314 results on '"Harry J. Flint"'
Search Results
252. Inhibitory activity of gut bacteria against Escherichia coli O157 mediated by dietary plant metabolites
- Author
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Sylvia H. Duncan, Colin S. Stewart, and Harry J. Flint
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Rumen ,Colon ,Swine ,Biology ,medicine.disease_cause ,Umbelliferone ,Escherichia coli O157 ,Microbiology ,chemistry.chemical_compound ,Bacteria, Anaerobic ,Coumarins ,Scopoletin ,Bone plate ,Antibiosis ,Genetics ,medicine ,Animals ,Humans ,heterocyclic compounds ,Anaerobiosis ,Umbelliferones ,Molecular Biology ,Escherichia coli ,chemistry.chemical_classification ,Sheep ,Hydrolysis ,Glycoside ,Plants ,equipment and supplies ,biology.organism_classification ,Fatty Acids, Volatile ,Enterobacteriaceae ,Aerobiosis ,Culture Media ,Diet ,Esculin ,chemistry ,Biochemistry ,Digestive System ,Bacteria - Abstract
Under both aerobic and anaerobic conditions, the growth of Escherichia coli O157 strain NCTC 12,900 was inhibited by the coumarins esculetin, umbelliferone and scopoletin, but not by the coumarin glycoside esculin. Esculin-hydrolysing bacteria from the rumen, the pig gut and the human gut inhibited growth of E. coli in an overlay-plate assay in the presence of esculin. The combined effect of esculetin and volatile fatty acids was greater than the effect of either factor alone suggesting that coumarin glycosides in the diet might reduce the growth or survival of E. coli O157 in the gut. Adding esculin to incubations of mixed rumen contents significantly reduced the survival of E. coli O157.
- Published
- 1998
253. A cysteine desulphurase gene from the cellulolytic rumen anaerobe Ruminococcus flavefaciens
- Author
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James Kirby, Harry J. Flint, and Frank Wright
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Rumen ,Molecular Sequence Data ,Biophysics ,Lyases ,Cellulase ,medicine.disease_cause ,Biochemistry ,Microbiology ,Gene product ,Evolution, Molecular ,Bacteria, Anaerobic ,Bacterial Proteins ,Structural Biology ,medicine ,Animals ,Amino Acid Sequence ,Cysteine ,Molecular Biology ,Escherichia coli ,Gene ,biology ,Sequence Homology, Amino Acid ,Cysteine desulfurase ,biology.organism_classification ,Gram-Positive Cocci ,Carbon-Sulfur Lyases ,Genes, Bacterial ,biology.protein ,bacteria ,Clostridium thermocellum ,Bacteria - Abstract
A gene whose predicted product shows 40–50% sequence identity with the products of nifS genes from nitrogen-fixing bacteria was found downstream from a cellulase gene in a DNA fragment from the cellulolytic rumen anaerobe, Ruminococcus flavefaciens 17. The R. flavefaciens gene product released sulphur from l -cysteine when expressed in Escherichia coli , indicating that the R. flavefaciens NifS enzyme may play a role in sulphuration, perhaps, as in nitrogen-fixing bacteria, supplying sulphur to FeS proteins. Sequences hybridising with the R. flavefaciens 17 nifS -like gene were also detected in R. flavefaciens 007 and in R. albus SY3.
- Published
- 1998
254. Plant cell wall degrading enzyme complexes from the cellulolytic rumen bacterium Ruminococcus flavefaciens
- Author
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Vincenzo Aurilia, Jennifer C. Martin, Sheila I. McCrae, James Kirby, and Harry J. Flint
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chemistry.chemical_classification ,Rumen ,biology ,Chemistry ,Ruminococcus ,Molecular Sequence Data ,Ruminococcus flavefaciens ,biology.organism_classification ,Gram-Positive Bacteria ,Biochemistry ,Microbiology ,Cell wall ,Xylan Endo-1,3-beta-Xylosidase ,Enzyme ,Xylosidases ,Cellulase ,Cell Wall ,Xylanase ,Animals ,Amino Acid Sequence ,Rumen microorganisms ,Cellulose ,Bacteria - Published
- 1998
255. Isolation and overexpression of a gene encoding an extracellular beta-(1,3-1,4)-glucanase from Streptococcus bovis JB1
- Author
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Sheila I. McCrae, Harry J. Flint, and M S Ekinci
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DNA, Bacterial ,Endo-1,3(4)-beta-Glucanase ,Glycoside Hydrolases ,Genetic Vectors ,Molecular Sequence Data ,Gene Expression ,Biology ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,Enterococcus faecalis ,Microbiology ,Gene expression ,medicine ,Extracellular ,Glycoside hydrolase ,Amino Acid Sequence ,Cloning, Molecular ,Escherichia coli ,Ecology ,Base Sequence ,Sequence Homology, Amino Acid ,Lactococcus lactis ,Glucanase ,Streptococcus bovis ,biology.organism_classification ,Molecular biology ,Recombinant Proteins ,Genes, Bacterial ,Extracellular Space ,Food Science ,Biotechnology ,Plasmids ,Research Article - Abstract
Streptococcus bovis JB1 was found to produce a 25-kDa extracellular enzyme active against beta-(1,3-1,4)-glucans. A gene was isolated encoding a specific beta-(1,3-1,4)-glucanase that corresponds to this size and belongs to glycoside hydrolase family 16. A 4- to 10-fold increase in supernatant beta-glucanase activity was obtained when the cloned beta-glucanase gene was reintroduced into S. bovis JB1 by use of constructs based on the plasmid vector pTRW10 or pIL253. The beta-(1,3-1,4)-glucanase gene was also expressed upon introduction of the pTRW10 construct pTRWL1R into Lactococcus lactis IL2661 and Enterococcus faecalis JH2-SS, although extracellular activity was 8- to 50-fold lower than that in S. bovis JB1. The beta-(1,3-1,4)-glucanase purified from the culture supernatant of S. bovis JB1 carrying pTRWL1R showed a K(m) of 2.8 mg per ml and a Vmax of 338 mumol of glucose equivalents per min per mg of protein with barley beta-glucan as the substrate. The S. bovis beta-(1,3-1,4)-glucanase may contribute to the ability of this bacterium to utilize starch by degrading structural polysaccharides present in endosperm cell walls.
- Published
- 1997
256. Degradation and utilization of xylans by the rumen anaerobe Prevotella bryantii (formerly P. ruminicola subsp. brevis) B(1)4
- Author
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Kohji Miyazaki, Romana Marinšek-Logar, Jennifer C. Martin, and Harry J. Flint
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chemistry.chemical_classification ,Arabinose ,animal structures ,biology ,Pentose ,Prevotella ruminicola ,Xylose ,biology.organism_classification ,Microbiology ,Xylan ,chemistry.chemical_compound ,Infectious Diseases ,Biochemistry ,chemistry ,Xylanase ,Energy source ,Prevotella bryantii - Abstract
Freshly harvested whole cells from cultures of P. bryantii B(1)4 grown with oat spelt xylan (OSX) as an energy source showed less than 25% of the enzyme activity against OSX, and less than 15% of the activity against birchwood xylan (BWX) and carboxymethylcellulose, that was detectable in sonicated cell preparations. This indicates that much of this hydrolytic activity is either periplasmic, membrane-associated or intracellular and may be concerned with the processing of transported oligosaccharides.P. bryantii B(1)4 cultures were able to utilise up to 45% and 51% of the total pentose present in OSX and BWX, respectively, after 24 h, but could utilize 84% of a water-soluble fraction of BWX. Analysis of the xylan left undegraded after incubation with P. bryantii showed that while xylose and arabinose were removed to a similar extent, uronic acids were utilized to a greater extent than xylose. Predigestion of xylans with two cloned xylanases from the cellulolytic rumen anaerobe Ruminococcus flavefaciens gave little increase in overall pentose utilization suggesting that external P. bryantii xylanases are as effective as the cloned R. flavefaciens enzymes in releasing products that can be utilised by P. bryantii cells. The xylanase system of P. bryantiiis able to efficiently utilise not only xylo-oligosaccharides but also larger water-soluble xylan fragments.
