Neurotransmitter Profiles Are Altered in the Gut and Brain of Mice Mono-Associated with Bifidobacterium dentium.

BACKGROUND: Accumulating evidence indicates that the gut microbiota can synthesize neurotransmitters as well as impact host-derived neurotransmitter levels. In the past, it has been challenging to decipher which microbes influence neurotransmitters due to the complexity of the gut microbiota. METHODS: To address whether a single microbe, Bifidobacterium dentium, could regulate important neurotransmitters, we examined Bifidobacteria genomes and explored neurotransmitter pathways in secreted cell-free supernatant using LC-MS/MS. To determine if B. dentium could impact neurotransmitters in vivo, we mono-associated germ-free mice with B. dentium ATCC 27678 and examined fecal and brain neurotransmitter concentrations. RESULTS: We found that B. dentium possessed the enzymatic machinery to generate γ-aminobutyric acid (GABA) from glutamate, glutamine, and succinate. Consistent with the genome analysis, we found that B. dentium secreted GABA in a fully defined microbial media and elevated fecal GABA in B. dentium mono-associated mice compared to germ-free controls. We also examined the tyrosine/dopamine pathway and found that B. dentium could synthesize tyrosine, but could not generate L-dopa, dopamine, norepinephrine, or epinephrine. In vivo, we found that B. dentium mono-associated mice had elevated levels of tyrosine in the feces and brain. CONCLUSIONS: These data indicate that B. dentium can contribute to in vivo neurotransmitter regulation.

FN3 protein fragment containing two type III fibronectin domains from B. longum GT15 binds to human tumor necrosis factor alpha in vitro.

Most species of the genus Bifidobacterium contain the gene cluster PFNA, which is presumably involved in the species-specific communication between bacteria and their hosts. The gene cluster PFNA consists of five genes including fn3, which codes for a protein containing two fibronectin type III domains. Each fibronectin domain contains sites similar to cytokine-binding sites of human receptors. Based on this finding we assumed that this protein would bind specifically to human cytokines in vitro. We cloned a fragment of the fn3 gene (1503 bp; 501 aa) containing two fibronectin domains, from the strain B. longum subsp. longum GT15. After cloning the fragment into the expression vector pET16b and expressing it in E. coli, the protein product was purified to a homogenous state for further analysis. Using the immunoferment method, we tested the purified fragment's ability to bind the following human cytokines: IL-1ß, IL-6, IL-10, TNFα. We developed a sandwich ELISA system to detect any specific interactions between the purified protein and any of the studied cytokines. We found that the purified protein fragment only binds to TNFα.

The impact of metabolites derived from the gut microbiota on immune regulation and diseases.

The gut microbiota strongly impacts the physiology and pathology in the host. To understand the complex interactions between host and gut microbiota, an 'integrated omics' approach has been employed, where exhaustive analyses for the different layers of cellular functions, such as epigenomics, transcriptomics and metabolomics, in addition to metagenomics, are combined. With this approach, the mechanisms whereby short-chain fatty acids (SCFAs) regulate host defense and the immune system have been elucidated. In a gnotobiotic mouse model of enterohemorrhagic Escherichia coli infection, Bifidobacterium-derived acetate can protect from infection-mediated death by changing the gene expression profile of colonic epithelial cells. It has also been shown that gut microbiota-derived butyrate enhances colonic regulatory T-cell differentiation through its epigenetic modulatory ability via histone deacetylase inhibition. SCFAs are involved in many other immunomodulatory effects as well as host pathophysiological conditions. Dysbiosis in the gut has been implicated in the pathogenesis of many diseases. Although the causal relationship of gut microbial dysbiosis and/or metabolites with pathogenesis is mostly unknown, mechanistic insights have been elucidated in some cases. Metabolism in the gut microbiota and host liver produces trimethylamine N-oxide, which is known to aggravate atherosclerosis, and a secondary bile acid deoxycholate, which reportedly induces non-alcoholic steatohepatitis-related hepatocellular carcinoma. It has been reported that secondary bile acids could also induce the differentiation of peripherally derived regulatory T cells in the gut. Further studies on the interactions between the host and gut microbiota could lead to the development of new therapeutic strategies as well as in preventive medicine.

