scholarly journals Polysaccharide Biosynthesis Locus Required for Virulence of Bacteroides fragilis

2001 ◽  
Vol 69 (7) ◽  
pp. 4342-4350 ◽  
Author(s):  
Michael J. Coyne ◽  
Arthur O. Tzianabos ◽  
Benjamin C. Mallory ◽  
Vincent J. Carey ◽  
Dennis L. Kasper ◽  
...  
Keyword(s):  
Free Amino ◽  
Animal Experiments ◽  
Bacteroides Fragilis ◽  
Minor Component ◽  
Amino Sugar ◽  
Abscess Formation ◽  
Pcr Analysis ◽  
Capsular Polysaccharides ◽  
Polysaccharide Biosynthesis ◽  
Abdominal Abscesses

ABSTRACT Bacteroides fragilis, though only a minor component of the human intestinal commensal flora, is the anaerobe most frequently isolated from intra-abdominal abscesses. B. fragilis 9343 expresses at least three capsular polysaccharides—polysaccharide A (PS A), PS B, and PS C. Purified PS A and PS B have been tested in animal models and are both able to induce the formation of intra-abdominal abscesses. Mutants unable to synthesize PS B or PS C still facilitate abscess formation at levels comparable to those of wild-type 9343. To determine the contribution of PS A to abscess formation in the context of the intact organism, the PS A biosynthesis region was cloned, sequenced, and deleted from 9343 to produce a PS A-negative mutant. Animal experiments demonstrate that the abscess-inducing capability of 9343 is severely attenuated when the organism cannot synthesize PS A, despite continued synthesis of the other capsular polysaccharides. The PS A of 9343 contains an unusual free amino sugar that is essential for abscess formation by this polymer. PCR analysis of the PS A biosynthesis loci of 50 B. fragilis isolates indicates that regions flanking each side of this locus are conserved in all strains. The downstream conserved region includes two terminal PS A biosynthesis genes that homology-based analyses predict are involved in the synthesis and transfer of the free amino sugar of PS A. Conservation of these genes suggests that this sugar is present in the PS A of all serotypes and may explain the abscessogenic nature of B. fragilis.

scholarly journals Genetic Diversity of the Capsular Polysaccharide C Biosynthesis Region of Bacteroides fragilis

2000 ◽  
Vol 68 (11) ◽  
pp. 6182-6188 ◽  
Author(s):  
Laurie E. Comstock ◽  
Annalisa Pantosti ◽  
Dennis L. Kasper
Keyword(s):  
Genetic Diversity ◽  
Capsular Polysaccharide ◽  
Bacteroides Fragilis ◽  
Hybridization Analysis ◽  
Clinical Isolates ◽  
Pcr Analysis ◽  
Capsular Polysaccharides ◽  
Genetic Approach ◽  
Polysaccharide Biosynthesis ◽  
Putative Aminotransferase

ABSTRACT A genetic approach was used to assess the heterogeneity of the capsular polysaccharide C (PS C) biosynthesis locus ofBacteroides fragilis and to determine whether distinct loci contain genes whose products are likely to be involved in conferring charged groups that enable the B. fragilis capsular polysaccharides to induce abscesses. A collection of 50 B. fragilis strains was examined. PCR analysis demonstrated that the genes flanking the PS C biosynthesis region are conserved, whereas the genes within the loci are heterogeneous. OnlycfiA + B. fragilis strains, which represent 3% of the clinical isolates of B. fragilis, displayed heterogeneity in the regions flanking the polysaccharide biosynthesis genes. Primers were designed in the conserved regions upstream and downstream of the PS C locus and were used to amplify the region from 45 of the 50 B. fragilis strains studied. Fourteen PS C genetic loci could be differentiated by a combination of PCR and extended PCR. These loci ranged in size from 14 to 26 kb. Hybridization analysis with genes from the PS C loci of strains 9343 and 638R revealed that the majority of strains contain homologs ofwcgC (N-acetylmannosamine dehydrogenase),wcfF (putative dehydrogenase), and wcgP(putative aminotransferase). The data suggest that the synthesis of polysaccharides that have zwitterionic characteristics rendering them able to induce abscesses is common in B. fragilis.

scholarly journals Analysis of a Capsular Polysaccharide Biosynthesis Locus of Bacteroides fragilis

