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Background/Objectives: The global emergence of antibiotic-resistant zooanthroponotic Escherichia coli strains, producing extended-spectrum beta-lactamases (ESBL-E) and persisting in the intestines of farm animals, has now led to the development of a pandemic of extra-intestinal infectious diseases in humans. The search for innovative probiotic microorganisms that eliminate ESBL-E from the intestines of humans and animals is relevant. Previously, we received three isolates of bifidobacteria: from milk of a calved cow (BLLT1), feces of a newborn calf (BLLT2) and feces of a three-year-old child who received fresh milk from this calved cow (BLLT3). Our goal was to evaluate the genetic identity of BLLT1, BLLT2, BLLT3 isolates using genomic DNA fingerprinting (GDF), to study the tolerance, adhesion, homeostatic and antibacterial activity of BLLT1 against ESBL-E. Methods: We used a complex of microbiological, molecular biological, and immunological methods, including next generation sequencing (NGS). Results: GDF showed that DNA fragments of BLLT2 and BLLT3 isolates were identical in number and size to DNA fragments of BLLT1. These data show for the first time the possibility of natural horizontal transmission of BLLT1 through with the milk of a calved cow into the intestines of a calf and the intestines of a child. BLLT1 was resistant to gastric and intestinal stresses and exhibited high adhesive activity to calf, pig, chicken, and human enterocytes. This indicates the unique ability of BLLT1 to inhabit the intestines of animals and humans. We are the first to show that BLLT1 has antibacterial activity against ESBL-E strains that persist in humans and animals. BLLT1 produced 145 ± 8 mM of acetic acid, which reduced the pH of the nutrient medium from 6.8 to 5.2. This had an antibacterial effect on ESBL-E. The genome of BLLT1 contains ABC-type carbohydrate transporter gene clusters responsible for the synthesis of acetic acid with its antibacterial activity against ESBL-E. BLLT1 inhibited TLR4 mRNA expression induced by ESBL-E in HT-29 enterocytes, and protected the enterocyte monolayers used in this study as a bio-model of the intestinal barrier. BLLT1 increased intestinal alkaline phosphatase (IAP) as one of the main molecular factors providing intestinal homeostasis. Conclusions: BLLT1 shows promise for the creation of innovative functional nutritional products for humans and feed additives for farm animals that will reduce the spread of ESBL-E strains in the food chain. [ABSTRACT FROM AUTHOR]

This study was conducted to isolate and identify Bifidobacterium from the feces of Kazakh school-age children in Yining, Xinjiang and evaluate the in vitro probiotic characteristics of B. longum subsp. longum isolates. By groEL gene sequencing and repetitive element sequence-based polymerase chain reaction (rep-PCR) fingerprinting, 416 Bifidobacterium strains were identified to belong to B. longum, B. bifidum, B. pseudocatenulatum, B. catenulatum and B. breve. According to the fingerprints of B. longum subsp. longum, 27 genotypes showed genetic differences between individual strains and the coexistence of multiple strains in the gut was found. The in vitro experimental results showed that out of 27 representative strains, strains 2B3-21, 1B23-11, 2B33-3, and 1B68-16 were optimal in acid and bile salt tolerance, strains 1B68-16, 2B13-5, 2B33-3, and 1B39-2 had broad-spectrum antibacterial properties, and strains 1B38-1, 2B33-3, 1B68-16, and 2B13-28 showed a strong antioxidant capacity. Considering the antibiotic resistance of all strains and their ability to utilize plant-derived glycans, strains 1B38-1 and 2B13-28 were selected to assess their in vivo probiotic potentials. This study may lay the foundation for the development of excellent probiotics and related products for populations from specific areas.

