Using whey, a by-product of the cheese-making process, is important for maximizing resource efficiency and promoting sustainable practices in the food industry. Reusing whey can help minimize environmental impact and produce bio-preservatives for foods with high bacterial loads, such as Mexican-style fresh cheeses. This research aims to evaluate the antimicrobial and physicochemical effect of CFS from Lactobacillus casei 21/1 produced in a conventional culture medium (MRS broth) and another medium using whey (WB medium) when applied in Mexican-style fresh cheese inoculated with several indicator bacteria (Escherichia coli, Salmonella enterica serovar Typhimurium, Staphylococcus aureus, and Listeria monocytogenes). The CFSs (MRS or WB) were characterized for organic acids concentration, pH, and titratable acidity. By surface spreading, CFSs were tested on indicator bacteria inoculated in fresh cheese. Microbial counts were performed on inoculated cheeses during and after seven days of storage at 4 ± 1.0 °C. Moreover, pH and color were determined in cheeses with CFS treatment. Lactic and acetic acid were identified as the primary antimicrobial metabolites produced by the Lb. casei 21/1 fermentation in the food application. A longer storage time (7 days) led to significant reductions (p < 0.05) in the microbial population of the indicator bacteria inoculated in the cheese when it was treated with the CFSs (MRS or WB). S. enterica serovar Typhimurium was the most sensitive bacteria, decreasing 1.60 ± 0.04 log10 CFU/g with MRS-CFS, whereas WB-CFS reduced the microbial population of L. monocytogenes to 1.67 log10 CFU/g. E. coli and S. aureus were the most resistant at the end of storage. The cheese\'s pH with CFSs (MRS or WB) showed a significant reduction (p < 0.05) after CFS treatment, while the application of WB-CFS did not show greater differences in color (ΔE) compared with MRS-CFS. This study highlights the potential of CFS from Lb. casei 21/1 in the WB medium as an ecological bio-preservative for Mexican-style fresh cheese, aligning with the objectives of sustainable food production and guaranteeing food safety.

It is widely agreed that microbial biofilms play a major role in promoting infection and delaying healing of chronic wounds. In the era of microbial resistance, probiotic strains or their metabolic products are emerging as an innovative approach for the treatment of hard-to-heal (chronic) wounds due to their antimicrobial, healing, and host immune-modulatory effects. In this study, we aimed to investigate the potential of cell-free supernatants (CFS) from Lacticaseibacillus rhamnosus GG against mono- and dual-species biofilms of wound pathogens in a 3D in vitro infection model. Mature biofilms of Pseudomonas aeruginosa and Staphylococcus aureus were obtained on collagen scaffolds in the presence of a simulant wound fluid (SWF) and treated with CFS at different doses and time intervals. At 1:4 dilution in SWF, CFS caused a marked reduction in the colony forming-unit (CFU) numbers of bacteria embedded in mono-species biofilms as well as bacteria released by the biofilms in the supernatant. CFU count and electron microscopy imaging also demonstrated a marked antibiofilm effect against dual-species biofilms starting from 8 h of incubation. Furthermore, CFS exhibited acceptable levels of cytotoxicity at 24 h of incubation against HaCaT cells and, differently from ciprofloxacin, failed to induce resistance after 15 passages at sub-inhibitory concentrations. Overall, the results obtained point to L. rhamnosus GG postbiotics as a promising strategy for the treatment of wound biofilms.

Chronic lung infections caused by Pseudomonas aeruginosa play a significant role in the mortality and morbidity of cystic fibrosis (CF) patients. The widespread bacterial resistance to conventional antimicrobials demands identifying new strategies to complement or replace current antibiotic therapies. In this study, we evaluated the antibacterial, antibiofilm, and antivirulence properties of cell-free supernatants (CFS) from several Lactobacillus probiotic strains against P. aeruginosa isolated from the sputum of CF patients. A strong and fast antibacterial activity of CFS from different strains of lactobacilli was observed at acidic pH towards P. aeruginosa, both in planktonic and biofilm mode of growth, in conditions mimicking CF lung. Interestingly, although when adjusted at pH 6.0, CFS lost most of their antibacterial potential, they retained some antivirulence activity towards P. aeruginosa, largely dependent on the dose, exposure time, and the Lactobacillus-P. aeruginosa strain combination. In vivo testing in the invertebrate Galleria mellonella model disclosed the lack of toxicity of acidic CFS and their ability to prevent P. aeruginosa infection. For the first time, the results revealed lactobacilli postbiotic activities in the context of the pulmonary environment, pointing to innovative postbiotics\' uses in anti-infective therapy.

