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Probiotics offer a promising prophylactic approach against various pathogens and represent an alternative strategy to combat biofilm-related infections. In this study, we isolated vaginal commensal microbiota from 54 healthy Indian women to investigate their probiotic traits. We primarily explored the ability of cell-free supernatant (CFS) from Lactobacilli to prevent Uropathogenic Escherichia coli (UPEC) colonization and biofilm formation. Our findings revealed that CFS effectively reduced UPEC's swimming and swarming motility, decreased cell surface hydrophobicity, and hindered matrix production by downregulating specific genes (fimA, fimH, papG, and csgA). Subsequent GC-MS analysis identified Tryptamine, a monoamine compound, as the potent bioactive substance from Lactobacilli CFS, inhibiting UPEC biofilms with an MBIC of 4 µg/ml and an MBEC of 8 µg/ml. Tryptamine induced significant changes in E. coli colony biofilm morphology, transitioning from the Red, Dry, and Rough (RDAR) to the Smooth and White phenotype, indicating reduced extracellular matrix production. Biofilm time-kill assays demonstrated a four-log reduction in UPEC viability when treated with Tryptamine, highlighting its potent antibacterial properties, comparable to CFS treatment. Biofilm ROS assays indicated a significant elevation in ROS generation within UPEC biofilms, suggesting a potential antibacterial mechanism. Gene expression studies with Tryptamine-treated samples showed a reduction in expression of curli gene (csgA), consistent with CFS treatment. This study underscores the potential of Tryptamine from probiotic Lactobacilli CFS as a promising antibiofilm agent against UPEC biofilms., (© 2024. The Author(s).)

Background: Probiotics and their derived postbiotics, as cell-free supernatants (CFS), are gaining a solid reputation owing to their prodigious health-promoting effects. Probiotics play a valuable role in the alleviation of various diseases among which are infectious diseases and inflammatory disorders. In this study, three probiotic strains, Lactiplantibacillus plantarum, Lacticaseibacillus rhamnosus, and Pediococcus acidilactici, were isolated from marketed dietary supplements. The antimicrobial activity of the isolated probiotic strains as well as their CFS was investigated. The neutralized CFS of the isolated probiotics were tested for their antibiofilm potential. The anti-inflammatory activity of the isolated Lactobacillus spp., together with their CFS, was studied in the carrageenan-induced rat paw edema model in male Wistar rats. To the best of our knowledge, such a model was not previously experimented to evaluate the anti-inflammatory activity of the CFS of probiotics. The histopathological investigation was implemented to assess the anti-inflammatory prospect of the isolated L. plantarum and L. rhamnosus strains as well as their CFS., Results: The whole viable probiotics and their CFS showed variable growth inhibition of the tested indicator strains using the agar overlay method and the microtiter plate assay, respectively. When tested for virulence factors, the probiotic strains were non-hemolytic lacking both deoxyribonuclease and gelatinase enzymes. However, five antibiotic resistance genes, blaZ, ermB, aac(6')- aph(2"), aph(3'')-III, and vanX, were detected in all isolates. The neutralized CFS of the isolated probiotics exhibited an antibiofilm effect as assessed by the crystal violet assay. This effect was manifested by hindering the biofilm formation of the tested Staphylococcus aureus and Pseudomonas aeruginosa clinical isolates in addition to P. aeruginosa PAO1 strain. Generally, the cell cultures of the two tested probiotics moderately suppressed the acute inflammation induced by carrageenan compared to indomethacin. Additionally, the studied CFS relatively reduced the inflammatory changes compared to the inflammation control group but less than that observed in the case of the probiotic cultures treated groups., Conclusions: The tested probiotics, along with their CFS, showed promising antimicrobial and anti-inflammatory activities. Thus, their safety and their potential use as biotherapeutics for bacterial infections and inflammatory conditions are worthy of further investigation., (© 2023. The Author(s).)

High-moisture mozzarella cheese (HMMC) is a highly perishable cheese with a short shelf life. In this study, the effects of UF cheese whey containing postbiotics from Lactobacillus acidophilus LA-5 (P-LA-5), Bifidobacterium animalis BB-12 (P-BB-12), and their combination on the microbial (i.e., psychrophiles, mesophiles, lactic acid bacteria, and mold-yeast population) and sensory properties of HMMC were investigated. Postbiotics were prepared in a cheese whey model medium as a novel growth media and used as a preserving liquid in HMMC. The results demonstrate that postbiotics reduced the growth of all microorganisms (1.5-2 log CFU/g reduction) and P-LA5 and P-BB12 had the highest antibacterial performance on mesophiles and psychrophiles, respectively. Mold and yeast had the highest susceptibility to the postbiotics. Postbiotics showed a significant effect on maintaining the microbial quality of HMMC during storage, proposing postbiotics as a new preserving liquid for HMMC.