Background/Objectives: The global emergence of antibiotic-resistant zooanthroponotic Escherichia coli strains, producing extended-spectrum beta-lactamases (ESBL-E) and persisting in the intestines of farm animals, has now led to the development of a pandemic of extra-intestinal infectious diseases in humans. The search for innovative probiotic microorganisms that eliminate ESBL-E from the intestines of humans and animals is relevant. Previously, we received three isolates of bifidobacteria: from milk of a calved cow (BLLT1), feces of a newborn calf (BLLT2) and feces of a three-year-old child who received fresh milk from this calved cow (BLLT3). Our goal was to evaluate the genetic identity of BLLT1, BLLT2, BLLT3 isolates using genomic DNA fingerprinting (GDF), to study the tolerance, adhesion, homeostatic and antibacterial activity of BLLT1 against ESBL-E. Methods: We used a complex of microbiological, molecular biological, and immunological methods, including next generation sequencing (NGS). Results: GDF showed that DNA fragments of BLLT2 and BLLT3 isolates were identical in number and size to DNA fragments of BLLT1. These data show for the first time the possibility of natural horizontal transmission of BLLT1 through with the milk of a calved cow into the intestines of a calf and the intestines of a child. BLLT1 was resistant to gastric and intestinal stresses and exhibited high adhesive activity to calf, pig, chicken, and human enterocytes. This indicates the unique ability of BLLT1 to inhabit the intestines of animals and humans. We are the first to show that BLLT1 has antibacterial activity against ESBL-E strains that persist in humans and animals. BLLT1 produced 145 ± 8 mM of acetic acid, which reduced the pH of the nutrient medium from 6.8 to 5.2. This had an antibacterial effect on ESBL-E. The genome of BLLT1 contains ABC-type carbohydrate transporter gene clusters responsible for the synthesis of acetic acid with its antibacterial activity against ESBL-E. BLLT1 inhibited TLR4 mRNA expression induced by ESBL-E in HT-29 enterocytes, and protected the enterocyte monolayers used in this study as a bio-model of the intestinal barrier. BLLT1 increased intestinal alkaline phosphatase (IAP) as one of the main molecular factors providing intestinal homeostasis. Conclusions: BLLT1 shows promise for the creation of innovative functional nutritional products for humans and feed additives for farm animals that will reduce the spread of ESBL-E strains in the food chain.

l-Arabinofuranosides with β-linkages are present in several plant molecules, such as arabinogalactan proteins (AGPs), extensin, arabinan, and rhamnogalacturonan-II. We previously characterized a β-l-arabinofuranosidase from Bifidobacterium longum subsp. longum JCM 1217, Bll1HypBA1, which was found to belong to the glycoside hydrolase (GH) family 127. This strain encodes two GH127 genes and two GH146 genes. In the present study, we characterized a GH146 β-l-arabinofuranosidase, Bll3HypBA1 (BLLJ_1848), which was found to constitute a gene cluster with AGP-degrading enzymes. This recombinant enzyme degraded AGPs and arabinan, which contain Araf-β1,3-Araf structures. In addition, the recombinant enzyme hydrolyzed oligosaccharides containing Araf-β1,3-Araf structures but not those containing Araf-β1,2-Araf and Araf-β1,5-Araf structures. The crystal structures of Bll3HypBA1 were determined at resolutions up to 1.7 Å. The monomeric structure of Bll3HypBA1 comprised a catalytic (α/α)6 barrel and two β-sandwich domains. A hairpin structure with two β-strands was observed in Bll3HypBA1, to extend from a β-sandwich domain and partially cover the active site. The active site contains a Zn2+ ion coordinated by Cys3-Glu and exhibits structural conservation of the GH127 cysteine glycosidase Bll1HypBA1. This is the first study to report on a β1,3-specific β-l-arabinofuranosidase. Key points: • β1,3-l-Arabinofuranose residues are present in arabinogalactan proteins and arabinans as a terminal sugar. • β-l-Arabinofuranosidases are widely present in intestinal bacteria. • Bll3HypBA1 is the first enzyme characterized as a β1,3-linkage-specific β-l-arabinofuranosidase. [ABSTRACT FROM AUTHOR]

