| 1. |
van den Broek LA,
Struijs K,
Verdoes JC,
Beldman G,
Voragen AG,
( 2003 ) Cloning and characterization of two alpha-glucosidases from Bifidobacterium adolescentis DSM20083. PMID : 12658515 : DOI : 10.1007/s00253-002-1179-1 Abstract >>
Two alpha-glucosidase encoding genes (aglA and aglB) from Bifidobacterium adolescentis DSM 20083 were isolated and characterized. Both alpha-glucosidases belong to family 13 of the glycosyl hydrolases. Recombinant AglA (EC 3.2.1.10) and AglB (EC 3.2.1.20), expressed in Escherichia coli, showed high hydrolytic activity towards isomaltose and pnp-alpha-glucoside. The K(m) for pnp-alpha-glucoside was 1.05 and 0.47 mM and the V(max) was 228 and 113 U mg(-1) for AglA and AglB, respectively. Using pnp-alpha-glucoside as substrate, the pH optimum for AglA was 6.6 and the temperature optimum was 37 degrees C. For AglB, values of pH 6.8 and 47 degrees C were found. AglA also showed high hydrolytic activity towards isomaltotriose and, to a lesser extent, towards trehalose. AglB has a high preference for maltose and less activity towards sucrose; minor activity was observed towards melizitose, low molecular weight dextrin, maltitol, and maltotriose. The recombinant alpha-glucosidases were tested for their transglucosylation activity. AglA was able to synthesize oligosaccharides from trehalose and sucrose. AglB formed oligosaccharides from sucrose, maltose, and melizitose.
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2. |
Requena T,
Burton J,
Matsuki T,
Munro K,
Simon MA,
Tanaka R,
Watanabe K,
Tannock GW,
( 2002 ) Identification, detection, and enumeration of human bifidobacterium species by PCR targeting the transaldolase gene. PMID : 11976117 : DOI : 10.1128/aem.68.5.2420-2427.2002 PMC : PMC127544 Abstract >>
Methods that enabled the identification, detection, and enumeration of Bifidobacterium species by PCR targeting the transaldolase gene were tested. Bifidobacterial species isolated from the feces of human adults and babies were identified by PCR amplification of a 301-bp transaldolase gene sequence and comparison of the relative migrations of the DNA fragments in denaturing gradient gel electrophoresis (DGGE). Two subtypes of Bifidobacterium longum, five subtypes of Bifidobacterium adolescentis, and two subtypes of Bifidobacterium pseudocatenulatum could be differentiated using PCR-DGGE. Bifidobacterium angulatum and B. catenulatum type cultures could not be differentiated from each other. Bifidobacterial species were also detected directly in fecal samples by this combination of PCR and DGGE. The number of species detected was less than that detected by PCR using species-specific primers targeting 16S ribosomal DNA (rDNA). Real-time quantitative PCR targeting a 110-bp transaldolase gene sequence was used to enumerate bifidobacteria in fecal samples. Real-time quantitative PCR measurements of bifidobacteria in fecal samples from adults correlated well with results obtained by culture when either a 16S rDNA sequence or the transaldolase gene sequence was targeted. In the case of samples from infants, 16S rDNA-targeted PCR was superior to PCR targeting the transaldolase gene for the quantification of bifidobacterial populations.
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3. |
Masco L,
Van Hoorde K,
De Brandt E,
Swings J,
Huys G,
( 2006 ) Antimicrobial susceptibility of Bifidobacterium strains from humans, animals and probiotic products. PMID : 16698847 : DOI : 10.1093/jac/dkl197 Abstract >>
The aim of this study was to assess the antimicrobial susceptibility of a taxonomically diverse set of Bifidobacterium strains to different classes of antimicrobial agents using a recently described medium. The susceptibility of 100 strains encompassing 11 bifidobacterial species originating from humans, animals and probiotic products to 12 antimicrobial agents was tested by agar overlay disc diffusion. Based on these results, one or two strains per species were selected for susceptibility testing to nine antibiotics by broth microdilution using the Lactic acid bacteria Susceptibility test Medium (LSM) supplemented with cysteine. The genotypic basis of atypical tetracycline resistance was further characterized using PCR, Southern blotting and partial sequencing. Based on the distribution of inhibition zone diameters and MIC values, all strains tested were susceptible to amoxicillin, chloramphenicol, erythromycin, quinupristin/dalfopristin, rifampicin and vancomycin. Our data also reinforce earlier observations indicating that bifidobacteria are intrinsically resistant to gentamicin, sulfamethoxazole and polymyxin B. Susceptibility to trimethoprim, trimethoprim/sulfamethoxazole, ciprofloxacin, clindamycin, tetracycline and minocycline was variable. The tet(W) gene was responsible for tetracycline resistance in 15 strains including 7 probiotic isolates belonging to the taxa Bifidobacterium animalis subsp. lactis and Bifidobacterium bifidum. This gene was present in a single copy on the chromosome and did not appear to be associated with the conjugative transposon TnB1230 previously found in tet(W)-containing Butyrivibrio fibrisolvens. The use of the LSM + cysteine medium allowed us to discriminate between intrinsic and atypical resistance properties of bifidobacteria and sets the scene for future definition of epidemiological cut-off values for all important Bifidobacterium species. The presence of an acquired tet(W) gene in several probiotic product isolates stresses the need for a minimal safety evaluation during the selection of Bifidobacterium strains for probiotic use.