Some endophyte bacteria can improve plant growth and suppress plant diseases. However, little is known about the potential of endophytes bacteria to promote wheat growth and suppress the Fusarium seedling blight pathogen Fusarium graminearum. This study was conducted to isolate and identify endophytic bacteria and evaluate their efficacy for the plant growth promotion and disease suppression of Fusarium seedling blight (FSB) in wheat. The Pseudomonas poae strain CO showed strong antifungal activity in vitro and under greenhouse conditions against F. graminearum strain PH-1. The cell-free supernatants (CFSs) of P. poae strain CO were able to inhibit the mycelium growth, the number of colonies forming, spore germination, germ tube length, and the mycotoxin production of FSB with an inhibition rate of 87.00, 62.25, 51.33, 69.29, and 71.08%, respectively, with the highest concentration of CFSs. The results indicated that P. poae exhibited multifarious antifungal properties, such as the production of hydrolytic enzymes, siderophores, and lipopeptides. In addition, compared to untreated seeds, wheat plants treated with the strain showed significant growth rates, where root and shoot length increased by about 33% and the weight of fresh roots, fresh shoots, dry roots, and dry shoots by 50%. In addition, the strain produced high levels of indole-3-acetic acid, phosphate solubilization, and nitrogen fixation. Finally, the strain demonstrated strong antagonistic properties as well as a variety of plant growth-promoting properties. Thus, this result suggest that this strain could be used as an alternate to synthetic chemicals, which can serve as an effective method of protecting wheat from fungal infection.

Soil salinity is a major constraint for soybean production worldwide, and the exploitation of plant growth-promoting bacteria (PGPB) and their bioactive metabolite(s) can improve plant salinity tolerance. With this objective, two experiments were performed, aiming to test 4 culture media (YEM(A), TYE(A), TS(A), and LB(A)) for growing a novel Devosia sp. (strain SL43), and then evaluating cell-free supernatants (CFS) from the Devosia sp. on germination of soybean (Glycine max L.) seeds under salinity stress. Soybean seeds were subjected to three salinity levels (0, 100, and 125 mM NaCl) and 6 levels of Devosia sp. CFS dilution (0, 1:1, 1:100, 1:250, 1:500, 1:1000). The results indicated that 125 mM NaCl concentration caused the greatest reduction in the total number of germinated seeds (15%), germination rate (43.6%), root length (55.2%), root weight (39.3%), and seed vigor (68%), and it also increased mean germination time by 71.9%. However, Devosia-CFS improved soybean germination, and the greatest effect was obtained at 1:1 dilution. Under the highest salinity level, application of CFS at 1:1 dilution increased final germination (17.6%), germination rate (18.6%), root length (162.2%), root weight (239.4%), seed vigor index (318.7%), and also shortening mean germination time by 19.2%. The results indicated that seed vigor index was positively correlated with other traits except for mean germination time. Our study suggested that the highest productivity of Devoisa sp. was obtained from the YEM medium. Results also suggested that CFS produced by the novel Devosia sp. (SL43 strain) can successfully alleviate salt stress effects on soybean seed germination and manipulating the chemical composition of the growth medium can influence the effectiveness of these bioactive metabolites.

Synthetic microbial communities (SynComs) could potentially enhance some functions of the plant microbiome and emerge as a promising inoculant for improving crop performance. Here, we characterized a collection of bacteria, previously isolated from the wheat rhizosphere, for their antifungal activity against soilborne fungal pathogens. Ten SynComs with different compositions from 14 bacterial strains were created. Seven SynComs protected wheat from Rhizoctonia solani AG8 infection, although SynComs were not more effective than single strains in reducing wheat root rot disease. Further, the mechanisms of interaction of the tested bacteria with each other and plants were explored. We found that nine bacteria and nine SynComs impacted the root growth of Arabidopsis. Nine bacteria and four SynComs significantly inhibited the growth of AG8 by producing volatiles. The cell-free supernatants from six bacteria inhibited the growth of AG8. Together, this study provided the potential for improving crop resilience by creating SynComs.

BACKGROUND: Probiotics can release bioactive substances known as postbiotics, which can inhibit pathogenic microorganisms, improve immunomodulation, reduce antioxidant production, and modulate the gut microbiota.
METHODS: In this study, we evaluated the in vitro antimicrobial effects, antioxidant activity, and anti-inflammatory potential of 10 lyophilized cell-free supernatants (LCFS) of Lactobacillus isolates. LCFS was obtained via centrifugation and subsequent lyophilization of the supernatant collected from the culture medium ofeach isolate. The antibacterial and antibiofilm activities of the LCFS were determined using broth microdilution. The antioxidant potential was evaluated by measuring the total phenolic and flavonoid contents and 2,2-Diphennyl-1-picrylhydrazyl (DPPH) and 2,2\'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) radical cation (ABTS+) radical scavenging activities.
RESULTS: All the isolates were able to inhibit the four tested pathogens. The isolates exhibited strong antibiofilm activity and eradicated the biofilms formed by Acinetobacter buamannii and Escherichia coli. All the prepared Lactobacillus LCFS contained phenols and flavonoids and exhibited antioxidant activities in the DPPH and ABTS+ radical scavenging assays. The MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) assay revealed that LCFS was not cytotoxic to RAW 264.7 cells. In addition, the ten Lactobacillus LCFS decreased the production of nitric oxide.
CONCLUSIONS: All the isolates have beneficial properties. This research sheds light on the role of postbiotics in functional fermented foods and pharmaceutical products. Further research to elucidate the precise molecular mechanisms of action of probiotics is warranted.

