Bioactive metabolites in functional and fermented foods and their role as immunity booster and anti-viral innate mechanisms.

Live microorganisms in the fermented foods termed probiotics and their secondary metabolites with bioactive potential were considered as potential anti-viral capabilities through various mechanisms. Given the importance of functional and fermented foods in disease prevention, there is a need to discuss the contextualization and deep understanding of the mechanism of action of these foods, particularly considering the appearance of coronavirus (COVID-19) pandemic, which is causing health concerns and increased social services globally. The mechanism of probiotic strains or their bioactive metabolites is due to stimulation of immune response through boosting T-lymphocytes, cytokines, and cell toxicity of natural killer cells. Proper consumption of these functional and fermented foods may provide additional antiviral approaches for public benefit by modulating the immune functions in the hosts. Supplementary Information: The online version contains supplementary material available at 10.1007/s13197-022-05528-8.

Lactococcus garvieae subsp. bovis subsp. nov., lactic acid bacteria isolated from wild gaur (Bos gaurus) dung, and description of Lactococcus garvieae subsp. garvieae subsp. nov.

A taxonomic investigation was performed on a Gram-stain-positive coccus, designated strain BSN307T, isolated from gaur (Indian bison, Bos gaurus) dung based on phenotypic and molecular approaches. Based on the biochemical tests, cellular morphology and 16S rRNA gene sequence similarity, this strain was found to be a member of the genus Lactococcus and closely related to Lactococcus garvieae ATCC 49156T (99.6 % 16S rRNA gene sequence similarity) and L.actococcus formosensis 516T (99.0 %). However, DNA-DNA hybridization studies showed that the level of relatedness between strain BSN307T and L. garvieae ATCC 49156T was 75.8 %, suggesting that it represented a novel subspecies of L. garvieae. The inability to grow in brain heart infusion (BHI) medium at pH 9.6, in tryptic soy agar (TSA) with 4 % (w/v) NaCl and at 42 °C (MRS agar) clearly differentiated BSN307T from L. garvieae ATCC 49156T. Rep-PCR fingerprint patterns, substantial differences in summed feature 8 (C18 : 1ω7c/C18 : 1ω6c), C19 : 0 cyclo ω8c and C16 : 0 also differentiated strain BSN307T from the reference strain of L. garvieae. Moreover, analysis of the housekeeping genes pheS and rpoA revealed sequence similarities that were at the limit for species differentiation (92.2 and 97.8 %, respectively). Combined genotypic and phenotypic data indicate that strain BSN307T represents a subspecies of L. garvieae for which the name Lactococcus garvieae subsp. bovis subsp. nov. is proposed. The type strain is BSN307T (=DSM 100577 T=MCC 2824T=KCTC 21083T).

Antifungal, Anticancer and Aminopeptidase Inhibitory Potential of a Phenazine Compound Produced by Lactococcus BSN307.

A bioactive compound was purified from the culture medium of a new strain of Lactococcus BSN307 by solvent extraction followed by chromatographic techniques. This bioactive compound was identified to belong to phenazine class of compounds by MS, NMR and FTIR. The phenazine compound showed antifungal activity against Aspergillus niger, Penicillium chrysogenum as well as Fusarium oxysporum by disc diffusion assay in addition to antioxidant potential as demonstrated by DPPH scavenging assay. The compound demonstrated selective cytotoxicity against cancer cell lines HeLa and MCF-7 where IC50 was achieved with 20 and 24 µg/mL respectively. At the same time no cytotoxicity was occurred in normal H9c2 cells. The bioactive found to be inhibitory to both leucine and proline aminopeptidases and thus revealed its potential as metalloenzyme inhibitor. This study, for the first time reports the production of phenazine class of compounds by lactic acid bacteria.

Identification of miRNAs that target Fcγ receptor-mediated phagocytosis during macrophage activation syndrome.