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4. |
Vaugien L,
Prevots F,
Roques C,
( 2002 ) Bifidobacteria identification based on 16S rRNA and pyruvate kinase partial gene sequence analysis. PMID : 16887679 : DOI : 10.1016/S1075-9964(03)00025-8 Abstract >>
The lack of a simple and rapid identification system for Bifidobacterium species makes them difficult to use in industrial applications. To obtain valuable discriminating factor, we studied different strains, and human isolates by two molecular taxonomy methods. First method was based on chrono-differentiation. A metabolic gene (pyruvate kinase) was chosen to be used as a systematic discriminating factor. A comparison of about 40 pyruvate kinase protein sequences allowed us to synthesize two oligonucleotides that were able to amplify a fragment of this corresponding gene in our strains. Based on these partial pyruvate kinase gene sequences, several clusters could be identified. The second method used in this study was based on 16S rRNA sequences analysis. We compared sequences present in GenBank database, and this allowed to separate bifidobacteria species into different clusters. They were different from those obtained with partial pyruvate kinase gene sequences analysis. So, by combining both methods, we were able to identify our isolates, when only 10% of them could be strictly identified using the 16S rRNA method. Moreover, pyruvate kinase analysis allowed to differentiate very ambivalent groups such as B. animalis/B. lactis or B. infantis/B. longum, but created different clusters for B. infantis species group, questioning on the homogeneity of this species.
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5. |
Rhim SL,
Park MS,
Ji GE,
( 2006 ) Expression and secretion of Bifidobacterium adolescentis amylase by Bifidobacterium longum. PMID : 16489493 : DOI : 10.1007/s10529-005-5330-9 Abstract >>
Bifidobacterium adolescentis Int-57 (INT57), isolated from human feces, secretes an amylase. We have shot-gun cloned, sequence analyzed and expressed the gene encoding this amylase in B. longum. The sequenced 2477 bp fragment was homologous to other extracellular amylases. The encoded protein was predicted to be composed of 595 amino acids with a molecular weight of 64 kDa, and was designated AmyB. Highly conserved amylase domains were found in AmyB. The signal sequence and cleavage site was predicted by sequence analysis. AmyB was subcloned into pBES2, a novel E. coli-Bifidobacterium shuttle vector, to construct pYBamy59. Subsequently, B. longum, with no apparent amylase activity, was transformed with pYBamy59. More than 90% of the amylase activity was detected in the culture broth. This approach may open the way for the development of more efficient expression and secretion systems for Bifidobacterium.
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6. |
Ventura M,
Canchaya C,
Bernini V,
Del Casale A,
Dellaglio F,
Neviani E,
Fitzgerald GF,
van Sinderen D,
( 2005 ) Genetic characterization of the Bifidobacterium breve UCC 2003 hrcA locus. PMID : 16332909 : DOI : 10.1128/AEM.71.12.8998-9007.2005 PMC : PMC1317471 Abstract >>
The bacterial heat shock response is characterized by the elevated expression of a number of chaperone complexes and transcriptional regulators, including the DnaJ and the HrcA proteins. Genome analysis of Bifidobacterium breve UCC 2003 revealed a second copy of a dnaJ gene, named dnaJ2, which is flanked by the hrcA gene in a genetic constellation that appears to be unique to the actinobacteria. Phylogenetic analysis using 53 bacterial dnaJ sequences, including both dnaJ1 and dnaJ2 sequences, suggests that these genes have followed a different evolutionary development. Furthermore, the B. breve UCC 2003 dnaJ2 gene seems to be regulated in a manner that is different from that of the previously characterized dnaJ1 gene. The dnaJ2 gene, which was shown to be part of a 2.3-kb bicistronic operon with hrcA, was induced by osmotic shock but not significantly by heat stress. This induction pattern is unlike those of other characterized dnaJ genes and may be indicative of a unique stress adaptation strategy by this commensal microorganism.
