Steel slag amendment impacts on soil microbial communities and activities of rice (Oryza sativa L.).

With the increase in iron/steel production, the higher volume of by-products (slag) generated necessitates its efficient recycling. Because the Linz-Donawitz (LD) slag is rich in silicon (Si) and other fertilizer components, we aim to evaluate the impact of the LD slag amendment on soil quality (by measuring soil physicochemical and biological properties), plant nutrient uptake, and strengthens correlations between nutrient uptake and soil bacterial communities. We used 16 S rRNA illumine sequencing to study soil bacterial community and APIZYM assay to study soil enzymes involved in C, N, and P cycling. The LD slag was applied at 2 Mg ha-1 to Japonica and Indica rice cultivated under flooded conditions. The LD slag amendment significantly improved soil pH, plant photosynthesis, soil nutrient availability, and the crop yield, irrespective of cultivars. It significantly increased N, P, and Si uptake of rice straw. The slag amendment enhanced soil microbial biomass, soil enzyme activities and enriched certain bacterial taxa featuring copiotrophic lifestyles and having the potential role for ecosystem services provided to the benefit of the plant. The study evidenced that the short-term LD slag amendment in rice cropping systems is useful to improve soil physicochemical and biological status, and the crop yield.

Novel reductive dehalogenases from the marine sponge associated bacterium Desulfoluna spongiiphila.

Desulfoluna spongiiphila strain AA1 is an organohalide respiring bacterium, isolated from the marine sponge Aplysina aerophoba, that can use brominated and iodinated phenols, in addition to sulfate and thiosulfate as terminal electron acceptors. The genome of Desulfoluna spongiiphila strain AA1 is approximately 6.5 Mb. Three putative reductive dehalogenase (rdhA) genes involved in respiratory metabolism of organohalides were identified within the sequence. Conserved motifs found in respiratory reductive dehalogenases (a twin arginine translocation signal sequence and two iron-sulfur clusters) were present in all three putative AA1 rdhA genes. Transcription of one of the three rdhA genes was significantly upregulated during respiration of 2,6-dibromophenol and sponge extracts. Strain AA1 appears to have the ability to synthesize cobalamin, the key cofactor of most characterized reductive dehalogenase enzymes. The genome contains genes involved in cobalamin synthesis and uptake and can grow without cobalamin supplementation. Identification of this target gene associated with debromination lays the foundation for understanding how dehalogenating bacteria control the fate of organohalide compounds in sponges and their role in a symbiotic organobromine cycle. In the sponge environment, D. spongiiphila strain AA1 may thus take advantage of both brominated compounds and sulfate as electron acceptors for respiration.

Identification of Bacteria Synthesizing Ribosomal RNA in Response to Uranium Addition During Biostimulation at the Rifle, CO Integrated Field Research Site.

Understanding which organisms are capable of reducing uranium at historically contaminated sites provides crucial information needed to evaluate treatment options and outcomes. One approach is determination of the bacteria which directly respond to uranium addition. In this study, uranium amendments were made to groundwater samples from a site of ongoing biostimulation with acetate. The active microbes in the planktonic phase were deduced by monitoring ribosomes production via RT-PCR. The results indicated several microorganisms were synthesizing ribosomes in proportion with uranium amendment up to 2 µM. Concentrations of U (VI) >2 µM were generally found to inhibit ribosome synthesis. Two active bacteria responding to uranium addition in the field were close relatives of Desulfobacter postgateii and Geobacter bemidjiensis. Since RNA content often increases with growth rate, our findings suggest it is possible to rapidly elucidate active bacteria responding to the addition of uranium in field samples and provides a more targeted approach to stimulate specific populations to enhance radionuclide reduction in contaminated sites.

Bradymonas sediminis gen. nov., sp. nov., isolated from coastal sediment, and description of Bradymonadaceae fam. nov. and Bradymonadales ord. nov.

A novel Gram-stain-negative, rod-shaped, gliding, facultatively anaerobic, oxidase-negative and catalase-positive bacterium, designated FA350(T), was isolated from coastal sediment from Xiaoshi Island, Weihai, China. Strain FA350(T) showed growth on modified nutrient agar supplemented with 0.1% d-(+)-trehalose and with distilled water replaced by seawater. Optimal growth occurred at 33 °C and pH 8.5 with 4% NaCl. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain FA350(T) belongs to a novel bacterial order in the class Deltaproteobacteria , and the most closely related type strains belong to the order Desulfuromonadales , with 85.1-85.6% 16S rRNA gene sequence similarity. The polar lipid profile of the novel strain consisted of phosphatidylethanolamine, phosphatidylglycerol and two unknown phospholipids. Major cellular fatty acids were iso-C15 : 0, iso-C17 : 0 and iso-C17 : 1ω10c and menaquinone MK-7 was the sole respiratory quinone. The DNA G+C content of strain FA350(T) was 60.3 mol%. The isolate and closely related environmental clones formed a novel order-level clade in the class Deltaproteobacteria . Comparative analysis of 16S rRNA gene sequences and characterization indicated that strain FA350(T) may represent a novel order of the Deltaproteobacteria . Here, we propose the name Bradymonas sediminis gen. nov., sp. nov. to accommodate strain FA350(T). The type strain of Bradymonas sediminis is FA350(T) ( =DSM 28820(T) =CICC 10904(T)); Bradymonadales ord. nov. and Bradymonadaceae fam. nov. are also proposed to accommodate the novel taxon.

Microbial carbon metabolism associated with electrogenic sulphur oxidation in coastal sediments.