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IntroductionThere are differences in the gut microbiome and metabolome when the host undergoes different physical or pathological conditions. However, the inter-relationship of microbiome and metabolome biomarkers to potentially promote the health of dairy cows needs to be studied. Further, the development of next-generation probiotics for dairy cattle health promotion has not been demonstrated.ObjectiveIn the present study, we identified the microbiome and metabolome biomarkers associated with healthy cows.MethodsWe analyzed the relationships of the ruminal microorganism profile and metabolites between healthy and mastitis lactating dairy cows. The roles of bacterial biomarker were further verified by in vitro fermentation and cow-to-mouse fecal microbiota transplantation (FMT).ResultsTwo species, Ruminococcus flavefaciens and Bifidobacterium longum subsp. longum, and six rumen metabolites were positively correlated with healthy cows by Spearman’s correlation analysis. Through in vitro ruminal fermentation, inoculating R. flavefaciens and B. longum subsp. longum showed the upregulation of the levels of putrescine, xanthurenic acid, and pyridoxal in the mastitis ruminal fluid, which confirmed the inter-relationships between these microbiota and metabolites associated with healthy cows. Further, we verified the role of R. flavefaciens and B. longum subsp. longum in promoting health by FMT. The administration of R. flavefaciens and B. longum subsp. longum reduced the death rate and recovered the bodyweight loss of germ-free mice caused by FMT mastitis feces.DiscussionWe provided evidence that the bacterial biomarkers alter downstream metabolites. This could indirectly indicate that the two bacterial biomarkers have the potential to be used as next-generation probiotics for dairy cattle, although it needs more evidence to support our hypothesis. Two species, R. flavefaciens and B. longum subsp. longum, with three metabolites, putrescine, xanthurenic acid, and pyridoxal, identified in the ruminal fluid, may point to a new health-promoting and disease-preventing approach for dairy cattle.

Abstract Background The importance of the gut microbiota at the early stage of life and their longitudinal effect on host health have recently been well investigated. In particular, Bifidobacterium longum subsp. longum, a common component of infant gut microbiota, appears in the gut shortly after birth and can be detected there throughout an individual’s lifespan. However, it remains unclear whether this species colonizes in the gut over the long term from early infancy. Here, we investigated the long-term colonization of B. longum subsp. longum by comparing the genotypes of isolates obtained at different time points from individual subjects. Strains were isolated over time from the feces of 12 subjects followed from early infancy (the first six months of life) up to childhood (approximately six years of age). We also considered whether the strains were transmitted from their mothers’ perinatal samples (prenatal feces and postnatal breast milk). Results Intra-species diversity of B. longum subsp. longum was observed in some subjects’ fecal samples collected in early infancy and childhood, as well as in the prenatal fecal samples of their mothers. Among the highlighted strains, several were confirmed to colonize and persist in single individuals from as early as 90 days of age for more than six years; these were classified as long-term colonizers. One of the long-term colonizers was also detected from the corresponding mother’s postnatal breast milk. Quantitative polymerase chain reaction data suggested that these long-term colonizers persisted in the subjects’ gut despite the existence of the other predominant species of Bifidobacterium. Conclusions Our results showed that several strains belonging to B. longum subsp. longum colonized in the human gut from early infancy through more than six years, confirming the existence of long-term colonizers from this period. Moreover, the results suggested that these strains persisted in the subjects’ gut while co-existing with the other predominant bifidobacterial species. Our findings also suggested the importance of microbial-strain colonization in early infancy relative to their succession and showed the possibility that probiotics targeting infants might have longitudinal effects. Trial Registration TRN: ISRCTN25216339. Date of registration: 11/03/2016. Prospectively registered.

(1) Background: We aimed to isolate and identify potential next-generation probiotics (NGP) by investigating the interrelationships between gastrointestinal microbiota and diarrhea in preruminant Holstein calves. (2) Material and methods: Twenty preruminant Holstein calves were divided into healthy and diarrheic groups after the combination outcomes of veterinary diagnosis and fecal scores. The fecal microbiome, plasma cytokines, plasma immunoglobulin (Ig) G and haptoglobin were analyzed. The potential probiotic bacteria were identified by comparing the microbiota difference between healthy and diarrheic calves and correlation analysis with fecal scores and inflammatory markers. The identified bacteria were also isolated for further evaluation for antimicrobial activities and immunoregulatory effects. (3) Results: Microbiota analysis suggested that Ruminococcaceae_UCG_014, Bifidobacterium and Pseudoflavonifractor positively correlated with bovine IgG and negatively correlated with fecal score; inflammatory factors, bovine HP, and IL-8 were classified as beneficial bacteria contributing to the health of the calves. The alternation of gut microbial composition also induced changes in the functional gene enrichment of gut microbiota in calves. The gathering of microbiomic data strongly indicated the possible beneficial effects of Bifidobacterium longum subsp. longum, expected to develop as NGP. After isolation and evaluation of the potential functionality in vitro, two specific bifidobacterial strains demonstrated antimicrobial activities and immunoregulatory effects. (4) Conclusions: The results provide a new probiotic searching approach for preventing gastrointestinal disorders in preruminant calves. Further animal study is necessary to verify the results.

