RESUMO
Studies on the physiological characteristics of two obligately anaerobic, rod-shaped bacteria from the human intestinal tract indicated that the organisms represented two previously undescribed species of Bacteroides, for which we propose the names Bacteroides pectinophilus (type strain, N3) and Bacteroides galacturonicus (type strain, N6). Both strains were pectinophilic; that is, they utilized as fermentable substrates for growth only pectin and a few related compounds. The two species differed significantly from each other in guanine plus cytosine content of the DNA, in substrate utilization patterns, and in other phenotypic characteristics. Both species deesterified pectin by means of an extracellular pectinesterase (EC 3.1.1.11) activity. Polygalacturonate (the main component of deesterified pectin) was depolymerized extracellularly with formation of unsaturated products by both species. The depolymerizing activity required Ca2+, functioned at a higher rate when polygalacturonate was the substrate as compared with pectin, and had an alkaline pH optimum. These data, as well as viscosity decrease studies and identification of products formed from polygalacturonate, indicated that the extracellular depolymerizing activity of either species was characteristic of an exopectate (exopolygalacturonate) lyase. The exopectate lyase activity had an unusual action pattern that resulted in terminal cleavage of unsaturated trigalacturonic acid units from polygalacturonate. An unsaturated trimer was the major product that accumulated in cell-free reaction mixtures, where it was not cleaved further. Growing cells of both Bacteroides species released the exopectate lyase into the external environment by processes that did not involve cell lysis to any significant extent.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Bacteroides/classificação , Intestinos/microbiologia , Pectinas/metabolismo , Bacteroides/metabolismo , Bacteroides/fisiologia , Bacteroides/ultraestrutura , DNA Bacteriano/análise , Fezes/microbiologia , Humanos , Microscopia Eletrônica , Fenótipo , Terminologia como AssuntoRESUMO
A large, obligately anaerobic spirochete (strain PB) was isolated from bovine rumen fluid by a procedure involving rifampin as a selective agent. The helical cells measured 0.6 to 0.7 micron by 12 to 20 micron and possessed approximately 16 periplasmic flagella inserted near each end of the protoplasmic cylinder. The periplasmic flagella were arranged in a bundle wound around the cell body. Strain PB utilized as fermentable substrates various plant polysaccharides (e.g., pectin, arabinogalactan, starch, and inulin) as well as pentoses, hexoses, disaccharides, and uronic acids. Glucose was fermented to acetate, formate, and ethanol, whereas the fermentation of pectin or glucuronic acid yielded only acetate and formate as major end products. Determinations of radioactivity in end products and assays of enzymatic activities indicated that strain PB catabolized glucose via the Embden-Meyerhof pathway. Extracts of cells grown in pectin-containing media possessed relatively high levels of phospho-2-keto-3-deoxygluconate aldolase activity, an enzymatic activity typical of the Entner-Doudoroff pathway. The guanine-plus-cytosine content of the DNA of strain PB (54 mol%) was considerably higher than that of known host-associated anaerobic spirochetes. This study indicates that strain PB represents a new species of Treponema, for which we propose the name Treponema saccharophilum.
Assuntos
Bovinos/microbiologia , Pectinas/metabolismo , Rúmen/microbiologia , Treponema/fisiologia , Animais , Metabolismo dos Carboidratos , Fermentação , Microscopia Eletrônica , Polissacarídeos/metabolismo , Terminologia como Assunto , Treponema/classificação , Treponema/citologia , Treponema/enzimologia , Treponema/isolamento & purificaçãoRESUMO
A selective procedure was used to isolate pectinolytic intestinal bacteria from human subjects. The three isolates with the greatest pectinolytic activity utilized pectin and a few related compounds as fermentable substrates for growth but did not utilize any other compound tested. Thus, their substrate utilization pattern was markedly different from that of previously described intestinal pectinolytic isolates. The three isolates are representatives of a nutritionally defined group of bacteria for which the term pectinophilic is proposed.
