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Integrated genome-based probiotic relevance and safety evaluation of Lactobacillus reuteri PNW1.

ABSTRACT: This study evaluates whole-genome sequence of Lactobacillus reuteri PNW1 and identifies its safety genes that may qualify it as a putative probiotic. It further extracted the bacteriocin produced by the strain and tested its effectiveness against pathogenic STEC E. coli O177. The genomic DNA was sequenced on illuminal Miseq instrument and the sequenced data was assessed for quality reads before assembled with SPAdes. The draft assembly was annotated with Prokaryotic Genome Annotation Pipeline (PGAP) and Rapid Annotations using Subsystems Technology (RAST). Further downstream analyses were carried out using appropriate bioinformatic tools. Production of biogenic amines was biochemically confirmed through HPLC analysis. The assembled genome was 2,430,215 bp long in 420 contigs with 39% G+C content. Among all known genes, putatively responsible for the production of toxic biochemicals, only arginine deiminase (EC3.5.3.6) was spotted. Coding sequences (CDS) putative for D-lactate dehydrogenase (EC1.1.1.28), L-lactate dehydrogenase (EC1.1.1.27) and bacteriocin helveticin J were found within the genome together with plethora of other probiotic important genes. The strain harbours only resistant genes putative for Lincosamide (lnuC) and Tetracycline resistant genes (tetW). There was no hit found for virulence factors and probability of the strain being a human pathogen was zero. Two intact prophage regions were detected within the genome of L. reuteri PNW1 and nine CDS were identified for insertion sequence by OASIS which are belong to seven different families. Five putative CDS were identified for the CRISPR, each associated with Cas genes. Maximum zone of inhibition exhibited by the bacteriocin produced L. reuteri PNW1 is 20.0±1.00 mm (crude) and 23.3±1.15 mm (at 0.25 mg/ml) after being partially purified. With the strain predicted as non-human pathogen, coupled with many other identified desired features, L. reuteri PNW1 stands a chance of making good and safe candidates for probiotic, though further in-vivo investigations are still necessary.

SUBMITTER: Alayande KA

PROVIDER: S-EPMC7371166 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Integrated genome-based probiotic relevance and safety evaluation of Lactobacillus reuteri PNW1.

Alayande Kazeem Adekunle KA Aiyegoro Olayinka Ayobami OA Nengwekhulu Thizwilondi Michael TM Katata-Seru Lebogang L Ateba Collins Njie CN

PloS one 20200720 7

This study evaluates whole-genome sequence of Lactobacillus reuteri PNW1 and identifies its safety genes that may qualify it as a putative probiotic. It further extracted the bacteriocin produced by the strain and tested its effectiveness against pathogenic STEC E. coli O177. The genomic DNA was sequenced on illuminal Miseq instrument and the sequenced data was assessed for quality reads before assembled with SPAdes. The draft assembly was annotated with Prokaryotic Genome Annotation Pipeline (P ...[more]

PMID: 32687505

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Project description:Probiotics and commensal intestinal microbes suppress mammalian cytokine production and intestinal inflammation in various experimental model systems. Limited information exists regarding potential mechanisms of probiotic-mediated immunomodulation in vivo. In this report, we demonstrate that specific probiotic strains of Lactobacillus reuteri suppress intestinal inflammation in a trinitrobenzene sulfonic acid (TNBS)-induced mouse colitis model. Only strains that possess the hdc gene cluster, including the histidine decarboxylase and histidine-histamine antiporter genes, can suppress colitis and mucosal cytokine (interleukin-6 [IL-6] and IL-1? in the colon) gene expression. Suppression of acute colitis in mice was documented by diminished weight loss, colonic injury, serum amyloid A (SAA) protein concentrations, and reduced uptake of [(18)F]fluorodeoxyglucose ([(18)F]FDG) in the colon by positron emission tomography (PET). The ability of probiotic L. reuteri to suppress colitis depends on the presence of a bacterial histidine decarboxylase gene(s) in the intestinal microbiome, consumption of a histidine-containing diet, and signaling via the histamine H2 receptor (H2R). Collectively, luminal conversion of l-histidine to histamine by hdc(+) L. reuteri activates H2R, and H2R signaling results in suppression of acute inflammation within the mouse colon.Probiotics are microorganisms that when administered in adequate amounts confer beneficial effects on the host. Supplementation with probiotic strains was shown to suppress intestinal inflammation in patients with inflammatory bowel disease and in rodent colitis models. However, the mechanisms of probiosis are not clear. Our current studies suggest that supplementation with hdc(+) L. reuteri, which can convert l-histidine to histamine in the gut, resulted in suppression of colonic inflammation. These findings link luminal conversion of dietary components (amino acid metabolism) by gut microbes and probiotic-mediated suppression of colonic inflammation. The effective combination of diet, gut bacteria, and host receptor-mediated signaling may result in opportunities for therapeutic microbiology and provide clues for discovery and development of next-generation probiotics.

