Search

Objectives: This paper reviews the principles of polymerase chain reaction (PCR) methodology, its application in identification of endodontic pathogens and the perspectives regarding the knowledge to be reached with the use of this highly sensitive, specific and accurate methodology as a microbial identification test., Data Sources: Studies published in the medical, dental and biological literature., Study Selection: Evaluation of published epidemiological studies examining the endodontic microbiota through PCR methodology., Conclusions: PCR technology has enabled the detection of bacterial species that are difficult or even impossible to culture as well as cultivable bacterial strains showing a phenotypically divergent or convergent behaviour. Moreover, PCR is more rapid, much more sensitive, and more accurate when compared with culture. Its use in endodontics to investigate the microbiota associated with infected root canals has expanded the knowledge on the bacteria involved in the pathogenesis of periradicular diseases. For instance, Tannerella forsythensis (formerly Bacteroides forsythus), Treponema denticola, other Treponema species, Dialister pneumosintes, and Prevotella tannerae were detected in infected root canals for the first time and in high prevalence when using PCR analysis. The diversity of endodontic microbiota has been demonstrated by studies using PCR amplification, cloning and sequencing of the PCR products. Moreover, other fastidious bacterial species, such as Porphyromonas endodontalis, Porphyromonas gingivalis and some Eubacterium spp., have been reported in endodontic infections at a higher prevalence than those reported by culture procedures.

A novel anaerobic, heterotrophic thermophile was isolated from a deep-sea hydrothermal vent chimney at the Suiyo Seamount in the Izu-Bonin Arc, Japan. The cells were bent, flexible rods, with a single polar flagellum. Growth was observed between 40 and 70 degrees C (optimum temperature: 60-65 degrees C; doubling time, 40 min) and between pH 5.0 and 7.5(optimum pH 6.5). The isolate was a strictly anaerobic heterotroph capable of using complex organic compounds (yeast extract, tryptone, peptone, casein and Casamino acids), ethanol and various organic acids as energy and carbon sources. Hydrogen could serve as a supplementary energy source. Elemental sulfur (S(0)), nitrate or arsenate was required for growth as an electron acceptor. The G + C content of the genomic DNA was 38.6 mol%. Phylogenetic analysis based on 16S rDNA sequences indicated that isolate SSM1(T) is closely related to Deferribacter thermophilus BMA(T) (98.1%). However, the novel isolate could be clearly differentiated from D. thermophilus BMA(T) on the basis of its physiological and genetic properties. The name Deferribacter desulfuricans sp. nov. (type strain SSM1(T) = JCM 11476(T) = DSM 14783(T)) is proposed.

A polyphasic taxonomic study was performed on a novel facultatively anaerobic, hydrogen- or sulfur/thiosulfate-oxidizing, thermophilic chemolithoautotroph recently isolated from subsurface hot aquifer water in a Japanese gold mine. The cells were straight to slightly curved rods, with a single polar flagellum. Growth was observed at 40-70 degrees C (optimum 60-65 degrees C; 80 min doubling time) and at pH 6.4-8.8 (optimum pH 7.5). The isolate was unable to use complex organic compounds, carbohydrates, amino acids or organic acids as sole energy and carbon sources. The G + C content of the genomic DNA was 31.3 mol%. Phylogenetic analysis based on 16S rDNA sequences indicated that the isolate was closely related to an uncultivated group of micro-organisms within the order Aquificales obtained from Icelandic and Japanese hot spring microbial mats, but distantly related to previously identified genera of the Aquificales such as Persephonella, Aquifex and Hydrogenobacter. The name Sulfurihydrogenibium subterraneum gen. nov., sp. nov. is proposed for this novel species; the type strain is HGMK1(T) (= JCM 11477(T) = ATCC BAA-562(T) = DSM 15120(T)).

Acute laminitis has been associated with the overgrowth of gram-positive bacteria within the equine hindgut, causing the release of factor(s) leading to ischemia-reperfusion of the digits. The products of fermentation which trigger acute laminitis are, as yet, unknown; however, vasoactive amines are possible candidates. The objectives of this study were to use an in vitro model of carbohydrate overload to study the change in populations of cecal streptococci and lactobacilli and to establish whether certain species of these bacteria were capable of producing vasoactive amines from amino acids. Cecal contents from 10 horses were divided into aliquots and incubated anaerobically with either corn starch or inulin (fructan; both at 1 g/100 ml). Samples were taken at 6-h intervals over a 24-h period for enumeration of streptococci, lactobacilli, and gram-negative anaerobes by a dilution method onto standard selective growth media. The effects of the antibiotic virginiamycin (1 mg/100 ml) and calcium hydrogen phosphate (CaHPO(4); 0.3 g/100 ml) were also examined. Fermentation of excess carbohydrate was associated with increases in numbers of streptococci and lactobacilli (2- to 3.5-log unit increases; inhibited by virginiamycin) but numbers of gram-negative anaerobes were not significantly affected. A screening agar technique followed by 16S rRNA gene sequence analysis enabled the identification of 26 different bacterial strains capable of producing one or more vasoactive amines. These included members of the species Streptococcus bovis and five different Lactobacillus spp. These data suggest that certain bacteria, whose overgrowth is associated with carbohydrate fermentation, are capable of producing vasoactive amines which may play a role in the pathogenesis of acute laminitis.

Two unknown, Gram-negative, catalase-negative and strictly anaerobic cocci were isolated from two independent human samples (strains AIP 49.01 and AIP 412.00T). Comparative 16S rRNA gene sequencing demonstrated that these two organisms displayed 99.8% sequence identity and that they are members of the Sporomusa sub-branch of the low-G + C Gram-positive bacteria. The most closely related 16S rDNA sequences were from Megasphaera sp. oral clone BU057 (99.8%) and from isolates of Megasphaera cerevisiae and Megasphaera elsdenii (94.5 and 93.8%, respectively). Phylogenetic analysis based on 16S rDNA sequences showed that these two strains were most closely related to M. elsdenii and belonged to the Megasphaera genus. Differences from previously described Megasphaera species in terms of size, biochemical tests (particularly the analysis of metabolic end products), gas production and DNA G+C content indicated that the two strains studied represent a novel species of anaerobic Gram-negative cocci. The name Megasphaera micronuciformis sp. nov. is proposed for these two isolates. It is also proposed that the uncultured organism previously deposited as Megasphaera sp. oral clone BU057 should be named 'Candidatus Megasphaera micronuciformis'. The type strain of Megasphaera micronuciformis is AIP 412.00T (=CIP 107280T =CCUG 45952T).

Recently developed molecular biology approaches make possible the detailed genetic, taxonomic and ecological examination of microorganisms from various habitats. Animal gut represents one of the most complex microbial ecosystems with a large degree of microbial biodiversity present. Bacteria inhabiting the gut usually play important roles in metabolic transformations of substrates and sometimes, e.g. in ruminants, they make the basis for an obligate symbiosis with the host. Here we discuss molecular microbiology as a strategy for examination of gut bacteria, concentrating on a typical and in such environment dominant group of strictly anaerobic Gram-negative bacteria from the phylogenetic group Cytophaga/Flexibacter/Bacteroides. The bacteria from the genus Prevotella are the most abundant Gram-negative bacteria in the rumen and form a distinctive phylogenetic cluster, clearly separated from prevotellas isolated from other ecological niches. They may represent a good choice for a model organism in genetic manipulation experiments and for studies of gene transfer mechanisms taking place in the gut. The molecular tools for detection and monitoring of ruminal prevotellas are discussed.