1999 ◽  
Vol 67 (7) ◽  
pp. 3525-3532 ◽  
Author(s):  
Laurie E. Comstock ◽  
Michael J. Coyne ◽  
Arthur O. Tzianabos ◽  
Annalisa Pantosti ◽  
Andrew B. Onderdonk ◽  
...  
Keyword(s):  
Capsular Polysaccharide ◽  
Bacteroides Fragilis ◽  
Open Reading Frames ◽  
Capsular Polysaccharides ◽  
Nucleotide Sugar ◽  
Polysaccharide Biosynthesis ◽  
Abdominal Abscesses ◽  
Intra Abdominal Infection ◽  
Reading Frames ◽  
Nucleotide Sugar Biosynthesis

ABSTRACT A major clinical manifestation of infection with Bacteroides fragilis is the formation of intra-abdominal abscesses, which are induced by the capsular polysaccharides of this organism. Transposon mutagenesis was used to locate genes involved in the synthesis of capsular polysaccharides. A 24,454-bp region was sequenced and found to contain a 15,379-bp locus (designated wcf) with 16 open reading frames (ORFs) encoding products similar to those encoded by genes of other bacterial polysaccharide biosynthesis loci. Four genes encode products that are similar to enzymes involved in nucleotide sugar biosynthesis. Seven genes encode products that are similar to sugar transferases. Two gene products are similar toO-acetyltransferases, and two products are probably involved in polysaccharide transport and polymerization. The product of one ORF, WcfH, is similar to a set of deacetylases of the NodB family. Deletion mutants demonstrated that the wcf locus is necessary for the synthesis of polysaccharide B, one of the two capsular polysaccharides of B. fragilis 9343. The virulence of the polysaccharide B-deficient mutant was comparable to that of the wild type in terms of its ability to induce abscesses in a rat model of intra-abdominal infection.

scholarly journals Binding and degradation of fibrinogen by Bacteroides fragilis and characterization of a 54 kDa fibrinogen-binding protein

Microbiology ◽  
2010 ◽  
Vol 156 (8) ◽  
pp. 2516-2526 ◽  
Author(s):  
Simon Houston ◽  
Garry W. Blakely ◽  
Andrew McDowell ◽  
Lorraine Martin ◽  
Sheila Patrick
Keyword(s):  
Binding Protein ◽  
Bacteroides Fragilis ◽  
Abscess Formation ◽  
Human Fibrinogen ◽  
Abdominal Abscesses ◽  
Fibrinogen Binding ◽  
Major Structural Component ◽  
Culture Isolate ◽  
Normal Human ◽  
Gastro Intestinal Tract

Bacteroides fragilis is a bacterium that resides in the normal human gastro-intestinal tract; however, it is also the most commonly isolated Gram-negative obligate anaerobe from human clinical infections, such as intra-abdominal abscesses, and the most common cause of anaerobic bacteraemia. Abscess formation is important in bacterial containment, limiting dissemination of infection and bacteraemia. In this study, we investigated B. fragilis binding and degradation of human fibrinogen, the major structural component involved in fibrin abscess formation. We have shown that B. fragilis NCTC9343 binds human fibrinogen. A putative Bacteroides fragilis fibrinogen-binding protein, designated BF-FBP, identified in the genome sequence of NCTC9343, was cloned and expressed in Escherichia coli. The purified recombinant BF-FBP bound primarily to the human fibrinogen Bβ-chain. In addition, we have identified fibrinogenolytic activity in B. fragilis exponential phase culture supernatants, associated with fibrinogenolytic metalloproteases in NCTC9343 and 638R, and cysteine protease activity in YCH46. All nine clinical isolates of B. fragilis examined degraded human fibrinogen; with eight isolates, initial Aα-chain degradation was observed, with varying Bβ-chain and γ-chain degradation. With one blood culture isolate, Bβ-chain and γ-chain degradation occurred first, followed by subsequent Aα-chain degradation. Our data raise the possibility that the fibrinogen-binding protein of B. fragilis, along with a variety of fibrinogenolytic proteases, may be an important virulence factor that facilitates dissemination of infection via reduction or inhibition of abscess formation.

scholarly journals A Family of Transcriptional Antitermination Factors Necessary for Synthesis of the Capsular Polysaccharides of Bacteroides fragilis

2009 ◽  
Vol 191 (23) ◽  
pp. 7288-7295 ◽  
Author(s):  
Maria Chatzidaki-Livanis ◽  
Michael J. Coyne ◽  
Laurie E. Comstock
Keyword(s):  
Mutational Analysis ◽  
Bacteroides Fragilis ◽  
Small Region ◽  
Capsular Polysaccharides ◽  
Single Strain ◽  
Conserved Residues ◽  
Polysaccharide Biosynthesis ◽  
Transcriptional Termination ◽  
Amino Acid Segment ◽  
By Products