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7. |
Yin X,
Chambers JR,
Barlow K,
Park AS,
Wheatcroft R,
( 2005 ) The gene encoding xylulose-5-phosphate/fructose-6-phosphate phosphoketolase (xfp) is conserved among Bifidobacterium species within a more variable region of the genome and both are useful for strain identification. PMID : 15899413 : DOI : 10.1016/j.femsle.2005.04.013 Abstract >>
The nucleotide sequence of the xfp-gene region in six known and two unknown species of Bifidobacterium was determined and compared with the published sequences of B. animalis subsp. lactis DSM10140 and B. longum biovar longum NCC2705. The xfp coding sequences were 73% identical and coded for 825 amino acids in all 10 sequences. Partial sequences of an adjacent gene, guaA, were 61% identical in six sequences for which data were available. The region between xfp and guaA was variable in both length and sequence. Oligonucleotide sequences from the conserved and variable xfp regions were used as PCR primers, in combinations of appropriate specificity, for the detection and identification of Bifidobacterium isolates.
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8. |
Ventura M,
Zhang Z,
Cronin M,
Canchaya C,
Kenny JG,
Fitzgerald GF,
van Sinderen D,
( 2005 ) The ClgR protein regulates transcription of the clpP operon in Bifidobacterium breve UCC 2003. PMID : 16321946 : DOI : 10.1128/JB.187.24.8411-8426.2005 PMC : PMC1317013 Abstract >>
Five clp genes (clpC, clpB, clpP1, clpP2, and clpX), representing chaperone- and protease-encoding genes, were previously identified in Bifidobacterium breve UCC 2003. In the present study, we characterize the B. breve UCC 2003 clpP locus, which consists of two paralogous genes, designated clpP1 and clpP2, whose deduced protein products display significant similarity to characterized ClpP peptidases. Transcriptional analyses showed that the clpP1 and clpP2 genes are transcribed in response to moderate heat shock as a bicistronic unit with a single promoter. The role of a clgR homologue, known to control the regulation of clpP gene expression in Streptomyces lividans and Corynebacterium glutamicum, was investigated by gel mobility shift assays and DNase I footprint experiments. We show that ClgR, which in its purified form appears to exist as a dimer, requires a proteinaceous cofactor to assist in specific binding to a 30-bp region of the clpP promoter region. In pull-down experiments, a 56-kDa protein copurified with ClgR, providing evidence that the two proteins also interact in vivo and that the copurified protein represents the cofactor required for ClgR activity. The prediction of the ClgR three-dimensional structure provides further insights into the binding mode of this protein to the clpP1 promoter region and highlights the key amino acid residues believed to be involved in the protein-DNA interaction.
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9. |
Kim GB,
Brochet M,
Lee BH,
( 2005 ) Cloning and characterization of a bile salt hydrolase (bsh) from Bifidobacterium adolescentis. PMID : 16086241 : DOI : 10.1007/s10529-005-6717-3 Abstract >>
A gene coding for bile salt hydrolase (BSH) from Bifidobacterium adolescentis was cloned and expressed in Escherichia coli, and the nucleotide sequence was determined. The BSH of E. coli transformants was produced intracellularly in the absence of bile salts. A unique bsh promoter (P(bsh)) sequence was identified by using a Neural Network Promoter Prediction (NNPP, version 2.2). In spite of their high-level sequence homology with other bsh genes in the Bifidobacterium species, their genetic organization surrounding the bsh gene and their promoter sequences are different depending on the species.
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10. |
van den Broek LA,
Lloyd RM,
Beldman G,
Verdoes JC,
McCleary BV,
Voragen AG,
( 2005 ) Cloning and characterization of arabinoxylan arabinofuranohydrolase-D3 (AXHd3) from Bifidobacterium adolescentis DSM20083. PMID : 15650848 : DOI : 10.1007/s00253-004-1850-9 Abstract >>
Arabinoxylan arabinofuranohydrolase-D3 (AXHd3) from Bifidobacterium adolescentis releases only C3-linked arabinose residues from double-substituted xylose residues. A genomic library of B. adolescentis DSM20083 was screened for the presence of the axhD3 gene. Two plasmids were identified containing part of the axhD3 gene. The nucleotide sequences were combined and three open reading frames (ORFs) were found. The first ORF showed high homology with xylanases belonging to family 8 of the glycoside hydrolases and this gene was designated xylA. The second ORF was the axhD3 gene belonging to glycoside hydrolase family 43. The third (partial) ORF coded for a putative carboxylesterase. The axhD3 gene was cloned and expressed in Escherichia coli. Several substrates were employed in the biochemical characterization of recombinant AXHd3. The enzyme showed the highest activity toward wheat arabinoxylan oligosaccharides. In addition, beta-xylanase from Trichoderma sp. was able to degrade soluble wheat arabinoxylan polymer to a higher extent, after pretreatment with recombinant AXHd3. Arabinoxylan oligosaccharides incubated with a combination of recombinant AXHd3 and an alpha-L-arabinofuranosidase from Aspergillus niger did not result in a higher maximal release of arabinose than incubation with these enzymes separately.