Because of access to 16S rDNA sequencing, changes in the taxonomy and nomenclature of anaerobic gram-negative bacteria have occurred lately. New genera and species have been described, and existing taxa have been reclassified. The present article compiles a list of clinically relevant anaerobes and provides synonyms as well as the old nomenclature used for these bacteria. Although names and classifications of anaerobic bacteria are changing quickly, it is important to keep track of new bacterial names to work toward better description and recognition of bacterium-disease associations.

A novel bacterium was isolated and characterized from the amniotic fluid of a woman who experienced intrauterine fetal demise in the second trimester of pregnancy. The bacterium was a slow-growing, gram-negative anaerobic coccobacillus belonging to the genus LEPTOTRICHIA: Unlike Leptotrichia sanguinegens, the isolate did not grow in chopped-meat glucose broth or on sheep blood agar upon subculturing. The isolate was characterized by sequencing and analyzing its 16S rRNA gene. The 1,493-bp 16S ribosomal DNA sequence had only 96% homology with L. sanguinegens. Several phylogenetic analyses indicated that L. amnionii is a distinct species and most closely related to L. sanguiegens.

Phenotypic and phylogenetic studies were performed on seven unidentified gram-negative, facultatively anaerobic, coccobacillus-shaped organisms isolated from human clinical specimens. Comparative 16S rRNA gene sequencing demonstrated that four of the strains corresponded to Dysgonomonas capnocytophagoides whereas the remaining three isolates represent a new sub-line within the genus Dysgonomonas, displaying greater than 5% sequence divergence with Dysgonomonas capnocytophagoides and Dysgonomonas gadei. The three novel isolates were readily distinguished from D.capnocytophagoides and D. gadei by biochemical tests. The DNA base composition of the novel species was consistent with its assignment to the genus Dysgonomonas. Based on phylogenetic and phenotypic evidence it is proposed that the unknown species, be classified as Dysgonomonas mossii sp. nov. The type strain of Dysgonomonas mossii is CCUG 43457T (= CIP 107079T).

A novel, anaerobic, non-spore-forming, mobile, Gram-negative, thermophilic bacterium, strain TMOT, was isolated from a thermophilic sulfate-reducing bioreactor operated at 65 C with methanol as the sole substrate. The G+C content of the DNA of strain TMOT was 39.2 mol%. The optimum pH, NaCl concentration, and temperature for growth were 7.0, 1.0%, and 65 degrees C, respectively. Strain TMOT was able to degrade methanol to CO2 and H2 in syntrophic culture with Methanothermobacter thermautotrophicus AH or Thermodesulfovibrio yellowstonii. Thiosulfate, elemental sulfur, Fe(III) and anthraquinone-2,6-disulfonate were able to serve as electron acceptors during methanol degradation. In the presence of thiosulfate or elemental sulfur, methanol was converted to CO2 and partly to alanine. In pure culture, strain TMOT was also able to ferment methanol to acetate, CO2 and H2. However, this degradation occurred slower than in syntrophic cultures or in the presence of electron acceptors. Yeast extract was required for growth. Besides growing on methanol, strain TMOT grew by fermentation on a variety of carbohydrates including monomeric and oligomeric sugars, starch and xylan. Acetate, alanine, CO2, H2, and traces of ethanol, lactate and alpha-aminobutyrate were produced during glucose fermentation. Comparison of 16S rDNA genes revealed that strain TMOT is related to Thermotoga subterranea (98%) and Thermotoga elfii (98%). The type strain is TMOT (= DSM 14385T = ATCC BAA-301T). On the basis of the fact that these organisms differ physiologically from strain TMOT, it is proposed that strain TMOT be classified as a new species, within the genus Thermotoga, as Thermotoga lettingae.

A new sulfate-reducing bacterium was isolated from marine sediment with phosphite as sole electron donor and CO(2) as the only carbon source. Strain FiPS-3 grew slowly, with doubling times of 3-4 days, and oxidized phosphite, hydrogen, formate, acetate, fumarate, pyruvate, glycine, glutamate, and other substrates nearly completely, with concomitant reduction of sulfate to sulfide. Acetate was formed as a side product to a small extent. Glucose, arabinose, and proline were partly oxidized and partly fermented to acetate plus propionate. Growth with phosphite, hydrogen, or formate was autotrophic. Also, in the presence of sulfate, CO dehydrogenase was present, and added acetate did not increase growth rates or growth yields. In the absence of sulfate, phosphite oxidation was coupled to homoacetogenic acetate formation, with growth yields similar to those in the presence of sulfate. Cells were small rods, 0.6 - 0.8 x 2-4 microm in size, and gram-negative, with a G+C content of 53.9 mol%. They contained desulforubidin, but no desulfoviridin. Based on sequence analysis of the 16S rRNA gene and the sulfite reductase genes dsrAB, strain FiPS-3 was found to be closely related to Desulfotignum balticum. However, physiological properties differed in many points from those of D. balticum. These findings justify the establishment of a new species, Desulfotignum phosphitoxidans.

It has recently been recognized that the ability to use Fe(III) as a terminal electron acceptor is a highly conserved characteristic in hyperthermophilic microorganisms. This suggests that it may be possible to recover as-yet-uncultured hyperthermophiles in pure culture if Fe(III) is used as an electron acceptor. As part of a study of the microbial diversity of the Obsidian Pool area in Yellowstone National Park, Wyo., hot sediment samples were used as the inoculum for enrichment cultures in media containing hydrogen as the sole electron donor and poorly crystalline Fe(III) oxide as the electron acceptor. A pure culture was recovered on solidified, Fe(III) oxide medium. The isolate, designated FW-1a, is a hyperthermophilic anaerobe that grows exclusively by coupling hydrogen oxidation to the reduction of poorly crystalline Fe(III) oxide. Organic carbon is not required for growth. Magnetite is the end product of Fe(III) oxide reduction under the culture conditions evaluated. The cells are rod shaped, about 0.5 microm by 1.0 to 1.2 microm, and motile and have a single flagellum. Strain FW-1a grows at circumneutral pH, at freshwater salinities, and at temperatures of between 65 and 100 degrees C with an optimum of 85 to 90 degrees C. To our knowledge this is the highest temperature optimum of any organism in the Bacteria. Analysis of the 16S ribosomal DNA (rDNA) sequence of strain FW-1a places it within the Bacteria, most closely related to abundant but uncultured microorganisms whose 16S rDNA sequences have been previously recovered from Obsidian Pool and a terrestrial hot spring in Iceland. While previous studies inferred that the uncultured microorganisms with these 16S rDNA sequences were sulfate-reducing organisms, the physiology of the strain FW-1a, which does not reduce sulfate, indicates that these organisms are just as likely to be Fe(III) reducers. These results further demonstrate that Fe(III) may be helpful for recovering as-yet-uncultured microorganisms from hydrothermal environments and illustrate that caution must be used in inferring the physiological characteristics of at least some thermophilic microorganisms solely from 16S rDNA sequences. Based on both its 16S rDNA sequence and physiological characteristics, strain FW-1a represents a new genus among the Bacteria. The name Geothermobacterium ferrireducens gen. nov., sp. nov., is proposed (ATCC BAA-426).

