RESUMO
Forty-six anaerobic gram-negative bacilli, including 26 members of the Bacteroides fragilis group (BFG), were examined for capsules by the India ink technic. Thirty-five were encapsulated, including all the BFG strains. As a follow-up, seven of these isolates and two previously studied reference strains (B. fragilis ATCC 23745 and Bacteroides vulgatus ATCC 8482) were examined for capsules by ultrastructural cytochemistry. Using the periodic acid thiocarbohydrazide silver proteinate (PATCSP) method of Thiéry, all the BFG examined were encapsulated. In addition to the reference strains, this included one strain of B. fragilis and four of Bacteroides thetaiotaomicron. One non-BFG strain showed no capsular material. Differences between these results and those reported previously with the ruthenium red technic may reflect species differences in the chemical composition of Bacteroides capsules.
Assuntos
Bacteroides fragilis/citologia , Bacteroides fragilis/análise , Bacteroides fragilis/ultraestrutura , Membrana Celular/análise , Histocitoquímica , Tinta , Polissacarídeos Bacterianos/análiseRESUMO
The occurrence of free ceramides at high concentrations was demonstrated in the chloroform-methanol extractable lipids of Bacteroides fragilis NCTC 9343. The long-chain bases were isolated from the free ceramides and identified as branched and normal saturated dihydroxy bases with carbon chains consisting of 17, 18, and 19 atoms. The major fatty acid was 3-hydroxy 15-methylhexadecanoic acid. The major molecular species of the ceramides were identified by gas chromatography-mass spectrometry and gas chromatography of the cleaved products as LCB-d-iso17: 0-3-OH iso17: 0 FA, LCB-d-anteiso17: 0-3-OH iso17: 0 FA, LCB-d-iso18: 0-3-OH iso17: 0 FA, and LCB-d-anteiso19: 0-3-OH iso17: 0 FA.
Assuntos
Bacteroides fragilis/análise , Ceramidas/análise , Cromatografia Gasosa , Cromatografia em Camada Fina , Ácidos Graxos/análise , Espectrometria de Massas , Concentração Osmolar , Espectrofotometria Infravermelho , Esfingolipídeos/análiseRESUMO
We analyzed the ability of 120 encapsulated strains of B. fragilis to agglutinate guinea pig and human red blood cells. Sixteen strains showed a strong hemagglutination (HA) ability, 21 strains a moderate HA ability, 7 strains a weak HA ability and 74 strains did not agglutinate the tested red blood cells. Six strains tested from each HA group were able to adhere to cheek epithelial cells and to a cultured human intestinal cell line. Hemagglutinating strains were the most adhesive. By electron microscopy, pilus-like structures were found in three of the encapsulated adhesive strains. Treatment of the bacterial cells with pronase E reduced both HA ability and adherence of piliated encapsulated, and of piliated non-encapsulated strains. Glucosidase treatment of cells reduced HA activity and adherence of piliated encapsulated and of non-piliated encapsulated strains. Finally, it was found that hemagglutinating strains are more frequently isolated from clinical specimens (55%) than from feces of healthy donors (20%).
Assuntos
Aderência Bacteriana , Bacteroides fragilis/fisiologia , Fímbrias Bacterianas/fisiologia , Hemaglutinação , Bacteroides fragilis/análise , Bacteroides fragilis/patogenicidade , Humanos , VirulênciaRESUMO
The outer membrane protein (OMP) profiles of four different strains of Bacteroides fragilis, as determined by Coomassie blue stained polyacrylamide gels, were compared after growth in broth culture and in the mouse peritoneal cavity. There was no induction of the expression of large quantities of novel OMP after growth in vivo. Mouse immunoglobulin G and albumin were associated with the bacterial OMP, but could be removed by washing.
