Bifidobacterium Bifidum PRL2010 Research Articles

GH136-encoding gene (perB) is involved in gut colonization and persistence by Bifidobacterium bifidum PRL2010.

Bifidobacteria are commensal microorganisms that typically inhabit the mammalian gut, including that of humans. As they may be vertically transmitted, they commonly colonize the human intestine from the very first day following birth and may persist until adulthood and old age, although generally at a reduced relative abundance and prevalence compared to infancy. The ability of bifidobacteria to persist in the human intestinal environment has been attributed to genes involved in adhesion to epithelial cells and the encoding of complex carbohydrate-degrading enzymes. Recently, a putative mucin-degrading glycosyl hydrolase belonging to the GH136 family and encoded by the perB gene has been implicated in gut persistence of certain bifidobacterial strains. In the current study, to better characterize the function of this gene, a comparative genomic analysis was performed, revealing the presence of perB homologues in just eight bifidobacterial species known to colonize the human gut, including Bifidobacterium bifidum and Bifidobacterium longum subsp. longum strains, or in non-human primates. Mucin-mediated growth and adhesion to human intestinal cells, in addition to a rodent model colonization assay, were performed using B. bifidum PRL2010 as a perB prototype and its isogenic perB-insertion mutant. These results demonstrate that perB inactivation reduces the ability of B. bifidum PRL2010 to grow on and adhere to mucin, as well as to persist in the rodent gut niche. These results corroborate the notion that the perB gene is one of the genetic determinants involved in the persistence of B. bifidum PRL2010 in the human gut.

Bifidobacterium bifidum PRL2010 alleviates intestinal ischemia/reperfusion injury.

Mesenteric ischemia/reperfusion is a clinical emergency with high morbidity and mortality due to the transient reduction of blood supply to the bowel. In recent years, the critical contribution of gut microbiome to human health and proper gastrointestinal functions has gradually emerged. In the current study, we investigated the protective effects of five days supplementation with Bifidobacterium bifidum PRL2010 in a murine model of gut ischemia/reperfusion. Our findings indicate that animals pretreated with B. bifidum PRL2010 showed lower neutrophil recruitment in the lungs, remarkably reduced bacterial translocation and decreased transcription levels of TNFalpha and IL-10 both in liver and kidneys, at the same time increasing those of IL-12 in kidneys. Inhibiting the adhesion of pathogenic bacteria and boosting host innate immunity responses are among the possible protective mechanisms enacted by the probiotic. These results demonstrate that short-period treatment with B. bifidum PRL2010 is a potential strategy to dampen remote organ injury due to mesenteric ischemia/reperfusion.

The Role of Bifidobacteria in Ulcerative Colitis: Preliminary Results

Introduction: Ulcerative colitis (UC) has a multifactorial pathogenesis, including genetic susceptibility, environmental triggers, gut microbiota and microbial imbalance due to its immunomodulatory function. Previous studies showed that Bifidobacterium bifidum PRL2010 cells can effectively activate signals in macrophages, modulate metabolic activities and influence immune responses in vitro and when tested in a murine colitis model. The aim of this study was to characterize the microbiome of UC patients compared to healthy controls (HS), in order to evaluate the functional role of bifidobacteria as microbial biomarkers.Figure 1Methods: A stool sample was obtained from UC patients, HS and processed for microbial DNA extraction. The microbiota taxonomic profile and the bacterial population diversity index were determined by using a metagenomic approach. Patient demographics, level of disease activity and therapy were collected and analyzed. Data extracted from stool samples of UC patients were compared with those of HS. Results: Between April 2015 and April 2016, 28 patients were enrolled: 14 UC patients (mean age 39y), and 14 HS (mean age 33y). Among UC patients, 6 had active disease (3 were under therapy with infliximab, 1 with adalimumab, 1 with azathioprine), and 8 showed disease remission (3 were under therapy with azathioprine, 1 with adalimumab; 1 with mesalamine, 3 no therapy). The fecal microbiome of UC patients, independently from the type of therapy, showed a marked reduction in bacterial population diversity index. Bifidobacteria were reduced in abundance of >30 % in UC group compared to HS group. Among UC patients, those with active disease showed a significant reduction (4 fold; P< 0.05) of the bifidobacteria compared to those in remission. Conclusion:Bifidobacterium bifidum is a reliable microbial biomarker for UC. The reduction in its overall abundance and the decreased microbial diversity index appear to be related to the presence of disease and may explain, at least in part, the role of intestinal microbiome in the pathogenesis of ulcerative colitis. Moreover, the significantly different abundance of Bifidobacterium bifidum between patients with active disease and in remission, highlights that Bifidobacterium bifidum may be considered an activity index of the disease.