- Published
- 1997
257. Interrupted catalytic domain structures in xylanases from two distantly related strains of Prevotella ruminicola
- Author
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Ales Gasparic, Terence R. Whitehead, Harry J. Flint, and Jennifer C. Martin
- Subjects
DNA, Bacterial ,animal structures ,Rumen ,Molecular Sequence Data ,Biophysics ,Prevotella ,Prevotella ruminicola ,Biochemistry ,Structural Biology ,Animals ,Amino Acid Sequence ,Binding site ,Molecular Biology ,Peptide sequence ,chemistry.chemical_classification ,Binding Sites ,biology ,Base Sequence ,Molecular Structure ,biology.organism_classification ,Amino acid ,Xylan Endo-1,3-beta-Xylosidase ,Enzyme ,Xylosidases ,chemistry ,Genes, Bacterial ,Xylanase ,Bacteria - Abstract
Two xylanases from the rumen anaerobic bacterium Prevotella ruminicola were found to possess highly unusual structures in which family 10 catalytic domains are interrupted by unrelated sequences. XynC from P. ruminicola B(1)4 carries a 160 amino-acid insertion, while a P. ruminicola D31d xylanase carries an unrelated region of 280 amino acids, containing an imperfect 130 amino-acid duplication. Both regions of family 10 similarity were shown to be essential for activity of the D31d enzyme.
- Published
- 1997
258. A 16S rDNA-based molecular profiling approach for studying relative changes in ruminal bacterial populations
- Author
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G. Avguštin, F. McIntosh, Harry J. Flint, Karen P. Scott, C. J. Newbold, J. Wood, and Revues Inra, Import
- Subjects
[SDV.AEN] Life Sciences [q-bio]/Food and Nutrition ,business.industry ,[SDV.BDD] Life Sciences [q-bio]/Development Biology ,Profiling (information science) ,Computational biology ,Biology ,16S ribosomal RNA ,business ,[SDV.BDLR] Life Sciences [q-bio]/Reproductive Biology ,Biotechnology - Published
- 1997
259. The rumen bacteria
- Author
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Marvin P. Bryant, Colin S. Stewart, and Harry J. Flint
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Rumen ,biology ,Obligate ,Microorganism ,Food science ,Selenomonas ruminantium ,Ribosomal RNA ,biology.organism_classification ,Desulfovibrio ,Bacteria ,Microbiology ,Butyrivibrio fibrisolvens - Abstract
This chapter will deal mainly with the characteristics of bacteria from the rumen that have been successfully cultivated in the laboratory. For some ecosystems, particularly those dominated by slow-growing or specialized microorganisms, it has become clear that only a very small fraction (often
- Published
- 1997
260. The fate of Escherichia coli 0157 isolates under simulated rumen conditions and the use of a gfp-labelled isolate for ecological studies
- Author
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S. H. Duncan, TH Yennington, C. S. Stewart, Harry J. Flint, F. Thompson-Carter, Karen P. Scott, and Revues Inra, Import
- Subjects
Rumen ,[SDV.AEN] Life Sciences [q-bio]/Food and Nutrition ,[SDV.BDD] Life Sciences [q-bio]/Development Biology ,medicine ,Biology ,medicine.disease_cause ,Escherichia coli ,[SDV.BDLR] Life Sciences [q-bio]/Reproductive Biology ,Microbiology ,Green fluorescent protein - Published
- 1997
261. Expression of Cellulosome Components and Type IV Pili within the Extracellular Proteome of Ruminococcus flavefaciens 007
- Author
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Raphael Lamed, Bryan A. White, Martin D. Reid, Keith Turner, Louise Cantlay, Romana Marinšek-Logar, Carl J. Yeoman, Julian Parkhill, Margret E. Berg Miller, Edward A. Bayer, Harry J. Flint, Sylvia H. Duncan, and Maša Vodovnik
- Subjects
animal structures ,Proteome ,Applied Microbiology ,Veterinary Microbiology ,lcsh:Medicine ,Library science ,Biology ,Biochemistry ,Protein Chemistry ,Microbiology ,7. Clean energy ,Bacterial protein ,03 medical and health sciences ,Bacterial Proteins ,Molecular Cell Biology ,Ruminococcus ,Cellulose ,lcsh:Science ,030304 developmental biology ,0303 health sciences ,Multidisciplinary ,Cellulose metabolism ,030306 microbiology ,Rubredoxins ,lcsh:R ,Proteins ,Agriculture ,Bacteriology ,Gene Expression Regulation, Bacterial ,Ruminococcus flavefaciens ,Sustainable Agriculture ,Hemerythrin ,Bacterial Biochemistry ,Cellulosomes ,Chemistry ,Scholarship ,Fimbriae, Bacterial ,Multigene Family ,Veterinary Science ,lcsh:Q ,Extracellular Space ,Research Article ,Biotechnology - Abstract
BACKGROUND: Ruminococcus flavefaciens is an important fibre-degrading bacterium found in the mammalian gut. Cellulolytic strains from the bovine rumen have been shown to produce complex cellulosome structures that are associated with the cell surface. R. flavefaciens 007 is a highly cellulolytic strain whose ability to degrade dewaxed cotton, but not Avicel cellulose, was lost following initial isolation in the variant 007S. The ability was recovered after serial subculture to give the cotton-degrading strain 007C. This has allowed us to investigate the factors required for degradation of this particularly recalcitrant form of cellulose. METHODOLOGY/PRINCIPAL FINDINGS: The major proteins associated with the bacterial cell surface and with the culture supernatant were analyzed for R. flavefaciens 007S and 007C grown with cellobiose, xylan or Avicel cellulose as energy sources. Identification of the proteins was enabled by a draft genome sequence obtained for 007C. Among supernatant proteins a cellulosomal GH48 hydrolase, a rubrerthyrin-like protein and a protein with type IV pili N-terminal domain were the most strongly up-regulated in 007C cultures grown on Avicel compared with cellobiose. Strain 007S also showed substrate-related changes, but supernatant expression of the Pil protein and rubrerythrin in particular were markedly lower in 007S than in 007C during growth on Avicel. CONCLUSIONS/SIGNIFICANCE: This study provides new information on the extracellular proteome of R. flavefaciens and its regulation in response to different growth substrates. Furthermore it suggests that the cotton cellulose non-degrading strain (007S) has altered regulation of multiple proteins that may be required for breakdown of cotton cellulose. One of these, the type IV pilus was previously shown to play a role in adhesion to cellulose in R. albus, and a related pilin protein was identified here for the first time as a major extracellular protein in R. flavefaciens.
- Published
- 2013
262. Tu1716 Evaluation of the Impact of Different Commercially Available DNA Extraction Kits and Laboratories for Assessing Bacterial Community Structure in Fecal Samples - Implications for IBD Research
- Author
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UK Ibd Microbiota Consortia, Georgina L. Hold, Susan H. Berry, Freda M. Farquharson, Harry J. Flint, Petra Louis, Jack Satsangi, UK Ibd Genetics Consortia, Charlie W. Lees, Nicholas A. Kennedy, Alan W. Walker, and Sylvia H. Duncan
- Subjects
Hepatology ,biology ,Sequence analysis ,Lachnospiraceae ,Gastroenterology ,biology.organism_classification ,DNA extraction ,Microbiology ,chemistry.chemical_compound ,chemistry ,Pyrosequencing ,Bacteroides ,DNA ,Feces ,Ruminococcaceae - Abstract
Introduction Determining faecal sample bacterial community structure through sequence analysis of DNA has become a very important facet of inflammatory bowel disease (IBD) research. The possible impact of different commercially available DNA extraction kits and the influence of different lab environments on the data generated has however received relatively little attention. The study compared bacterial communities in faecal samples extracted using commercial DNA kits in established gastrointestinal research laboratories. Methods Faecal samples from 2 healthy volunteers and 2 IBD patients with relapse were investigated. DNA extraction was undertaken using MoBio and Fastprep DNA extraction kits in two established labs. Two protocols were followed for processing samples using the Fastprep kit. Each DNA sample was then split and an aliquot transferred to the other lab. Pyrosequencing PCR of bacterial 16S rRNA genes was performed in both labs on all samples. Quantitative PCR analysis (q-PCR) to validate sequencing data was also performed. Hierarchical clustering was done using the Jaccard and Theta Yue & Clayton similarity coefficients on the pyrosequencing data. Results DNA extracted using methods FastPrep1, FastPrep2 and the MoBio kit yielded median DNA concentrations of 476 (interquartile range 290–518), 453 (IQR 228–689) and 22 (IQR 9–36) ng/µL respectively. Those obtained with MoBio were significantly lower than FastPrep (p Ruminococcaceae and Bacteroidetes were significantly increased in MoBio extracted samples, while Lachnospiraceae and Enterobacteriaceae were significantly reduced (p Enterobacteriaceae , Bacteroides , Ruminococcaceae and Lachnospiraceae respectively. Conclusion This study demonstrates significant differences in DNA yield and bacterial DNA composition seen when comparing DNA extracted from the same faecal sample with different extraction kits. This highlights the importance of ensuring that all samples to be analysed together are prepared with the same DNA extraction method, and the need for caution when comparing studies that have used different methods. Disclosure of Interest None Declared
- Published
- 2013
263. P705 Evaluation of the impact of different commercially available DNA extraction kits and laboratories for assessing bacterial community structure in fecal samples – implications for IBD research
- Author
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Harry J. Flint, Georgina L. Hold, Charlie W. Lees, Susan H. Berry, Alan W. Walker, Nicholas A. Kennedy, F. Farquarson, Petra Louis, Sylvia H. Duncan, and Jack Satsangi
- Subjects
medicine.medical_specialty ,business.industry ,Gastroenterology ,General Medicine ,Stool specimen ,medicine.disease ,Inflammatory bowel disease ,DNA extraction ,Internal medicine ,Immunology ,medicine ,business ,Feces ,Irritable bowel syndrome - Published
- 2013
264. Apoplastic expression of the xylanase and ?(1?3, 1?4) glucanase domains of the xyn D gene from Ruminococcus flavefaciens leads to functional polypeptides in transgenic tobacco plants
- Author
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Uwe Sonnewald, Harry J. Flint, and Karin Herbers
- Subjects
Expression vector ,Transgene ,fungi ,food and beverages ,Plant physiology ,Plant Science ,Genetically modified crops ,Biology ,Glucanase ,Apoplast ,Biochemistry ,Genetics ,Xylanase ,Agronomy and Crop Science ,Molecular Biology ,Gene ,Biotechnology - Abstract
We are investigating the possibilities of transgenic plants as bioreactors for the production of industrial enzymes using cell wall-hydrolysing enzymes as first examples. Within the frame work of this work two distinct domains of the xynD gene from Ruminococcus flavefaciens encoding a xylanase (XYLD-A) and a β(1–3, 1–4)glucanase (XYLD-C) were separately cloned into a plant expression vector which would target the proteins into the apoplast. Transgenic tobacco plants were obtained expressing xylan-hydrolysing as well as lichenan-hydrolysing activities. Despite similar steady-state levels of the respective mRNAs xylan hydrolysis rates were between 40 and 170 μmol min−1 m−2 leaf area depending on the transgenic plant while β(1–3, 1–4)glucan degradation was much more effective ranging between 200 and 2000 μmol min−1 m−2. The high activity levels of the XYLD-C expressing plants were reflected on the protein level. XYLD-C accumulated in the intercellular space and was one of the most prominent bands in protein gels. Despite their apoplastic location as confirmed by activity measurements using intercellular fluids the transgenic plants had not undergone any phenotypic alteration.