Assuntos
Proteínas da Membrana Bacteriana Externa/biossíntese , Bacteroides fragilis/metabolismo , Animais , Proteínas da Membrana Bacteriana Externa/análise , Bacteroides fragilis/análise , Bacteroides fragilis/crescimento & desenvolvimento , Meios de Cultura , Cultura em Câmaras de Difusão , Eletroforese em Gel de Poliacrilamida , Immunoblotting , Imunoglobulina G/análise , Camundongos , Cavidade Peritoneal , Albumina Sérica/análiseRESUMO
The growth of capsulate and non- capsulate Bacteroides fragilis in chambers implanted in the mouse peritoneal cavity was compared. Capsulate and essentially non- capsulate (less than 1% capsulate ) populations of B. fragilis strains NCTC9343 and NCTC10584 consistently grew exponentially to greater than 10(9) cfu/ml within 24 h in vivo, and low numbers of capsulate bacteria were maintained in the essentially non- capsulate population; however, the degree of capsulation of the capsulate population decreased by more than 60%. B. fragilis ATCC23745 differed from strains NCTC9343 and NCTC10584 in that growth was unpredictable and only occurred in some of the implanted chambers. Capsule production by cells of strain ATCC23745 varied from chamber to chamber: sometimes the proportion of capsulate cells increased after prolonged implantation. This could occur with either an increase or decrease in viable numbers in vivo and also after in-vitro incubation of this strain in chambers. The survival of capsulate and non- capsulate B. fragilis strains NCTC9343 and ATCC23745 was compared in aerobic and anaerobic conditions in vitro. In anaerobic conditions, capsulate and non- capsulate strain NCTC9343 survived equally well, whereas capsulate ATCC23745 survived better than its non- capsulate variants. Capsulate populations of both strains survived better than non- capsulate in aerobic conditions.
Assuntos
Bacteroides fragilis/crescimento & desenvolvimento , Polissacarídeos Bacterianos/fisiologia , Aerobiose , Anaerobiose , Animais , Bacteroides fragilis/análise , Bacteroides fragilis/citologia , Feminino , Camundongos , Cavidade Peritoneal , Polissacarídeos Bacterianos/análiseRESUMO
The determinant responsible for the ability of Bacteroides spp. to inhibit polymorph phagocytic killing of aerobic organisms has not yet been identified. Therefore, the roles of lipopolysaccharide and capsular polysaccharide of B. fragilis were investigated. Serum-resistant and serum-sensitive strains of Proteus mirabilis were used to indicate inhibition of phagocytic killing and serum killing of aerobes. Whole organisms of B. fragilis, purified lipopolysaccharide and capsular polysaccharide were added to an in-vitro phagocytosis system. Results showed that greater than 10(7) bacteroides/ml inhibited both serum and phagocytic killing. Concentrations below 10(7)/ml had little effect on either process. Purified capsular polysaccharide (10 or 100 micrograms/ml), either alone in the system or in combination with sub-inhibitory concentrations of B. fragilis also markedly inhibited serum and phagocytic killing. Lipopolysaccharide (9 micrograms/ml) appeared relatively inert. B. ovatus, reputedly non-capsulated, produced identical results to those obtained with B. fragilis, but an encapsulated strain of Streptococcus pneumoniae did not inhibit serum or phagocytic killing.
Assuntos
Bacteroides fragilis/fisiologia , Atividade Bactericida do Sangue , Lipopolissacarídeos/fisiologia , Neutrófilos/imunologia , Fagocitose , Polissacarídeos Bacterianos/fisiologia , Bacteroides fragilis/análise , Humanos , Neutrófilos/microbiologia , Proteus mirabilis/imunologia , Proteus mirabilis/ultraestrutura , Vacúolos/microbiologiaRESUMO
The effect of capsulation on the ability of Bacteroides fragilis, B. asaccharolyticus and anaerobic gram-positive cocci to induce bacteraemia and seeding to various organs was investigated. The test species were injected into mice subcutaneously alone, or mixed with other aerobic or facultative organisms. Capsulate anaerobes were isolated more frequently from the blood, spleen, liver, and kidneys of infected animals than were non-capsulate organisms. After injection of single anaerobic strains, capsulate organisms were recovered from 163 (39%) of 420 animals; non-capsulate anaerobes were recovered from only 14 (3%) of 420 animals. After injection of B. fragilis mixed with aerobic or facultative organisms, the capsulate B. fragilis strain was isolated more often and for longer periods than the non-capsulate strain. Capsulate B. fragilis was also recovered more often 5 days after injection with other organisms, than when injected alone. These data demonstrate that capsulate Bacteroides spp. and anaerobic gram-positive cocci are more virulent than non-capsulate strains in single and mixed infections.