The strictly anaerobic gram-negative bacterium strain Fru22T grows at 50 degrees C in media containing up to 75 g NaCl l(-1). Hexoses and pentoses are fermented to equal molar amounts of acetate, propionate and succinate, and no CO2 is formed. An orange-red pigment similar to flexirrubin is produced during stationary phase upon exposure to light for several days. Cells also produce a surface-active extracellular compound which lowers the surface tension of the medium. This tenside is heat-tolerant up to 70 degrees C and is destroyed by treatment with proteinase K or trypsin, but not by lipase. Comparative 16S rDNA sequence analysis confirmed a phylogenetic affiliation of strain Fru22T to the phylum Bacteroides (Cytophaga/Flavobacterium/Bacteroides), moderately related to the genus Marinilabilia. Therefore, on the basis of phylogenetic, phenotypic and physiological evidence, a new genus, Anaerophaga, is proposed to harbour strain Fru22T (DSM 12881T, OCM 798T) which is described as the type strain of a new species, Anaerophaga thermohalophila gen. nov., sp. nov.

A novel anaerobic, moderately thermophilic, peptide-fermenting bacterium, strain NGA(T), was isolated from an anaerobic wool-scouring wastewater treatment lagoon. The cells were gram-negative, straight rods of 0.5-1.0 x 2.0-4.0 microm, motile by means of a single flagellum. The DNA G+C content was 51.5 mol%. The optimum pH and temperature range for growth were 6.6-7.3 and 55-60 degrees C, respectively. The optimum NaCl concentration was 0.08 g l(-1). The bacterium fermented organic acids (malate, tartrate, pyruvate, glycerol and fumarate), a few carbohydrates (starch, glucose, fructose and gluconate), Casamino acids, tryptone and yeast extract. Carbohydrates and organic acids were converted to acetate, hydrogen and CO2. The bacterium oxidized leucine to isovalerate with crotonate as an electron acceptor, but not in co-culture with Methanothermobacter thermoautotrophicus DSM 3720T. Thiosulfate, sulfur and cystine were reduced to sulfide and crotonate was reduced to butyrate with glucose and tryptone-yeast extract as electron donors. Phylogenetic analysis of the 16S rRNA gene indicated that strain NGA(T) was related to Anaerobaculum thermoterrenum (98% similarity), the only described species of the genus. The DNA-DNA hybridization value for strain NGA(T) and A. thermoterrenum ACM 5076T was 40.8%. On the basis of these results, strain NGA(T) is proposed as a novel species of the genus Anaerobaculum, namely Anaerobaculum mobile sp. nov. The type strain is NGA(T) (= DSM 13181T =ATCC BAA-54T).

Some Gram-negative anaerobes are associated with the incidence and progression of periodontal disease. In periodontal pockets, however, the localization of those bacteria is unknown. We investigated the localization of 5 bacterial species in human periodontal pockets. Fifteen teeth with a part of periodontal pockets from 10 adult periodontitis patients were obtained, and the localization of bacteria was examined immunohistochemically. Positive reactions with anti-Prevotella nigrescens antibody were located at the epithelium-associated plaque area in the middle pocket zones. In the middle and deep pocket zones, Fusobacterium nucleatum and Treponema denticola were especially localized in the unattached plaque area, but Eikenella corrodens was observed in the tooth-attached plaque area. Actinobacillus actinomycetemcomitans, detected in 2 of 15 samples examined, was found in the unattached plaque area, in the middle pocket zone. The present findings indicated that the 5 bacterial species examined localized at distinct regions in human periodontal pockets.

A total of 206 gram negative anaerobic bacilli were isolated from 2591 routine clinical specimens. Majority of them (78.2%) were part of a mixed flora. A high incidence of gram negative anaerobic bacilli was found in post-operative wound infections and diabetic foot infections (30.1%), followed by necrotizing fasciitis and crush injuries (18%), pus of empyema and lung abscesses (14.5%) and aspirates from other deep abscesses (10.7%). Bacteroides fragilis was the most frequently recognized species (42.2%), followed by Fusobacterium sp. (17.4%), Prevotella sp. (15.1%) and Porphyromonas sp. (11.2%). Neomycin blood agar plate yielded the maximum number (72.8%) of strains in pure culture. Gram negative anaerobic bacilli in infective processes are significantly on the rise and so suspected cultures should be processed for both aerobic and anaerobic organisms.

Long-term exposure to 2-bromoethanesulfonate (BES), an agent known to inhibit methanogenesis, altered the bacterial community structure of an anaerobic enrichment culture that reductively dechlorinated trichloroethene (TCE). BES did not hinder the dechlorination of TCE or other chlorinated ethenes as previously reported, although different intermediates and end products were observed.

Results of a polyphasic taxonomic study on an unknown Gram-negative, facultatively anaerobic, coccobacillus-shaped organism isolated from an infected human gall bladder are presented. Phenotypic and molecular taxonomic studies revealed the organism to be close to, but distinct from, organisms designated CDC (Centers for Disease Control and Prevention) group DF-3. The unknown bacterium was readily distinguished from reference strains of Bacteroides, Prevotella, Porphyromonas and related taxa by 16S rRNA gene sequencing, biochemical tests, analysis of cellular long-chain fatty acids and electrophoretic analysis of whole-cell proteins. Based on the results of the present study, it is proposed that the unknown bacterium be classified in a new genus, Dysgonomonas, as Dysgonomonas gadei sp. nov. (type strain CCUG 42882T = CIP 106420T). In addition, a new species, Dysgonomonas capnocytophagoides sp. nov., is proposed to accommodate strains previously belonging to CDC group DF-3. The type species of the genus Dysgonomonas is Dysgonomonas gadei.

The skin microflora of patients with chronic dermatoses (atopic dermatitis and psoriasis) have been studied by the original "Bactotests" method. The data thus obtained indicate that the clinical picture of the disease is related to the severity of skin dysbacteriosis. The electron-microscopic study of 2 staphylococcal strains isolated from patients has revealed the presence of the immunoglobulin cover (capsule-like outer sheath consisting of immunoglobulins and other humoral protective factors) on the cell wall of these bacteria.

