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N R Krieg

Publications and source records attributed to N R Krieg.

At least 19 recordsLinked to original sources

Cytochrome composition and oxygen-dependent respiration-driven proton translocation in Wolinella curva, Wolinella recta, Bacteroides ureolyticus, and Bacteroides gracilis.

The membrane fractions of the microaerobically grown type strains of Wolinella curva, Wolinella recta, Bacteroides ureolyticus, and Bacteroides gracilis contained membrane-bound cytochrome b, cytochrome c, and CO-binding cytochrome c. Soluble cytochrome c and CO-binding cytochrome c were also present. Although B. gracilis is oxidase negative, it possessed cytochrome c. With H2 or formate as the electron donor, proton efflux from anaerobic cells occurred upon addition of a pulse of oxygen. With formate as the electron donor, the H+/O ratios of W. curva, W. recta, B. ureolyticus, and B. gracilis were 0.75, 1.66, 2.06, and 2.04, respectively. With H2 as the electron donor, the H+/O ratios of W. curva, B. ureolyticus, and B. gracilis were 1.25, 1.97, and 2.36, respectively. Proton translocation was inhibited by the protonophore carbonylcyanide m-chlorophenylhydrazone. The results confirm that the organisms are not anaerobes but are microaerophiles capable of respiring with oxygen.

Bacteroides

Wolinella recta, Wolinella curva, Bacteroides ureolyticus, and Bacteroides gracilis are microaerophiles, not anaerobes.

Although the nonfermentative, asaccharolytic, putative anaerobes Wolinella curva, Wolinella recta, Bacteroides ureolyticus, and Bacteroides gracilis are phylogenetically related to the true campylobacters, the type strains of these species exhibited O2-dependent microaerophilic growth in brucella broth and on brucella agar. The optimum O2 levels for growth of these strains ranged from 4 to 14% in brucella broth and from 2 to 8% on brucella agar, when H2 was provided as the electron donor. No growth occurred under 21% O2, and scant or no growth occurred under anaerobic conditions unless fumarate or nitrate was provided as a terminal electron acceptor. Aspartate, asparagine, and malate also served as apparent electron acceptors. The organisms were catalase negative and, except for B. gracilis, oxidase positive. Catalase added to brucella broth enhanced growth. O2 uptake by all species was inhibited by cyanide and 2-heptyl-4-hydroxyquinoline N-oxide. We concluded that these organisms are not anaerobes but instead are microaerophiles, like their campylobacter relatives.

Anaerobiosis

Catalase activity in Campylobacter jejuni: comparison of a wild-type strain with an aerotolerant variant.

A comparison of Campylobacter jejuni VPI strain H840 (ATCC 29428), which can grow at O2 levels up to 15%, with variant strain MC711-01 (which can grow at O2 levels up to 21-26%) indicated that the specific activity of catalase in crude cell extracts was higher in the variant by a factor of 1.6 to 2.5, depending on cultural conditions. Smaller differences occurred with superoxide dismutase activity, while peroxidase activities were invariably lower in the variant strain. The variant strain was much more resistant than the wild type to the bactericidal effects of H2O2. The results suggest that catalase activity might be one of the factors associated with the greater tolerance of O2 by the variant strain. However, both strains became more susceptible to H2O2 when cultures were initially grown at 6% O2 and then shifted to 21% O2; thus the role of catalase in the oxygen tolerance of C. jejuni is probably minor.

Campylobacter fetus

Bacterial classification: an overview.

Classification of bacteria evolved from limited subjective groupings to general, more objective arrangements based on overall phenotypic similarities. However, classifications based on phenotypic characters lack stability, whereas those based on genetic relatedness tend to be stable. DNA-DNA hybridization has proven to be extremely useful in resolving taxonomic problems at the species level. Broad relationships among bacteria have been identified by comparing ribosomal RNA cistrons; however, many groups based on ribosomal RNA analysis are not easily definable in terms of phenotypic similarities. Unless resolved, these problems could lead to the establishment of two separate classification systems, one phylogenetic and the other practical.

Bacteria

Effect of incubation temperature, ageing, and bisulfite content of unsupplemented Brucella agar on aerotolerance of Campylobacter jejuni.

