Intracellular pathogens including Staphylococcus aureus contribute to the non‐healing phenotype of chronic wounds. Lactobacilli, well known as beneficial bacteria, are also reported to modulate the immune system, yet their role in cutaneous immunity remains largely unknown. We explored the therapeutic potential of bacteria‐free postbiotics, bioactive lysates of lactobacilli, to reduce intracellular S. aureus colonization and promote healing. Fourteen postbiotics derived from various lactobacilli species were screened, and Latilactobacillus curvatus BGMK2‐41 was selected for further analysis based on the most efficient ability to reduce intracellular infection by S. aureus diabetic foot ulcer clinical isolate and S. aureus USA300. Treatment of both infected keratinocytes in vitro and infected human skin ex vivo with BGMK2‐41 postbiotic cleared S. aureus . Keratinocytes treated in vitro with BGMK2‐41 upregulated expression of antimicrobial response genes, of which DEFB4 , ANG , and RNASE7 were also found upregulated in treated ex vivo human skin together with CAMP exclusively upregulated ex vivo. Furthermore, BGMK2‐41 postbiotic treatment has a multifaceted impact on the wound healing process. Treatment of keratinocytes stimulated cell migration and the expression of tight junction proteins, while in ex vivo human skin BGMK2‐41 increased expression of anti‐inflammatory cytokine IL‐10, promoted re‐epithelialization, and restored the epidermal barrier via upregulation of tight junction proteins. Together, this provides a potential therapeutic approach for persistent intracellular S. aureus infections.

Pomegranate peel extract (PoPEx) has been shown to have antioxidant and anti-inflammatory properties, but its effect on the adaptive immune system has not been sufficiently investigated. In this study, the treatment of human peripheral blood mononuclear cells (PBMC) with PoPEx (range 6.25–400 µg/mL) resulted in cytotoxicity at concentrations of 100 µg/mL and higher, due to the induction of apoptosis and oxidative stress, whereas autophagy was reduced. At non-cytotoxic concentrations, the opposite effect on these processes was observed simultaneously with the inhibition of PHA-induced PBMC proliferation and a significant decrease in the expression of CD4. PoPEx differently modulated the expression of activation markers (CD69, CD25, ICOS) and PD1 (inhibitory marker), depending on the dose and T-cell subsets. PoPEx (starting from 12.5 µg/mL) suppressed the production of Th1 (IFN-γ), Th17 (IL-17A, IL-17F, and IL-22), Th9 (IL-9), and proinflammatory cytokines (TNF-α and IL-6) in culture supernatants. Lower concentrations upregulated Th2 (IL-5 and IL-13) and Treg (IL-10) responses as well as CD4+CD25hiFoxp3+ cell frequency. Higher concentrations of PoPEx increased the frequency of IL-10- and TGF-β-producing T-cells (much higher in the CD4+ subset). In conclusion, our study suggested for the first time complex immunoregulatory effects of PoPEx on T cells, which could assist in the suppression of chronic inflammatory and autoimmune diseases.

Over-activated myeloid cells and disturbance in gut microbiota composition are critical factors contributing to the pathogenesis of Multiple Sclerosis (MS). Myeloid-derived suppressor cells (MDSCs) emerged as promising regulators of chronic inflammatory diseases, including autoimmune diseases. However, it remained unclear whether MDSCs display any therapeutic potential in MS, and how this therapy modulates gut microbiota composition. Here, we assessed the potential of in vitro generated bone marrow-derived MDSCs to ameliorate experimental autoimmune encephalomyelitis (EAE) in Dark Agouti rats and investigated how their application associates with the changes in gut microbiota composition. MDSCs differentiated with prostaglandin (PG)E2 (MDSC-PGE2) and control MDSCs (differentiated without PGE2) displayed strong immunosuppressive properties in vitro, but only MDSC-PGE2 significantly ameliorated EAE symptoms. This effect correlated with a reduced infiltration of Th17 and IFN-γ-producing NK cells, and an increased proportion of regulatory T cells in the CNS and spleen. Importantly, both MDSCs and MDSC-PGE2 prevented EAE-induced reduction of gut microbiota diversity, but only MDSC-PGE2 prevented the extensive alterations in gut microbiota composition following their early migration into Payer’s patches and mesenteric lymph nodes. This phenomenon was related to the significant enrichment of gut microbial taxa with potential immunoregulatory properties, as well as higher levels of butyrate, propionate, and putrescine in feces. This study provides new insights into the host–microbiota interactions in EAE, suggesting that activated MDSCs could be potentially used as an efficient therapy for acute phases of MS. Considering a significant association between the efficacy of MDSC-PGE2 and gut microbiota composition, our findings also provide a rationale for further exploring the specific microbial metabolites in MS therapy.