Assuntos
Bactérias Anaeróbias/isolamento & purificação , Bacteroides/isolamento & purificação , Intestinos/microbiologia , Pectinas/metabolismo , Bactérias Anaeróbias/enzimologia , Bactérias Anaeróbias/crescimento & desenvolvimento , Bactérias Anaeróbias/metabolismo , Bacteroides/enzimologia , Bacteroides/crescimento & desenvolvimento , Bacteroides/metabolismo , Hidrolases de Éster Carboxílico/metabolismo , Fezes/microbiologia , Fermentação , Humanos , Polissacarídeo-Liases/metabolismoRESUMO
Five strains of obligately anaerobic, pectin-fermenting spirochetes were isolated from the subgingival plaque of humans. The strains produced two extracellular enzymatic activities that functioned in pectin degradation. One of these enzymatic activities was pectin methylesterase (EC 3.1.1.11), and the other was pectate lyase (EC 4.2.2.2) of the endo type. The data indicate that the cumulative action of these two enzymatic activities brought about depolymerization of pectin in spirochete cultures. Pectin- or polygalacturonate-degrading hydrolases were not detected. A cell-associated lyase activity that catalyzed polygalacturonate breakdown was present in one of the spirochete strains. In addition to pectin, the isolates utilized polygalacturonic, glucuronic, or galacturonic acid as fermentable substrate but did not neutral sugars, amino acids, or other substrates tested. Although the oral spirochetes did not ferment hyaluronic acid, one of the strains grew in coculture with a hyaluronidase-producing Peptostreptococcus strain in a medium containing hyaluronic acid as fermentable substrate. Two of the isolates were identified as Treponema pectinovorum strains on the basis of their substrate utilization pattern, end products of fermentation, other phenotypic characteristics, and the guanine-plus-cytosine content of their DNA. Even though the pectinolytic isolates were specialized with respect to the fermentable substrates they utilized, they appeared to compete successfully with other microorganisms in their habitat.
Assuntos
Hidrolases de Éster Carboxílico/isolamento & purificação , Placa Dentária/microbiologia , Polissacarídeo-Liases/isolamento & purificação , Spirochaetales/enzimologia , Hidrolases de Éster Carboxílico/metabolismo , Meios de Cultura/metabolismo , Gengiva/microbiologia , Humanos , Microscopia Eletrônica , Microscopia de Contraste de Fase , Pectinas/metabolismo , Peptostreptococcus/enzimologia , Polissacarídeo-Liases/metabolismo , Spirochaetales/isolamento & purificação , Spirochaetales/ultraestrutura , Treponema/enzimologia , Treponema/isolamento & purificaçãoRESUMO
The pathways of carbohydrate metabolism in Spirochaeta stenostrepta, a free-living, strictly anaerobic spirochete, were studied. The organism fermented glucose to ethyl alcohol, acetate, lactate, CO(2), and H(2). Assays of enzymatic activities in cell extracts, and determinations of radioactivity distribution in products formed from (14)C-labeled glucose indicated that S. stenostrepta degraded glucose via the Embden-Meyerhof pathway. The spirochete utilized a clostridial-type clastic reaction to metabolize pyruvate to acetyl-coenzyme A, CO(2), and H(2), without production of formate. Acetyl-coenzyme A was converted to ethyl alcohol by nicotinamide adenine dinucleotide-dependent acetaldehyde and alcohol dehydrogenase activities. Phosphotransacetylase and acetate kinase catalyzed the formation of acetate from acetyl-coenzyme A. Hydrogenase and lactate dehydrogenase activities were detected in cell extracts. A rubredoxin was isolated from cell extracts of S. stenostrepta. Preparations of this rubredoxin stimulated acetyl phosphate formation from pyruvate by diethylaminoethyl cellulose-treated extracts of S. stenostrepta, an indication that rubredoxin may participate in pyruvate cleavage by this spirochete. Nutritional studies showed that S. stenostrepta fermented a variety of carbohydrates, but did not ferment amino acids or other organic acids. An unidentified growth factor present in yeast extract was required by the organism. Exogenous supplements of biotin, riboflavin, and vitamin B(12) were either stimulatory or required for growth.