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  • 1
    Electronic Resource
    Electronic Resource
    Springer
    Archives of microbiology 150 (1988), S. 477-481 
    ISSN: 1432-072X
    Keywords: Methanogenic bacteria ; Alcohols ; Trace elements ; Methylreductase ; Taxonomy ; Methanogenium thermophilum ; Methanogenium, organophilum ; Methanospirillum hungatei
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A thermophilic coccoid methanogenic bacterium, strain TCI, that grew optimally around 55° C was isolated with 2-propanol as hydrogen donor for methanogenesis from CO2. H2, formate or 2-butanol were used in addition. Each secondary alcohol was oxidized to its ketone. Growth occurred in defined freshwater as well as salt (2% NaCl, w/v) medium. Acetate was required as carbon source, and 4-aminobenzoate and biotin as growth factors. A need for molybdate or alternatively tungstate was shown. Strain TCI was further characterized together with two formerly isolated mesophilic secondary alcohol-utilizing methanogens, the coccoid strain CV and the spirilloid strain SK. The guanine plus cytosine content of the DNA of the three strains was 55,47, and 39 mol%, respectively. Determination of the molecular weights of the methylreductase subunits and sequencing of ribosomal 16S RNA of strains TCI and CV revealed close relationships to the genus Methanogenium. The new isolate TCI is classified as a strain of the existing species, Methanogenium thermophilum (thermophilicum). For strain CV, that uses ethanol or 1-propanol in addition, a classification as new species, Methanogenium organophilum, is proposed. Strain SK is affiliated with the existing species, Methanospirillum hungatei. The ability to use secondary alcohols was also tested with described species of methanogens. Growth with secondary alcohols was observed with Methanogenium marisnigri, Methanospirillum hungatei strain GP1 and Methanobacterium bryantii, but not with Methanospirillum strains JF1 and M1h, Methanosarcina barkeri, Methanococcus species or thermophilic strains or species other than the new isolate TCI.
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  • 2
    ISSN: 1432-072X
    Keywords: Interspecies hydrogen transfer ; Interspecies formate transfer ; Alanine ; Serine ; Glycine fermentation ; Selenium ; Glycine reductase ; Sarcosine reduction ; Betaine reduction ; Eubacterium acidaminophilum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract An obligately anaerobic, rod-shaped bacterium was isolated on alanine in co-culture with H2-scavenging Desulfovibrio and obtained in pure culture with glycine as sole fermentation substrate. The isolated strain, al-2, was motile by a polar to subpolar flagellum and stained Gram-positive. The guanine plus cytosine content of the DNA was 44.0 mol%. Strain al-2 grew in defined, reduced glycine media supplemented with biotin. The pure culture fermented 4 mol glycine to 3 mol acetate, 4 mol ammonia and 2 mol CO2. Under optimum conditions (34°C, pH 7.3), the doubling time on glycine was 60 min and the molar growth yield 7.6 g cell dry mass. Serine was fermented to acetate, ethanol, CO2, H2 and ammonia. In addition, betaine, sarcosine or creatine served as substrates for growth and acetate production if H2, formate or e.g. valine were added as H-donors. In pure culture on alanine under N2, strain al-2 grew very poorly and produced H2 up to a partial pressure of 3.6 kPa (0.035 atm). Desulfovibrio species, Methanospirillum hungatei and Acetobacterium woodii served as H2-scavengers that allowed good syntrophic growth on alanine. The co-cultures also grew on aspartate, leucine, valine or malate. Alanine and aspartate were stoichiometrically degraded to acetate and ammonia, whereas the reducing equivalents were recovered as H2S, CH4 or newly synthetized acetate, respectively. Growth of strain al-2 in co-culture with the hydrogenase-negative, formate-utilizing Desulfovibrio baarsii indicated that a syntrophy was also possible by interspecies formate transfer. Growth on glycine, or on betaine, sarcosine or creatine (plus H-donors) depended strictly on the addition of selenite (≥0.1 μM); selenite was not required for fermentation of serine, or for degradation of alanine, aspartate or valine by the co-cultures. Cell-free extracts of glycine-grown cells contained active glycine reductase, glycine decarboxylase and reversible methyl viologen-dependent formate dehydrogenase in addition to the other enzymes necessary for an oxidation to CO2. In all reactions NADP was the preferred H-carrier. Both formate and glycine could be synthesized from bicarbonate. Serine-grown cells did not contain serine hydroxymethyl transferase but serine dehydratase and other enzymes commonly involved in pyruvate metabolism to acetate, CO2 and H2. The enzymes involved in glycine metabolism were repressed during growth on serine. By its morphology and physiology, strain al-2 did not resemble described amino acid-degrading species. Therefore, the new isolate is proposed as type strain of a new species, Eubacterium acidaminophilum.