Lactic acid bacteria (LAB) are distinguished by their ability to produce lactic acid, among other interesting metabolites with antimicrobial activity. A cell-free supernatant (CFS) is a liquid containing the metabolites resulting from microbial growth and the residual nutrients of the medium used. CFS from LAB can have antimicrobial activity due to organic acids, fatty acids, and proteinaceous compounds, among other compounds. This review aims to summarize the information about CFS production, CFS composition, and the antimicrobial (antibacterial and antifungal) activity of CFS from LAB in vitro, on foods, and in active packaging. In addition, the mechanisms of action of CFS on cells, the stability of CFS during storage, CFS cytotoxicity, and the safety of CFS are reviewed. The main findings are that CFS\'s antibacterial and antifungal activity in vitro has been widely studied, particularly in members of the genus Lactobacillus. CFS has produced strong inhibition of bacteria and molds on foods when applied directly or in active packaging. In most studies, the compounds responsible for antimicrobial activity are identified. A few studies indicate that CFSs are stable for 1 to 5 months at temperatures ranging from 4 to 35°C. The cytotoxicity of CFS on human cells has not been well studied. However, the studies that have been performed reported no toxicity of CFS. Therefore, it is necessary to investigate novel growth mediums for CFS preparation that are compatible with food sensory properties. More studies into CFS stability and cytotoxic effects are also needed.

Tyramine is one of the most toxic biogenic amines and it is produced commonly by lactic acid bacteria in fermented food products. In present study, we investigated the influence of selected nisin-producing Lactococcus lactis subsp. lactis strains and their cell-free supernatants (CFSs) on tyramine production by four Lactobacillus and two Lactiplantibacillus strains isolated from cheese and beer. Firstly, we examined the antimicrobial effect of the CFSs from twelve Lactococcus strains against tested tyramine producers by agar-well diffusion assay. Six Lactococcus strains whose CFSs showed the highest antimicrobial effect on tyramine producers were further studied. Secondly, we investigated the influence of the selected six Lactococcus strains and their respective CFSs on tyramine production by tested Lactobacillus and Lactiplantibacillus strains in MRS broth supplemented with 2 g.L-1 of l-tyrosine. Tyramine production was monitored by HPLC-UV. The tyramine formation of all tested Lactobacillus and Lactiplantibacillus strains was not detected in the presence of Lc. lactis subsp. lactis CCDM 71 and CCDM 702, and their CFSs. Moreover, the remainder of the investigated Lactococcus strains (CCDM 670, CCDM 686, CCDM 689 and CCDM 731) and their CFSs decreased tyramine production significantly (P < 0.05) - even suppressing it completely in some cases - in four of the six tested tyramine producing strains.

Clostridium difficile infection (CDI) is a common cause of morbidity and mortality in hospitalized patients worldwide. The major problem facing current treatment is multiple recurrences, prompting the need for alternative therapies. In this study we isolated bacterial species, from Egyptian individuals\' stool, with antimicrobial activity against clinical isolates of C. difficile and tried to examine the nature of the produced antimicrobials. In vitro antibacterial activity against C. difficile was initially screened in 123 fecal samples cultures using an agar overlay method. The isolates with antimicrobial activity against C. difficile in addition to Clostridium isolates were identified using partial 16S rDNA gene sequencing analysis. The isolates acting against C. difficile belonged to Lactobacillus, Enterococcus and Clostridium genera. The concentrated cell-free supernatants (CFSs) from these bacterial isolates were examined for antimicrobial activity against C. difficile growth by broth dilution method. 10 x concentrated CFSs of five isolates showed inhibition for C. difficile growth which was significantly different (p < 0.001) from control. Lactobacillus agilis T99A and Clostridium butyricum T58A isolates were selected for further evaluation of the produced antimicrobials. The antimicrobial activity of 10x CFSs of the two isolates was stable after enzymatic treatment with proteinase K or heating treatments up to 90 °C or neutralizing pH. The spectrum of activity of the two isolates was evaluated using different gram-positive and gram-negative bacterial species and did not show antimicrobial activity against these species. Our results showed two unconventional bacterial isolates: L. agilis T99A and C. butyricum T58A producing extracellular thermo stable antimicrobial agents against C. difficile clinical isolates.