Background: Lactobacillus rhamnosus SD11 is a probiotic derived from the human oral cavity and has potential being used for dental prophylaxis. The cell free supernatant (CFS) of L. rhamnosus SD11 has good antimicrobial and antioxidant effects., Aim: This study aimed to incorporate CFS of the probiotic into thermoresponsive copolymers to create a sol-gel formulation., Methods: The sol-gel formulation was developed using Poloxamer 407 as the main polymer, which was mixed with natural polymers such as gellan gum, sodium alginate, and xyloglucan in different proportions. The sol-gel formulations were characterized based on their physicochemical parameters such as appearance, pH, viscosity, flow-ability in low temperature, antioxidant and antibacterial activity. An in vitro release study was performed using Franz's diffusion cell and the stability was determined under freeze-thaw cycle conditions., Results: The combination of 15% w/v of poloxamer 407 with 0.5% w/v of sodium alginate was the best sol-gel formulation to deliver the CFS of the probiotic., Conclusion: This study was successful in creating a sol-gel formulation using a thermoresponsive copolymer, that could efficiently deliver CFS of the probiotic L. rhamnosus SD11., Competing Interests: No potential conflict of interest was reported by the author(s)., (© 2021 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group.)

Biogenics are compounds produced by living organisms such as animals, plants, bacteria, etc. Probiotics and their biogenics are known for their antimicrobial potential. Therefore, the present study was designed to evaluate the antibiofilm potential of probiotic-derived biogenics in conjunction with zingerone against the Pseudomonas aeruginosa PAO1 biofilm. Cell-free supernatant (CFS) of potential probiotics Pediococcus acidilactici BNS5B and Lactobacillus fermentum PUM was found to inhibit the growth of Ps. aeruginosa PAO1 maximally among the nineteen isolated lactic acid bacteria. L. fermentum PUM produced precipitated protein fraction (PP), organic acids (OAs), exopolysaccharides (EPSs), biosurfactants (BSs) and various volatile antimicrobial compounds, while Ped. acidilactici BNS5B was found to produce PP, OA, EPS, BS and fewer volatile antimicrobial compounds only. More specifically, CFS and selected biogenics (OA and PP from L. fermentum PUM; OA from Ped. acidilactici BNS5B) of both potential probiotics showed synergy with zingerone against Ps. aeruginosa growth as observed by FIC index (< 0.5). Interestingly, CFS of both potential probiotics in combination with zingerone led to the formation of a more distorted biofilm compared with OA of L. fermentum PUM and zingerone, OA of Ped. acidilactici BNS5B and zingerone, PP of L. fermentum PUM and zingerone as well as their individual counterparts. Similarly, both confocal laser scanning microscopy and XTT assay showed an increased number of dead and impaired cells along with the decreased viability of biofilm cells. Thus, it can be reckoned that a combination of probiotic-derived biogenics and zingerone can have therapeutic application against Ps. aeruginosa infections which needs to be validated clinically., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Postbiotics may be defined as soluble metabolites released by food-grade microorganisms during the growth and fermentation in complex microbiological culture, food or gut. It is rich in high and low molecular weight biologically active metabolites. There are still gaps concerning these substances, mainly how to use them for food applications. Although the most recent work on preparation and application of postbiotics from several probiotics are very encouraging, the suitability of postbiotics to combat microorganisms that deal with food safety should be tested mainly by analyzing the chemical composition and conducting antagonistic tests. Consequently, foods can effectively benefit from an identified postbiotic with a defined effect. This review approached the recent advances in relation to the preparation of postbiotics from lactic acid bacteria. The function of different instrumental analysis techniques and factors affecting the chemical composition of postbiotics were also comprehensively reviewed., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The aim of the work is to assess the antimicrobial activities of Cell Free Supernatants (CFS) and Membrane Vesicles (MVs), produced by Lactobacillus reuteri DSM 17938, versus Gram-positive and Gram-negative bacteria and investigate their metabolic profiles. The Minimum Inhibitory Concentration was determined through the broth microdilution method and cell proliferation assay and the Minimum Bactericidal Concentration was determined by Colony Forming Units counts. The characteristics of the antimicrobial compounds were evaluated by pH adjustments, proteinase treatment, and size fractionation of the CFS. The cytotoxicity of CFS was tested on two human cell lines. A detailed snapshot of the L . reuteri metabolism was attained through an untargeted metabolic profiling by means of high resolution Fourier Transform Ion Cyclotron Resonance Mass Spectrometry (FT-ICR MS) coupled with Electrospray Ionization Source (ESI). The results showed (i) a greater efficacy of CFS and its fractions towards Gram-negative compared to Gram-positive bacteria; (ii) an antimicrobial effect related to pH-dependent compounds but not to MVs; (iii) a molecular weight < 3 KDa as well as an a non-proteinaceous nature of the antimicrobial compounds; and (iv) more than 200 and 500 putative metabolites annotated in MVs and supernatants, covering several classes of metabolites, including amino acids, lipids, fatty and organic acids, polyalcohols, nucleotides, and vitamins. Some putative compounds were proposed not only as characteristic of specific fractions, but also possibly involved in antimicrobial activity., Competing Interests: Authors S.R. and H.B. are currently employed by the company BioGaia AB. Author R.G. has received a research grant from Company BioGaia which supported the microbiological part of the funded project. The remaining authors declare no competing interests.