- Published
- 1996
265. Heterologous expression of an endoglucanase gene (endA) from the ruminal anaerobe Ruminococcus flavefaciens 17 in Streptococcus bovis and Streptococcus sanguis
- Author
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Terence R. Whitehead and Harry J. Flint
- Subjects
Gene Expression ,medicine.disease_cause ,Microbiology ,Shuttle vector ,Bacterial Proteins ,Cellulase ,Genetics ,medicine ,Molecular Biology ,Peptococcaceae ,Escherichia coli ,Endodeoxyribonucleases ,biology ,Streptococcus ,Ruminococcus ,Gene Transfer Techniques ,Membrane Proteins ,biology.organism_classification ,Streptococcus bovis ,Streptococcaceae ,Molecular biology ,Gram-Positive Cocci ,Heterologous expression ,Streptococcus sanguis - Abstract
The heterologous expression of a cloned endoglucanase gene (endA) from the ruminal bacterium Ruminococcus flavefaciens 17 was demonstrated in the Streptococcus species S. bovis JB1 and S. sanguis DL1. The endA gene was introduced into S. bovis and S. sanguis using the Escherichia coli/Streptococcus shuttle vector pVA838. Expression of the gene was detected by clearing zones around the recombinant colonies on agar plates containing carboxymethylcellulose stained with Congo red. S. bovis JB1 containing the endA gene was capable of utilizing cellotetraose at a faster rate than the parent strain. This is the first demonstration that Streptococcus species can express a gene from a Ruminococcus flavefaciens strain.
- Published
- 1995
266. Transfer of plasmids between strains of Escherichia coli under rumen conditions
- Author
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Harry J. Flint and Karen P. Scott
- Subjects
animal structures ,Rumen ,Tetracycline ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,Microbiology ,Plasmid ,medicine ,Escherichia coli ,Animals ,Anaerobiosis ,Sheep ,biology ,Genetic transfer ,food and beverages ,biology.organism_classification ,Enterobacteriaceae ,Aerobiosis ,Culture Media ,Conjugation, Genetic ,Anaerobic exercise ,Bacteria ,medicine.drug ,Plasmids - Abstract
Strains of Escherichia coli originally isolated from the rumen of sheep were shown to be capable of exchanging a 60kb plasmid, conferring resistance to tetracycline and ampicillin, at low frequencies (below 10(-6) per recipient) under anaerobic conditions in the presence of (a) autoclaved and clarified rumen fluid, (b) raw clarified rumen fluid, or (c) whole rumen fluid. Under anaerobic conditions the two rumen strains showed no inhibition of growth rate when 50 mmol l-1 volatile fatty acids were added to LB medium at pH 7, although significant inhibition resulted with 100 mmol l-1 VFA. The two rumen strains, and four strains from the pig gut, showed less inhibition of anaerobic growth by volatile fatty acids than did three laboratory strains examined for comparison. These findings indicate that plasmid transfer between certain E. coli strains can occur under conditions that closely simulate an anaerobic but environment.
- Published
- 1995
267. Identification of non-catalytic conserved regions in xylanases encoded by the xynB and xynD genes of the cellulolytic rumen anaerobe Ruminococcus flavefaciens
- Author
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Harry J. Flint, Jun-Xian Zhang, and Jennifer C. Martin
- Subjects
Rumen ,Molecular Sequence Data ,Sequence alignment ,Bacillus subtilis ,Homology (biology) ,Bacteria, Anaerobic ,Bacterial Proteins ,Species Specificity ,Genetics ,Escherichia coli ,Animals ,Amino Acid Sequence ,Molecular Biology ,Peptide sequence ,biology ,Base Sequence ,Sequence Homology, Amino Acid ,Ruminococcus ,biology.organism_classification ,Gram-Positive Cocci ,Xylan Endo-1,3-beta-Xylosidase ,Xylosidases ,Biochemistry ,Genes, Bacterial ,Codon usage bias ,Xylanase ,Anaerobic bacteria ,Sequence Alignment - Abstract
xynB is one of at least four genes from the cellulolytic rumen anaerobe Ruminococcus flavefaciens 17 that encode xylanase activity. The xynB gene is predicted to encode a 781-amino acid product starting with a signal peptide, followed by an amino-terminal xylanase domain which is identical at 89% and 78% of residues, respectively, to the amino-terminal xylanase domains of the bifunctional XynD and XynA enzymes from the same organism. Two separate regions within the carboxy-terminal 537 amino acids of XynB also show close similarities with domain B of XynD. These regions show no significant homology with cellulose- or xylan-binding domains from other species, or with any other sequences, and their functions are unknown. In addition a 30 to 32-residue threonine-rich region is present in both XynD and XynB. Codon usage shows a consistent pattern of bias in the three xylanase genes from R. flavefaciens that have been sequenced.
- Published
- 1994
268. Molecular genetics of obligate anaerobes from the rumen
- Author
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Harry J. Flint
- Subjects
animal structures ,Rumen ,biology ,Glycoside Hydrolases ,Microorganism ,Genetic transfer ,Gene Transfer Techniques ,food and beverages ,Obligate anaerobe ,Genetic Variation ,Fungus ,Ribosomal RNA ,biology.organism_classification ,Microbiology ,Bacteria, Anaerobic ,Biochemistry ,Genes, Bacterial ,Genetics ,Animals ,Anaerobic bacteria ,Molecular Biology ,Bacteria ,Phylogeny - Abstract
The rumen is inhabited by a highly specialised microflora consisting of obligately anaerobic bacteria, fungi and protozoa. Rumen bacteria belong to many different phylogenetic groupings and many species exhibit a high degree of rRNA gene sequence diversity, whereas the rumen fungi are monophyletic. At least 21 genes concerned with the degradation and utilisation of plant cell wall polysaccharides, from five species of rumen bacteria and from rumen fungi, have been isolated and sequenced. In general, the catalytic domains of the encoded enzymes belong to enzyme families identified among non-rumen microorganisms, but some show unusual organisation, consisting of multiple catalytic domains. Several bacterial species have been used as recipients for gene transfer by electrotransformation or by conjugation, allowing development of methods for genetic analysis. The rumen is also considered as a potential site for natural gene transfer.