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11. |
Ventura M,
Zink R,
Fitzgerald GF,
van Sinderen D,
( 2005 ) Gene structure and transcriptional organization of the dnaK operon of Bifidobacterium breve UCC 2003 and application of the operon in bifidobacterial tracing. PMID : 15640225 : DOI : 10.1128/AEM.71.1.487-500.2005 PMC : PMC544267 Abstract >>
The incorporation and delivery of bifidobacterial strains as probiotic components in many food preparations expose these microorganisms to a multitude of environmental insults, including heat and osmotic stresses. We characterized the dnaK gene region of Bifidobacterium breve UCC 2003. Sequence analysis of the dnaK locus revealed four genes with the organization dnaK-grpE-dnaJ-ORF1, whose deduced protein products display significant similarity to corresponding chaperones found in other bacteria. Northern hybridization and real-time LightCycler PCR analysis revealed that the transcription of the dnaK operon was strongly induced by osmotic shock but was not induced significantly by heat stress. A 4.4-kb polycistronic mRNA, which represented the transcript of the complete dnaK gene region, was detected. Many other small transcripts, which were assumed to have resulted from intensive processing or degradation of this polycistronic mRNA, were identified. The transcription start site of the dnaK operon was determined by primer extension. Phylogenetic analysis of the available bifidobacterial grpE and dnaK genes suggested that the evolutionary development of these genes has been similar. The phylogeny derived from the various bifidobacterial grpE and dnaK sequences is consistent with that derived from 16S rRNA. The use of these genes in bifidobacterial species as an alternative or complement to the 16S rRNA gene marker provides sequence signatures that allow a high level of discrimination between closely related species of this genus.
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12. |
Ventura M,
Canchaya C,
Zink R,
Fitzgerald GF,
van Sinderen D,
( 2004 ) Characterization of the groEL and groES loci in Bifidobacterium breve UCC 2003: genetic, transcriptional, and phylogenetic analyses. PMID : 15466567 : DOI : 10.1128/AEM.70.10.6197-6209.2004 PMC : PMC522111 Abstract >>
The bacterial heat shock response is characterized by the elevated expression of a number of chaperone complexes, including the GroEL and GroES proteins. The groES and groEL genes are highly conserved among eubacteria and are typically arranged as an operon. Genome analysis of Bifidobacterium breve UCC 2003 revealed that the groES and groEL genes are located in different chromosomal regions. The heat inducibility of the groEL and groES genes of B. breve UCC 2003 was verified by slot blot analysis. Northern blot analyses showed that the cspA gene is cotranscribed with the groEL gene, while the groES gene is transcribed as a monocistronic unit. The transcription initiation sites of these two mRNAs were determined by primer extension. Sequence and transcriptional analyses of the region flanking the groEL and groES genes of various bifidobacteria revealed similar groEL-cspA and groES gene units, suggesting a novel genetic organization of these chaperones. Phylogenetic analysis of the available bifidobacterial groES and groEL genes suggested that these genes evolved differently. Discrepancies in the phylogenetic positioning of groES-based trees make this gene an unreliable molecular marker. On the other hand, the bifidobacterial groEL gene sequences can be used as an alternative to current methods for tracing Bifidobacterium species, particularly because they allow a high level of discrimination between closely related species of this genus.
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13. |
Ventura M,
Canchaya C,
van Sinderen D,
Fitzgerald GF,
Zink R,
( 2004 ) Bifidobacterium lactis DSM 10140: identification of the atp (atpBEFHAGDC) operon and analysis of its genetic structure, characteristics, and phylogeny. PMID : 15128574 : DOI : 10.1128/aem.70.5.3110-3121.2004 PMC : PMC404453 Abstract >>
The atp operon is highly conserved among eubacteria, and it has been considered a molecular marker as an alternative to the 16S rRNA gene. PCR primers were designed from the consensus sequences of the atpD gene to amplify partial atpD sequences from 12 Bifidobacterium species and nine Lactobacillus species. All PCR products were sequenced and aligned with other atpD sequences retrieved from public databases. Genes encoding the subunits of the F(1)F(0)-ATPase of Bifidobacterium lactis DSM 10140 (atpBEFHAGDC) were cloned and sequenced. The deduced amino acid sequences of these subunits showed significant homology with the sequences of other organisms. We identified specific sequence signatures for the genus Bifidobacterium and for the closely related taxa Bifidobacterium lactis and Bifidobacterium animalis and Lactobacillus gasseri and Lactobacillus johnsonii, which could provide an alternative to current methods for identification of lactic acid bacterial species. Northern blot analysis showed that there was a transcript at approximately 7.3 kb, which corresponded to the size of the atp operon, and a transcript at 4.5 kb, which corresponded to the atpC, atpD, atpG, and atpA genes. The transcription initiation sites of these two mRNAs were mapped by primer extension, and the results revealed no consensus promoter sequences. Phylogenetic analysis of the atpD genes demonstrated that the Lactobacillus atpD gene clustered with the genera Listeria, Lactococcus, Streptococcus, and Enterococcus and that the higher G+C content and highly biased codon usage with respect to the genome average support the hypothesis that there was probably horizontal gene transfer. The acid inducibility of the atp operon of B. lactis DSM 10140 was verified by slot blot hybridization by using RNA isolated from acid-treated cultures of B. lactis DSM 10140. The rapid increase in the level of atp operon transcripts upon exposure to low pH suggested that the ATPase complex of B. lactis DSM 10140 was regulated at the level of transcription and not at the enzyme assembly step.