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  • 3
    ISSN: 1432-072X
    Keywords: Autotrophic growth ; Sulfate-reducing bacteria ; Carbon dioxide ; Hydrogen ; Formate ; Homoacetogenic bacteria ; Desulfobacterium autotrophicum ; Desulfovibrio
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The capacity of mesophilic sulfate-reducing bacteria to grow lithoautotrophically with H2, sulfate and CO2 was investigated with enrichment cultures and isolated species. (a) Enrichments in liquid mineral media with H2, sulfate and CO2 consistently yielded mixed cultures of nonautotrophic, acetate-requiring Desulfovibrio species and autotrophic, acetate-producing Acetobacterium species (cell ratio approx. 20:1). (b) By direct dilution of mud samples in agar, various non-sporing sulfate reducers were isolated in pure cultures that did grow autotrophically. Two oval cell types (strains HRM2, HRM4) and one curved cell type (strain HRM6) from marine sediment were studied in detail. The strains grew in mineral medium supplemented only with vitamins (biotin, p-aminobenzoate, nicotinate). Carbon autotrophy was evident (i) from comparative growth experiments with non-autotrophic, acetate-requiring species, (ii) from high cell densities ruling out a cell synthesis from organic impurities in the mineral media, and (iii) by demonstrating that 96–99% of the cell carbon was derived from 14C-labelled CO2. Autotrophic growth occurred with a doubling time of 16–20 h at 24–28°C. Formate, fatty acids up to palmitate, ethanol, lactate, succinate, fumarate, malate and other organic acids were also used and completely oxidized. The three strains possessed cytochromes of the b-and c-type, but no desulfoviridin. Strain HRM2 is described as a new species of a new genus, Desulfobacterium autotrophicum. (c) The capacity for autotrophic growth was also tested with sulfate-reducing bacteria that originally had been isolated on organic substrates. The incompletely oxidizing, non-sporing types such as Desulfovibrio and Desulfobulbus species and Desulfomonas pigra were confirmed to be obligate heterotrophs that required acetate for growth with H2 and sulfate. In contrast, several of the completely oxidizing sulfate reducers were facultative autotrophs, such as Desulfosarcina variabilis, Desulfonema limicola, Desulfococcus niacini, and the newly isolated Desulfobacterium vacuolatum and Desulfobacter hydrogenophilus. The only incompletely oxidizing sulfate reducer that could grow autotrophically was the sporing Desulfotomaculum orientis, which obtained 96% of its cell carbon from 14C-labelled CO2. Desulfovibrio baarsii and Desulfococcus multivorans may also be regarded as types of facultative autotrophs; they could not oxidize H2, but grew on sulfate with formate as the only organic substrate.
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  • 4
    Electronic Resource
    Electronic Resource
    Springer
    Archives of microbiology 148 (1987), S. 286-291 
    ISSN: 1432-072X
    Keywords: Sulfate-reducing bacteria ; Desulfobacter species ; Acetate ; Hydrogen ; Autotrophic growth ; Nitrogen fixation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Sulfate-reducing bacteria with oval to rod-shaped cells (strains AcRS1, AcRS2) and vibrio-shaped cells (strains AcRM3, AcRM4, AcRM5) differing by size were isolated from anaerobic marine sediment with acetate as the only electron donor. A vibrio-shaped type (strain AcKo) was also isolated from freshwater sediment. Two strains (AcRS1, AcRM3) used ethanol and pyruvate in addition to acetate, and one strain (AcRS1) grew autotrophically with H2, sulfate and CO2. Higher fatty acids or lactate were never utilized. All isolates were able to grow in ammonia-free medium in the presence of N2. Nitrogenase activity under such conditions was demonstrated by the acetylene reduction test. The facultatively lithoautotrophic strain (AcRS1), a strain (AcRS2) with unusually large cells (2×5 μm), and a vibrio-shaped strain (AcRM3) are described as new Desulfobacter species, D. hydrogenophilus, D. latus, and D. curvatus, respectively.