- Published
- 1994
269. Multiple lactate dehydrogenase activities of the rumen bacterium Selenomonas ruminantium
- Author
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Martin Gilmour, Wilfrid J. Mitchell, and Harry J. Flint
- Subjects
Gram-Negative Anaerobic Bacteria ,Rumen ,L-Lactate Dehydrogenase ,Metabolism ,Biology ,NAD ,Microbiology ,Isoenzymes ,chemistry.chemical_compound ,chemistry ,Biochemistry ,Bacterial Proteins ,Species Specificity ,Lactate dehydrogenase ,Fermentation ,D-lactate dehydrogenase ,Lactates ,Animals ,NAD+ kinase ,Anaerobic bacteria ,Selenomonas ruminantium ,Lactic Acid ,Oxidation-Reduction - Abstract
The lactate utilizing strain of Selenomonas ruminantium 5934e was found to contain three lactate dehydrogenase (LDH) activities in sonicated cell extracts. One activity, an NAD dependent L-LDH (L-nLDH) was measured at 15-fold greater levels in extracts of cells grown to mid-exponential phase on glucose compared to cells grown to the equivalent growth stage on DL-lactate. A second nLDH activity specific for D-lactate (D-nLDH) was detected at similar levels in both lactate-grown cell extracts and glucose-grown cell extracts. The third activity, an NAD independent DLDH (D-iLDH) was very low in cells grown on glucose but was induced more than 10-fold when DL-lactate was used as the carbon source. The three LDH activities could be separated by gel filtration. Recovery of the activities was low due to the apparent instability of the enzymes at 4 degrees C, which was most pronounced in the case of the D-iLDH. A Km for lactate of 0.5 mM was estimated for the D-iLDH and this was considerably lower than the values of 45 mM and 70 mM measured for L-nLDH and D-nLDH respectively. It is proposed that the D-iLDH may be largely responsible for the formation of pyruvate in lactate-grown cells of S. ruminantium strain 5934e. Three other lactate utilizing strains of S. ruminantium, HD4, 5521C1 and JW13 exhibited a similar profile of LDH activities to strain 5934e when grown on glucose and DL-lactate.
- Published
- 1994
270. Gut Environmental Factors May Shape the Persistence of Faecalibacterium Prausnitzii in the Healthy and Diseased Large Intestine
- Author
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Xavier Aldeguer Mante, L. Jesús Garcia-Gil, Hermie J. M. Harmsen, Tanweer M. Khan, Mireia Lopez-Siles, Sylvia H. Duncan, and Harry J. Flint
- Subjects
Programmed cell death ,Pathology ,medicine.medical_specialty ,Hepatology ,biology ,Autophagy ,Gastroenterology ,Faecalibacterium prausnitzii ,Ileum ,bacterial infections and mycoses ,medicine.disease ,biology.organism_classification ,digestive system diseases ,Proinflammatory cytokine ,Pathogenesis ,medicine.anatomical_structure ,Necrotizing enterocolitis ,Immunology ,medicine ,Helicobacter hepaticus - Abstract
Background: Exposure to Helicobacter hepaticus, common enterohepatic bacteria, increases the incidence of experimental colitis and promotes colonic tumorigenesis. However, the role of H. hepaticus in pathogenesis of necrotizing enterocolitis (NEC) is not known. Neonatal NEC is a devastating disease of prematurely born babies. The cause of NEC is not clear, but inappropriate bacterial colonization plays an important role in NEC pathogenesis. Autophagy protects organisms against diverse pathologies, such as bacterial infections. However, uncontrolled activation of autophagy may lead to cellular injury. Aim: To test the effect of H. hepaticus on development of intestinal injury and expression of inflammatory and autophagic regulators in the rat model of NEC. Methods: Pregnant rats and newborn pups were kept either in H. hepaticus-free or in the H. hepaticus-contaminated environment. Prematurely born rat pups were hand-fed with formula and exposed to asphyxia/cold stress to develop NEC. After 96 hours, all animals were sacrificed and ileal tissue was collected. Incidence of NEC injury, expression of inflammatory cytokines (IL-6, IL-8, IL-18 & TNF-α) and protein levels of autophagy regulators (Beclin 1, LC3II & p62) were evaluated in the ileum. Results: Results: The incidence of NEC was significantly increased to 71% (25/35) in rats exposed to H. hepaticus compared with the H. hepaticus-free group with NEC incidence of 33% (13/39; p < 0.001). The survival rates were not affected by H. hepaticus. The presence of H. hepaticus was detected in the GI tract of the H. hepaticus group. IL-8 and IL-18 gene expression was significantly altered in the H. hepaticus group, whereas IL-6 and TNF-α levels were not changed. Protein levels of Beclin 1 and LC3II were markedly increased, whereas p62 levels were significantly decreased in the ileum from the H. hepaticus group. Conclusions: Helicobacter hepaticus exacerbates intestinal injury and induces intestinal inflammation in the rat NEC model. Exposure to H. hepaticus is also associated with a strong induction of the autophagy pathway in the site of injury the terminal ileum. Inappropriate activation of intestinal autophagy by bacterial insult may facilitate massive cell death, leading to severe mucosal injury. Supported by the NIH Grant HD-39657 (to B.D.)
- Published
- 2011
271. Mode of action, kinetic properties and physicochemical characterization of two different domains of a bifunctional (1-->4)-beta-D-xylanase from Ruminococcus flavefaciens expressed separately in Escherichia coli
- Author
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Vicenta Garcia-Campayo, J X Zhang, Sheila I. McCrae, Harry J. Flint, and Thomas M. Wood
- Subjects
Arabinose ,Rumen ,Glycoside Hydrolases ,Molecular Sequence Data ,Gene Expression ,Oligosaccharides ,Biochemistry ,chemistry.chemical_compound ,Bacteria, Anaerobic ,Arabinoxylan ,Xylobiose ,Escherichia coli ,Animals ,Cloning, Molecular ,Molecular Biology ,Binding Sites ,Endo-1,4-beta Xylanases ,Molecular mass ,Chromatofocusing ,Cell Biology ,Hydrogen-Ion Concentration ,Chromatography, Ion Exchange ,Xylan ,Recombinant Proteins ,Gram-Positive Cocci ,Molecular Weight ,Kinetics ,Isoelectric point ,chemistry ,Carbohydrate Sequence ,Genes, Bacterial ,Xylanase ,Electrophoresis, Polyacrylamide Gel ,Research Article - Abstract
Two catalytic domains, A and C, of xylanase A (XYLA) from Ruminococcus flavefaciens were expressed separately as truncated gene products from lacZ fusions in Escherichia coli. The fusion products, referred to respectively as XYLA-A1 and XYLA-C2, were purified to homogeneity by anion-exchange chromatography and chromatofocusing. XYLA-A1 was isoelectric at pH 5.0 and had a molecular mass of 30 kDa, whereas XYLA-C2 had a pI of 5.4 and a molecular mass of 44 kDa. The catalytic activity shown by both domains was optimal at 50 degrees C, but XYLA-A1 was more sensitive than XYLA-C2 to temperatures higher than the optimum. XYLA-A1 showed a higher sensitivity to pH than XYLA-C2. The enzyme activity of both domains was completely inactivated in the presence of copper or silver ions and partially inactivated by iron or zinc ions. Neither domain was active on xylo-oligosaccharides shorter than xylopentaose: the rate of degradation of longer xylo-oligosaccharides (degree of polymerization 5-10) increased as the chain length increased. Analysis of the products of hydrolysis of xylo-oligosaccharides and xylan (arabinoxylan) polysaccharide showed that the two domains differed in their modes of action: xylobiose was the shortest product of the hydrolysis. With oat spelt xylan as substrate, XYLA-A1 activity was apparently restricted to regions where xylopyranosyl residues did not carry arabinofuranosyl substituents, whereas XYLA-C2 was able to release hetero-oligosaccharides carrying arabinofuranosyl residues. Neither domain was able to release arabinose from oat spelt xylan.