Anaerobic enrichment cultures, with erythritol as substrate, resulted in the isolation of a strain with properties not yet found in an existing genus in this combination. The strain, FKBS1, was strictly anaerobic, stained gram-negative and formed spores. Cells were small motile vibrios with flagella inserted at the concave side of the cell. Spores were located terminally and caused only slight swelling of the cells if compared to related spore-forming genera. FKBS1 fermented fructose, mannitol, sorbitol, xylitol and erythritol to propionic acid, acetic acid, CO2 and small amounts of H2 to balance the difference in the oxidation-reduction value between substrate and cell mass. The 16S rDNA sequence revealed relationship to the Sporomusa-Pectinatus-Selenomonas group. However, the phylogenetic distance to any of its members was too great to allow it to be placed in one of the existing genera. Morphologically the strain resembled Sporomusa, which, however, performs an acetogenic type of fermentation. The propionic-acid-forming genera of the group are either not spore-formers or, in the case of Dendrosporobacter quercicolus (syn. Clostridium quercicolum), morphologically different. It is therefore proposed to classify strain FKBS1 as a new genus and species, Propionispora vibrioides.

Dialister pneumosintes is a nonfermentative, anaerobic, gram-negative rod that grows with small, circular, transparent, shiny, smooth colonies on blood agar. Even though D. pneumosintes has been recovered from deep periodontal pockets, little is known about the relationship between the organism and destructive periodontal disease. This study describes a rapid PCR method to identify D. pneumosintes in periodontal samples. The PCR identification method detected as little as 10 pg of D. pneumosintes DNA or about 1 to 10 cells without nonspecific amplification of various periodontopathic bacteria. Twelve of 22 subgingival samples from adult periodontitis lesions yielded D. pneumosintes either by culture or by PCR identification. In culture-positive samples, D. pneumosintes averaged 3.9% (0.001 to 10.8%) of total isolates. Studies are needed to delineate virulence factors of D. pneumosintes pertinent to periodontal disease.

A competitive polymerase chain reaction assay targeting the 16S rDNA was developed for quantitating the rumen bacterium Butyrivibrio fibrisolvens OB156. A competitor DNA, serving as an internal control in the competitive polymerase chain reaction reaction, was constructed by polymerase chain reaction using a looped oligo longer than the normal primer. Coamplification of the target DNA with known amounts of the competitor DNA allowed quantitation of the target DNA in both pure culture and mixed culture systems, where minimum quantifiable level of OB156 was 1.7x10(2) and 5.6x10(4) cells, respectively. When an erythromycin-resistant recombinant derived from OB156 was inoculated into a rumen fluid culture, its numbers decreased with time. The rate of decrease measured by the competitive polymerase chain reaction assay was much slower than the rate determined by culture enumeration using erythromycin selection. The competitive polymerase chain reaction assay also showed 48 h persistence of the recombinant at 10(4) ml(-1) even after disappearance of culturable recombinant, suggesting maintenance of the target DNA from uncultivable cells. In an in vivo tracking trial, the recombinant became undetectable within 72 h with either assay, indicating rapid hydrolysis and/or outflow of the cells from the rumen.

A novel dissimilatory, nitrate-reducing bacterium, designated strain N2460T, was isolated from an oil reservoir model column. Strain N2460T is a mesophilic, obligately anaerobic, marine, gram-negative bacterium. The cells are vibrio-shaped and motile by a bipolar flagellum. Strain N2460T reduces nitrate to ammonia in a mineral medium supplied by acetate. The presence of a 2-oxoglutarate dehydrogenase activity indicates that acetate is oxidized via the citric acid cycle. No growth is obtained on formate, higher fatty acids, malate, fumarate, benzoate, alcohols, sugar, yeast extract, crude oil, alkanes, proline, hydrogen, sulfur or thiosulfate with nitrate as electron acceptor. Oxygen, sulfate, thiosulfate and sulfur are not utilized as alternative electron acceptors. Strain N2460T grows fermentatively on fumarate, but not on pyruvate. The G+C content of the DNA is 42.6 mol%. 16S rRNA gene analysis shows that strain N2460T belongs to the Bacteria and that the closest relative is 'Geovibrio ferrireducens' (sequence similarity 86.9%). On the basis of phylogenetic as well as phenotypic data, it is proposed that strain N2460T represents the type strain of a new genus and species, Denitrovibrio acetiphilus gen. nov., sp. nov.

A strain of strictly anaerobic and moderately halophilic bacteria isolated from salted puffer fish ovaries was studied phenotypically, genotypically and phylogenetically. On the basis of its physiological and morphological characteristics, the new isolate is considered to be a member of the genus Haloanaerobium. It is a motile, rod-shaped, non-spore-forming, gram-negative, obligate anaerobe that grows in the presence of 25% (w/v) NaCl. The optimum salt concentration for growth is 10% (w/v). It grows well at 15 and 45 degrees C, but not at 10 or 50 degrees C. The optimum temperature for growth is 35 degrees C. It grows at pH 6.0-9.0 and the optimum pH for growth is 7.5. It ferments N-acetylglucosamine, cellobiose, fructose, galactose, D-glucose, lactose, maltose, D-mannose, raffinose, D-ribose, sucrose and D-xylose. It ferments D-glucose with the production of hydrogen, carbon dioxide, ethanol and organic acids such as acetate, formate and lactate. 16S rRNA gene sequence information confirmed the phylogenetic position of the new isolate, strain R-9T, as a member of the genus Haloanaerobium. DNA-DNA hybridization data revealed that isolate R-9T exhibited low levels of reassociation (less than 30%) with previously described Haloanaerobium species. Based on these results, the new isolate appears to represent a new Haloanaerobium species, for which the name Haloanaerobium fermentans sp. nov. is proposed. The type strain is R-9T (= JCM 10494T).

Two novel strains of Propionigenium maris able to reductively debrominate 2,4,6-tribromophenol (TBP) to monobromophenols were isolated from marine hemichordate and polychaete burrows. These two strains, DSL-1 and ML-1, were anaerobic, non-motile rods that stained Gram-negative and required 0.05% yeast extract for growth. Strain DSL-1 fermented pyruvate and succinate to predominantly butyrate and strain ML-1 fermented glucose and succinate primarily to propionate. No inorganic terminal electron acceptors were identified. The pH and temperature optima for growth were 7.6 and 30 degrees C for strain DSL-1 and 7.0 and 32 degrees C for strain ML-1, respectively; doubling times for strains DSL-1 and ML-1 were 0.32 h and 0.30 h, respectively. Both strains required 2-3% (w/v) NaCl for optimal growth. Morphological and physiological features, as well as the results of 16S rDNA sequence analysis, showed these to be new strains of Propionigenium maris. Because they differ from the P. maris type strain (DSM 9537T) in a number of respects, including their ability to rapidly debrominate di- and tribromophenols, and in their specific habitats, the species description is amended to include these ecologically important properties.