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14. |
Ventura M,
Canchaya C,
Meylan V,
Klaenhammer TR,
Zink R,
( 2003 ) Analysis, characterization, and loci of the tuf genes in lactobacillus and bifidobacterium species and their direct application for species identification. PMID : 14602655 : DOI : 10.1128/aem.69.11.6908-6922.2003 PMC : PMC262312 Abstract >>
We analyzed the tuf gene, encoding elongation factor Tu, from 33 strains representing 17 Lactobacillus species and 8 Bifidobacterium species. The tuf sequences were aligned and used to infer phylogenesis among species of lactobacilli and bifidobacteria. We demonstrated that the synonymous substitution affecting this gene renders elongation factor Tu a reliable molecular clock for investigating evolutionary distances of lactobacilli and bifidobacteria. In fact, the phylogeny generated by these tuf sequences is consistent with that derived from 16S rRNA analysis. The investigation of a multiple alignment of tuf sequences revealed regions conserved among strains belonging to the same species but distinct from those of other species. PCR primers complementary to these regions allowed species-specific identification of closely related species, such as Lactobacillus casei group members. These tuf gene-based assays developed in this study provide an alternative to present methods for the identification for lactic acid bacterial species. Since a variable number of tuf genes have been described for bacteria, the presence of multiple genes was examined. Southern analysis revealed one tuf gene in the genomes of lactobacilli and bifidobacteria, but the tuf gene was arranged differently in the genomes of these two taxa. Our results revealed that the tuf gene in bifidobacteria is flanked by the same gene constellation as the str operon, as originally reported for Escherichia coli. In contrast, bioinformatic and transcriptional analyses of the DNA region flanking the tuf gene in four Lactobacillus species indicated the same four-gene unit and suggested a novel tuf operon specific for the genus Lactobacillus.
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15. |
Omori T,
Ueno K,
Muramatsu K,
Kikuchi M,
Onodera S,
Shiomi N,
( 2010 ) Characterization of recombinant beta-fructofuranosidase from Bifidobacterium adolescentis G1. PMID : 20380746 : DOI : 10.1186/1752-153X-4-9 PMC : PMC2873357 Abstract >>
We have previously reported on purification and characterization of beta-fructofuranosidase (beta-FFase) from Bifidobacterium adolescentis G1. This enzyme showed high activity of hydrolysis on fructo-oligosaccharides with a low degree of polymerization. Recently, genome sequences of B. longum NCC2705 and B. adolescentis ATCC 15703 were determined, and cscA gene in the both genome sequences encoding beta-FFase was predicted. Here, cloning of cscA gene encoding putative beta-FFase from B. adolescentis G1, its expression in E. coli and properties of the recombinant protein are described. Using the information of cscA gene from Bifidobacterium adolescentis ATCC 15703, cscA gene from B. adolescentis G1 was cloned and sequenced. The N-terminal amino acid sequence of purified beta-FFase from B. adolescentis G1 was identical to the deduced amino acid sequences of cscA gene from B. adolescentis G1. To confirm the translated product of the cscA gene, the recombinant protein was expressed in Escherichia coli. Molecular mass of the purified recombinant enzyme was estimated to be about 66,000 by SDS-PAGE and 60,300 by MALDI TOF-MS. The optimum pH of the enzyme was 5.7 and the enzyme was stable at pH 5.0-8.6. The thermostability of the enzyme was up to 50 degrees C. The K(m) (mM), Vmax (micromol/mg of protein/min), k0 (sec(-1)) and k0/K(m)(mM(-1) sec(-1)) for 1-kestose, neokestose, nystose, fructosylnystose, sucrose and inulin were 1.7, 107, 107.5, 63.2, and 1.7, 142, 142.7, 83.9, and 3.9, 152, 152.8, 39.2, and 2.2, 75, 75.4, 34.3, and 38, 79, 79.4, 2.1, and 25.9, 77, 77.4, 3.0, respectively. The hydrolytic activity was strongly inhibited by AgNO3, SDS, and HgCl2. The recombinant enzyme had similar specificity to the native enzyme, high affinity for 1-kestose, and low affinity for sucrose and inulin, although properties of the recombinant enzyme showed slight difference from those of the native one previously described.