Deciphering bifidobacterial-mediated metabolic interactions and their impact on gut microbiota by a multi-omics approach.

The intricacies of cooperation and competition between microorganisms are poorly investigated for particular components of the gut microbiota. In order to obtain insights into the manner by which different bifidobacterial species coexist in the mammalian gut, we investigated possible interactions between four human gut commensals, Bifidobacterium bifidum PRL2010, Bifidobacterium adolescentis 22L, Bifidobacterium breve 12L and Bifidobacterium longum subsp. infantis ATCC15697, in the intestine of conventional mice. The generated information revealed various ecological/metabolic strategies, including glycan-harvesting, glycan-breakdown and cross-feeding behavior, adopted by bifidobacteria in the highly competitive environment of the mammalian intestine. Introduction of two or multiple bifidobacterial strains caused a clear shift in the microbiota composition of the murine cecum. Whole-genome transcription profiling coupled with metagenomic analyses of single, dual or multiple associations of bifidobacterial strains revealed an expansion of the murine gut glycobiome toward enzymatic degradation of plant-derived carbohydrates, such as xylan, arabinoxylan, starch and host-derived glycan substrates. Furthermore, these bifidobacterial communities evoked major changes in the metabolomic profile of the microbiota as observed by shifts in short chain fatty acid production and carbohydrate availability in the murine cecum. Overall, these data support an ecological role of bifidobacteria acting directly or through cross-feeding activities in shaping the gut murine microbiome to instigate an enrichment of saccharolytic microbiota.

Glycan cross-feeding activities between bifidobacteria under in vitro conditions.

Bifidobacteria colonize the gut of various mammals, including humans, where they may metabolize complex, diet-, and host-derived carbohydrates. The glycan-associated metabolic features encoded by bifidobacteria are believed to be strongly influenced by cross-feeding activities due to the co-existence of strains with different glycan-degrading properties. In this study, we observed an enhanced growth yield of Bifidobacterium bifidum PRL2010 when co-cultivated with Bifidobacterium breve 12L, Bifidobacterium adolescentis 22L, or Bifidobacterium thermophilum JCM1207. This enhanced growth phenomenon was confirmed by whole genome transcriptome analyses, which revealed co-cultivation-associated transcriptional induction of PRL2010 genes involved in carbohydrate metabolism, such as those encoding for carbohydrate transporters and associated energy production, and genes required for translation, ribosomal structure, and biogenesis, thus supporting the idea that co-cultivation of certain bifidobacterial strains with B. bifidum PRL2010 causes enhanced metabolic activity, and consequently increased lactate and/or acetate production. Overall, these data suggest that PRL2010 cells benefit from the presence of other bifidobacterial strains.

Insights into teichoic acid biosynthesis by Bifidobacterium bifidum PRL2010.

Bifidobacteria are colonizers of the human gut, where they are interacting with their host as well as with other members of the intestinal microbiota. Teichoic acids (TAs) have previously been shown to play an important role in modulating microbe-host interactions in the human gut. However, so far, there is a paucity of information regarding the presence of TAs in the cell envelope of bifidobacteria. In silico analyses targeting the chromosomes of all 48 (sub)species that currently represent the genus Bifidobacterium revealed the presence of genes responsible for TA biosynthesis, suggesting that bifidobacteria contain both wall TAs and lipoteichoic acids. Transcriptome analyses of the infant gut commensal Bifidobacterium bifidum PRL2010 highlighted that the transcription of the presumptive TA biosynthetic loci is modulated in response to environmental conditions reflecting those of the human gut. Furthermore, chemical characterization of TAs produced by PRL2010 indicates the presence of lipoteichoic acids.

