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There is growing body of evidences that the gut microbiota composition is associated with host health making the human gut a unique environment to explore commensal microorganisms with probiotic properties. The emerging subgroup of probiotics named Next-Generation Probiotics represent a novel group of gut bacteria which are strict anaerobes, whose safety status is still not fully determined, but could confer the specific healthy effect on the host. For several microbial strains that are part of the gut ecosystem have been shown to improve behaviour-related functions in the host. Therefore, the aim of this study was to evaluate the neuromodulating properties of gut anaerobic bacteria isolated from healthy individuals in Caenorhabditis elegans model system through assessment of the expression of the genes important for proper neural function of the worms. Gut anaerobic bacteria including Dorea longicatena NGB204, Dorea formicigenerans NGB229, Bacteroides xylanisolvens NGB220, Bacteroides cellulosilyticus NGB202, Roseburia intestinalis NGB215 and Lancefieldella pravula NGB231 were isolated from human fecal material in BACTEC media followed by serial dilutions spreading on Columbia Blood Agar supplemented with cysteine and sodium thioglycolate in Whitley Anaerobic Workstation. All of tested gut anaerobes showed capability to differently regulate the expression of the genes involved in serotonin, dopamine and GABA synthesis (tph-1, cat-2, unc-25), neurotransmitters’ vesicle release (unc-64, snb-1, snt-1), neuropeptide Y receptor homolog synthesis (npr-1), and different classes of neuropeptide Y homologs (flp-18, flp-21) in C. elegans in comparison to worms fed with standard laboratory food Escherichia coli OP50. These results imply that selected bacterial species could be considered as potential novel Neurobiotics, the Next-Generation Probiotics with capability to target neural function and modulate different neurodegenerative and psychiatric disorders.

Psychobiotics are live bacterial strains impacting the central nervous system, producing neuroactive substances like GABA. GABA from microbiota influences neural signals, affecting neurological parameters, sleep, appetite, mood, and cognition, traversing the intestinal barrier to bind to receptors on enteric neurons and the vagus nerve. Lactobacillus and Bifidobacterium species can synthesize GABA from dietary glutamate, with Lactobacillus rhamnosus shown to reduce anxiety and depressive behavior, elevating hippocampal GABA. Limited knowledge exists about anaerobic GABA producers, warranting further research for a comprehensive understanding. Material for isolation comprised fecal samples from healthy donors, with isolation conducted in an anaerobic chamber within a maximum of 1 hour after sampling. Isolated bacteria were identified through sequencing the 16S rRNA gene. For bacterial cultivation, different types of media were used. PYG medium contains hematine and vitamin K, essential supplements for the cultivation of anaerobic bacteria. All media included 0.1% L-cysteine, playing a role in oxygen reduction, and 0.5% glutamate, a precursor for GABA production. After identification, the presence of GABA in 8 tested bacterial species was determined using the TLC method. Quantification of GABA was performed using the HPLC method. Furthermore, the positive effects observed in Caco2 cells with induced inflammation, after treatment with certain anaerobic postbiotics producing GABA, indicate the potential anti-inflammatory effects of these postbiotics. The study implies anti-inflammatory effects of anaerobic GABA producers, offering insights into the complex interplay among gut microbiota, immune function, and mental health. Recognizing inflammation’s role in depressive symptoms, targeting anaerobic bacteria involved in GABA synthesis could modulate neurotransmitters and inflammatory responses, presenting a comprehensive approach to mental well-being. Advancing research i

Introduction: Gut-brain axis has been identified as an important target for prevention of neurodegenerative and psychiatric disease. To date, specific microbial strains resident in the intestinal ecosystem have been described to modulate several behaviour-related functions in the host. Faecalimonas sp. is anaerobic bacteria affiliated with the family Lachnospiraceae, which represents a highly prevalent beneficial bacteria in the human gut and have potential to be used as next generation probiotic. Methods: Faecalimonas sp. NGB245 was isolated from human fecal material by pre-inoculation in BACTEC media followed by serial dilutions spreading on Columbia Blood Agar supplemented with cysteine and sodium thioglycolate in Whitley Anaerobic Workstation. Production of short chain fatty acid (SCFA) was detected after bacterial growth in Columbia broth supplemented with cellobiose by HPLC. Host response was followed on Caenorhabditis elegans model by evaluated expression of the genes involved in neurosignaling by qPCR. Results: We showed that Faecalimonassp. NGB245 exhibits high capacity of production of SCFA including acetate (12,17 mM), propionate (3,02 mM) and butyrate (10,33 mM). Moreover, C. elegansfed with Faecalimonas sp. NGB245 showed higher expression of the genes involved in neurotransmitter synthesis (tph-1, cat-2), neurotransmitter release (unc-64, snb-1, snt-1), neurotransmitter receptor (npr-1) and different classes of neuropeptides(flp-18, flp-21, nlp-28, nlp-29) in comparison to wormsfed with Escherichia coli OP50, as a standard laboratory food. Conclusion: The obtained results imply that Faecalimonas sp. NGB245 isolate could be considered as next generation probiotic to be used in prevention and treatment of neurodegenerative and psychiatric diseases.