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  • 5
    Electronic Resource
    Electronic Resource
    Springer
    Archives of microbiology 163 (1995), S. 96-103 
    ISSN: 1432-072X
    Keywords: Key words Anaerobic degradation ; Aromatic ; hydrocarbons ; Alkylbenzenes ; Ethylbenzene ; Crude oil ; Denitrifying bacteria ; Phylogeny ; Thauera selenatis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Anaerobic degradation of alkylbenzenes with side chains longer than that of toluene was studied in freshwater mud samples in the presence of nitrate. Two new denitrifying strains, EbN1 and PbN1, were isolated on ethylbenzene and n-propylbenzene, respectively. For comparison, two further denitrifying strains, ToN1 and mXyN1, were isolated from the same mud with toluene and m-xylene, respectively. Sequencing of 16S rDNA revealed a close relat ionship of the new isolates to Thauera selenatis. The strains exhibited different specific capacities for degradation of alkylbenzenes. In addition to ethylbenzene, strain EbN1 utilized toluene, but not propylbenzene. In contrast, propylbenzene-degrading strain PbN1 did not grow on toluene, but was able to utilize ethylbenzene. Strain ToN1 used toluene as the only hydrocarbon substrate, whereas strain mXyN1 utilized both toluene and m-xylene. Measurement of the degradation balance demonstr ated complete oxidation of ethylbenzene to CO2 by strain EbN1. Further characteristic substrates of strains EbN1 and PbN1 were 1-phenylethanol and acetophenone. In contrast to the other isolates, strain mXyN1 did not grow on benzyl alcohol. Benzyl alcohol (also m-methylbenzyl alcohol) was even a specific inhibitor of toluene and m-xylene utilization by strain mXyN1. None of the strains was able to grow on any of the alkylbenzenes with oxygen as electron acceptor. However, polar aromatic compounds such as benzoate were utilized under both oxic and anoxic conditions. All four isolates grew anaerobically on crude oil. Gas chromatographic analysis of crude oil after growth of strain ToN1 revealed specific depletion of toluene.
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  • 6
    ISSN: 1432-072X
    Keywords: Dissimilatory sulfate reduction ; Indole ; Pyridine ; Quinoline ; Complete anaerobic degradation ; Desulfobacterium indolicum ; Species description
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Indole (1.5 mmol/l) added to suflate-rich marine mud or sulfate-free sewage digestor sludge was anaerobically degraded within one week. Enrichments from sludge samples in defined indole-containing media with or without sulfate were selective for sulfate-reducing bacteria or mixed methanogenic associations, respectively. Other enrichments of sulfate-reducing bacteria were obtained with skatole, indoleacetate, indolepropionate, quinoline, and pyridine. From a marine enrichment with indole as sole electron donor and carbon source, an oval to rod-shaped, Gram-negative, nonsporing sulfate-reducing bacterium (strain In04) was isolated. Growth occurred in defined bicarbonate-buffered, sulfide-reduced media supplemented with vitamin B12. Furthen aromatic compounds utilized as electron donors and carbon sources were anthranilic acid and quinoline. Nonaromatic compounds used as substrates were formate, acetate, propionate, ethanol, propanol, butanol, pyruvate, malate, fumarate, and succinate. However, growth with substrates other than indole was rather slow. Thiosulfate served as an alternative electron acceptor. Complete oxidation of indole to CO2 was shown by stoichiometric measurements in batch culture with sulfate as electron acceptor. An average growth yield of 31.3 g cell dry weight was obtained per mol of indole oxidized. Pigment analysis revealed that cytochromes and menaquinone MK-7 (H2) were present. Desulfoviridin could not be detected. Strain In04 is described as new species of the new genus Desulfobacterium indolicum.
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  • 7
    ISSN: 1432-072X
    Keywords: Desulfobacter hydrogenophilus ; Sulfate-reducing bacteria ; Autotrophy ; Citric acid cycle ; ATP-citrate lyase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The strict anaerobe Desulfobacter hydrogenophilus is able to grow autotrophically with CO2, H2, and sulfate as sole carbon and energy sources. The generation time at 30°C under autotrophic conditions in a pure mineral medium was 15 h, the growth yield was 8 g cell dry mass per mol sulfate reduced to H2S. Enzymes of the autotrophic CO2 assimilation pathway were investigated. Key enzymes of the Calvin cycle and of the acetyl CoA pathway could not be found. All enzymes of a reductive citric acid cycle were present at specific activities sufficient to account for the observed growth rate. Notably, an ATP-citrate lyase (1.3 μmol · min-1 · mg cell protein-1) was present both in autotrophically and in heterotrophically grown cells, which was rapidly inactivated in the absence of ATP. The data indicate that in D. hydrogenophilus a reductive citric acid cycle is operating in autotrophic CO2 fixation. Since other autotrophic sulfate reducers possess an acetyl CoA pathway for CO2 fixation, two different autotrophic pathways occur in the same physiological group.