- Published
- 1993
272. Expression of Escherichia coli homoserine kinase in mouse 3T3 cells
- Author
-
Harry J. Flint, Susan Hay, and W D Rees
- Subjects
Homoserine kinase ,Homoserine ,Gene Expression ,medicine.disease_cause ,Transfection ,Biochemistry ,chemistry.chemical_compound ,Mice ,Plasmid ,medicine ,Escherichia coli ,Animals ,Threonine ,Cloning, Molecular ,Phosphorylation ,Molecular Biology ,biology ,Phosphotransferases ,Cell Biology ,3T3 Cells ,biology.organism_classification ,Blotting, Northern ,Enterobacteriaceae ,Molecular biology ,Kinetics ,Phosphotransferases (Alcohol Group Acceptor) ,Homoserine kinase activity ,chemistry ,RNA ,Research Article ,Plasmids - Abstract
The Escherichia coli gene for homoserine kinase (thrB) has been cloned into a simian-virus-40-based eukaryotic expression vector which also includes a neomycin-resistance gene. Mouse 3T3 cells transfected with this plasmid were selected for resistance and screened for homoserine kinase activity. It has thus been possible to isolate clones which are capable of accumulating homoserine O-phosphate when supplied with homoserine. In broken-cell preparations the kinetic constants for the production of homoserine O-phosphate were similar to those of the wild-type E. coli enzyme. These experiments demonstrate that E. coli homoserine kinase can be expressed in an animal cell and that it can successfully phosphorylate L-homoserine in the intact cell utilizing endogenous ATP.
- Published
- 1992
273. S1212 The Intestinal Microbiota of Irritable Bowel Syndrome Patients Is Characterized By Functional Dysbiosis
- Author
-
Christophe Chassard, Claude Dubray, Alain Eschalier, Annick Bernalier-Donadille, Christophe Del'Homme, Karen P. Scott, Harry J. Flint, and Michel Dapoigny
- Subjects
medicine.medical_specialty ,Hepatology ,business.industry ,Internal medicine ,Gastroenterology ,Medicine ,business ,medicine.disease ,Dysbiosis ,Irritable bowel syndrome - Published
- 2009
274. Expression of two xylanase genes from the rumen cellulolytic bacterium Ruminococcus flavefaciens 17 cloned in pUC13
- Author
-
Carol A. McPherson, Jennifer C. Martin, and Harry J. Flint
- Subjects
DNA, Bacterial ,Rumen ,Glycoside Hydrolases ,Restriction Mapping ,Gene Expression ,Cellobiose ,Xylose ,Microbiology ,Substrate Specificity ,chemistry.chemical_compound ,Bacteria, Anaerobic ,Polysaccharides ,Xylobiose ,Animals ,Cloning, Molecular ,biology ,Ruminococcus ,Hydrolysis ,biology.organism_classification ,Xylan ,Xylan Endo-1,3-beta-Xylosidase ,Biochemistry ,chemistry ,Genes, Bacterial ,Xylanase ,Clostridium thermocellum ,Energy source ,Plasmids - Abstract
Two distinct xylanase genes (designated xynA and xynB) were subcloned in pUC13 from non-homologous restriction fragments of Ruminococcus flavefaciens 17 DNA originally isolated in λ EMBL3. The products of the two genes showed similar pH optima for hydrolysis of oat spelt xylan (around 5·5) and had little or no activity against carboxymethylcellulose. Trace activities against p-nitrophenyl (pNP) cellobioside and pNP-xyloside were detected in clones containing xynA, but not in one harbouring xynB. The xylanase associated with clones carrying xynA produced mainly xylobiose and xylose from xylan and did not give hydrolysis of xylobiose, while that encoded by xynB produced mainly xylobiose and higher xylo-oligosaccharides from xylan. There was evidence of increased expression, at the RNA level, of these two genes, and of another cloned region encoding multiple activities including xylanase, in R. flavefaciens 17 grown with xylan, as compared with cellobiose, as energy source. Total cell-associated xylanase and β-xylosidase activities, and supernatant xylanase activity, were shown to be similarly induced in xylan-grown R. flavefaciens, 17.
- Published
- 1991
275. Deoxyribonuclease activity in rumen bacteria
- Author
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Harry J. Flint and A.M. Thomson
- Subjects
DNA, Bacterial ,animal structures ,Deoxyribonucleases ,Rumen ,biology ,Bacteria ,food and beverages ,Deoxyribonuclease ,biology.organism_classification ,Deoxyribonuclease activity ,Applied Microbiology and Biotechnology ,Microbiology ,Bacteriophage ,Biochemistry ,Species Specificity ,Animals ,Bacteroides ,Deoxyribonuclease I ,Bacteroidaceae - Abstract
Deoxyribonuclease activity was surveyed in 22 strains belonging to 12 species of rumen bacteria, with lambda bacteriophage DNA as substrate. Activity was readily detected in broken cell preparations from 15 of these strains. Particularly high levels of activity were present in cells and culture supernatant of all 5 strains of Bacteroides succinogenes, and 2 out of 6 strains of Bacteroides ruminicola, examined.
- Published
- 1990
276. A novel mobile chromosomal element conferring TcR in rumen Butyrivibrio species
- Author
-
Harry J. Flint, Teresa M. Barbosa, K. Forbes, and Karen P. Scott
- Subjects
Genetics ,Butyrivibrio species ,Rumen ,T-cell receptor ,Biology - Published
- 1997
277. Evolutionary relationships and the diversity of the rumen bacteria belonging to the Cytophaga-Flexibacter-Bacteroides phylum
- Author
-
Harry J. Flint, G. Avguštin, M. Peterka, FV Nekrep, A. Ramšak, and Revues Inra, Import
- Subjects
[SDV.AEN] Life Sciences [q-bio]/Food and Nutrition ,Rumen ,Cytophaga-Flexibacter-Bacteroides phylum ,media_common.quotation_subject ,[SDV.BDD] Life Sciences [q-bio]/Development Biology ,Biology ,biology.organism_classification ,[SDV.BDLR] Life Sciences [q-bio]/Reproductive Biology ,Bacteria ,Diversity (politics) ,media_common ,Microbiology - Published
- 1997
278. Some genetic and biochemical aspects of Prevotella bryantii B 14 xylanases
- Author
-
F.V. Nekrep, R. Marinsek-Logar, A. Gasparic, and Harry J. Flint
- Subjects
biology ,biology.organism_classification ,Prevotella bryantii ,Microbiology - Published
- 1997
279. USE OF AN ILEAL EXPLANT MODEL TO STUDY THE CONTRIBUTION OF S.ENTERITIDIS FIMBRIAL LECTINS TO INFECTION IN THE RAT GUT
- Author
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Emma Allen-Vercoe, J. M. C. Robertson, Harry J. Flint, George Grant, and M.J. Woodward
- Subjects
S. enteritidis ,Biology ,Biochemistry ,Microbiology ,Explant culture - Published
- 1999
280. Nutritional influences on the gut microbiota and the consequences for gastrointestinal health.
- Author
-
Karen P. Scott, Sylvia H. Duncan, Petra Louis, and Harry J. Flint
- Subjects
NUTRITION ,GASTROINTESTINAL system ,HEALTH ,MICROBIAL metabolites ,MOLECULAR biology ,BACTERIA ,COLONIZATION (Ecology) ,HOST-bacteria relationships ,DIET - Abstract
The human colonic microbiota degrades dietary substrates that are indigestible in the upper GIT (gastrointestinal tract), releasing bacterial metabolites, some of which are important for gut health. Advances in molecular biology techniques have facilitated detailed analyses of the composition of the bacterial community resident in the lower GIT. Such analyses have indicated that more than 500 different bacterial species colonize an individual, and that, although there is much functional consistency in the resident bacterial groups, there is considerable inter-individual variation at the species/strain level. The bacterial community develops during early childhood until it reaches an adult-like composition. Whereas colonization and host factors influence the species composition, dietary factors also have an important impact, with specific bacterial groups changing in response to specific dietary interventions. Since bacterial species have different metabolic activities, specific diets have various consequences for health, dependent on the effect exerted on the bacterial population. [ABSTRACT FROM AUTHOR]
- Published
- 2011
- Full Text
- View/download PDF
281. Electroporation induced transformation ofBacteroides ruminicolaandBacteroides uniformisby plasmid DNA
- Author
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Andrew M. Thomson and Harry J. Flint
- Subjects
Transformation (genetics) ,Plasmid dna ,Chemistry ,Electroporation ,Genetics ,Molecular Biology ,Microbiology ,Molecular biology - Published
- 1989
282. Use of antibiotic resistance mutations to track strains of obligately anaerobic bacteria introduced into the rumen of sheep
- Author
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Jacqueline Bisset, Jeanette Webb, and Harry J. Flint
- Subjects
Rumen ,animal structures ,Bacteroidaceae ,Applied Microbiology and Biotechnology ,Microbiology ,chemistry.chemical_compound ,Animals ,Bacteroides ,Sorbitol ,Selenomonas ruminantium ,Sheep ,biology ,food and beverages ,Drug Resistance, Microbial ,biology.organism_classification ,In vitro ,chemistry ,Mutation ,Anaerobic bacteria ,Rifampin ,Anaerobic exercise ,Bacteria - Abstract
Selective plating procedures were used to follow the fate of rifampicin-resistant mutant strains of the obligately anaerobic species Bacteroides multiacidus and Selenomonas ruminantium after their introduction at numbers around 10(7)/ml into the rumen of sheep. Bacteroides multiacidus strain F100 showed an initially rapid rate of loss (49%/h) but subsequently numbers declined more gradually approaching the limits of detection (less than 10(3)/ml) after 100 h. Viable cell numbers also decreased in vitro upon addition of F100 cells to whole rumen contents, but remained stable upon addition to cell-free rumen fluid, suggesting protozoal predation. F100 cells were able to grow in vitro in whole rumen contents in the presence of an added utilizable substrate such as sorbitol, but addition of sorbitol to the rumen failed to enhance survival in vivo. In the case of S. ruminantium, introduced rifR strains persisted in the rumen at levels around 10(6) ml for at least 30 days.