A moderately thermophilic, organotrophic bacterium with vibrioid cells was isolated from a sample of a cyanobacterial mat from caldera Uzon, Kamchatka, Russia, and designated strain Z-9701T. Cells of strain Z-9701T were curved, Gram-negative rods, 0.5-0.7 x 2.5-5.0 microm in size, with tapering ends and with fast, wavy movement by means of lateral flagella located on the concave side of the cell. Colonies were small, white, irregular or round, 0.2 mm in diameter, and with even edges. Strain Z-9701T was an obligate anaerobe with a temperature optimum at 60-65 degrees C and a pH optimum at 7.3. It fermented glucose, fructose, mannose, N-acetyl-D-glucosamine, adonite, arginine, serine, peptone, yeast extract and Casamino acids. The fermentation products formed during growth on glucose were acetate, lactate, H2, CO2 and ethanol. Strain Z-9701T reduced elemental sulfur to H2S during organotrophic growth with glucose or peptides as energy and carbon sources. In the presence of S0, strain Z-9701T was capable of lithotrophic growth with molecular hydrogen as energy substrate and 0.1 g yeast extract l(-1) as carbon source. Sulfate, thiosulfate, nitrate, Fe(III) and sulfite were not reduced and did not stimulate growth. The G+C content of strain Z-9701T DNA was 54.6 mol%. The results of 16S rDNA sequence analyses revealed that strain Z-9701T belongs to the cluster within the Clostridium group formed by Thermanaerovibrio acidaminovorans, Dethiosulfovibrio peptidovorans, Anaerobaculum thermoterrenum and Aminobacterium colombiense, but the level of sequence similarity with the members of this cluster was not very high (87.6-92.2%). Among these organisms, Thermanaerovibrio acidaminovorans is phenotypically close to strain Z-9701T. However, the two organisms showed a relatively low level of similarity of their 16S rRNA sequences (92.2%) and of DNA-DNA hybridization (15 +/- 1%). Nevertheless, on the basis of the similar morphology and physiology of the new isolate and Thermanaerovibrio acidaminovorans, strain Z-9701T was placed in the genus Thermanaerovibrio and a new species, Thermanaerovibrio velox, proposed for it. The type strain is Z-9701T (= DSM 12556T).

The strictly anaerobic, Gram-negative, spore-forming bacterium strain WoGl3T had been enriched and isolated in mineral medium with glutarate as the sole source of energy and organic carbon. Glutarate was fermented to a mixture of butyrate, isobutyrate, CO2 and small amounts of acetate. Strain WoGl3T grew only with the dicarboxylates glutarate, methylsuccinate and succinate. 16S rDNA sequence analysis revealed an affiliation of strain WoGl3T to the family Syntrophomonadaceae. This monophyletic group is comprised of strain WoGl3T and the genera Syntrophomonas, Syntrophospora and Thermosyntropha, within the phylum of Gram-positive bacteria with a low DNA G + C content. Overall intra-group 16S rRNA sequence similarities of 89.2-93.9% document a separate phylogenetic status for strain WoGl3T. Strain WoGl3T (= DSM 6652T) is described as the type strain of a new species within a new genus, Pelospora glutarica gen. nov., sp. nov.

A new anaerobic, thermophilic, syntrophic, fatty-acid-oxidizing bacterium designated strain TGB-C1T was isolated from granular sludge in a thermophilic upflow anaerobic sludge blanket (UASB) reactor. The cells were slightly curved rods and were weakly motile. Spore formation was not observed. The optimal temperature for growth was around 55 degrees C and growth occurred in the range 45 to 60 degrees C. The pH range for growth was 5.8-7.5, and the optimum pH was 6.5-7.0. Crotonate was the only substrate that allowed the strain to grow in pure culture. However, in co-culture with the thermophilic, hydrogenotrophic Methanobacterium thermoautotrophicum strain delta H, the isolate could syntrophically oxidize saturated fatty acids with 4-10 carbon atoms, including isobutyrate. During the degradation of isobutyrate by the co-culture, isobutyrate was isomerized to butyrate, which was then oxidized. The strain was not able to utilize sulfate, sulfite, thiosulfate, nitrate, fumarate or Fe(III) as electron acceptor. The DNA base composition was 51.0 mol%. 16S rDNA sequence analysis revealed that the strain belongs to the family Syntrophomonadaceae, but it was only distantly related to other known species of beta-oxidizing syntrophs. Hence, the name Syntrophothermus lipocalidus is proposed for TGB-C1T as a new species of a new genus.

A novel, curved (0.3 x 4.0-5.0 microm), Gram-negative, non-sporulating, mesophilic bacterium, designated strain ILE-3T (T = type strain), was isolated from an anaerobic lagoon in a dairy wastewater treatment plant. Optimal growth occurred at 37 degrees C and pH 7.4 on a medium containing serine as an energy source and yeast extract. The strain was motile by means of one or two lateral flagella. It required yeast extract for growth on serine, glycine, threonine and pyruvate. Poor growth was obtained on cysteine, Casamino acids, biotrypcase, peptone and 2-oxoglutarate. In the presence of Methanobacterium formicicum, strain ILE-3T oxidized alanine, glutamate, leucine, isoleucine, valine and aspartate to a minor extent. The G+C content of the DNA was 44 mol%. Phylogenetic analysis of the 16S rRNA gene of strain ILE-3T indicated that it was related to Aminobacterium colombiense (95% similarity value). On the basis of the phenotypic and phylogenetic characteristics, strain ILE-3T is designated as a new species of the genus Aminobacterium, namely Aminobacterium mobile sp. nov. (= DSM 12262T).

A novel cellulosomal scaffoldin gene, termed cipV, was identified and sequenced from the mesophilic cellulolytic anaerobe Acetivibrio cellulolyticus. Initial identification of the protein was based on a combination of properties, including its high molecular weight, cellulose-binding activity, glycoprotein nature, and immuno-cross-reactivity with the cellulosomal scaffoldin of Clostridium thermocellum. The cipV gene is 5,748 bp in length and encodes a 1,915-residue polypeptide with a calculated molecular weight of 199,496. CipV contains an N-terminal signal peptide, seven type I cohesin domains, an internal family III cellulose-binding domain (CBD), and an X2 module of unknown function in tandem with a type II dockerin domain at the C terminus. Surprisingly, CipV also possesses at its N terminus a catalytic module that belongs to the family 9 glycosyl hydrolases. Sequence analysis indicated the following. (i) The repeating cohesin domains are very similar to each other, ranging between 70 and 90% identity, and they also have about 30 to 40% homology with each of the other known type I scaffoldin cohesins. (ii) The internal CBD belongs to family III but differs from other known scaffoldin CBDs by the omission of a 9-residue stretch that constitutes a characteristic loop previously associated with the scaffoldins. (iii) The C-terminal type II dockerin domain is only the second such domain to have been discovered; its predicted "recognition codes" differ from those proposed for the other known dockerins. The putative calcium-binding loop includes an unusual insert, lacking in all the known type I and type II dockerins. (iv) The X2 module has about 60% sequence homology with that of C. thermocellum and appears at the same position in the scaffoldin. (v) Unlike the other known family 9 catalytic modules of bacterial origin, the CipV catalytic module is not accompanied by a flanking helper module, e.g., an adjacent family IIIc CBD or an immunoglobulin-like domain. Comparative sequence analysis of the CipV functional modules with those of the previously sequenced scaffoldins provides new insight into the structural arrangement and phylogeny of this intriguing family of microbial proteins. The modular organization of CipV is reminiscent of that of the CipA scaffoldin from C. thermocellum as opposed to the known scaffoldins from the mesophilic clostridia. The phylogenetic relationship of the different functional modules appears to indicate that the evolution of the scaffoldins reflects a collection of independent events and mechanisms whereby individual modules and other constituents are incorporated into the scaffoldin gene from different microbial sources.