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  • 8
    ISSN: 1432-072X
    Keywords: Methanogenic bacteria ; Alcohols ; Ketones ; Aldehydes ; Alcohol dehydrogenase ; Enzyme expression ; Methanogenium thermophilum ; Factor F420
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In four species of methanogens able to grow with secondary alcohols as hydrogen donors the expression and properties of secondary alcohol dehydrogenase (sec-ADH) were investigated. Cells grown with 2-propanol and CO2 immediately started to oxidize secondary alcohols to ketones if transferred to new media. In the presence of H2, such cells reduced ketones or aldehydes to alcohols. In the absence of H2, aldehydes were dismutated (without growth) to primary alcohols and fatty acids. None of these reactions was catalyzed by cells grown with only H2 and CO2 at non-limiting concentration. This indicated an induction or derepression of sec-ADH by its substrate. Apparently, sec-ADH in all strains enabled not only the reduction of ketones or aldehydes, but also the dismutation of the latter. Sec-ADH was also expressed if strains were grown on H2 and CO2 in the presence of non-oxidizable, tertiary alcohols. Methanogenium thermophilum expressed sec-ADH even without added alcohol when H2 became limiting. From this species, an F420-specific sec-ADH was purified; the final gel filtration chromatography yielded a single protein peak that coincided with the activity. The enrichment was 12-fold, the activity recovery 26%. SDS polyacrylamide gel electrophoresis indicated that the enzyme was a homodimer with an apparent M r of 79,000. At the pH optimum around 4.2, the specific activity for oxidation of 2-propanol (130 mM) and reduction of acetone (20 mM) was 176 and 110 μmol/ min·mg, respectively (40°C). The apparent K m for 2-propanol and acetone (with 15 μM F420) was 2.5 and 0.25 mM, respectively. Aldehydes also were reduced.
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  • 9
    Electronic Resource
    Electronic Resource
    Springer
    Archives of microbiology 173 (2000), S. 58-64 
    ISSN: 1432-072X
    Keywords: Key wordsn-Alkanes ; Anaerobic hydrocarbon ¶oxidation ; Denitrifying bacteria ; Isolation ; Degradation ¶balance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The capacity of denitrifying bacteria for anaerobic utilization of saturated hydrocarbons (alkanes) was investigated with n-alkanes of various chain lengths and with crude oil in enrichment cultures containing nitrate as electron acceptor. Three distinct types of denitrifying bacteria were isolated in pure culture. A strain (HxN1) with oval-shaped, nonmotile cells originated from a denitrifying enrichment culture with crude oil and was isolated with n-hexane (C6H14). Another strain (OcN1) with slender, rod-shaped, motile cells was isolated from an enrichment culture with n-octane (C8H18). A third strain (HdN1) with oval, somewhat pleomorphic, partly motile cells originated from an enrichment culture with aliphatic mineral oil and was isolated with n-hexadecane (C16H34). Cells of hexane-utilizing strain HxN1 grew homogeneously in the growth medium and did not adhere to the alkane phase, in contrast to the two other strains. Quantification of substrate consumption and cell growth revealed the capacity for complete oxidation of alkanes under strictly anoxic conditions, with nitrate being reduced to dinitrogen.
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  • 10
    ISSN: 1432-072X
    Keywords: Key words Sulfate-reducing bacteria ; Gliding bacteria ; Desulfonema ; Isolation ; Oligonucleotide probing ; Sediments ; Microbial mats
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Microscopy of organic-rich, sulfidic sediment samples of marine and freshwater origin revealed filamentous, multicellular microorganisms with gliding motility. Many of these neither contained sulfur droplets such as the Beggiatoa species nor exhibited the autofluorescence of the chlorophyll-containing cyanobacteria. A frequently observed morphological type of filamentous microorganism was enriched under anoxic conditions in the dark with isobutyrate plus sulfate. Two strains of filamentous, gliding sulfate-reducing bacteria, Tokyo 01 and Jade 02, were isolated in pure cultures. Both isolates oxidized acetate and other aliphatic acids. Enzyme assays indicated that the terminal oxidation occurs via the anaerobic C1 pathway (carbon monoxide dehydrogenase pathway). The 16S rRNA genes of the new isolates and of the two formerly described filamentous species of sulfate-reducing bacteria, Desulfonema limicola and Desulfonema magnum, were analyzed. All four strains were closely related to each other and affiliated with the δ-subclass of Proteobacteria. Another close relative was the unicellular Desulfococcus multivorans. Based on phylogenetic relationships and physiological properties, Strains Tokyo 01 and Jade 02 are assigned to a new species, Desulfonema ishimotoi. A new, fluorescently labeled oligonucleotide probe targeted against 16S rRNA was designed so that that it hybridized specifically with whole cells of Desulfonema species. Filamentous bacteria that hybridized with the same probe were detected in sediment samples and in association with the filamentous sulfur-oxidizing bacterium Thioploca in its natural habitat. We conclude that Desulfonema species constitute an ecologically significant fraction of the sulfate-reducing bacteria in organic-rich sediments and microbial mats.
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