Ulcerative colitis (UC) is a major disease that has endangered human health. Our previous study demonstrated that Bifidobacterium longum subsp. longum YS108R, a ropy exopolysaccharide (EPS)-producing bacterium, could alleviate UC in mice, but it is unclear whether EPS is the key substance responsible for its action. In this study, we proposed to investigate the remitting effect of EPS from B. longum subsp. longum YS108R on UC in a DSS-induced UC mouse model. Water extraction and alcohol precipitation were applied to extract EPS from the supernatant of B. longum subsp. longum YS108R culture. Then the animal trial was performed, and the results indicated that YS108R EPS ameliorated colonic pathological damage and the intestinal barrier. YS108R EPS suppressed inflammation via NF-κB signaling pathway inhibition and attenuated oxidative stress via the Nrf2 signaling pathway activation. Remarkably, YS108R EPS regulated gut microbiota, as evidenced by an increase in short-chain fatty acid (SCFA)-producing bacteria and a decline in Gram-negative bacteria, resulting in an increase of propionate and butyrate and a reduction of lipopolysaccharide (LPS). Collectively, YS108R EPS manipulated the intestinal microbiota and its metabolites, which further improved the intestinal barrier and inhibited inflammation and oxidative stress, thereby alleviating UC.

Bifidobacterium longum subsp. longum is a prevalent group in the human gut microbiome. Its persistence in the intestinal microbial community suggests a close host-microbe relationship according to age. The subspecies adaptations are related to metabolic capabilities and genomic and functional diversity. In this study, 154 genomes from public databases and four new Chilean isolates were genomically compared through an in silico approach to identify genomic divergence in genes associated with carbohydrate consumption and their possible adaptations to different human intestinal niches. The pangenome of the subspecies was open, which correlates with its remarkable ability to colonize several niches. The new genomes homogenously clustered within subspecies longum, as observed in phylogenetic analysis. B. longum SC664 was different at the sequence level but not in its functions. COG analysis revealed that carbohydrate use is variable among longum subspecies. Glycosyl hydrolases participating in human milk oligosaccharide use were found in certain infant and adult genomes. Predictive genomic analysis revealed that B. longum M12 contained an HMO cluster associated with the use of fucosylated HMOs but only endowed with a GH95, being able to grow in 2-fucosyllactose as the sole carbon source. This study identifies novel genomes with distinct adaptations to HMOs and highlights the plasticity of B. longum subsp. longum to colonize the human gut microbiota.

Background: The importance of the gut microbiota at the early stage of life and their longitudinal effect on host health have recently been well investigated. In particular, Bifidobacterium longum subsp. longum, a common component of infant gut microbiota, appears in the gut shortly after birth and can be detected there throughout an individual's lifespan. However, it remains unclear whether this species colonizes in the gut over the long term from early infancy. Here, we investigated the long-term colonization of B. longum subsp. longum by comparing the genotypes of isolates obtained at different time points from individual subjects. Strains were isolated over time from the feces of 12 subjects followed from early infancy (the first six months of life) up to childhood (approximately six years of age). We also considered whether the strains were transmitted from their mothers' perinatal samples (prenatal feces and postnatal breast milk). Results: Intra-species diversity of B. longum subsp. longum was observed in some subjects' fecal samples collected in early infancy and childhood, as well as in the prenatal fecal samples of their mothers. Among the highlighted strains, several were confirmed to colonize and persist in single individuals from as early as 90 days of age for more than six years; these were classified as long-term colonizers. One of the long-term colonizers was also detected from the corresponding mother's postnatal breast milk. Quantitative polymerase chain reaction data suggested that these long-term colonizers persisted in the subjects' gut despite the existence of the other predominant species of Bifidobacterium. Conclusions: Our results showed that several strains belonging to B. longum subsp. longum colonized in the human gut from early infancy through more than six years, confirming the existence of long-term colonizers from this period. Moreover, the results suggested that these strains persisted in the subjects' gut while co-existing with the other predominant bifidobacterial species. Our findings also suggested the importance of microbial-strain colonization in early infancy relative to their succession and showed the possibility that probiotics targeting infants might have longitudinal effects. Trial Registration: TRN: ISRCTN25216339. Date of registration: 11/03/2016. Prospectively registered. [ABSTRACT FROM AUTHOR]