- Published
- 1989
283. Control of the Flux in the Arginine Pathway of Neurospora crassa: Effects of Co-ordinate Changes of Enzyme Concentration
- Author
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Fiona Stuart, Harry J. Flint, David J. Porteous, and Henrik Kacser
- Subjects
chemistry.chemical_classification ,biology ,Arginine ,Mutant ,biology.organism_classification ,Microbiology ,Neurospora crassa ,chemistry.chemical_compound ,Enzyme ,Biochemistry ,chemistry ,biology.protein ,Citrulline ,Flux (metabolism) ,Derepression ,Acetylglutamate kinase - Abstract
Summary: The flux to arginine was determined in growing mycelium of Neurospora crassa carrying the aga mutation, by measuring the exponential growth rate and the arginine content of the free pool and that in protein. Derepression of the enzymes in the pathway by a factor of about three resulted in a 14% increase in the flux. Using a mutant (cpc-1), which affected the cross-pathway control, the pathway enzymes were found to have about threefold lower activities. This resulted in a 16% decrease in the measured flux. The effect of arginine acting as a feedback inhibitor of acetylglutamate kinase was estimated in an arg-12 mutant by varying the steady-state arginine concentration using histidine as a competitive inhibitor of the citrulline uptake. It is concluded that the feedback loop is mainly responsible for the small response of the flux to the large coordinate changes in the pathway enzymes. The results are discussed in terms of control analysis of metabolic systems.
- Published
- 1986
284. Alterations in the pattern of polypeptide synthesis resulting from amino acid limitation in Neurospora crassa
- Author
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Harry J. Flint and Janis McKormick
- Subjects
chemistry.chemical_classification ,Strain (chemistry) ,Arginine ,Immunology ,General Medicine ,Biology ,biology.organism_classification ,Applied Microbiology and Biotechnology ,Microbiology ,Amino acid ,Neurospora crassa ,Electrophoresis ,chemistry.chemical_compound ,Biochemistry ,chemistry ,Biosynthesis ,Genetics ,Molecular Biology ,Mycelium - Abstract
Two-dimensional electrophoresis was used to examine the pattern of polypeptide synthesis in Neurospora crassa mycelium of an arg-6 strain grown in the presence of [14C]arginine. Reduction in the arginine supply rate led to amino acid limited growth, and to major alterations in the pattern of polypeptide synthesis. Strains carrying wild-type (cpc-1+) or mutant (cpc-1) alleles at a locus governing cross-pathway control of amino acid synthetic enzymes differed markedly with respect to their pattern of polypeptide synthesis under limitation conditions, but differed little during arginine sufficient growth. Among 160 abundant polypeptide species classified for their response to limitation in two dimensional fluorographs, 31 were induced by limitation only in a cpc-1+ strain, 13 only in a cpc-1 strain, and 9 showed induction in both strains.
- Published
- 1988
285. Control of the flux in the arginine pathway of Neurospora crassa. The flux from citrulline to arginine
- Author
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Harry J. Flint, Henrik Kacser, and David J. Porteous
- Subjects
Growth medium ,Neurospora crassa ,biology ,Arginine ,Endogeny ,Cell Biology ,Ornithine ,biology.organism_classification ,Models, Biological ,Biochemistry ,Kinetics ,Neurospora ,chemistry.chemical_compound ,chemistry ,Glycine ,Citrulline ,Urea ,Molecular Biology ,Histidine ,Research Article - Abstract
The arginine pathway is a complex one, having many branch points and effector interactions. In order to assess the quantitative role of the various mechanisms that influence the flux in the pathway, the system was divided experimentally into two moieties by the introduction of a genetic block abolishing ornithine carbamoyltransferase activity. This normally produces citrulline from ornithine within the mitochondria. The endogenous citrulline supply was replaced by citrulline in the growth medium, and control of the influx rate was achieved by using glycine or histidine as uptake inhibitors. By modulating the influx rate over a large range of values, the importance of such factors as reversibility, saturation, inhibition and induction in affecting the flux and the sizes of intermediate pools between citrulline and arginine was assessed. The role of expansion fluxes as important controls in the exponentially growing system was established.
- Published
- 1980
286. Characteristics of isolates of Lactobacillus fermentum from the rumen of sheep
- Author
-
Harry J. Flint, Sylvia H. Duncan, and Colin S. Stewart
- Subjects
biology ,Lactobacillus fermentum ,food and beverages ,Lactobacillaceae ,biology.organism_classification ,Applied Microbiology and Biotechnology ,Microbiology ,chemistry.chemical_compound ,Rumen ,chemistry ,Lactobacillus ,Fermentation ,Lactose ,Polyacrylamide gel electrophoresis ,Bacteria - Abstract
A detailed characterization of four representative Lactobacillus isolates from the rumen was undertaken after tests with API CH50 kits had indicated that they differed from currently recognized species. These isolates are shown to be related to Lactobacillus fermentum, although they differ from the type and reference strains of this species in sodium dodecyl sulphate polyacrylamide gel electrophoresis patterns and electrophoretic mobility of LDH. It seemed that certain heterofermentative lactobacilli could give negative results for some fermentation tests (particularly fructose and lactose) in API kits, whilst showing good growth on the same substrates in conventional tests.
- Published
- 1988
287. Transmissible antibiotic resistance in strains of Escherichia coli isolated from the ovine rumen
- Author
-
Harry J. Flint, Sylvia H. Duncan, and Colin S. Stewart
- Subjects
animal structures ,Tetracycline ,Genetic transfer ,biochemical phenomena, metabolism, and nutrition ,Biology ,medicine.disease_cause ,biology.organism_classification ,Applied Microbiology and Biotechnology ,Enterobacteriaceae ,Microbiology ,chemistry.chemical_compound ,Rumen ,chemistry ,Streptomycin ,Ampicillin ,medicine ,TetR ,Escherichia coli ,medicine.drug - Abstract
Transmissible multiple resistance to tetracycline, ampicillin and streptomycin was found to be associated with transfer of the larger of the two plasmids (˜ 80 and ˜ 45 kbp) present in a group of tetracycline-resistant Escherichia coli strains isolated from the rumen of sheep. A second group of rumen tetR strains, which differed in the ability to utilize several sugars, showed non-transmissible resistances to tetracycline and streptomycin.
- Published
- 1987
288. The isolation of tetracycline-resistant strains of strictly anaerobic bacteria from the rumen
- Author
-
Sylvia H. Duncan, Jacqueline Bisset, Colin S. Stewart, and Harry J. Flint
- Subjects
animal structures ,biology ,Tetracycline ,medicine.drug_class ,Antibiotics ,biology.organism_classification ,Applied Microbiology and Biotechnology ,Microbiology ,Rumen ,Plasmid ,medicine ,Anaerobic bacteria ,Selenomonas ruminantium ,Bacteria ,Butyrivibrio fibrisolvens ,medicine.drug - Abstract
Tetracycline resistant (TcR) strains of three of the major species of strictly anaerobic rumen bacteria Megasphaera elsdenii, Selenomonas ruminantium and Butyrivibrio fibrisolvens, were recovered with an isolation medium containing 20 μg/ml tetracycline. Only two of 14 strains of these species from other sources, isolated without antibiotic selection, showed tetracycline resistance. Evidence was found for the presence of plasmids in two tetracycline-resistant strains of M. elsdenii, and in some strains of S. ruminantium.