In an attempt to understand better the micro-organisms involved in anaerobic degradation of aromatic hydrocarbons in the Fe(III)-reducing zone of petroleum-contaminated aquifers, Fe(III)-reducing micro-organisms were isolated from contaminated aquifer material that had been adapted for rapid oxidation of toluene coupled to Fe(III) reduction. One of these organisms, strain H-5T, was enriched and isolated on acetate/Fe(III) medium. Strain H-5T is a Gram-negative strict anaerobe that grows with various simple organic acids such as acetate, propionate, lactate and fumarate as alternative electron donors with Fe(III) as the electron acceptor. In addition, strain H-5T also oxidizes long-chain fatty acids such as palmitate with Fe(III) as the sole electron acceptor. Strain H-5T can also grow by fermentation of citrate or fumarate in the absence of an alternative electron acceptor. The primary end-products of citrate fermentation are acetate and succinate. In addition to various forms of soluble and insoluble Fe(III), strain H-5T grows with nitrate, Mn(IV), fumarate and the humic acid analogue 2,6-anthraquinone disulfonate as alternative electron acceptors. As with other organisms that can oxidize organic compounds completely with the reduction of Fe(III), cell suspensions of strain H-5T have absorbance maxima indicative of a c-type cytochrome(s). It is proposed that strain H-5T represents a novel genus in the Holophaga-Acidobacterium phylum and that it should be named Geothrix fermentans sp. nov., gen. nov.

Chemotaxonomic, electron microscopic and 16S rRNA gene sequence analyses of the three described species of the genus Anaerovibrio demonstrated only remote similarities to each other. The 16S rRNA gene sequence similarities between Anaerovibrio lipolytica, Anaerovibrio glycerini and Anaerovibrio burkinabensis and the derived phylogenetic relationships of the three species studied fell below genus level. All three species clustered within the Sporomusa-Pectinatus-Selenomonas phyletic group. Each species showed a distinct phospholipid pattern and whole-cell fatty acid distribution. Several isoprenologues of the lipoquinone 'lipid F' were found to differ in their quantitative distribution in the Anaerovibrio species. On the basis of these results, the new genera Anaerosinus gen. nov. and Anaeroarcus gen. nov. are proposed. The type species of Anaerosinus is Anaerosinus glycerini comb. nov., and the type species of Anaeroarcus is Anaeroarcus burkinensis [corrig.] comb. nov. The genus Anaerovibrio is consequently restricted to a single species, namely Anaerovibrio lipolyticus [corrig.]

A cellulolytic anaerobic bacterium, strain I77R1BT, was isolated from a biomat sample of an Icelandic, slightly alkaline, hot spring (78 degrees C). Strain I77R1BT was rod-shaped, non-spore-forming, non-motile and stained Gram-negative at all stages of growth. It grew at 45-82 degrees C, with an optimum growth temperature around 78 degrees C. At 70 degrees C, growth occurred at pH 5.8-8.0, with an optimum near pH 7.0. At the optimum temperature and pH, with 2 g cellobiose l-1 as substrate, strain I77R1BT had a generation time of 2 h. During growth on Avicel, strain I77R1BT produced acetate, hydrogen and carbon dioxide as major fermentation products together with small amounts of lactic acid and ethanol. The strain fermented many substrates, including cellulose, xylan, starch and pectin, but did not grow with casein peptone, pyruvate, D-ribose or yeast extract and did not reduce thiosulfate to H2S. The G+C ratio of the cellular DNA was 35 mol%. Comparative 16S rDNA analysis placed strain I77R1BT among species of Caldicellulosiruptor. The closest relative was Caldicellulosiruptor lactoaceticus. Hybridization of total DNA showed 42% hybridization to C. lactoaceticus and 22% hybridization to Caldicellulosiruptor saccharolyticus. A new species, Caldicellulosiruptor kristjanssonii sp. nov. (I77R1BT) is proposed.

A novel, asaccharolytic, amino-acid-degrading bacterium, designated strain GLU-3T, was isolated from an anaerobic lagoon of a dairy wastewater treatment plant. Strain GLU-3T stained Gram-negative and was an obligately anaerobic, non-spore-forming, slightly curved, rod-shaped bacterium (0.3 x 4.0-6.0 microns) which existed singly or in pairs. The DNA G+C content was 43 mol%. Optimum growth occurred at 35 degrees C and pH 7.5 on arginine with a generation time of 16 h. Good growth was obtained on arginine, histidine, threonine and glycine. Acetate was the end-product formed from all these substrates, but in addition, a trace of formate was detected from arginine and histidine, and ornithine was produced from arginine. Strain GLU-3T grew slowly on glutamate and produced acetate, carbon dioxide, formate, hydrogen and traces of propionate as the end-products. In syntrophic association with Methanobacterium formicicum, strain GLU-3T oxidized arginine, histidine and glutamate to give propionate as the major product; acetate, carbon dioxide and methane were also produced. Strain GLU-3T did not degrade alanine and the branched-chain amino acids valine, leucine and isoleucine either in pure culture or in association with M. formicicum. The nearest phylogenetic relative of strain GLU-3T was the thermophile Selenomonas acidaminovorans (similarity value of 89.5%). As strain GLU-3T is phylogenetically, physiologically and genotypically different from other amino-acid-degrading genera, it is proposed that it should be designated a new species of a new genus Aminomonas paucivorans gen. nov., sp. nov. (DSM 12260T).

Three strains, designated VS-751T, VS-511 and VS-732, of a strictly anaerobic, moderately halophilic, Gram-negative, rod-shaped bacterium were isolated from a highly saline (15-20%) brine from an oil reservoir in central Oklahoma, USA. The optimal concentration of NaCl for growth of these three strains was 2 M (12%), and the strains also grew in the presence of an additional 1 M MgCl2. The strains were mesophilic and grew at a pH range of 6-8. Carbohydrates used by all three strains included glucose, fructose, arabinose, galactose, maltose, mannose, cellobiose, sucrose and inulin. Glucose fermentation products included ethanol, acetate, H2 and CO2, with formate produced by two of the three strains. Differences were noted among strains in the optimal temperature and pH for growth, the maximum and minimum NaCl concentration that supported growth, substrate utilization and cellular fatty acid composition. Despite the phenotypic differences among the three strains, analysis of the 16S rRNA gene sequences and DNA-DNA hybridizations showed that these three strains were members of the same genospecies which belonged to the genus Haloanaerobium. The phenotypic and genotypic characteristics of strains VS-751T, VS-511 and VS-732 are different from those of previously described species of Haloanaerobium. It is proposed that strain VS-751T (ATCC 700103T) be established as the type strain of a new species, Haloanaerobium kushneri.