Bifidobacteria are one of the major components in human gut microbiota and well-known as beneficial microbes. However, clarification of commensal mechanisms of bifidobacteria in the intestines is still ongoing, especially in the presence of the gut microbiota. Here, we applied recombinase-based in vivo expression technology (R-IVET) using the bacteriophage P1 Cre/loxP system to Bifidobacterium longum subsp. longum 105-A (B. longum 105-A) to identify genes that are specifically expressed in the gastrointestinal tract of conventionally raised mice. Oral administration of the genomic DNA library of B. longum 105-A to conventionally raised mice resulted in the identification of 73 in vivo-induced genes. Four out of seven tested genes were verified in vivo-specific induction at least in the cecum by quantitative reverse transcription PCR. Although there is still room for improvement of the system, our findings can contribute to expanding our understanding of the commensal behavior of B. longum in the gut ecosystem.

Introduction: There are differences in the gut microbiome and metabolome when the host undergoes different physical or pathological conditions. However, the inter-relationship of microbiome and metabolome biomarkers to potentially promote the health of dairy cows needs to be studied. Further, the development of next-generation probiotics for dairy cattle health promotion has not been demonstrated., Objective: In the present study, we identified the microbiome and metabolome biomarkers associated with healthy cows., Methods: We analyzed the relationships of the ruminal microorganism profile and metabolites between healthy and mastitis lactating dairy cows. The roles of bacterial biomarker were further verified by in vitro fermentation and cow-to-mouse fecal microbiota transplantation (FMT)., Results: Two species, Ruminococcus flavefaciens and Bifidobacterium longum subsp. longum , and six rumen metabolites were positively correlated with healthy cows by Spearman's correlation analysis. Through in vitro ruminal fermentation, inoculating R. flavefaciens and B. longum subsp. longum showed the upregulation of the levels of putrescine, xanthurenic acid, and pyridoxal in the mastitis ruminal fluid, which confirmed the inter-relationships between these microbiota and metabolites associated with healthy cows. Further, we verified the role of R. flavefaciens and B. longum subsp. longum in promoting health by FMT. The administration of R. flavefaciens and B. longum subsp. longum reduced the death rate and recovered the bodyweight loss of germ-free mice caused by FMT mastitis feces., Discussion: We provided evidence that the bacterial biomarkers alter downstream metabolites. This could indirectly indicate that the two bacterial biomarkers have the potential to be used as next-generation probiotics for dairy cattle, although it needs more evidence to support our hypothesis. Two species, R. flavefaciens and B. longum subsp. longum , with three metabolites, putrescine, xanthurenic acid, and pyridoxal, identified in the ruminal fluid, may point to a new health-promoting and disease-preventing approach for dairy cattle., Competing Interests: S-HC was employed by Biotools Co. Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Hsieh, Chuang, Hsu, Ho, Li, Chou and Chen.)