Bifidobacterial Dialogue With Its Human Host and Consequent Modulation of the Immune System.

Since bifidobacteria are among the pioneering colonizers of the human infant gut, their interaction with their host is believed to start soon following birth. Several members of the Bifidobacterium genus are purported to exert various health-promoting effects at local and systemic levels, e.g., limiting pathogen colonization/invasion, influencing gut homeostasis, and influencing the immune system through changes in innate and/or adaptive immune responses. This has promoted extensive research efforts to shed light on the precise mechanisms by which bifidobacteria are able to stimulate and interact with the host immune system. These studies uncovered a variety of secreted or surface-associated molecules that act as essential mediators for the establishment of a bifidobacteria-host immune system dialogue, and that allow interactions with mucosa-associated immune cells. Additionally, the by-products generated from bifidobacterial carbohydrate metabolism act as vectors that directly and indirectly trigger the host immune response, the latter by stimulating growth of other commensal microorganisms such as propionate- or butyrate-producing bacteria. This review is aimed to provide a comprehensive overview on the wide variety of strategies employed by bifidobacteria to engage with the host immune system.

Sharing of human milk oligosaccharides degradants within bifidobacterial communities in faecal cultures supplemented with Bifidobacterium bifidum.

Gut microbiota of breast-fed infants are generally rich in bifidobacteria. Recent studies show that infant gut-associated bifidobacteria can assimilate human milk oligosaccharides (HMOs) specifically among the gut microbes. Nonetheless, little is known about how bifidobacterial-rich communities are shaped in the gut. Interestingly, HMOs assimilation ability is not related to the dominance of each species. Bifidobacterium longum susbp. longum and Bifidobacterium breve are commonly found as the dominant species in infant stools; however, they show limited HMOs assimilation ability in vitro. In contrast, avid in vitro HMOs consumers, Bifidobacterium bifidum and Bifidobacterium longum subsp. infantis, are less abundant in infant stools. In this study, we observed altruistic behaviour by B. bifidum when incubated in HMOs-containing faecal cultures. Four B. bifidum strains, all of which contained complete sets of HMO-degrading genes, commonly left HMOs degradants unconsumed during in vitro growth. These strains stimulated the growth of other Bifidobacterium species when added to faecal cultures supplemented with HMOs, thereby increasing the prevalence of bifidobacteria in faecal communities. Enhanced HMOs consumption by B. bifidum-supplemented cultures was also observed. We also determined the complete genome sequences of B. bifidum strains JCM7004 and TMC3115. Our results suggest B. bifidum-mediated cross-feeding of HMOs degradants within bifidobacterial communities.

In vitro evaluation of the prebiotic effect of red and white grape polyphenolic extracts

Nowadays, the strong relationship between diet and health is well known. Although the primary role of diet is to provide nutrients to fulfill metabolic requirements, the use of foods to improve health and the state of well-being is an idea increasingly accepted by society in the last three decades. During the last years, an important number of scientific advances have been achieved in this field and, although in some situations, it is difficult to establish a distinction between "harmful" and "good" bacteria, experts agree in classifying the genera Bifidobacterium and Lactobacillus as beneficial bacteria. Thus, several strategies can be used to stimulate the proliferation of these beneficial intestinal bacteria, being one of them the consumption of prebiotics. The development of new prebiotics, with added functionality, is one of the most serious challenges shared not only by the scientific community but also by the food industry. The objective of this work was to evaluate the potential prebiotic effect of red and white grape residues, both obtained during the winemaking process. For such purpose, an in vitro study with pure cultures of Lactobacillus and Bifidobacterium was first conducted. Secondly, a study with mixed cultures using human fecal inocula was carried out in a simulator of the distal part of the colon. The obtained results showed an increase in the Lactobacillus and Bifidobacterium population, indicating that these ingredients are serious candidates to be considered as prebiotics