Cell free supernatants (CFS) obtained from probiotic species are routinely used to preliminary investigate the antimicrobial activity of potential probiotic isolates by the agar well diffusion and broth culture. Both methods have documented limitations. In this work, the potential of isothermal microcalorimetry (IMC), a technique based on the measurement of heat produced by growing bacteria was used to investigate the antimicrobial effects of two commercial probiotic species (Lactobacillus acidophilus LA-5 ® and Bifidobacterium lactis BB-12 ® ) and one reference strain (Bifidobacterium bifidum ATCC 11863) against Pseudomonas aeruginosa NCIMB 8628 using unmodified, neutralised and heat-treated CFS. P. aeruginosa was inhibited in growth by the unmodified CFS of all the species. No inhibitory activity was recorded for neutralised CFS of all the species using the agar well diffusion assay. Plate count during co-incubation of P. aeruginosa with the neutralised CFS of all the species showed no inhibition. However, IMC data showed significant inhibition with neutralised CFS obtained from the two Bifidobacterium species suggesting presence of other non-acidic inhibitory compounds in the CFS. The results in this work demonstrated that IMC has potential in probiotic bioassay as it has the capability to record in real-time and capture even subtle effects, which could be unnoticed with traditional assays.

Probiotics have beneficial effects on management of type 2 diabetes (T2D). The major hallmarks of T2D are insulin deficiency and insulin resistance which emphasize insulin therapy in onset of disease. Lactobacilli such as Lactobacillus acidophilus ( L. acidophilus ) have well known properties on prevention of T2D and insulin resistance but not on insulin degradation. Insulin-degrading enzyme (IDE) degrades insulin in the human body. We studied the effects of cell-free supernatant (CFS) and ultraviolet (UV)-killed L. acidophilus (ATCC 314) on IDE activity and insulin degradation in vitro. Cell growth inhibition by CFS and UV-killed L. acidophilus (ATCC 314) was studied and Western blotting and a fluoregenic assay was performed to determine IDE expression and its activity, respectively. Insulin degradation was evaluated by sandwich enzyme-linked immunosorbent assay(ELISA). IDE expression and activity was reduced by CFS and UV-killed L. acidophilus (ATCC 314). Although, decreased enzyme expression and activity was not significant for CFS in contrast to MRL (MRS with same pH as CFS). Also, reduction in IDE activity was not statistically considerable when compared to IDE expression. Insulin degradation was increased by CFS but decreased by UV-killed L. acidophilus (ATCC 314)., Competing Interests: The authors declare no conflict of interest.

The enterovirus Coxsackievirus B4 (CV-B4) can infect different human tissues and provoke abnormal function or destruction of various organs and cells. Moreover, its infections have been linked to the onset of type 1 diabetes. Coxsackievirus B4 is classified as a "challenging virus", due to the intense yet vain efforts to find effective prevention and therapeutic agents, especially within biological compounds. Lactobacillus plantarum is a lactic acid bacterium that is endowed with probiotic properties, and holds great potential for applications in medical and food industry sectors. Several compounds produced by this microorganism have been associated with various benefits including antimicrobial activity. In this work, we investigated the possible antiviral abilities of two Lb. plantarum strains and their derivatives against CV-B4. The different assays carried out (e.g. pre-incubation, competition and post-infection, using HEp-2 cells as human cell model) suggest that the tested microorganisms and their derivatives have an in vitro inhibiting activity against CV-B4. This is the first report showing the anti-CVB4 activity of Lb. plantarum strains and their derivatives., (Copyright © 2018 Elsevier GmbH. All rights reserved.)