Simple Summary: The present study investigated the relationship between gastrointestinal microbiota and diarrhea in preruminant calves by using immune-related markers and further isolating specific bacterial strains, enriched in clinically healthy individuals, for potential next-generation probiotics. The gathering of microbiomic data strongly indicated the possible beneficial effects of Bifidobacterium longum subsp. longum. With further screening and isolating with immunomodulatory and antagonistic effects, two Bifidobacterium longum subsp. longum strains might be expected to emerge as next-generation probiotics. The finding here might provide a solution for preventing gastrointestinal disorders for preruminant calves without sustained periods of administration through inhibiting the infectious bacteria, immunomodulatory effect and possible modulating microbiota. (1) Background: We aimed to isolate and identify potential next-generation probiotics (NGP) by investigating the interrelationships between gastrointestinal microbiota and diarrhea in preruminant Holstein calves. (2) Material and methods: Twenty preruminant Holstein calves were divided into healthy and diarrheic groups after the combination outcomes of veterinary diagnosis and fecal scores. The fecal microbiome, plasma cytokines, plasma immunoglobulin (Ig) G and haptoglobin were analyzed. The potential probiotic bacteria were identified by comparing the microbiota difference between healthy and diarrheic calves and correlation analysis with fecal scores and inflammatory markers. The identified bacteria were also isolated for further evaluation for antimicrobial activities and immunoregulatory effects. (3) Results: Microbiota analysis suggested that Ruminococcaceae_UCG_014, Bifidobacterium and Pseudoflavonifractor positively correlated with bovine IgG and negatively correlated with fecal score; inflammatory factors, bovine HP, and IL-8 were classified as beneficial bacteria contributing to the health of the calves. The alternation of gut microbial composition also induced changes in the functional gene enrichment of gut microbiota in calves. The gathering of microbiomic data strongly indicated the possible beneficial effects of Bifidobacterium longum subsp. longum, expected to develop as NGP. After isolation and evaluation of the potential functionality in vitro, two specific bifidobacterial strains demonstrated antimicrobial activities and immunoregulatory effects. (4) Conclusions: The results provide a new probiotic searching approach for preventing gastrointestinal disorders in preruminant calves. Further animal study is necessary to verify the results. [ABSTRACT FROM AUTHOR]

Abstract: The taxonomic positions of the subspecies of Bifidobacterium longum (B. longum subsp. longum, subsp. infantis, and subsp. suis) have been controversial. A current proposal is that the former two species “B. infantis” and “B. suis” be unified with B. longum and all three reclassified as three subspecies. To test this proposal, ribosomal protein profiling as observed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was applied to the classification of 17 strains of B. longum, including three subspecies. Among 41 different kinds of ribosomal proteins selected as biomarkers whose masses were calculated from their amino acid sequences, 31–41 ribosomal proteins were observed in sample strains with the same masses as the references. The high matching rate indicates high conservation of ribosomal proteins within the sample strains, and therefore strongly supports the unification of the former species. However, the masses of some ribosomal proteins varied within species. The phylogenetic tree constructed from the profiles of ribosomal proteins matched the references, showing a clear cluster of the subsp. longum and the subsp. infantis strains. This result supports the proposal to reclassify B. longum into subsp. longum and subsp. infantis. The subsp. suis strains formed an individual sub-cluster within the infantis cluster. However, their ribosomal proteins have both characters of longum and infantis types. This result suggests that the taxonomic position of the subsp. suis should be reconsidered. [Copyright &y& Elsevier]

Bifidobacterium longum subsp. longum is a prevalent group in the human gut microbiome. Its persistence in the intestinal microbial community suggests a close host-microbe relationship according to age. The subspecies adaptations are related to metabolic capabilities and genomic and functional diversity. In this study, 154 genomes from public databases and four new Chilean isolates were genomically compared through an in silico approach to identify genomic divergence in genes associated with carbohydrate consumption and their possible adaptations to different human intestinal niches. The pangenome of the subspecies was open, which correlates with its remarkable ability to colonize several niches. The new genomes homogenously clustered within subspecies longum, as observed in phylogenetic analysis. B. longum SC664 was different at the sequence level but not in its functions. COG analysis revealed that carbohydrate use is variable among longum subspecies. Glycosyl hydrolases participating in human milk oligosaccharide use were found in certain infant and adult genomes. Predictive genomic analysis revealed that B. longum M12 contained an HMO cluster associated with the use of fucosylated HMOs but only endowed with a GH95, being able to grow in 2-fucosyllactose as the sole carbon source. This study identifies novel genomes with distinct adaptations to HMOs and highlights the plasticity of B. longum subsp. longum to colonize the human gut microbiota. [ABSTRACT FROM AUTHOR]