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16. |
Kim MS,
Roh SW,
Bae JW,
( 2010 ) Bifidobacterium stercoris sp. nov., isolated from human faeces. PMID : 20081020 : DOI : 10.1099/ijs.0.019943-0 DOI : 10.1099/ijs.0.019943-0 Abstract >>
Strain Eg1(T), an anaerobic, Gram-stain-positive, non-motile and non-spore-forming bacterium, was isolated from human faeces. The optimal temperature for growth was 37 �XC and tests for oxidase and catalase activities gave negative results. Fructose-6-phosphate phosphoketolase activity was detected. Acid was produced during fermentation of several substrates, including glucose. The end products of glucose fermentation were acetic acid and lactic acid, which were produced in a molar ratio of 1.76 : 1 (approximately 3 : 2). The G+C content was 57.8 mol%. Comparative analysis of 16S rRNA gene sequences showed that strain Eg1(T) was closely related to Bifidobacterium adolescentis YIT 4011(T) (98.36 % 16S rRNA gene sequence similarity) and Bifidobacterium ruminantium JCM 8222(T) (97.93 %) and analysis of hsp60 sequences showed that strain Eg1(T) was closely related to B. adolescentis JCM 1275(T) (99.35 % hsp60 sequence similarity) and B. ruminantium JCM 8222(T) (92.13 %). However, despite these degrees of similarity being high enough for strain Eg1(T) to be included at the same species level as B. adolescentis and B. ruminantium (96.5-100 % for the genus Bifidobacterium), the isolate could be distinguished from B. adolescentis KCTC 3216(T) and B. ruminantium KCTC 3425(T) by low levels of DNA-DNA relatedness (41 and 17 %, respectively). Based on phenotypic, genotypic and phylogenetic analyses, we propose that strain Eg1(T) is classified in a novel species, Bifidobacterium stercoris sp. nov. The type strain is Eg1(T) (=KCTC 5756(T) =JCM 15918(T)).
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17. |
Vitali B,
Turroni S,
Serina S,
Sosio M,
Vannini L,
Candela M,
Guerzoni ME,
Brigidi P,
( 2008 ) Molecular and phenotypic traits of in-vitro-selected mutants of Bifidobacterium resistant to rifaximin. PMID : 18462927 : DOI : 10.1016/j.ijantimicag.2008.02.002 Abstract >>
Nucleotide mutations inside a core region of the rpoB gene, encoding the beta subunit of RNA polymerase, were found in rifaximin-resistant mutants of Bifidobacterium. Five different missense mutations of codons 513, 516, 522 and 529 were identified. Further aspects of rifaximin resistance were investigated, using Bifidobacterium infantis BI07 as a model strain. Partial resistance of RNA polymerase of a BI07 mutant at a rifaximin concentration >10 microg/mL was observed by cell-free transcription assay. Mass spectrometry detection of rifaximin in the cellular pellet of the BI07 resistant mutant, as well as changes in biosynthesis of saturated and cyclopropane fatty acids during growth, suggested a reduction in membrane permeability for the antibiotic moiety.
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18. |
Mättö J,
Maukonen J,
Alakomi HL,
Suihko ML,
Saarela M,
( 2008 ) Influence of oral doxycycline therapy on the diversity and antibiotic susceptibility of human intestinal bifidobacterial population. PMID : 18397263 : DOI : 10.1111/j.1365-2672.2008.03792.x Abstract >>
To evaluate the influence of doxycycline therapy on the composition and antibiotic susceptibility of intestinal bifidobacteria. Faecal samples were collected from nine subjects receiving doxycycline therapy and ten control subjects, and analysed for bifidobacteria by culturing and PCR-DGGE (denaturing gradient gel electrophoresis). A marked decrease in the diversity (average number of amplicons detected by PCR-DGGE 0.8 in the antibiotic vs 4.3 in the control group) of Bifidobacterium populations was observed during doxycycline therapy. The proportion of a tetracycline-resistant bifidobacterial population was higher in the antibiotic group than in the control group (83%vs 26%). Based on the tet gene PCR, resistance could be associated with the presence of tet(W). In two subjects, strains representing highly similar genetic fingerprints but different tetracycline susceptibilities were detected. A mutation causing lack of functionality in the tet(W) was observed in one of the susceptible strains. Doxycycline therapy had a drastic effect on the diversity and tetracycline susceptibility of intestinal Bifidobacterium populations. The use of broad-spectrum antibiotics increased the pool of tetracycline-resistant commensal bacteria in the intestine. The detection of resistance genes alone is not sufficient for the evaluation of bacterial antibiotic resistance.