ABSTRACT A single strain of Bacteroides fragilis synthesizes eight distinct capsular polysaccharides, designated PSA to PSH. These polysaccharides are synthesized by-products encoded by eight separate polysaccharide biosynthesis loci. The genetic architecture of each of these eight loci is similar, including the fact that the first gene of each locus is a paralog of the first gene of each of the other PS loci. These proteins are designated the UpxY family, where x is replaced by a to h, depending upon the polysaccharide locus from which it is produced. Mutational analysis of three separate upxY genes demonstrated that they are necessary and specific for transcription of their respective polysaccharide biosynthesis operon and that they function in trans. Transcriptional reporter constructs, reverse transcriptase PCR, and deletion analysis demonstrated that the UpxYs do not affect initiation of transcription, but rather prevent premature transcriptional termination within the 5′ untranslated region between the promoter and the upxY gene. The UpxYs have conserved motifs that are present in NusG and NusG-like proteins. Mutation of two conserved residues within the conserved KOW motif abrogated UpaY activity, further confirming that these proteins belong to the NusG-like (NusGSP) family. Alignment of highly similar UpxYs led to the identification of a small region of these proteins predicted to confer specificity for their respective loci. Construction of an upaY-upeY hybrid that produced a protein in which a 17-amino-acid segment of UpaY was changed to that of UpeY altered UpaY's specificity, as it was now able to function in transcriptional antitermination of the PSE biosynthesis operon.

scholarly journals Efficacy of trovafloxacin for treatment of experimental Bacteroides infection in young and senescent mice.

1997 ◽  
Vol 41 (9) ◽  
pp. 1933-1936 ◽  
Author(s):  
H Thadepalli ◽  
S K Chuah ◽  
U Reddy ◽  
N Hanna ◽  
R Clark ◽  
...  
Keyword(s):  
Adverse Effect ◽  
Therapeutic Response ◽  
Bacteroides Fragilis ◽  
Therapeutic Outcome ◽  
Abscess Formation ◽  
Abdominal Abscesses ◽  
Old Mice ◽  
Bacteroides Infection ◽  
Young Mice

We investigated the efficacy of trovafloxacin, a new quinolone, in comparison with that of clindamycin in the treatment of intra-abdominal abscesses caused by Bacteroides fragilis in young and senescent mice. The development of abscess formation, the number of viable organisms, and antibiotic concentrations were measured, and the values for young and old mice were compared. Trovafloxacin was well distributed to the tissues in both young and old animals. Although the pharmacokinetics and concentrations of trovafloxacin in serum were similar between young and old mice, the levels in tissue were higher in senescent mice than in young mice. Trovafloxacin therapy sterilized abscesses in 94% of young mice and in 73% of old mice, but this difference was not significant. This therapeutic response to trovafloxacin was similar to that seen with clindamycin. These results suggest that aging may not have any adverse effect on the therapeutic outcome for intra-abdominal abscesses caused by B. fragilis.

scholarly journals Mutational analysis of genes implicated in LPS and capsular polysaccharide biosynthesis in the opportunistic pathogen Bacteroides fragilis

Microbiology ◽  
2009 ◽  
Vol 155 (4) ◽  
pp. 1039-1049 ◽  
Author(s):  
Sheila Patrick ◽  
Simon Houston ◽  
Zubin Thacker ◽  
Garry W. Blakely
Keyword(s):  
Capsular Polysaccharide ◽  
Mutational Analysis ◽  
Bacteroides Fragilis ◽  
Gene Clusters ◽  
Opportunistic Pathogen ◽  
Extracellular Polysaccharides ◽  
Phase Variable ◽  
Multiple Gene ◽  
Polysaccharide Biosynthesis ◽  
Group 1

The obligate anaerobe Bacteroides fragilis is a normal resident of the human gastrointestinal tract. The clinically derived B. fragilis strain NCTC 9343 produces an extensive array of extracellular polysaccharides (EPS), including antigenically distinct large, small and micro- capsules. The genome of NCTC 9343 encodes multiple gene clusters potentially involved in the biosynthesis of EPS, eight of which are implicated in production of the antigenically variable micro-capsule. We have developed a rapid and robust method for generating marked and markerless deletions, together with efficient electroporation using unmodified plasmid DNA to enable complementation of mutations. We show that deletion of a putative wzz homologue prevents production of high-molecular-mass polysaccharides (HMMPS), which form the micro-capsule. This observation suggests that micro-capsule HMMPS constitute the distal component of LPS in B. fragilis. The long chain length of this polysaccharide is strikingly different from classical enteric O-antigen, which consists of short-chain polysaccharides. We also demonstrate that deletion of a putative wbaP homologue prevents expression of the phase-variable large capsule and that expression can be restored by complementation. This suggests that synthesis of the large capsule is mechanistically equivalent to production of Escherichia coli group 1 and 4 capsules.