This study aimed to explore the probiogenomic characteristics of artisanal bacteriocin-producing Enterococcus faecium BGZLM1-5 and its potential application in reducing Listeria monocytogenes in a milk model. The BGZLM1-5 strain was isolated from raw cow’s milk from households in the Zlatar Mountain region. The whole genome sequencing approach and bioinformatics analyses reveal that the strain BGZLM1-5 is non-pathogenic to humans. Bacteriocin-containing supernatant was thermally stable and antimicrobial activity retained 75% of the initial activity compared with that of the control after treatment at 90 °C for 30 min. Antimicrobial activity maintained relative stability at pH 3–11 and retained 62.5% of the initial activity compared with that of the control after treatment at pH 1, 2, and 12. The highest activity of the partially purified bacteriocin was obtained after precipitation at 40% saturation with ammonium sulfate and further purification by mixing with chloroform. Applying 3% and 5% (v/v) of the bacteriocin-containing supernatant and 0.5% (v/v) of the partially purified bacteriocin decreased the viable number of L. monocytogenes ATCC19111 after three days of milk storage by 23.5%, 63.5%, and 58.9%, respectively.

This study aimed to investigate the probiogenomic features of artisanal bacteriocin-producing Enterococcus faecium BGPAS1-3 and the use of the improved pMALc5HisEk expression vector for overexpressing class II bacteriocins and the application of purified bacteriocin 31 in a milk model as a preservative against L. monocytogenes. The BGPAS1-3 strain was isolated from traditional fresh soft cheese manufactured in households on a small scale in rural locations surrounding Pale Mountain City in Bosnia and Herzegovina. The whole-genome sequencing approach and bioinformatics analyses revealed that the strain BGPAS1-3 was non-pathogenic to humans. The presence of bacteriocin operons suggested the ability of the isolate to suppress the growth of pathogens. Coding regions for three maturated bacteriocins (bacteriocin 31, bacteriocin 32, and enterocin P) produced by BGPAS1-3 were amplified and expressed in Escherichia coli ER2523 using the pMALc5HisEk system. All three bacteriocins were successfully overexpressed and purified after enterokinase cleavage but showed different antimicrobial activity. Bacteriocin 31 showed significantly stronger antimicrobial activity compared with bacteriocin 32. It was the only one that proved to be suitable for use as a food preservative against L. monocytogenes in a milk model.

Yogurt represent one of the oldest fermented foods containing viable lactic acid bacteria and many bioactive compounds that could exhibit beneficial effects on human health and train our immune system to better respond to invading pathogens. Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus are commonly used for yogurt preparation under controlled temperature and environmental conditions. In this study, we investigated probiotic features of S. thermophilus BGKMJ1-36 and L. bulgaricus BGVLJ1-21 strains isolated from artisanal sour milk and yogurt by using Caenorhabditis elegans as an in vivo model system. Further, we evaluated content of total fat, saturated fatty acids, proteins, and lactose, as well as vitamins and AA of yogurt prepared from above-mentioned starter cultures during 21 d of storage at 4°C to get insights of final product stability. We showed that S. thermophilus BGKMJ1-36 and L. bulgaricus BGVLJ1-21 strains applied in combination upregulated the expression of autophagy-related genes in C. elegans. Beside autophagy, we observed activation of TIR-1-dependent transcription of lysozyme-like antimicrobial genes involved in the immune defense of C. elegans. Upregulation of these genes strongly correlates with an increase in the longevity of the worms fed with yogurt culture bacteria. Further, we showed that yogurt prepared with S. thermophilus BGKMJ1-36 and L. bulgaricus BGVLJ1-21, as a final product, is rich with vitamin B2 and dominant AA known by their prolongevity properties. Taken together, our study pointed to the beneficial features of the tested starter cultures and yogurt and highlighted their potential to be used as a fermented food with added-value properties.