Evidence for cholesterol-lowering activity by Bifidobacterium bifidum PRL2010 through gut microbiota modulation.

Bifidobacteria are members of the human gut microbiota, which are known to influence the metabolic abilities of their host. Here, we investigated the capabilities of bifidobacteria to reduce cholesterol levels in synthetic growth media, clearly demonstrating assimilation of this molecule by particular bifidobacterial strains, including Bifidobacterium bifidum PRL2010 (LMG S-28692). The transcriptomic analysis of PRL2010 cells cultivated in the presence of cholesterol revealed a significantly increased transcription level of genes encoding putative transporters and reductases, indicative of specific mechanisms for cholesterol assimilation as well as cholesterol conversion to coprostanol. Cholesterol lowering activity of B. bifidum PRL2010 cells was further evaluated by means of an in vivo murine model, showing that the fecal microbiota of mice is modified toward those bacteria involved in the metabolism of cholesterol.

Cross-feeding by Bifidobacterium breve UCC2003 during co-cultivation with Bifidobacterium bifidum PRL2010 in a mucin-based medium.

BackgroundBifidobacteria constitute a specific group of commensal bacteria that commonly inhabit the mammalian gastrointestinal tract. Bifidobacterium breve UCC2003 was previously shown to utilize a variety of plant/diet/host-derived carbohydrates, including cellodextrin, starch and galactan, as well as the mucin and HMO-derived monosaccharide, sialic acid. In the current study, we investigated the ability of this strain to utilize parts of a host-derived source of carbohydrate, namely the mucin glycoprotein, when grown in co-culture with the mucin-degrading Bifidobacterium bifidum PRL2010.ResultsB. breve UCC2003 was shown to exhibit growth properties in a mucin-based medium, but only when grown in the presence of B. bifidum PRL2010, which is known to metabolize mucin. A combination of HPAEC-PAD and transcriptome analyses identified some of the possible monosaccharides and oligosaccharides which support this enhanced co-cultivation growth/viability phenotype.ConclusionThis study describes the potential existence of a gut commensal relationship between two bifidobacterial species. We demonstrate the in vitro ability of B. breve UCC2003 to cross-feed on sugars released by the mucin-degrading activity of B. bifidum PRL2010, thus advancing our knowledge on the metabolic adaptability which allows the former strain to colonize the (infant) gut by its extensive metabolic abilities to (co-)utilize available carbohydrate sources.

Expression of sortase-dependent pili of Bifidobacterium bifidum PRL2010 in response to environmental gut conditions.

Cell surface pili have recently been found in many different bifidobacterial species, including the infant gut commensal Bifidobacterium bifidum PRL2010. Pili produced by PRL2010 have been shown to be important molecular mediators for bacterial interaction with its human host. However, nothing is known about the modulation of their expression in response to cues that reflect the gastro intestinal environment, such as thermal, acidic, and osmotic challenges, or the presence of other gut microorganisms. Here, we investigated how different stress conditions that simulate the gastrointestinal niche influence the expression of PRL2010 sortase-dependent pili, and how this may impact on the coexistence and interaction with other human gut commensals.

Metabolism of sialic acid by Bifidobacterium breve UCC2003.

Bifidobacteria constitute a specific group of commensal bacteria that inhabit the gastrointestinal tracts of humans and other mammals. Bifidobacterium breve UCC2003 has previously been shown to utilize several plant-derived carbohydrates that include cellodextrins, starch, and galactan. In the present study, we investigated the ability of this strain to utilize the mucin- and human milk oligosaccharide (HMO)-derived carbohydrate sialic acid. Using a combination of transcriptomic and functional genomic approaches, we identified a gene cluster dedicated to the uptake and metabolism of sialic acid. Furthermore, we demonstrate that B. breve UCC2003 can cross feed on sialic acid derived from the metabolism of 3'-sialyllactose, an abundant HMO, by another infant gut bifidobacterial strain, Bifidobacterium bifidum PRL2010.