Assuntos
Abscesso/microbiologia , Infecções por Bacteroides/microbiologia , Bacteroides/patogenicidade , Peptostreptococcus/patogenicidade , Sepse/microbiologia , Animais , Bacteroides/análise , Bacteroides/isolamento & purificação , Bacteroides fragilis/análise , Bacteroides fragilis/isolamento & purificação , Bacteroides fragilis/patogenicidade , Rim/microbiologia , Fígado/microbiologia , Masculino , Camundongos , Peptostreptococcus/análise , Peptostreptococcus/isolamento & purificação , Polissacarídeos Bacterianos , Baço/microbiologia , VirulênciaRESUMO
Living, heat or formalin killed Bacteroides fragilis and a crude preparation of their cell walls were examined by the Boyden technique for chemotactic activity upon guinea pig peritoneal exudate cells. Their relative chemotactic activity ranged from 3.0 to 5.2 compared to an average value of 6.4 for the positive control, an endotoxic culture filtrate of Escherichia coli. A culture filtrate of B. fragilis and an index of 3.7. Miocrogram quantities of cytoplasmic preparations obtained by ammonium sulphate precipitation had chemotactic indices ranging from 2.8 to 6.4, the highest value being displayed by the precipitate formed between 50 and 75% saturation with ammonium sulphate. This fraction retained leucotactic activity after exposure to strong acid and heat. The leucotactic potency of these fractions did not correlate directly with their protein content. Further precipitation of the most active fraction with 80% ethanol revealed that there was little chemotactic activity attributable to polysaccharides. Gas liquid chromatography of a chloroform-methanol extract of the cells which had a chemotactic index of 6.1 revealed the presence of more than thirty fatty acids ranging in carbon length from C8 to C25. These results suggest a role of lipids as initiators of the leucotactic response associated with infections caused by B. fragilis.
Assuntos
Bacteroides fragilis/análise , Quimiotaxia , Lipídeos/fisiologia , Animais , Bacteroides fragilis/fisiologia , Ácidos Graxos/fisiologia , CobaiasRESUMO
Previous nucleic acid association studies have identified at least nine deoxyribonucleic acid (DNA) homology classes of the Bacteroides fragilis group of organisms. Using these classes as a taxonomic framework, we have screened representative strains of the B. fragilis group for the presence of extrachromosomal (plasmid) DNA. [3H]thymidine-labeled cell lysates were subjected to sodium dodecyl sulfate-salt precipitation, and supernatant fractions from such preparations were analyzed using cesium chloride-ethidium bromide equilibrium centrifugation. One strain from each group was examined in this fashion. Five of the strains were judged to contain no detectable plasmid DNA; however, four strains were observed to yield satellite bands corresponding to covalently closed circular plasmid DNA. Plasmid DNA from such gradients was subjected to velocity sedimentation through both neutral and alkaline sucrose gradients to determine molecular size. A 23 X 10(6)-molecular-weight plasmid was found in a B. fragilis strain representing one DNA homology group of this species, whereas a 3 X 10(6)-molecular-weight plasmid was found in a B. fragilis strain representing a second homology group. Similarly, a 31 X 10(6)-molecular-weight plasmid was found in a Bacteroides thetaiotaomicron strain representing one DNA homology group of this species, whereas a 3 X 10(6)-molecular-weight plasmid was found in a B. thetaiotaomicron strain representing a second homology group. In all instances, the small-molecular weight plasmids were present to the extent of about 15 copies per chromosomal equivalent, whereas the large plasmids were present to the extent of approximately 1 copy per chromosomal equivalent. The biological function of these plasmids is unknown.
Assuntos
Bacteroides fragilis/análise , Herança Extracromossômica , Plasmídeos , Bacteroides fragilis/classificação , Centrifugação com Gradiente de Concentração , DNA Bacteriano/análise , Peso MolecularRESUMO
Bacteroides fragilis was analysed by pyrolysis gas chromatography (PGC). The influence of temperature rise time, pyrolysis temperature and pyrolysis time on the yield of volatile organic pyrolysis products was studied. The use of short pyrolysis time (8 msec) and a high pyrolysis end temperature (1300 degrees C) was found to provide a high yield. In consecutive analyses of the test organism, under the pyrolysis conditions indicated, the standard deviation of the total yield of volatile organic products was +/- 7%. The pyrolysis technique described suggests that few secondary reactions of the pyrolysis fragments occur.