In dentistry antimicrobials are used in the treatment of progressive periodontitis and odontogenic abscesses, therefore the susceptibility to commonly used antibiotics of capnophilic and anaerobic species causing these diseases should be investigated. The activity of penicillin, amoxycillin, cefoxitin, clindamycin, doxycycline, metronidazole and ciprofloxacin was investigated. One hundred and sixty four isolates from subgingival plaque samples of 66 patients with progressive periodontitis and 192 bacterial strains from pus of 74 patients with odontogenic abscesses were included in this study. The majority of species tested were gram-negative anaerobes (Prevotella spp., Porphyromonas spp., Fusobacterium spp.), and were highly susceptible to clindamycin and metronidazole. Nearly 6% of the periodontal isolates and 22% of the bacteria obtained from pus samples produced beta-lactamases. With the exception of the periodontopathogenic species Actinobacillus actinomycetemcomitans and Eikenella corrodens, clindamycin seemed to be a useful antibiotic and could be recommended for empirical antimicrobial treatment.

The phylogenetic position of Gram-negative, strictly anaerobic, non-spore-forming bacteria, representing four different genera, was determined by analysis of their 16S rDNA sequences. Formivibrio citricus and Propionivibrio dicarboxylicus are members of the beta-subclass of the class Proteobacteria. While Formivibrio citricus stands phylogenetically isolated, Propionivibrio dicarboxylicus is moderately related to members of the genus Rhodocyclus. Succinimonas amylolytica and Succinivibrio dextrinosolvens are members of the gamma-subclass of the class Proteobacteria in which they, together with members of the genus Anaerobiospirillum and Ruminobacter amylophilus, form a separate line of descent. This phylogenetic group is described as Succinivibrionaceae fam. nov.

The aim of this study was to report the microbiological and clinical effects of repeated subgingival administration of a 1% Chlorhexidine-gel in periodontal pockets from 10 patients with adult periodontitis. Results showed that the experimental treatment significantly improved clinical parameters (Plaque Index, Gingival Bleeding Index, and Pocket Probing Depth). Direct subgingival administration of Chlorhexidine-gel also produced a remarkable modification in the proportions of putative periodontopathic microorganisms, such as Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, Veillonella parvula, Fusobacterium nucleatum, and Peptostreptococcus micros, in subgingival bacterial plaque from periodontitis patients.

We have previously reported high-frequency transfer of tetracycline resistance between strains of the rumen anaerobic bacterium Butyrivibrio fibrisolvens. Donor strains were postulated to carry two TcR genes, one of which is transferred on a novel chromosomal element. It is shown here that coding sequences within the non-transmissible gene in B. fibrisolvens 1.230 are identical to those of the Streptococcus pneumoniae tet(O) gene. This provides the first evidence for genetic exchange between facultatively anaerobic bacteria and rumen obligate anaerobes. In contrast, the product of the transmissible TcR gene shares only 68% amino acid sequence identity with the TetO and TetM proteins and represents a new class of ribosome protection tetracycline resistance determinant, designated Tet W. The tet(W) coding region shows a higher DNA G + C content (53%) than other B. fibrisolvens genes or other ribosome protection-type tet genes, suggesting recent acquisition from a high G + C content genome. Tet(W) genes with almost identical sequences are also shown to be present in TcR strains of B. fibrisolvens from Australian sheep and in TcR strains of two other genera of rumen obligate anaerobes, Selenomonas ruminantium and Mitsuokella multiacidus. This provides compelling evidence for recent intergeneric transfer of resistance genes between ruminal bacteria. Tet(W) is not restricted to ruminal bacteria, as it was also present in a porcine strain of M. multiacidus.

The anaerobic spiral-shaped bacterium Anaerobiospirillum succiniciproducens was isolated from the blood of an AIDS patient for the first time in Europe. Electron-microscopical methods, especially negative staining, allowed rapid morphological classification and differentiation from Campylobacter species. While A. succiniciproducens revealed lophotriche flagellation all the investigated Campylobacter species showed monotriche flagellation. The cell diameter of A. succiniciproducens was at least double that of the investigated Campylobacter species. Other ultrastructural features, such as a ring-like structure underneath the flagellar area and fibrils arranged parallel along the axis, were also specific to A. succiniciproducens.

Strain SBT is a new, strictly anaerobic, gram-negative, nonmotile, non-sporeforming, rod-shaped bacterium that degrades benzoate and certain fatty acids in syntrophic association with hydrogen/formate-using microorganisms. Strain SBT produced approximately 3 mol of acetate and 0.6 mol of methane per mol of benzoate in coculture with Methanospirillum hungatei strain JF1. Saturated fatty acids, some unsaturated fatty acids, and methyl esters of butyrate and hexanoate also supported growth of strain SBT in coculture with Desulfovibrio strain G11. Strain SBT grew in pure culture with crotonate, producing acetate, butyrate, caproate, and hydrogen. The molar growth yield was 17 +/- 1 g cell dry mass per mol of crotonate. Strain SBT did not grow with fumarate, iron(III), polysulfide, or oxyanions of sulfur or nitrogen as electron acceptors with benzoate as the electron donor. The DNA base composition of strain SBT was 43.1 mol% G+C. Analysis of the 16 S rRNA gene sequence placed strain SBT in the delta-subdivision of the Proteobacteria, with sulfate-reducing bacteria. Strain SBT was most closely related to members of the genus Syntrophus. The clear phenotypic and genotypic differences between strain SBT and the two described species in the genus Syntrophus justify the formation of a new species, Syntrophus aciditrophicus.

Dissimilatory Fe(III) reduction by Shewanella putrefaciens and related species has generated considerable interest in biochemical characterization of the pathways for anaerobic electron transfer in this organism. Two strains, MR-1 and NCIMB 400, have been extensively used, and several respiratory enzymes have been isolated from each. It has become apparent that significant sequence differences exist between homologous proteins from these strains. The 16S rRNA from NCIMB 400 was sequenced and compared to the sequences from MR-1 and other Shewanella strains. The results indicate that NCIMB 400 is significantly more closely related to the newly identified Shewanella frigidimarina than to the S. putrefaciens type strain. It is therefore proposed that NCIMB 400 should be reclassified as S. frigidimarina.

The group of planctomycetes represents a separate line of descent within the domain Bacteria. Two phylum-specific 16S rRNA-targeted oligonucleotide probes for planctomycetes have been designed, optimized for in situ hybridization and used in different habitats to detect members of the group in situ. The probes, named PLA46 and PLA886, are targeting all or nearly all members of the planctomycete line of descent. Planctomycetes could be detected in almost all samples examined, e.g. a brackish water lagoon, activated sludge, and other wastewater habitats. In situ probing revealed quite uniform morphology and spatial arrangement of the detected cells but profound differences in abundance ranging from less than 0.1% to several percentage of the total cells. Single coccoid cells with diameters between 1 and 2.5 microm were dominating in most samples with the exception of the lagoon, in which rosettes of pear-shaped cells were abundant. The planctomycetes showed generally no hybridization signals with the bacterial probe EUB338, which is in accordance with base changes in their 165 rRNA sequences. A discrete ultrastructure of planctomycete cells was suggested by double staining with rRNA-targeted probes and the DNA-binding dye 4',6-diamidino-2-phenylindole (DAPI). The probe-conferred fluorescence was distributed in a ring-shaped manner around a central DAPI spot. The two probes developed extend the existing set of group-specific rRNA-targeted probes and help to elucidate the basic composition of bacterial communities in a first step of differential analysis. In situ hybridization of environmental samples indicated widespread presence of planctomycetes in different ecosystems.