Project description:Lactobacillus reuteri (L. reuteri) is a probiotic that inhibits the severity of enteric infections and modulates the immune system. Human-derived L. reuteri strains DSM17938, ATCC PTA4659, ATCC PTA 5289, and ATCC PTA 6475 have demonstrated strain-specific immunomodulation in cultured monocytoid cells, but information about how these strains affect inflammation in intestinal epithelium is limited. We determined the effects of the four different L. reuteri strains on lipopolysaccharide (LPS)-induced inflammation in small intestinal epithelial cells and in the ileum of newborn rats. IPEC-J2 cells (derived from the jejunal epithelium of a neonatal piglet) and IEC-6 cells (derived from the rat crypt) were treated with L. reuteri. Newborn rat pups were gavaged cow milk formula supplemented with L. reuteri strains in the presence or absence of LPS. Protein and mRNA levels of cytokines and histological changes were measured. We demonstrate that even though one L. reuteri strain (DSM 17938) did not inhibit LPS-induced IL-8 production in cultured intestinal cells, all strains significantly reduced intestinal mucosal levels of KC/GRO (∼IL-8) and IFN-γ when newborn rat pups were fed formula containing LPS ± L. reuteri. Intestinal histological damage produced by LPS plus cow milk formula was also significantly reduced by all four strains. Cow milk formula feeding (without LPS) produced mild gut inflammation, evidenced by elevated mucosal IFN-γ and IL-13 levels, a process that could be suppressed by strain 17938. Other cytokines and chemokines were variably affected by the different strains, and there was no toxic effect of L. reuteri on intestinal cells or mucosa. In conclusion, L. reuteri strains differentially modulate LPS-induced inflammation. Probiotic interactions with both epithelial and nonepithelial cells in vivo must be instrumental in modulating intrinsic anti-inflammatory effects in the intestine. We suggest that the terms anti- and proinflammatory be used only to describe the effects of a probiotic in the living host.

Project description:The comparative genomic analysis of Lactiplantibacillus plantarum YW11 (L. plantarum YW11) isolated from Tibetan kefir involves comparison of the complete genome sequences of the isolated strain with other closely related L. plantarum strains. This type of analysis can be used to identify the genetic diversity among strains and to explore the genetic characteristics of the YW11 strain. The genome of L. plantarum YW11 was found to be composed of a circular single chromosome of 4,597,470 bp with a G + C content of 43.2%. A total of 4,278 open reading frames (ORFs) were identified in the genome and the coding density was found to be 87.8%. A comparative genomic analysis was conducted using two other L. plantarum strains, L. plantarum C11 and L. plantarum LMG21703. Genomic comparison revealed that L. plantarum YW11 shared 72.7 and 75.2% of gene content with L. plantarum C11 and L. plantarum LMG21703, respectively. Most of the genes shared between the three L. plantarum strains were involved in carbohydrate metabolism, energy production and conversion, amino acid metabolism, and transcription. In this analysis, 10 previously sequenced entire genomes of the species were compared using an in-silico technique to discover genomic divergence in genes linked with carbohydrate intake and their potential adaptations to distinct human intestinal environments. The subspecies pan-genome was open, which correlated with its extraordinary capacity to colonize several environments. Phylogenetic analysis revealed that the novel genomes were homogenously grouped among subspecies of l Lactiplantibacillus. L. plantarum was resistant to cefoxitin, erythromycin, and metronidazole, inhibited pathogens including Listeria monocytogenes, Clostridium difficile, Vibrio cholera, and others, and had excellent aerotolerance, which is useful for industrial operations. The comparative genomic analysis of L. plantarum YW11 isolated from Tibetan kefir can provide insights into the genetic characteristics of the strain, which can be used to further understand its role in the production of kefir.

Dataset Information

Integrated genome-based probiotic relevance and safety evaluation of Lactobacillus reuteri PNW1.

Publications

Integrated genome-based probiotic relevance and safety evaluation of Lactobacillus reuteri PNW1.

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