Assuntos
Bacteroides fragilis/análise , Cromatografia Gasosa/métodos , Cromatografia Gasosa/instrumentação , Temperatura , Fatores de TempoRESUMO
1. Endotoxins isolated from six serotype specific reference strains of the Bacteroides fragilis group were dephosphorylated by treatment with aqueous 50% hydrofluoric acid. 2. Mild acidic hydrolysis of the dephosphorylated endotoxins released 2-keto-3-deoxyaldonic acid, the presence of which was demonstrated by the colorimetric thiobarbituric acid assay (TBA). 3. Thin layer chromatography of the dephosphorylated lipopolysaccharide of B. fragilis IPL E 323 (serotype E2), after acidic hydrolysis, revealed a TBA-positive substance with the same Rf-value as authentical 2-keto-3-deoxyoctolusonic acid (KDO). 4. Quantification of 2-keto-3-deoxyoctonate-in the lipopolysaccharide of B. fragilis IPL E 323 by means of the TBA resulted in a KDO content of 15 nM mg-1 lipopolysaccharide.
Assuntos
Bacteroides fragilis/análise , Endotoxinas/análise , Açúcares Ácidos/análise , Cromatografia em Camada Fina , Endotoxinas/isolamento & purificação , Endotoxinas/metabolismo , Concentração de Íons de Hidrogênio , Hidrólise , Lipopolissacarídeos/isolamento & purificação , Fosforilação , TiobarbitúricosRESUMO
The main fatty acids present in lipopolysaccharides from Bacteroides fragilis NCTC 9343 were identified as 13-methyl-tetradecanoic, D-3-hydroxypentadecanoic, D-3-hydroxyhexadecanoic, D-3-hydroxy-15-methyl-hexadecanoic, and D-3-hydroxyheptadecanoic acids. Of these, 13-methyl-tetradecanoic acid is exclusively ester bound, and 3-hydroxy-15-methyl-hexadecanoic acid is exclusively involved in amide linkage. The other 3-hydroxy fatty acids are both ester and amide bound. All 3-hydroxy fatty acids possess the D configuration, and the 3-hydroxyl group of ester-linked 3-hydroxy fatty acids is not substituted. Lipopolysaccharides of related Bacteroides species (B. thetaiotaomicron, B. ovatus, B. distasonis, and B. vulgatus) showed a fatty acid spectrum with both similar and distinct features compared to that of B. fragilis lipopolysaccharides.
Assuntos
Bacteroides fragilis/análise , Bacteroides/análise , Ácidos Graxos/análise , Lipopolissacarídeos/análise , Fenômenos Químicos , Química , EstereoisomerismoRESUMO
Two strains of Bacteroides asaccharolyticus and two strains of Bacteroides fragilis were analyzed for total fatty acid, total lipid fatty acid, and total bound fatty acid profiles. Extracted lipids and defatted cell residues were subjected to sequential alkaline and acid methanolyses to distinguish ester- and amide-linked fatty acids in each fraction. In the lipid fractions, all the ester-linked fatty acids were nonhydroxylated, whereas all of the amide-linked fatty acids were hydroxylated. In the nonextractable fractions, both hydroxy and nonhydroxy fatty acids were found in both ester and amide linkage, although hydroxy acids predominated. The fatty acid profiles of the bound fractions differed widely from those of the lipid fractions. Bound fatty acid represented approximately 10% of the total cellular fatty acids.