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19. |
Duranti S,
Milani C,
Lugli GA,
Mancabelli L,
Turroni F,
Ferrario C,
Mangifesta M,
Viappiani A,
Sánchez B,
Margolles A,
van Sinderen D,
Ventura M,
( 2016 ) Evaluation of genetic diversity among strains of the human gut commensal Bifidobacterium adolescentis. PMID : 27035119 : DOI : 10.1038/srep23971 PMC : PMC4817515 Abstract >>
Bifidobacteria are members of the human gut microbiota, being numerically dominant in the colon of infants, while also being prevalent in the large intestine of adults. In this study, we determined and analyzed the pan-genome of Bifidobacterium adolescentis, which is one of many bacteria found in the human adult gut microbiota. In silico analysis of the genome sequences of eighteen B. adolescentis strains isolated from various environments, such as human milk, human feces and bovine rumen, revealed a high level of genetic variability, resulting in an open pan-genome. Compared to other bifidobacterial taxa such as Bifidobacterium bifidum and Bifidobacterium breve, the more extensive B. adolescentis pan-genome supports the hypothesis that the genetic arsenal of this taxon expanded so as to become more adaptable to the variable and changing ecological niche of the gut. These increased genetic capabilities are particularly evident for genes required for dietary glycan-breakdown.
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20. |
Duranti S,
Turroni F,
Lugli GA,
Milani C,
Viappiani A,
Mangifesta M,
Gioiosa L,
Palanza P,
van Sinderen D,
Ventura M,
( 2014 ) Genomic characterization and transcriptional studies of the starch-utilizing strain Bifidobacterium adolescentis 22L. PMID : 25063659 : DOI : 10.1128/AEM.01993-14 PMC : PMC4178675 Abstract >>
Bifidobacteria are members of the gut microbiota, but the genetic basis for their adaptation to the human gut is poorly understood. The analysis of the 2,203,222-bp genome of Bifidobacterium adolescentis 22L revealed a nutrient acquisition strategy that targets diet/plant-derived glycans, in particular starch and starch-like carbohydrates. Starch-like carbohydrates were shown to support the growth of B. adolescentis 22L. Transcriptome profiling of 22L cultures grown under in vitro conditions or during colonization of the murine gut by RNA sequencing and quantitative real-time PCR assays revealed the expression of a set of chromosomal loci responsible for starch metabolism as well as for pilus production. Such extracellular structures include so-called sortase-dependent and type IVb pili, which may be involved in gut colonization of 22L through adhesion to extracellular matrix proteins.
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21. |
Killer J,
Sedlá?ek I,
Rada V,
Havlík J,
Kope?ný J,
( 2013 ) Reclassification of Bifidobacterium stercoris Kim et al. 2010 as a later heterotypic synonym of Bifidobacterium adolescentis. PMID : 24187022 : DOI : 10.1099/ijs.0.054957-0 DOI : 10.1099/ijs.0.054957-0 Abstract >>
The taxonomic position of Bifidobacterium stercoris Eg1(T) (= JCM 15918(T)) based on comparative 16S rRNA gene and hsp60 sequence analyses was found to be controversial, as the strain showed high similarity to the type strain of Bifidobacterium adolescentis, CCUG 18363(T). Therefore, the relationship between the two species was investigated by a taxonomic study that included, in addition to re-evaluation of the 16S rRNA gene sequence, determination of DNA-DNA binding and multilocus sequence analysis (MLSA) of housekeeping genes encoding the DNA-directed RNA polymerase B subunit (rpoC), putative xylulose-5-phosphate/fructose-6-phosphate phosphoketolase (xfp), elongation factor EF-G (fusA), 50S ribosomal protein L2 (rplB) and DNA gyrase B subunit (gyrB). Comparative 16S rRNA gene sequence analysis showed relatively high similarity (98.9 %) between B. stercoris KCTC 5756(T) and B. adolescentis ATCC 15703(T). MLSA revealed close relatedness between B. stercoris KCTC 5756(T) and B. adolescentis CCUG 18363(T), with 99.3-100 % similarity between the rpoC, xfp, fusA, rplB and gyrB gene sequences. In addition, relatively high dnaJ1 gene sequence similarity of 97.7 % was found between the strains. Similar phenotypes and a high DNA-DNA binding value (78.9 %) confirmed that B. stercoris and B. adolescentis are synonymous. Based on these results, it is proposed that the species Bifidobacterium stercoris Kim et al. 2010 should be reclassified as a later heterotypic synonym of Bifidobacterium adolescentis Reuter 1963 (Approved Lists 1980).