Kefir fermented milk and kefiran promote growth of Bifidobacterium bifidum PRL2010 and modulate its gene expression

Bifidobacteria constitute one of the dominant groups of microorganisms colonizing the human gut of infants. Their ability to utilize various host-derived glycans as well as dietary carbohydrates has received considerable scientific attention. However, very little is known about the role of fermented foods, such as kefir, or their constituent glycans, such as kefiran, as substrates for bifidobacterial growth and for the modulation of the expression of bifidobacterial host-effector molecules. Here, we show that Bifidobacterium bifidum PRL2010 exhibits high growth performance among the bifidobacterial strains tested when cultivated on kefir and/or kefiran polymer. Furthermore, a 16S rRNA metagenomic approach revealed that the microbiota of kefir is modified upon the addition of PRL2010 cells to the kefir matrix. Finally, our results show that kefir and kefiran are able to influence the transcriptome of B. bifidum PRL2010 causing increased transcription of genes involved in the metabolism of dietary glycans as well as genes that act as host–microbe effector molecules such as pili. Altogether, these data support the use of kefir as a valuable means for the delivery of effective microbial cells in probiotic therapy.

Role of sortase-dependent pili ofBifidobacterium bifidumPRL2010 in modulating bacterium–host interactions

Bifidobacteria represent one of the dominant groups of microorganisms colonizing the human infant intestine. Commensal bacteria that interact with a eukaryotic host are believed to express adhesive molecules on their cell surface that bind to specific host cell receptors or soluble macromolecules. Whole-genome transcription profiling of Bifidobacterium bifidum PRL2010, a strain isolated from infant stool, revealed a small number of commonly expressed extracellular proteins, among which were genes that specify sortase-dependent pili. Expression of the coding sequences of these B. bifidum PRL2010 appendages in nonpiliated Lactococcus lactis enhanced adherence to human enterocytes through extracellular matrix protein and bacterial aggregation. Furthermore, such piliated L. lactis cells evoked a higher TNF-α response during murine colonization compared with their nonpiliated parent, suggesting that bifidobacterial sortase-dependent pili not only contribute to adherence but also display immunomodulatory activity.

Evaluation of adhesion properties and antibacterial activities of the infant gut commensal Bifidobacterium bifidum PRL2010

Bifidobacteria are extensively exploited by the food industry as health-promoting microorganisms. However, very little is known about the molecular mechanisms responsible for these beneficial activities, or the molecular players that sustain their ability to colonize and persist within the human gut. Here, we have investigated the enteric adaptation features of the gut commensal Bifidobacterium bifidum PRL2010, originally isolated from infant feces. This strain was able to survive under gastrointestinal challenges, while it was shown to adhere to human epithelial intestinal cell monolayers (Caco 2 and HT-29), thereby inhibiting adhesion of pathogenic bacteria such as Escherichia coli and Cronobacter sakazakii.

An efficient and reproducible method for transformation of genetically recalcitrant bifidobacteria

This study describes an efficient transformation system for the introduction of plasmid DNA into Bifidobacterium bifidum PRL2010 and Bifidobacterium asteroides PRL2011, for which to the best of our knowledge no transformation data have been reported previously. The method is based on electroporation of bifidobacterial cells, which were made competent by an optimized methodology based on varying media and growth conditions. Furthermore, the transformation protocol was applied in order to design a PRL2010-derivative, which carries antibiotic resistance against chloramphenicol and which was used to monitor PRL2010 colonization in a murine model.

Genetic strategies for mucin metabolism in Bifidobacterium bifidum PRL2010: An example of possible human-microbe co-evolution

Recently, we decoded the genome sequence of Bifidobacterium bifidum PRL2010, an isolate originally recovered from a stool sample of a three month old infant. In silico analysis revealed that the the 2,214,650 bp genome of B. bifidum PRL2010 encoded a rich gene repertoire dedicated to the metabolism of host-derived glycans, such as those present in mucin. Whole proteome profiling as well as transcriptomic investigations were used to identify the genetic players and mechanisms responsible for mucin breakdown, revealing a set of chromosomal loci that allow degradation of various O-linked glycans in mucin (such as core 1, core 2, core 3 and core 4 O-glycans). Comparative genomic analyses involving different B. bifidum genomes demonstrated the ubiquitous presence of these genetic regions in genomes of members of this bifidobacterial taxon, and suggest specific B. bifidum metabolic traits that underpin this case of host-microbe co-evolution.