Assuntos
Bacteroides fragilis/análise , Bacteroides/análise , Ácidos Graxos/análise , Amidas , Fenômenos Químicos , Química , Lipídeos/análiseRESUMO
Whole-cell hydrolysates of Bacteroides fragilis, the type species of the genus Bacteroides Castellani and Chalmers 1919, and the genetical closely related species B. vulgatus, B. ovatus, B. eggerthii, B. distasonis, B. uniformis, B. thetaiotaomicron, B. stercoris, B. merdae, and B. caccae were used to determine characteristic carbohydrate patterns by capillary gas chromatography. On the basis of the chemical derivatization of the carbohydrates seven characteristic peaks for peracetylated aldononitriles and nine characteristic peaks for peracetylated o-methyloximes were selected from the carbohydrate fingerprints of the reference strains to prepare a dichotomous identification key. The classification of an unknown strain supposed to belong to the formerly called 'Bacteroides fragilis group' is possible with this key. Some of the advantages of the technique were that the identification of Bacteroides fragilis-like strains requires only 4-5 h after primary isolation and that the bacteria can be exposed to oxygen because viability of the organisms is not necessary. Sophisticated anaerobic techniques can therefore be avoided for identification.
Assuntos
Bacteroides fragilis/classificação , Bacteroides/classificação , Carboidratos/análise , Técnicas de Tipagem Bacteriana , Bacteroides/análise , Bacteroides fragilis/análise , Cromatografia GasosaRESUMO
An immunochemical comparison of outer membrane antigens obtained from five select and biochemically defined strains indicated that there are several serotypes of Bacteroides fragilis. Each strain was serologically defined by individual or by combinations of determinant groups composed of carbohydrates in the form of polysaccharides or glycoproteins. The carbohydrate constituents were tentatively identified as glucose, galactose, fucose, rhamnose, glucosamine, galactosamine, and traces of mannose. Strains were observed to have minor qualitative and major quantitative variations in carbohydrate composition.
Assuntos
Antígenos de Bactérias/análise , Bacteroides fragilis/classificação , Bacteroides fragilis/análise , Carboidratos/análise , Parede Celular/análise , Parede Celular/imunologia , Epitopos , Imunodifusão , SorotipagemRESUMO
2-Methylpentanoic acid and benzoic acid are suggested for use as routine internal standards for gas chromatographic analysis of microbial end products.
Assuntos
Ácidos/análise , Bacteroides fragilis/análise , Benzoatos/normas , Cromatografia Gasosa , Ácidos Pentanoicos/normas , Valeratos/normas , AnaerobioseRESUMO
Changes in cell macromolecular composition during different phases of growth correlated with alterations in morphology of the obligately anaerobic bacterium, Bacteroides fragilis, grown in a complex medium.
Assuntos
Bacteroides fragilis/crescimento & desenvolvimento , Bacteroides fragilis/análise , Meios de Cultura , DNA Bacteriano/metabolismo , Substâncias Macromoleculares , RNA Bacteriano/metabolismo , Análise de RegressãoRESUMO
Sixty-four strains of Bacteroides fragilis isolated from clinical specimens were tested for their bacteriocin production. It showed that 65.6% of them produced bacteriocin(s) and gave different patterns of inhibition. Among them 9 with clear inhibition zones were chosen as the indicator strains. With this indicator s,et 75.2% of 109 clinically isolated B. fragilis strains could be typed, with type 482 as the predominant type. The colonial dissociants gave the same bacteriocin type, but small and opaque colonies tend to lose their bactericidal ability.
Assuntos
Bacteriocinas/análise , Bacteroides fragilis/classificação , Bacteriocinas/farmacologia , Bacteroides fragilis/análise , Bacteroides fragilis/efeitos dos fármacosRESUMO
Three strains of B. thetaiotaomicron of different origin were investigated. Lipopolysaccharides were extracted from the studied strains using a phenol-water method. The best purification of LPS was achieved by digestion with nuclease and subsequent ultracentrifugation. Capsular material (CPS) was obtained from the most heavily encapsulated strain. The preparation were analyzed chemically, and their serological activity was determined. All antigens were active with homologous antibacterial sera in immunodiffusion, crossed immunoelectrophoresis, and passive hemagglutination tests. In the CPS equal amounts of saccharides and proteins were detected. All antigens were analyzed by polyacrylamide gel electrophoresis with SDS. Capsular antigen slowly migrated in the gel in the form of a single band. Migration pattern of lipopolysaccharides of the studied B. thetaiotaomicron strain was characteristic for S-type LPS.