A new anaerobic, proteolytic, moderately thermophilic bacterium, strain 3RT, was isolated from a methanogenic mesophilic reactor treating protein-rich wastewater. The cells were Gram-negative, non-spore-forming, non-motile rods. The DNA base composition was 43 mol% G + C. The optimum pH and temperature for growth were 7.0 and 55 degrees C respectively. The bacterium fermented gelatin, casein, bovine albumin, peptone and yeast extract. Glucose, fructose, sucrose, maltose and starch were poorly fermented. The major fermentation products from glucose were acetate, CO2 and H2 and, from gelatin, propionate was also detected. Growth on glucose was stimulated by thiosulfate, which was reduced to sulfide. Sulfate and nitrate were not reduced. 16S rRNA gene analysis revealed that the isolated bacterial strain was phylogenetically related to Coprothermobacter proteolyticus (96.3% sequence similarity), the only known species within the genus. DNA-DNA hybridization analysis demonstrated a very low level of homology, indicating that the isolated strain and C. proteolyticus were not related at species level. Therefore, it is proposed to classify the described strain in the genus Coprothermobacter as a new species, Coprothermobacter platensis. The type strain of C. platensis is strain 3RT (= DSM 11748T).

A syntrophic propionate-oxidizing bacterium, strain MPOBT, was isolated from a culture enriched from anaerobic granular sludge. It oxidized propionate syntrophically in co-culture with the hydrogen- and formate-utilizing Methanospirillum hungateii, and was able to oxidize propionate and other organic compounds in pure culture with sulfate or fumarate as the electron acceptor. Additionally, it fermented fumarate. 16S rRNA sequence analysis revealed a relationship with Syntrophobacter wolinii and Syntrophobacter pfennigii. The G + C content of its DNA was 60.6 mol%, which is in the same range as that of other Syntrophobacter species. DNA-DNA hybridization studies showed less than 26% hybridization among the different genomes of Syntrophobacter species and strain MPOBT. This justifies the assignment of strain MPOBT to the genus Syntrophobacter as a new species. The name Syntrophobacter fumaroxidans is proposed; strain MPOBT (= DSM 10017T) is the type strain.

Ferric iron was produced anaerobically from ferrous iron through the metabolic activity of recently described ferrous iron-oxidizing, nitrate-reducing bacteria. It was identified as poorly crystallized 2-line ferrihydrite with a particle size of 1-2 nm. This biologically produced ferrihydrite was shown to be a suitable electron acceptor for dissimilatory ferric iron-reducing bacteria in freshwater enrichment cultures, and was completely reduced to the ferrous state; no magnetite formation occurred. Geobacter metallireducens was also able to completely reduce the biologically produced ferrihydrite. These results indicate the possibility of an anaerobic, microbial cycling of iron. Using the biologically produced ferric iron, two isolates of obligately anaerobic, dissimilatory ferric iron-reducing bacteria, strains Dfr1 and Dfr2, were obtained from freshwater enrichment cultures. Analysis of 16S rRNA gene sequences revealed an affiliation with the Geobacter cluster within the family Geobacteraceae. The sequence similarity between strains Dfr1 and Dfr2 is 92.5%. The closest relative of strain Dfr1 is Geobacter sulfurreducens with 92.9%, and of strain Dfr2 Geobacter chapelleii with 93.7% sequence similarity. In addition, strains Dfr1 and Dfr2 are both able to grow by dissimilatory reduction of Mn(IV), S degree, and fumarate. Furthermore, strain Dfr2 is able to reduce akaganeite (beta-FeOOH), a more crystallized type of ferric iron oxide.

Malonomonas rubra is a microaerotolerant fermenting bacterium which can maintain its energy metabolism for growth by decarboxylation of malonate to acetate. 16S rRNA sequence analysis revealed that M. rubra is closely related to the cluster of mesophilic sulfur-reducing bacteria within the delta subclass of the Proteobacteria, with the fermenting bacterium Pelobacter acidigallici and the sulfur reducers Desulfuromusa kysingii, D. bakii and D. succinoxidans as closest relatives. The cells contain high amounts (up to 12% of the total cell protein content) of a c-type cytochrome which is present mainly (> 60%) in the cytoplasm and to minor parts in the periplasm (> 20%) and associated with the membrane fraction (> 10%), independent of the growth substrate. This cytochrome is a tetraheme cytochrome of 13,700 Da molecular mass with a midpoint redox potential of -0.210 V.M. rubra does not reduce sulfur or ferric iron compounds. Since this cytochrome appears not to be involved in the energy metabolism it is concluded that it is a remnant of sulfur-reducing ancestors of this bacterium, without a conceivable physiological function in its present energy metabolism.

A thermophilic, anaerobic, strictly autotrophic, sulphur-reducing bacterium, designated BSAT (T = type strain), was isolated from a deep-sea hydrothermal chimney sample collected at the mid-Atlantic ridge. Gram-negative cells occurred singly or in pairs as small highly motile rods. Spores were not observed. The temperature range for growth was 40 to 75 degrees C, with an optimum at 70 degrees C. The pH range for growth at 70 degrees C was from 4.4 to 7.5, with an optimum around 6.0. The sea salt concentration range for growth was 15-70 gI(-1) with an optimum at 35 gI(-1). Elemental sulphur, thiosulphate and sulphite were reduced to hydrogen sulphide. Sulphate and cystine were not reduced. The G+C content of the genomic DNA was 35 mol%. Phylogenetic analyses of the 16S rRNA gene indicated that the strain was a member of the domain Bacteria and formed a branch that was almost equidistant from members of the orders Aquificales and Thermotogales. The new organism possesses phenotypic and phylogenetic traits that do not allow its classification as a member of any previously described genus; therefore, it is proposed that this isolate should be described as a member of a novel species of a new genus, Desulfurobacterium gen. nov., of which Desulfurobacterium thermolithotrophum sp. nov. is the type species. The type strain is BSAT (= DMS 11699T).

A strictly anaerobic, nitrate-reducing bacterium, strain LuFRes1, was isolated using resorcinol as sole source of carbon and energy. The strain reduced nitrate to dinitrogen gas and was not able to use oxygen as an alternative electron acceptor. Cells were catalase-negative but superoxide-dismutase-positive. Resorcinol was completely oxidized to CO2. 16S rRNA sequence analysis revealed a high similarity with sequences of Azoarcus evansii and Azoarcus tolulyticus. Strain LuFRes1T (= DSM 12081T) is described as a new species of the genus Azoarcus, Azoarcus anaerobius.