Effects of wall inhibitors on thymineless Streptococcus faecalis

1974 ◽  
Vol 20 (8) ◽  
pp. 1185-1187 ◽  
Author(s):  
J. G. Bisaillon ◽  
J. de Repentigny ◽  
L. G. Mathieu
Keyword(s):  
Free Amino ◽  
Death Rate ◽  
Sugar Content ◽  
Amino Sugar ◽  
Streptococcus Faecalis ◽  
Inhibition Of Growth ◽  
Thymineless Death

Thymine-requiring (thy−) mutants of Streptococcus faecalis and S. faecalis var. zymogenes were isolated by selection with aminopterin. D-Cycloserine increased the thymineless death rate of a S. faecalis thy− strain, whereas bacitracin, penicillin, and vancomycin decreased it. A S. faecalis var. zymogenes thy− strain behaved similarly, except that penicillin increased its thymineless death rate. D-Alanine reversed the effect of D-cycloserine on both strains. Because of the specificity of the inhibition, our observations suggest that mucopeptide synthesis occurs in S. faecalis not only during growth but also during thymineless death. The variations in free amino sugar content resulting from D-cycloserine inhibition of growth and thymineless death also support this conclusion.

scholarly journals Bacteroides fragilis strains express multiple capsular polysaccharides.

1993 ◽  
Vol 31 (7) ◽  
pp. 1850-1855 ◽  
Author(s):  
A Pantosti ◽  
A O Tzianabos ◽  
B G Reinap ◽  
A B Onderdonk ◽  
D L Kasper
Keyword(s):  
Bacteroides Fragilis ◽  
Capsular Polysaccharides

Capsular Polysaccharides and Lipopolysaccharides from Two Strains of Bacteroides fragilis

1984 ◽  
Vol 6 (Supplement_1) ◽  
pp. S25-S29 ◽  
Author(s):  
Dennis L. Kasper ◽  
Alf A. Lindberg ◽  
Andrej Weintraub ◽  
Andrew B. Onderdonk ◽  
Jörgen Lönngren
Keyword(s):  
Bacteroides Fragilis ◽  
Capsular Polysaccharides

scholarly journals Immunochemical and Biological Characterization of Three Capsular Polysaccharides from a Single Bacteroides fragilisStrain

2001 ◽  
Vol 69 (4) ◽  
pp. 2339-2344 ◽  
Author(s):  
Wiltrud M. Kalka-Moll ◽  
Ying Wang ◽  
L. E. Comstock ◽  
Sylvia E. Gonzalez ◽  
Arthur O. Tzianabos ◽  
...  
Keyword(s):  
Capsular Polysaccharide ◽  
Basic Structure ◽  
Structural Features ◽  
Capsular Polysaccharides ◽  
Polysaccharide Complex ◽  
Abdominal Abscesses ◽  
Colonic Flora ◽  
Surface Polysaccharides ◽  
Normal Human ◽  
Net Charges

ABSTRACT Although Bacteroides fragilis accounts for only 0.5% of the normal human colonic flora, it is the anaerobic species most frequently isolated from intra-abdominal and other infections with an intestinal source. The capsular polysaccharides of B. fragilis are part of a complex of surface polysaccharides and are the organism's most important virulence factors in the formation of intra-abdominal abscesses. Two capsular polysaccharides from strain NCTC 9343, PS A1 and PS B1, have been characterized structurally. Their most striking feature is a zwitterionic charge motif consisting of both positively and negatively charged substituent groups on each repeating unit. This zwitterionic motif is essential for abscess formation. In this study, we sought to elucidate structural features of the capsular polysaccharide complex of a commonly studied B. fragilisstrain, 638R, that is distinct from strain 9343. We sought a more general picture of the species to establish basic structure-activity and structure-biosynthesis relationships among abscess-inducing polysaccharides. Strain 638R was found to have a capsular polysaccharide complex from which three distinct carbohydrates could be isolated by a complex purification procedure. Compositional and immunochemical studies demonstrated a zwitterionic charge motif common to all of the capsular polysaccharides that correlated with their ability to induce experimental intra-abdominal abscesses. Of interest is the range of net charges of the isolated polysaccharides—from positive (PS C2) to balanced (PS A2) to negative (PS 3). Relationships among structural components of the zwitterionic polysaccharides and their molecular biosynthesis loci were identified.

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