Abstract: A multilocus variable number of tandem repeat analysis (MLVA) scheme was developed and 44 isolates of Bifidobacterium longum subsp. longum were typed, including 5 isolates recovered during a clinical trial. The MLVA scheme generated 19 profiles and proved to be a fast, reliable and relatively cheap typing method. [Copyright &y& Elsevier]

Bifidobacteria are one of the major components in human gut microbiota and well-known as beneficial microbes. However, clarification of commensal mechanisms of bifidobacteria in the intestines is still ongoing, especially in the presence of the gut microbiota. Here, we applied recombinase-based in vivo expression technology (R-IVET) using the bacteriophage P1 Cre/loxP system to Bifidobacterium longum subsp. longum 105-A (B. longum 105-A) to identify genes that are specifically expressed in the gastrointestinal tract of conventionally raised mice. Oral administration of the genomic DNA library of B. longum 105-A to conventionally raised mice resulted in the identification of 73 in vivo-induced genes. Four out of seven tested genes were verified in vivo-specific induction at least in the cecum by quantitative reverse transcription PCR. Although there is still room for improvement of the system, our findings can contribute to expanding our understanding of the commensal behavior of B. longum in the gut ecosystem. [ABSTRACT FROM AUTHOR]

Bifidobacterium longum is considered as a potential supplement in antiobesity treatment; however, the underlying molecular mechanism has rarely been studied. To understand the contributions of B. longum subsp. longum (BL21) in the prevention of obesity, we investigated alterations in the liver metabonomic phenotype and gut microbiota by ultraperformance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry and 16S ribosomal RNA gene sequencing in C57BL/6J male mice orally administered with BL21 for 8 weeks [high-fat diet (HFD)]. BL21 at 1 × 109 CFU·day-1 per mouse reduced the weight of mice by 16.9% relative to that of the mice fed with HFD and significantly lowered the serum levels of total cholesterol, triglycerides, and low-density lipoprotein cholesterol. BL21 also ameliorated fat vacuolization in liver cells and epididymal fat accumulation. BL21 also lowered the Firmicutes/Bacteroidetes ratio, regulated liver remodeling in glycerophospholipids, and alleviated the levels of d-tryptophan. A positive correlation between the butyrate-producing strain Roseburia and the cell membrane component phosphatidylserine was found for the first time. Thus, BL21 can potentially prevent mice from being obese by rebalancing the gut microbiota and glycerophospholipid metabolism. BL21 can be a promising dietary supplement for weight control.

In our previous study, we found that the open reading frame bl0675 in the genome of Bifidobacterium longum subsp. longum isolated from human feces encoded a novel putative fimbrial protein, was highly polymorphic, and had five variants (A, B, C, D, and E types). The aim of this study was to evaluate the affinity of these variants to porcine colonic mucins (PCMs). Protein-binding properties were examined using the recombinant BL0675 protein containing a C-terminal 6 × His tag (His-BL0675). Surface plasmon resonance analysis demonstrated that the His-BL0675 A type had strong affinity to PCMs (KD = 9.82 × 10(-8) M), whereas the B, C, D, and E types exhibited little or no binding. In a competitive enzyme-linked immunosorbent assay, His-BL0675 A type binding was reduced by addition of mucin oligosaccharides, suggesting that the binding occurs via carbohydrate chains of PCMs. The localization of BL0675 to the B. longum subsp. longum cell surface was confirmed by western blot analysis using A type polyclonal antibodies. Bacterial adhesion of B. longum subsp. longum to PCMs was also blocked by A type-specific antibodies; however, its adhesion properties were strain specific. Our results suggest that the BL0675 variants significantly contribute to the adhesion of B. longum subsp. longum strains. The expression and the adhesive properties of this protein are affected by genetic polymorphisms and are specific for B. longum subsp. longum strains. However, further studies are required on the properties of binding of these putative fimbrial proteins to the human gastrointestinal tract.