Introduction: Hydrogen cyanide (HCN) is a volatile secondary metabolite synthesized by some bacteria, and this ability enablestheir activity against various pathogens. The aim of thisstudy wasto identify HCN-producing bacteria and investigate their biocontrol potential. Methods: Three HCN-producing strains were detected in a collection of bell pepper plant isolates using a semi-quantitative assay with picric acid. The presence of hcnABC operon genes was confirmed by PCR. The biological control potential of the HCN-producing strains wastested against three fungal (Fusarium oxysporum, Rhizoctonia solani, Verticillium dahliae) and eight bacterial (genera Xanthomonas, Pseudomonas and Clavibacter) pathogens of bell pepper plants in a split-section Petri dish experiment. The potential nematocidal activity was demonstrated by using the Caenorhabditis elegans AU37 strain, with temperature-sensitive sterility and enhanced sensitivity to pathogens. Results: Detailed characterization of 300 isolates from our collection revealed that we have three different HCN-producing strains identified as Bacillussubtilis, Pseudomonas moraviensis, and P. putida, with P. putida A32 being the most potent. Thisstrain is used for the deletion of the hcnB gene to confirm HCN as a biocontrol agent. Conclusion: The HCN-producing strains showed biocontrol potential against bacteria, fungi, and nematodes. It is concluded that the biological control activity isthe result of a volatile metabolite diffusing through the air. Our future exp

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.

Introduction: New theories of ageing recognized gut microbiota as one of twelve hallmarks of ageing (1). Recent data conducted on Caenorhabditis elegans imply a potential role of Lactobacillus species and other commensal bacteria in regulation of ageing highlighting TFEB/HLH-30-dependent autophagy, p38 MAPK/PMK-1 signalling and mitochondrial function as activated longevity-associated mechanisms (2,3). Here, we explore the potential of bacterial polysaccharides loosely attached to bacterial cell wall (exopolysaccharides), considering it is still unknown which bacterial molecules could activate longevity signalling pathways. Materials & Methods: Caenorhabditis elegans was used as ageing model. Evaluation of worm’s lifespan and locomotion rate were performed by feeding worms with six exopolysaccharide-producing lactobacilli. Worms fed with two selected strains were subjected to RNAseq analysis. Identified upregulated genes were confirmed by qPCR and expression of their mammalian orthologs checked in human HepG2 cell. Results: Two strains of lactobacilli showed the most pronounced effect on worms’ lifespan. RNAseq analysis identified core gene signature associate with exopolysaccharide-induced longevity. The expression of identified fmo-2, gsto-1, nlp-29, and clec-47 genes were confirmed by qPCR, while upregulation of FMO-5 was confirmed in HepG2 cells. Conclusion: Overall, our results imply that bacteria-derived exopolysaccharides could stimulate longevity- promoting flavin-containing monooxygenase 2 to regulate lifespan in Caenorhabditis elegans

Introduction: Microbial community established in the gut has been recognized as an important factor which influence host aging. Bacteria from the gut co-evolved with the host resulting in mutually beneficial interactions essential for host’s wellbeing. This complex crosstalk reflects mainly through the interaction between bacterial macromolecules (e.g., exopolysaccharides) and the host receptors leading to the activation of various cellular pathways. Here, we explore the potential of different lactobacilli, commonly used as probiotics, to activate longevity signalling in Caenorhabditis elegans. Methods: Evaluation of C. elegans lifespan and aging parameters (locomotion rate and pharyngeal pumping) were performed by feeding N2 wild-type worms with different Lactobacillus species. Worms fed with selected strains were subjected to RNAseq analysis, qPCR and Western blot to evaluate activation of autophagy, immunity, antioxidative response and mitochondrial function. Activation of autophagy was confirmed in DA2123 GFP-labelled LGG-1 transgenic strain and JIN1375 hlh-30 (tm1978) mutant, while immunity activation was evaluated by using KU25 pmk-1 (km25) mutant and through nematode killing assays. Results: Selected strains of lactobacilli promoted health and lifespan of worms through activation of TFEB/HLH-30 dependent autophagy and p38 MAPK/PMK-1 dependent immune response which provided resistance of worms exposed to pathogens. Moreover, RNAseq analysis identified core gene signature associate with exopolysaccharide-induced longevity highlighting involvement of fmo-2, gsto-1, nlp-29, and clec-47 genes in increased lifespan of the worms. Conclusion: Analyzed lactobacilli showed potential to promote healthy aging and could be further investigated in order to better understand application of lactobacilli as pro-longevity probiotics.