Macrophage activation syndrome (MAS) is a life-threatening complication of systemic juvenile arthritis, accompanied by cytokine storm and hemophagocytosis. In addition, COVID-19-related hyperinflammation shares clinical features of MAS. Mechanisms that activate macrophages in MAS remain unclear. Here, we identify the role of miRNA in increased phagocytosis and interleukin-12 (IL-12) production by macrophages in a murine model of MAS. MAS significantly increased F4/80+ macrophages and phagocytosis in the mouse liver. Gene expression profile revealed the induction of Fcγ receptor-mediated phagocytosis (FGRP) and IL-12 production in the liver. Phagocytosis pathways such as High-affinity IgE receptor is known as Fc epsilon RI -signaling and pattern recognition receptors involved in the recognition of bacteria and viruses and phagosome formation were also significantly upregulated. In MAS, miR-136-5p and miR-501-3p targeted and caused increased expression of Fcgr3, Fcgr4, and Fcgr1 genes in FGRP pathway and consequent increase in phagocytosis by macrophages, whereas miR-129-1-3p and miR-150-3p targeted and induced Il-12. Transcriptome analysis of patients with MAS revealed the upregulation of FGRP and FCGR gene expression. A target analysis of gene expression data from a patient with MAS discovered that miR-136-5p targets FCGR2A and FCGR3A/3B, the human orthologs of mouse Fcgr3 and Fcgr4, and miR-501-3p targets FCGR1A, the human ortholog of mouse Fcgr1. Together, we demonstrate the novel role of miRNAs during MAS pathogenesis, thereby suggesting miRNA mimic-based therapy to control the hyperactivation of macrophages in patients with MAS as well as use overexpression of FCGR genes as a marker for MAS classification.

Role of Gut Microbiota in Cannabinoid-Mediated Suppression of Inflammation.

Cannabinoids and the endocannabinoid system have been well established to play a crucial role in the regulation of the immune response. Also, emerging data from numerous investigations unravel the imperative role of gut microbiota and their metabolites in the maintenance of immune homeostasis and gut barrier integrity. In this review, we concisely report the immunosuppressive mechanisms triggered by cannabinoids, and how they are closely associated with the alterations in the gut microbiome and metabolome following exposure to endogenous or exogenous cannabinoids. We discuss how cannabinoid-mediated induction of microbial secondary bile acids, short chain fatty acids, and indole metabolites, produced in the gut, can suppress inflammation even in distal organs. While clearly, more clinical studies are necessary to establish the cross talk between exo- or endocannabinoid system with the gut microbiome and the immune system, the current evidence opens a new avenue of cannabinoid-gut-microbiota-based therapeutics to regulate immunological disorders.

Accomplishment of probiotics in human health pertaining to immunoregulation and disease control.

It is a well-established fact that the microbiome harboring the human body plays a critical role in maintaining human health and can influence treatments against various ailments. Human microbiome-based research contemplates the possibility of selecting and administering specific commensal bacterial strains to modulate the gut microbiota to attain favorable outcomes to the therapies. Consumption of probiotics and probiotic-based dietary supplements as functional foods has been a promising treatment strategy against various diseases. Clinical studies demonstrate that probiotic administration alters gut microbiota composition and instigates immune modulation in the host. The benefits of probiotics are reported to be strain-specific and depend on the host's baseline immune competence. This review explores the role of probiotics in alleviating symptoms of allergy, cancer, cardio vascular (CV) diseases, diabetes mellitus (DM), bowel diseases (IBD and IBS), periodontal disease, diseases affecting liver and kidney, neuroinflammatory diseases, and viral infections. Also, it surveyed the broad spectrum bioactive compounds produced by probiotics and possible mechanisms that trigger the immune system.

Acute toxicity analysis of an inhibitor of BCL2, Disarib, in rats.

Apoptosis or programmed cell death is a highly regulated process, which eliminates unwanted and damaged cells. Inhibition of apoptosis is a hallmark of cancer cells. BCL2 family proteins are known to play a vital role in the regulation of apoptosis. Overexpression of BCL2, an antiapoptotic protein, provides the advantage of prolonged survival to cancer cells. Over the years, several BCL2 inhibitors have been investigated extensively for their anticancer potential. However, most of them were abolished before clinical use due to their side effects. Previously, we had identified and characterized a novel BCL2 inhibitor, Disarib, with the potential to eliminate tumor cells in a BCL2 specific manner leading to reduction in tumor burden in multiple mouse models. Notably, a head-to-head comparison of Disarib to ABT199, the only FDA approved BCL2 inhibitor revealed that Disarib is as potent as ABT199. Recent studies using mice revealed that Disarib did not invoke significant side effects in mice. In the present study, we have investigated the acute toxicity of Disarib in Wistar rats. The bioavailability studies following exposure of Disarib in Wistar rats revealed its maximum availability in serum at 24 h following oral administration. Acute toxicity analysis revealed that even a dose as high as 2000 mg/kg of Disarib did not cause significant toxicity in rats. There was no significant variation in blood parameters or kidney and liver functions following administration of Disarib. Histological analysis of different tissues from Disarib treated groups revealed standard architecture with no observable cellular damage. Importantly, exposure to Diasrib did not result in genotoxicity as determined by micronucleus assay. Further, solubility assays revealed that besides DMSO, Disarib is also soluble in alcohol. While the high acidic condition can increase the solubility of Disarib, even a lower percentage of alcohol with acidic conditions can improve its solubility. Thus, the toxicological profile in the current study revealed no significant side effects when Disarib was administered orally to rats.