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22. |
( 1997 ) Evaluation of using a short region of the recA gene for rapid and sensitive speciation of dominant bifidobacteria in the human large intestine. PMID : 9311137 : DOI : 10.1111/j.1574-6968.1997.tb12670.x Abstract >>
The feasibility of intragenerically characterizing bifidobacteria by a comparison of a short region within the recA gene was tested. An approximately 300 bp fragment of the recA gene was PCR-amplified from six species from the genus Bifidobacterium using primers directed to two universally conserved regions of the recA gene. A phylogenetic analysis of the sequenced recA products compared favorably to classification based on the 16S rRNA sequences of the species tested. To apply this rapid methodology to unknown human intestinal bifidobacteria, 46 isolates were randomly chosen from the feces of four subjects and initially characterized by RFLP analysis of a PCR-amplified region of their 16S RNA genes. From a representative of the dominant RFLP family in each of the subjects, the recA segment was PCR-amplified, sequenced and phylogenetically analyzed. All four isolates were found to be related to one another and to B. longum and B. infantis. These results illustrate that the recA gene may be useful for intrageneric phylogenetic analysis as well as for the identification of unknown fecal bifidobacteria.
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23. |
Kawai Y,
Konishi H,
Horitsu H,
Sakurai H,
Takamizawa K,
Suzuki T,
Kawai K,
( 1994 ) Purification and characterization of D-xylose isomerase from Bifidobacterium adolescentis. PMID : 7764860 : DOI : 10.1271/bbb.58.691 Abstract >>
D-Xylose isomerase was purified to homogeneity from cell-free extracts of Bifidobacterium adolescentis by ammonium sulfate fractionation and chromatographies on DEAE-cellulose and Butyl-Toyopearl. The molecular weight of the purified enzyme was estimated to be 168,000 by gel filtration on TSKgel G-3000SW, and 53,000 on SDS-polyacrylamide gel electrophoresis. The optimum pH was around 7 and the enzyme was stable at pH 7-8. The enzyme required bivalent cations, Mg2+, Co2+, or Mn2+ for the activity, particularly Mn2+ to be best. The enzyme had a pI of 4.3, and the Km for D-xylose was 4 mM. The N-terminal amino acid sequence of the enzyme was not similar to those of D-xylose isomerases from other sources such as Clostridium thermosulfurogenes, Escherichia coli, or Bacillus subtilis.
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( 2013 ) Genetic diversity of bile salt hydrolases among human intestinal bifidobacteria. PMID : 23591474 : DOI : 10.1007/s00284-013-0362-1 PMC : PMC3722454 Abstract >>
This study analyzes the application of degenerative primers for the screening of bile salt hydrolase-encoding genes (bsh) in various intestinal bifidobacteria. In the first stage, the design and evaluation of the universal PCR primers for amplifying the partial coding sequence of bile salt hydrolase in bifidobacteria were performed. The amplified bsh gene fragments were sequenced and the obtained sequences were compared to the bsh genes present in GenBank. The determined results showed the utility of the designed PCR primers for the amplification of partial gene encoding bile salt hydrolase in different intestinal bifidobacteria. Moreover, sequence analysis revealed that bile salt hydrolase-encoding genes may be used as valuable molecular markers for phylogenetic studies and identification of even closely related members of the genus Bifidobacterium.
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25. |
( 2013 ) Bifidobacterial succession and correlation networks in a large unselected cohort of mothers and their children. PMID : 23124244 : DOI : 10.1128/AEM.02359-12 PMC : PMC3553782 Abstract >>
Bifidobacteria are a major microbial component of infant gut microbiota, which is believed to promote health benefits for the host and stimulate maturation of the immune system. Despite their perceived importance, very little is known about the natural development of and possible correlations between bifidobacteria in human populations. To address this knowledge gap, we analyzed stool samples from a randomly selected healthy cohort of 87 infants and their mothers with >90% of vaginal delivery and nearly 100% breast-feeding at 4 months. Fecal material was sampled during pregnancy, at 3 and 10 days, at 4 months, and at 1 and 2 years after birth. Stool samples were predicted to be rich in the species Bifidobacterium adolescentis, B. bifidum, B. dentium, B. breve, and B. longum. Due to high variation, we did not identify a clear age-related structure at the individual level. Within the population as a whole, however, there were clear age-related successions. Negative correlations between the B. longum group and B. adolescentis were detected in adults and in 1- and 2-year-old children, whereas negative correlations between B. longum and B. breve were characteristic for newborns and 4-month-old infants. The highly structured age-related development of and correlation networks between bifidobacterial species during the first 2 years of life mirrors their different or competing nutritional requirements, which in turn may be associated with specific biological functions in the development of healthy gut.
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