The aim of the present study was to evaluate the dietary administration of Bacillus aerophilus KADR3 on enhancing the immunity and disease resistance against Aeromonas hydrophila infection in Labeo rohita. B. aerophilus culture supernatant exhibited antagonistic activity against fish pathogenic bacteria in an agar well diffusion assay. Four fish groups were fed with either of following diets containing different concentrations of B. aerophilus KADR3 (cfu g -1 ): 0 (control), 1 × 10 7 (DI), 1 × 10 8 (DII), and 1 × 10 9 (DIII) -cfu g -1 . Various innate immune parameters were measured at - 3rd and 6th - week post-feeding. At the end of 6th week, fish were challenged intraperitoneally with A. hydrophila and survival percentage was recorded over 10 days post-challenge. Studied immunological parameters viz. serum lysozyme, phagocytosis, serum total protein, respiratory bursts, serum IgM levels, superoxide dismutase and alternative complement pathway activities were significantly enhanced (P < 0.05) in fish groups fed with B. aerophilus KADR3 supplemented diets, with the highest values were observed in DII (10 8  cfu g -1 ) fed group. Further, B. aerophilus supplementation at 10 8  cfu g -1 exhibited highest post-challenge survivability i.e. 72.83%, followed by DIII (64.19%) and DI (41.97%). Our results collectively suggest that B. aerophilus can potentially be used as probiotic strain in aquaculture to enhance the immunity and disease resistance with an optimal dietary supplementation of 10 8  cfu g -1 ., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

In view of emerging drug resistance in pathogens, there is a need to explore alternative strategies to combat infections. Use of probiotics is one such option. In this regard, efficacy of Lactobacillus plantarum has been reported against Staphylococcus aureus . Here, we propose that cell free supernatant (CFS) of Lactobacillus paraplantarum when used in combination with conventional antibiotics viz. ampicillin and oxacillin [to which the methicillin resistant Staphylococcus aureus (MRSA) strains were originally resistant] reduce the minimum inhibitory concentrations of these antibiotics, rendering the combination either synergistic or additive against the tested MRSA strain. The anti-staphylococcal activity was observed to be due to organic acids (acetic acid and lactic acid as confirmed by HPLC analysis) present in the CFS, as neutralization of the CFS with an alkali, sodium hydroxide (NaOH), caused the complete abrogation of its activity. The role of H 2 O 2 and bacteriocin present in the CFS was also ruled out. The findings of this study suggest that cell free supernatant and ampicillin/oxacillin combination(s) might help in rejuvenating the use of conventional anti-staphylococcal antibiotics for the treatment of multi-drug resistant strains.

This study was undertaken to characterize a lactic acid bacterium 4I1, isolated from the freshwater fish, Zacco koreanus . Morphological, biochemical, and molecular characterization of 4I1 revealed it to be Pediococcus pentosaceus 4I1. The cell free supernatant (CFS) of P. pentosaceus 4I1 exhibited significant ( p < 0.05) antibacterial effects (inhibition zone diameters: 16.5-20.4 mm) against tested foodborne pathogenic bacteria with MIC and MBC values of 250-500 and 500-1,000 μg/mL, respectively. Further, antibacterial action of CFS of P. pentosaceus 4I1 against two selected bacteria Staphylococcus aureus KCTC-1621 and Escherichia coli O157:H7 was determined in subsequent assays. The CFS of P. pentosaceus 4I1 revealed its antibacterial action against S. aureus KCTC-1621 and E. coli O157:H7 on membrane integrity as confirmed by a reduction in cell viability, increased potassium ion release (900 and 800 mmol/L), reduced absorption at 260-nm (3.99 and 3.77 OD), and increased relative electrical conductivity (9.9 and 9.7%), respectively. Gas chromatography-mass spectrometry (GC-MS) analysis of the CFS of P. pentosaceus 4I1 resulted in the identification of seven major compounds, which included amino acids, fatty acids and organic acids. Scanning electron microscopic-based morphological analysis further confirmed the antibacterial effect of CFS of P. pentosaceus 4I1 against S. aureus KCTC-1621 and E. coli O157:H7. In addition, the CFS of P. Pentosaceus 4I1 displayed potent inhibitory effects on biofilms formation by S. aureus KCTC-1621 and E. coli O157:H7. The study indicates the CFS of P. pentosaceus 4I1 offers an alternative means of controlling foodborne pathogens.