteria and lays the foundation for its application in the field of synthetic biology. Full article

On behalf of a circular economy, regular plastics have been replaced by biodegradable packagings. Besides, active films have been applied to improve the shelf-life and quality of foods. In this work, blends were developed using starch as a low-cost natural polymer, mixed with poly(vinyl alcohol) due to its physical-chemical and biodegradable properties. Moreover, maleic anhydride (MaAh), cellulose-nanocrystal (CN), and nisin-z (N-Z) were added, respectively, as a compatibilizer, a mechanical-reinforce, and antimicrobial agents. The thermal stability of the films was analyzed, which blends’ melting temperature occurred around 200–207 °C, and it was influenced by CN, N-Z, and MaAh amounts. N-Z and MaAh acted against S. aureus and P. aeruginosa by compound diffusion (inhibition-halo around 1.85 and 2.18 cm); while S. Choleraesuis and E. coli were inhibited by contact. Therefore, these blends presented the potential to be used as active biodegradable packaging in the food industry. Full article

Nisin Z, an amphipathic peptide, with a significant antibacterial activity against Gram-positive bacteria and low toxicity in humans, has been studied for food preservation applications. Thus far, very little research has been done to explore its potential in biomedicine. Here, we report the modification of sodium alginate (SA) and gelatin (GN) blended microfibers, produced via the wet-spinning technique, with Nisin Z, with the purpose of eradicating Staphylococcus aureus-induced infections. Wet-spun SAGN microfibers were successfully produced at a 70/30% v/v of SA (2 wt%)/GN (1 wt%) polymer ratio by extrusion within a calcium chloride (CaCl₂) coagulation bath. Modifications to the biodegradable fibers’ chemical stability and structure were then introduced via crosslinking with CaCl₂ and glutaraldehyde (SAGNCL). Regardless of the chemical modification employed, all microfibers were labelled as homogeneous both in size (≈246.79 µm) and shape (cylindrical and defect-free). SA-free microfibers, with an increased surface area for peptide immobilization, originated from the action of phosphate buffer saline solution on SAGN fibers, were also produced (GNCL). Their durability in physiological conditions (simulated body fluid) was, however, compromised very early in the experiment (day 1 and 3, with and without Nisin Z, respectively). Only the crosslinked SAGNCL fibers remained intact for the 28 day-testing period. Their thermal resilience in comparison with the unmodified and SA-free fibers was also demonstrated. Nisin Z was functionalized onto the unmodified and chemically altered fibers at an average concentration of 178 µg/mL. Nisin Z did not impact on the fiber’s morphology nor on their chemical/thermal stability. However, the peptide improved the SA fibers (control) structural integrity, guaranteeing its stability for longer, in physiological conditions. Its main effect was detected on the time-kill kinetics of the bacteria S. aureus. SAGNCL and GNCL loaded with Nisin Z were capable of progressively eliminating the bacteria, reaching an inhibition superior to 99% after 24 h of culture. The peptide-modified SA and SAGN were not as effective, losing their antimicrobial action after 6 h of incubation. Bacteria elimination was consistent with the release kinetics of Nisin Z from the fibers. In general, data revealed the increased potential and durable effect of Nisin Z (significantly superior to its free, unloaded form) against S. aureus-induced infections, while loaded onto prospective biomedical wet-spun scaffolds. Full article