- Published
- 1988
289. Molecular cloning of genes from Ruminococcus flavefaciens encoding xylanase and beta(1-3,1-4)glucanase activities
- Author
-
Harry J. Flint, C A McPherson, and J Bisset
- Subjects
DNA, Bacterial ,Rumen ,Glycoside Hydrolases ,Restriction Mapping ,Molecular cloning ,Biology ,Applied Microbiology and Biotechnology ,Beta-1 adrenergic receptor ,Bacteriophage ,Restriction map ,Animals ,Cloning, Molecular ,Gene ,Glucan ,chemistry.chemical_classification ,Ecology ,Nucleic Acid Hybridization ,Glucanase ,biology.organism_classification ,Molecular biology ,Xylan Endo-1,3-beta-Xylosidase ,Biochemistry ,chemistry ,Gene Expression Regulation ,Peptococcaceae ,Xylanase ,Cattle ,Xylans ,Food Science ,Biotechnology ,Research Article - Abstract
Clones expressing activity against xylan or beta(1-3,1-4)glucan (lichenan) were isolated from a library of Ruminococcus flavefaciens 17 DNA made in bacteriophage lambda EMBL3. Hybridization analyses indicated the recovery of four separate genes encoding xylanases that showed no detectable associated carboxylmethylcellulase activity. One of these genes was associated with clones that also expressed beta(1-3,1-4)glucanase and beta-xylosidase activities.
- Published
- 1989
290. General control of arginine biosynthetic enzymes in Neurospora crassa
- Author
-
Harry J. Flint and Barbara F. Kemp
- Subjects
chemistry.chemical_classification ,Arginine ,Neurospora crassa ,Biology ,biology.organism_classification ,Microbiology ,Amino acid ,Neurospora ,Enzyme ,Biochemistry ,chemistry ,Ornithine Carbamoyltransferase ,Gene Expression Regulation ,Histidine ,Enzyme Repression ,Amino acid synthesis ,Derepression - Abstract
SUMMARY: The response of six amino acid synthetic enzymes, including four concerned with arginine synthesis, one with histidine synthesis and one with lysine synthesis, to conditions of histidine and arginine limitation and to exogenously provided amino acids is described in Neurospora crassa. The activities of all these enzymes increased in response to lowered levels of histidine or arginine, but showed little or no repression in wild-type cultures supplemented with casein hydrolysate. The activity of glucose-6-phosphate dehydrogenase (an enzyme not concerned with amino acid synthesis) remained unaffected by any of these conditions. In the case of the four arginine synthetic enzymes evidence is presented that suggests the ‘general’ system implicated in the ‘cross-pathway’ response to histidine is also entirely responsible for the derepression occurring under conditions of arginine limitation. Further investigations into the control of one enzyme, ornithine carbamoyltransferase, are also reported. These show that the enzyme is stable, and suggest that its derepression in response to histidine limitation entails new protein synthesis and involves control at a stage prior to translation.
- Published
- 1981
291. Bacteroides (Fibrobacter) succinogenes, a cellulolytic anaerobic bacterium from the gastrointestinal tract
- Author
-
Colin S. Stewart and Harry J. Flint
- Subjects
Gastrointestinal tract ,Fibrobacter succinogenes ,Enzymatic digestion ,biology ,Energy metabolism ,food and beverages ,Anaerobic bacterium ,General Medicine ,biology.organism_classification ,Applied Microbiology and Biotechnology ,Bacteroides succinogenes ,Microbiology ,Bacteroides ,Bacteroidaceae ,Biotechnology - Abstract
Mini-revue sur la bacterie cellulolytique Bacteroides succinogenes. Rappel sur son isolement a partir du rumen de Bovin, sur ses caracteristiques phytogenetiques et taxonomiques, sur ses proprietes de digestion de la paroi cellulaire des plantes. Enfin, description des applications de cette bacterie en biotechnologie
- Published
- 1989
292. Cloning of the arg-12 gene of Neurospora crassa and regulation of its transcript via cross-pathway amino acid control
- Author
-
Johanna Wilkening and Harry J. Flint
- Subjects
Arginine ,Transcription, Genetic ,Genes, Fungal ,Molecular cloning ,Neurospora ,Neurospora crassa ,chemistry.chemical_compound ,Transcription (biology) ,Genetics ,Cloning, Molecular ,Molecular Biology ,Gene ,Ornithine Carbamoyltransferase ,chemistry.chemical_classification ,biology ,Nucleic Acid Hybridization ,DNA Restriction Enzymes ,Ornithine ,biology.organism_classification ,Molecular biology ,Amino acid ,Kinetics ,Biochemistry ,chemistry ,Genes - Abstract
The arg-12 locus of Neurospora crassa encodes ornithine carbamoyl transferase, which is one of many amino acid synthetic enzymes whose activity is regulated through cross-pathway (or general) amino acid control. We report here the use of probes derived from the functionally equivalent arg-B gene of Aspergillus nidulans to identify and clone a 10 kb Neurospora DNA fragment carrying the arg-12 gene. Short Neurospora DNA probes derived from this fragment were used to identify a 1.5 kb polyA+ transcript of the arg-12 region. Arg-12 transcript levels increased approximately 20 fold under conditions of arginine or histidine limitation in strains having normal cross-pathway regulation (cpc-1+) but showed no such response in a cpc-1 mutant strain impaired in this regulation. Time course studies in cpc-1+ strains revealed a rapid response (within 10 m) of arg-12 transcript levels following inhibition of histidine synthesis by 3 amino 1,2,4 triazole, but a delayed response following arginine deprivation of an arginine requiring strain. In contrast to the behaviour of arg-12 mRNA, the level of the Neurospora am gene transcript (specifying NADP dependent glutamate dehydrogenase) was unaffected either by amino acid limitation or by the cpc-1 mutation. A possible role for the cpc-1+ product as a positive regulator of transcription of genes subject to cross-pathway control is discussed.
- Published
- 1986
293. Changes in gene expression elicited by amino acid limitation in Neurospora crassa strains having normal or mutant cross-pathway amino acid control
- Author
-
Harry J. Flint
- Subjects
Arginine ,Mutant ,Genes, Fungal ,macromolecular substances ,Neurospora crassa ,Fungal Proteins ,Species Specificity ,Gene expression ,Genetics ,RNA, Messenger ,Amino Acids ,Molecular Biology ,Histidine ,Derepression ,chemistry.chemical_classification ,biology ,technology, industry, and agriculture ,biology.organism_classification ,Molecular biology ,Amino acid ,Neurospora ,Enzyme ,chemistry ,Biochemistry ,Genes ,Protein Biosynthesis ,Mutation ,RNA ,Electrophoresis, Polyacrylamide Gel ,Poly A - Abstract
The effects of amino acid limitation on gene expression have been investigated in Neurospora crassa strains carrying normal (cpc-1+) or mutant (cpc-1) alleles at a locus implicated in cross-pathway amino acid control. Electrophoresis and fluorography were used to reveal the patterns of label incorporation into polypeptides in vivo, or after in vitro translation of extracted mRNAs. In a cpc-1+ strain at least 20% of detectable in vitro translation products showed relative increases in incorporation when RNA was obtained from mycelium grown under conditions of arginine limitation, by comparison with conditions of arginine sufficiency. A cpc-1 mutation, which impairs derepression of a variety of amino acid synthetic enzymes following amino acid limitation, had little detectable effect on in vivo polypeptide synthesis during amino acid sufficient growth or following pyrimidine limitation. However the mutation substantially altered the response to arginine or histidine limitation. The majority of in vitro translation products that showed increased expression in arginine limited cpc-1+ failed to increase in cpc-1 strains, but arginine limitation of cpc-1 also resulted in increases that did not occur in cpc-1+ strains. This may reflect both direct and indirect consequences of the impairment of cross-pathway control.
- Published
- 1985
294. Plasmid-associated transfer of tetracycline resistance in Bacteroides ruminicola
- Author
-
Harry J. Flint, J Bisset, and A M Thomson
- Subjects
animal structures ,Ecology ,biology ,Strain (chemistry) ,Tetracycline ,R Factors ,Genetic transfer ,Mutant ,Tetracycline Resistance ,DNA Restriction Enzymes ,biology.organism_classification ,Applied Microbiology and Biotechnology ,Microbiology ,Plasmid ,medicine ,Bacteroides ,Anaerobiosis ,Bacteroidaceae ,Bacteria ,Food Science ,Biotechnology ,medicine.drug ,Research Article - Abstract
Tetracycline resistance was transferred at frequencies between 10(-7) and 10(-6) per recipient cell in anaerobic matings between two strains of the strictly anaerobic rumen bacterium Bacteroides ruminicola. The donor strain, 223/M2/7, was a multiple-plasmid-bearing tetracycline-resistant strain from the ovine rumen, and the recipient, F101, was a rifampin-resistant mutant of B14, a bovine strain belonging to B. ruminicola subsp. brevis. Resistance transfer could occur in the presence of DNase, but not in dummy mating mixtures in which filtrate from a donor culture replaced donor cells. Acquisition of tetracycline resistance by the recipient was accompanied by the appearance of a 19.5-kilobase pair plasmid (designated pRRI4) which was homologous with a plasmid of similar size and restriction pattern present in the donor strain. A transconjugant (F115) carrying pRRI4 was also able to act as a donor of tetracycline resistance and plasmid DNA in matings with another recipient. Derivatives of F115 that had spontaneously lost tetracycline resistance lacked detectable plasmid DNA. It is concluded that pRRI4 mediated the transfer of tetracycline resistance. Transfer of resistance was not detectably enhanced by pregrowth of the donor in medium containing tetracycline. Transfer of tetracycline resistance was not detected from 223/M2/7 to a strain, 23 belonging to B. ruminicola subsp. ruminicola.