A mutant strain of Campylobacter jejuni ATCC 29428 was isolated that grows on unsupplemented Brucella agar at O2 levels as high as 21% at 37 degrees C. While measuring the degree of aerotolerance of this mutant on unsupplemented Brucella medium and comparing it with that of the wild type, we found considerable variation among our estimates. As measured by colony counts on unsupplemented Brucella agar incubated at various oxygen levels, the degree of aerotolerance was affected by incubation temperature and the age of the medium. Aerotolerance was consistently higher on plates incubated at 42 degrees C than at 37 degrees C. When the commercial dehydrated Brucella medium that was used to prepare the Brucella agar was stored in a beaker for 2.5 months, the degree of aerotolerance of C. jejuni was decreased. Addition of 0.01% sodium bisulfite reversed this inhibition. Storage of bottles of hydrated Brucella agar for 1.5 months also resulted in a decreased aerotolerance; again, in addition of 0.01% bisulfite reversed the effect. Aerotolerance was greatly decreased when Brucella agar was prepared from all its individual components except 0.01% bisulfite. The results indicate that the bisulfite component of Brucella agar deteriorates during storage of the dehydrated and hydrated media, and that this deterioration can affect measurements of aerotolerance.

Aerobiosis

DNA homology studies of the catalase-negative campylobacters and "Campylobacter fecalis," an emended description of Campylobacter sputorum, and proposal of the neotype strain of Campylobacter sputorum.

Twenty-three strains of catalase-negative campylobacters and five strains of "Campylobacter fecalis," which is catalase-positive, were examined by DNA hybridization experiments. These organisms formed four distinct DNA homology groups corresponding to Campylobacter sputorum, Campylobacter mucosalis, Campylobacter concisus, and a currently unnamed group referred to as the "catalase-negative or weak" (CNW) strains. The strains were further characterized to determine which phenotypic characteristics provide the most reliable identification at the species level. Campylobacter sputorum ssp. sputorum, C. sputorum ssp. bubulus, and "C. fecalis" could not be distinguished by DNA homology; consequently, it is proposed that these three taxa be considered as biovars of C. sputorum. The description of C. sputorum is emended accordingly. ATCC strain 35980 is proposed as the neotype strain of C. sputorum.

Campylobacter

Studies of the microaerophilic nature of Campylobacter fetus subsp. jejuni. II. Role of exogenous superoxide anions and hydrogen peroxide.

The addition of bovine superoxide dismutase to Brucella broth or Brucellar agar greatly echanced the oxygen tolerance of Campylobacter fetus subsp. jejuni strain H840 (ATCC 29428). Catalase also enhanced oxygen tolerance, but to a lesser extent. These enzymes must act externally to the bacteria. All of the diverse compounds which enhance oxygen tolerance of C. fetus, including nor-epinephrine and a combination of ferrous sulfate, sodium metabisulfite, and sodium pyruvate, share the ability to quench either superoxide anions or hydrogen peroxide. On the basis of these and other data, we propose that C. fetus is more sensitive to exogenous superoxide anions and hydrogen peroxide than are aerotolerant bacteria, despite the occurrence of superoxide dismutase and catalse activities in C. fetus. Compounds that enhance oxygen tolerance in C. fetus appear to act by quenching superoxide anions and hydrogen peroxide which occur spontaneously in the culture medium.

Campylobacter

Improved media for growth and aerotolerance of Campylobacter fetus.

The microaerophilic nature of Campylobacter fetus has complicated its recovery from human and animal sources. In this study, modifications of brucella agar and broth were tested for enhancement of growth and aerotolerance of 64 strains of C. fetus, representing each subspecies. Brucella agar supplemented with 0.025% each FeSO4 7H2O, sodium metabisulfite, and sodium pyruvate, supported growth of 98, 77, and 63% of the strains at 6% O2, 17% O2, and 21% O2, respectively. Unsupplemented brucella agar supported growth of 94, 48, and 20% of the strains. Brucella broth supplemented with 0.2% FeSO4.7H2O, 0.025% sodium metabisulfite, and 0.05% sodium pyruvate supported growth of 98% of the strains at 21% O2, compared to 75% with unsupplemented brucella broth. With both the supplemented agar and broth, growth responses occurred 1 to 2 days earlier than usual. Growth and aerotolerance of three strains of Campylobacter sputorum subsp. bubulus were not enhanced by the supplements.

Campylobacter

A taxonomic study of the Spirillum lipoferum group, with descriptions of a new genus, Azospirillum gen. nov. and two species, Azospirillum lipoferum (Beijerinck) comb. nov. and Azospirillum brasilense sp. nov.