Acute toxicity analysis of Disarib, an inhibitor of BCL2.

Small molecule inhibitors targeting BCL2 are explored as anticancer therapeutics. Previously, we have reported identification and characterization of a novel BCL2 inhibitor, Disarib. Disarib induced cancer cell death in a BCL2 dependent manner in different cancer cell lines and mouse tumor models when it was administered intraperitoneally. In the present study, using two syngeneic mouse models, breast adenocarcinoma (EAC) and Dalton's lymphoma (DLA), we show that oral administration of Disarib resulted in significant tumor regression in a concentration dependent manner. Importantly, tumor developed in both female and male mice were equally sensitive to Disarib. Further, we have investigated the toxicity of Disarib in normal cells. Single dose toxicity analysis of Disarib in male and female mice after oral administration revealed no significant variations compared to control group for parameters such as body weight, food and water consumption and behavioural changes which were analysed for the entire period of study. Haematological and histopathological analyses also did not show any significant difference from the control groups. Thus, our results reveal safe use of Disarib as a small molecule inhibitor and provide the foundation for investigation of other preclinical studies.

2,4-Di-tert-butyl phenol as the antifungal, antioxidant bioactive purified from a newly isolated Lactococcus sp.

The volatile organic compound 2,4-di-tert-butyl phenol (2,4 DTBP) was purified from the cell free supernatant of a newly isolated Lactococcus sp. by solvent extraction and chromatographic techniques. Molecular characterization of the compound by ESI-MS, (1)H NMR and FTIR analysis revealed the structure, C14H22O. Fungicidal activity was demonstrated against Aspergillus niger, Fusarium oxysporum and Penicillium chrysogenum by disc diffusion assay. Among the cell lines tested for cytotoxicity of this compound (normal cell line H9c2 and cancer cell lines HeLa and MCF-7), a remarkable cytotoxicity against HeLa cells with an IC50 value of 10 µg/mL was shown. A biocontrol experiment with 2,4 DTBP supplemented fraction prevented growth of the abovementioned fungi on wheat grains. The study further strengthens the case for development of biopreservatives and dietary antioxidants from lactic acid bacteria for food applications.

Control of spoilage fungi by protective lactic acid bacteria displaying probiotic properties.

Thirty-six lactic acid bacteria belong to Lactococcus, Lactobacillus, Enterococcus, and Pediococcus were isolated, and the spectrum of antifungal activity was verified against Fusarium oxysporum (KACC 42109), Aspergillus niger (KACC 42589), Fusarium moniliforme (KACC 08141), Penicillium chrysogenum (NII 08137), and the yeast Candida albicans (MTCC 3017). Three isolates, identified as Pediococcus pentosaceus (TG2), Lactobacillus casei (DY2), and Lactococcus (BSN) were selected further, and their antifungal compounds were identified by ESI-MS and HPLC analysis as a range of carboxylic acids along with some unidentified, higher molecular weight compounds. An attempt to check out the shelf life extension of wheat bread without fungal spoilage was performed by fermenting the dough with the Lactococcus isolate. Apart from growth in low pH and tolerance to bile salts, probiotic potential of these three isolates was further substantiated by in vitro screening methods that include transit tolerance to the conditions in the upper human gastrointestinal tract and bacterial adhesion capacity to human intestinal cell lines.

Newly isolated lactic acid bacteria with probiotic features for potential application in food industry.

Five newly isolated lactic acid bacteria were identified as Weissella cibaria, Enterococcus faecium, and three different strains of Lactobacillus plantarum by 16S rRNA sequencing. Essential probiotic requirements of these isolates such as tolerance to phenol, low pH, high sodium chloride, and bile salt concentration were checked. Efficiency in adherence to mucin and hydrophobicity of the bacterial cell were also evaluated by in vitro studies. Antimicrobial activities against some pathogens were tried, and the sensitivity of these strains against 25 different antibiotics was also checked. Further studies revealed Weissella and Enterococcus as substantial producers of folic acid. Folate is involved as a cofactor in many metabolic reactions, and it has to be an essential component in the human diet. The folate level in the fermented samples was determined by microbiological assay using Lactobacillus casei NCIM 2364 as indicator strain. The three strains of L. plantarum showed significant inhibitory activity against various fungi that commonly contaminate food stuffs indicating their potential as a biopreservative of food material.