- Published
- 1988
295. Cross-pathway control of ornithine carbamoyltransferase synthesis in Neurospora crassa
- Author
-
Harry J. Flint and Barbara F. Kemp
- Subjects
chemistry.chemical_classification ,Methionine ,biology ,Arginine ,Neurospora crassa ,fungi ,biology.organism_classification ,Microbiology ,Neurospora ,Amino acid ,chemistry.chemical_compound ,chemistry ,Biochemistry ,Ornithine Carbamoyltransferase ,Valine ,Mutation ,Amino Acids ,Enzyme Repression ,Derepression - Abstract
The pattern of cross-pathway regulation of the arginine synthetic enzyme ornithine carbamoyltransferase was investigated in Neurospora crassa, using single and double mutant auxotrophic strains starved for their required amino acids. These experiments show that starvation for histidine, tryptophan, isoleucine, valine or arginine can result in derepression of ornithine carbamoyltransferase. Methionine starvation also gave slight derepression, but starvation for lysine or leucine gave little or no effect.
- Published
- 1982
296. Organisation of genes involved in xylan utilisation and xylan debranching in the rumen cellulolytic bacterium Ruminococcus flavefaciens
- Author
-
Harry J. Flint, V. Aurilia, J. Kirby, J. Martin, S. Ekinci, and Revues Inra, Import
- Subjects
[SDV.AEN] Life Sciences [q-bio]/Food and Nutrition ,Rumen ,Biochemistry ,[SDV.BDD] Life Sciences [q-bio]/Development Biology ,Biology ,Ruminococcus flavefaciens ,biology.organism_classification ,Gene ,Xylan ,[SDV.BDLR] Life Sciences [q-bio]/Reproductive Biology ,Bacteria ,Microbiology
297. Organisation and Variable Incidence of Genes Concerned with the Utilization of Xylans in the Rumen Cellulolytic Bacterium Ruminococcus flavefaciens
- Author
-
Aurilia, Vincenzo, Martin, Jennifer C, Scott, Karen P, Mercer, Derry K, Johnston, Moira E.A, and Flint, Harry J
- Published
- 2000
- Full Text
- View/download PDF
298. Faecalibacterium prausnitzii Strain HTF-F and Its Extracellular Polymeric Matrix Attenuate Clinical Parameters in DSS-Induced Colitis.
- Author
-
Oriana Rossi, M Tanweer Khan, Martin Schwarzer, Tomas Hudcovic, Dagmar Srutkova, Sylvia H Duncan, Ellen H Stolte, Hana Kozakova, Harry J Flint, Janneke N Samsom, Hermie J M Harmsen, and Jerry M Wells
- Subjects
Medicine ,Science - Abstract
A decrease in the abundance and biodiversity of intestinal bacteria within the Firmicutes phylum has been associated with inflammatory bowel disease (IBD). In particular, the anti-inflammatory bacterium Faecalibacterium prausnitzii, member of the Firmicutes phylum and one of the most abundant species in healthy human colon, is underrepresented in the microbiota of IBD patients. The aim of this study was to investigate the immunomodulatory properties of F. prausnitzii strain A2-165, the biofilm forming strain HTF-F and the extracellular polymeric matrix (EPM) isolated from strain HTF-F. For this purpose, the immunomodulatory properties of the F. prausnitzii strains and the EPM were studied in vitro using human monocyte-derived dendritic cells. Then, the capacity of the F. prausnitzii strains and the EPM of HTF-F to suppress inflammation was assessed in vivo in the mouse dextran sodium sulphate (DSS) colitis model. The F. prausnitzii strains and the EPM had anti-inflammatory effects on the clinical parameters measured in the DSS model but with different efficacy. The immunomodulatory effects of the EPM were mediated through the TLR2-dependent modulation of IL-12 and IL-10 cytokine production in antigen presenting cells, suggesting that it contributes to the anti-inflammatory potency of F. prausnitzii HTF-F. The results show that F. prausnitzii HTF-F and its EPM may have a therapeutic use in IBD.
- Published
- 2015
- Full Text
- View/download PDF
299. Gut microbiota signatures predict host and microbiota responses to dietary interventions in obese individuals.
- Author
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Katri Korpela, Harry J Flint, Alexandra M Johnstone, Jenni Lappi, Kaisa Poutanen, Evelyne Dewulf, Nathalie Delzenne, Willem M de Vos, and Anne Salonen
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Medicine ,Science - Abstract
BackgroundInteractions between the diet and intestinal microbiota play a role in health and disease, including obesity and related metabolic complications. There is great interest to use dietary means to manipulate the microbiota to promote health. Currently, the impact of dietary change on the microbiota and the host metabolism is poorly predictable and highly individual. We propose that the responsiveness of the gut microbiota may depend on its composition, and associate with metabolic changes in the host.MethodologyOur study involved three independent cohorts of obese adults (n = 78) from Belgium, Finland, and Britain, participating in different dietary interventions aiming to improve metabolic health. We used a phylogenetic microarray for comprehensive fecal microbiota analysis at baseline and after the intervention. Blood cholesterol, insulin and inflammation markers were analyzed as indicators of host response. The data were divided into four training set - test set pairs; each intervention acted both as a part of a training set and as an independent test set. We used linear models to predict the responsiveness of the microbiota and the host, and logistic regression to predict responder vs. non-responder status, or increase vs. decrease of the health parameters.Principal findingsOur models, based on the abundance of several, mainly Firmicute species at baseline, predicted the responsiveness of the microbiota (AUC = 0.77-1; predicted vs. observed correlation = 0.67-0.88). Many of the predictive taxa showed a non-linear relationship with the responsiveness. The microbiota response associated with the change in serum cholesterol levels with an AUC of 0.96, highlighting the involvement of the intestinal microbiota in metabolic health.ConclusionThis proof-of-principle study introduces the first potential microbial biomarkers for dietary responsiveness in obese individuals with impaired metabolic health, and reveals the potential of microbiota signatures for personalized nutrition.
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- 2014
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300. Rumen cellulosomics: divergent fiber-degrading strategies revealed by comparative genome-wide analysis of six ruminococcal strains.
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Bareket Dassa, Ilya Borovok, Vered Ruimy-Israeli, Raphael Lamed, Harry J Flint, Sylvia H Duncan, Bernard Henrissat, Pedro Coutinho, Mark Morrison, Pascale Mosoni, Carl J Yeoman, Bryan A White, and Edward A Bayer
- Subjects
Medicine ,Science - Abstract
BackgroundA complex community of microorganisms is responsible for efficient plant cell wall digestion by many herbivores, notably the ruminants. Understanding the different fibrolytic mechanisms utilized by these bacteria has been of great interest in agricultural and technological fields, reinforced more recently by current efforts to convert cellulosic biomass to biofuels.Methodology/principal findingsHere, we have used a bioinformatics-based approach to explore the cellulosome-related components of six genomes from two of the primary fiber-degrading bacteria in the rumen: Ruminococcus flavefaciens (strains FD-1, 007c and 17) and Ruminococcus albus (strains 7, 8 and SY3). The genomes of two of these strains are reported for the first time herein. The data reveal that the three R. flavefaciens strains encode for an elaborate reservoir of cohesin- and dockerin-containing proteins, whereas the three R. albus strains are cohesin-deficient and encode mainly dockerins and a unique family of cell-anchoring carbohydrate-binding modules (family 37).Conclusions/significanceOur comparative genome-wide analysis pinpoints rare and novel strain-specific protein architectures and provides an exhaustive profile of their numerous lignocellulose-degrading enzymes. This work provides blueprints of the divergent cellulolytic systems in these two prominent fibrolytic rumen bacterial species, each of which reflects a distinct mechanistic model for efficient degradation of cellulosic biomass.
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- 2014
- Full Text
- View/download PDF
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