Sixty-one strains of the root-associated nitrogen fixer Spirillum lipoferum exhibited a similar morphology in peptone--succinate salts medium: vibrioid cells having a diameter of 1.0 micrometer. When grown in broth the cells had a single polar flagellum, but when grown on agar at 30 degrees C lateral flagella of shorter wavelength were also formed. The DNA base composition was 69--71 mol% guanine + cytosine when determined by thermal denaturation. DNA homology experiments indicated the occurrence of two distinct but related homology groups: 46 strains were in group I and 15 strains were in group II. Group II strains were distinguished by their ability to use glucose as a sole carbon source for growth in nitrogen-free medium, by their production of an acidic reaction in a peptone-based glucose medium, by their requirement for biotin, and by their formation of wider, longer, S-shaped or helical cells in semisolid nitrogen-free malate medium. The results indicate that two species exist, and on the basis of their characteristics it is proposed that they be assigned to a new genus, Azospirillum. Strians belonging to group II are named A. lipoferum (Beijerinck) comb. nov., while those belonging to group I are named A. brasilense sp. nov. Strain Sp 59b (ATCC29707) is proposed as the neotype strain for A. lipoferum, and strain Sp 7 (ATCC 29145) is proposed as the type strain for A. brasilense.

Culture Media

Stimulatory effect of dihydroxyphenyl compounds on the aerotolerance of Spirillum volutans and Campylobacter fetus subspecies jejuni.

The aerotolerance of the microaerophilic bacterium Spirillum volutans was greatly stimulated in a defined medium by the presence of dihydroxyphenyl ferric iron-binding compounds such as nor-epinephrine at 10(-5) to 10(-6) M. Dihydroxyphenyl compounds at 2 X 10(-4) M, or iron salts (ferrous or ferric) at high concentration, greatly increased the aerotolerance of a strain of Campylobacter fetus subsp. jejuni when grown on streak plates of Brucella agar. The results suggest that the microaerophilism of these organisms might in part be caused by a failure to synthesize microbial ferric iron-binding compounds at sufficient levels to support aerobic growth.

Aerobiosis

Transport and catabolism of D-fructose by Spirillum itersomii.

Spirillum itersonii ATCC 12639 utilized d-fructose but neither d-glucose nor d-gluconate as a sole source of carbon and energy. The substrate saturation kinetics for d-fructose and d-glucose uptake by whole cells indicated the presence of a carrier-mediated transport system for d-fructose but not for d-glucose. The d-fructose uptake activity was induced (10- to 12-fold increase) during growth on d-fructose-Casamino Acids (CA) or d-glucose-CA medium, but not CA alone. d-Fructose uptake activity was stimulated by Na(+) or Li(+), but was inhibited by KCN, NaN(3), 2,4-dinitrophenol, and p-chloromercuribenzoate. High specific activities of glucokinase, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydratase, and 2-keto-3-deoxy-6-phosphogluconate aldolase were detected in extracts of cells cultured on d-fructose-CA medium. These enzymatic activities were undetectable in extracts of cells grown in CA or succinate-CA medium. No decrease in the maximally induced specific activities of these enzymes occurred after the addition of succinate to cells during exponential growth on d-fructose-CA. Fructose 1,6-diphosphate aldolase and glucose-6-phosphate isomerase specific activities were approximately the same irrespective of cultural conditions. These results indicated that d-glucose was not utilized by cells of S. itersonii because this bacterium was impermeable to this hexose.

Alcohol Oxidoreductases

Sugar catabolism in Aquaspirillum gracile.

Aquaspirillum (Spirillum) gracile is one of the few spirilla that cause acidification of the medium when cultured with sugars. Acidic reactions have been reported only for d-glucose, d-galactose, and l-arabinose, and the mode of attack of these sugars has not been previously investigated. The soluble portion of extracts of glucose-cultured cells of A. gracile ATCC 19624 was found by spectrophotometric methods to contain enzyme activities characteristic of the Entner-Doudoroff and Embden-Meyerhof-Parnas pathways. No activity for 6-phosphogluconate dehydrogenase (EC 1.1.1.44) was detected. Pyridine nucleotide-linked dehydrogenase activities for l-arabinose and d-galactose (EC 1.1.1.46 and EC 1.1.1.48) occurred in the soluble fraction of cells cultured with either sugar. Glucose-cultured cells contained not only glucokinase (EC 2.7.1.2) and glucose-6-phosphate dehydrogenase (EC 1.1.1.49) activities but also glucose dehydrogenase (EC 1.1.1.47) activity. Enzymes capable of oxidizing gluconate were not detectable, but gluconokinase (EC 2.7.1.12) activity was present. Paper chromatographic analysis of the spent culture supernatant media from glucose-cultured cells indicated an accumulation of gluconic acid, and this was confirmed by enzymatic methods. Evidence is presented for the production of d-galactonic and l-arabonic acids in cultures containing d-galactose or l-arabinose, respectively.

Alcohol Oxidoreductases