Recently, a novel electrogenic type of sulphur oxidation was documented in marine sediments, whereby filamentous cable bacteria (Desulfobulbaceae) are mediating electron transport over cm-scale distances. These cable bacteria are capable of developing an extensive network within days, implying a highly efficient carbon acquisition strategy. Presently, the carbon metabolism of cable bacteria is unknown, and hence we adopted a multidisciplinary approach to study the carbon substrate utilization of both cable bacteria and associated microbial community in sediment incubations. Fluorescence in situ hybridization showed rapid downward growth of cable bacteria, concomitant with high rates of electrogenic sulphur oxidation, as quantified by microelectrode profiling. We studied heterotrophy and autotrophy by following (13)C-propionate and -bicarbonate incorporation into bacterial fatty acids. This biomarker analysis showed that propionate uptake was limited to fatty acid signatures typical for the genus Desulfobulbus. The nanoscale secondary ion mass spectrometry analysis confirmed heterotrophic rather than autotrophic growth of cable bacteria. Still, high bicarbonate uptake was observed in concert with the development of cable bacteria. Clone libraries of 16S complementary DNA showed numerous sequences associated to chemoautotrophic sulphur-oxidizing Epsilon- and Gammaproteobacteria, whereas (13)C-bicarbonate biomarker labelling suggested that these sulphur-oxidizing bacteria were active far below the oxygen penetration. A targeted manipulation experiment demonstrated that chemoautotrophic carbon fixation was tightly linked to the heterotrophic activity of the cable bacteria down to cm depth. Overall, the results suggest that electrogenic sulphur oxidation is performed by a microbial consortium, consisting of chemoorganotrophic cable bacteria and chemolithoautotrophic Epsilon- and Gammaproteobacteria. The metabolic linkage between these two groups is presently unknown and needs further study.

Analyses of n-alkanes degrading community dynamics of a high-temperature methanogenic consortium enriched from production water of a petroleum reservoir by a combination of molecular techniques.

Despite the knowledge on anaerobic degradation of hydrocarbons and signature metabolites in the oil reservoirs, little is known about the functioning microbes and the related biochemical pathways involved, especially about the methanogenic communities. In the present study, a methanogenic consortium enriched from high-temperature oil reservoir production water and incubated at 55 °C with a mixture of long chain n-alkanes (C(15)-C(20)) as the sole carbon and energy sources was characterized. Biodegradation of n-alkanes was observed as methane production in the alkanes-amended methanogenic enrichment reached 141.47 µmol above the controls after 749 days of incubation, corresponding to 17 % of the theoretical total. GC-MS analysis confirmed the presence of putative downstream metabolites probably from the anaerobic biodegradation of n-alkanes and indicating an incomplete conversion of the n-alkanes to methane. Enrichment cultures taken at different incubation times were subjected to microbial community analysis. Both 16S rRNA gene clone libraries and DGGE profiles showed that alkanes-degrading community was dynamic during incubation. The dominant bacterial species in the enrichment cultures were affiliated with Firmicutes members clustering with thermophilic syntrophic bacteria of the genera Moorella sp. and Gelria sp. Other represented within the bacterial community were members of the Leptospiraceae, Thermodesulfobiaceae, Thermotogaceae, Chloroflexi, Bacteroidetes and Candidate Division OP1. The archaeal community was predominantly represented by members of the phyla Crenarchaeota and Euryarchaeota. Corresponding sequences within the Euryarchaeota were associated with methanogens clustering with orders Methanomicrobiales, Methanosarcinales and Methanobacteriales. On the other hand, PCR amplification for detection of functional genes encoding the alkylsuccinate synthase α-subunit (assA) was positive in the enrichment cultures. Moreover, the appearance of a new assA gene sequence identified in day 749 supported the establishment of a functioning microbial species in the enrichment. Our results indicate that n-alkanes are converted to methane slowly by a microbial community enriched from oilfield production water and fumarate addition is most likely the initial activation step of n-alkanes degradation under thermophilic methanogenic conditions.

High-density PhyloChip profiling of stimulated aquifer microbial communities reveals a complex response to acetate amendment.

There is increasing interest in harnessing the functional capacities of indigenous microbial communities to transform and remediate a wide range of environmental contaminants. Information about which community members respond to stimulation can guide the interpretation and development of remediation approaches. To comprehensively determine community membership and abundance patterns among a suite of samples associated with uranium bioremediation experiments, we employed a high-density microarray (PhyloChip). Samples were unstimulated, naturally reducing, or collected during Fe(III) (early) and sulfate reduction (late biostimulation) from an acetate re-amended/amended aquifer in Rifle, Colorado, and from laboratory experiments using field-collected materials. Deep community sampling with PhyloChip identified hundreds-to-thousands of operational taxonomic units (OTUs) present during amendment, and revealed close similarity among highly enriched taxa from drill core and groundwater well-deployed column sediment. Overall, phylogenetic data suggested that stimulated community membership was most affected by a carryover effect between annual stimulation events. Nevertheless, OTUs within the Fe(III)- and sulfate-reducing lineages, Desulfuromonadales and Desulfobacterales, were repeatedly stimulated. Less consistent, co-enriched taxa represented additional lineages associated with Fe(III) and sulfate reduction (e.g. Desulfovibrionales; Syntrophobacterales; Peptococcaceae) and autotrophic sulfur oxidation (Sulfurovum; Campylobacterales). Data implies complex membership among highly stimulated taxa and, by inference, biogeochemical responses to acetate, a nonfermentable substrate.

Performance and microbial diversity of palm oil mill effluent microbial fuel cell.

AIM: To evaluate the bioenergy generation and the microbial community structure from palm oil mill effluent using microbial fuel cell. METHODS AND RESULTS: Microbial fuel cells enriched with palm oil mill effluent (POME) were employed to harvest bioenergy from both artificial wastewater containing acetate and complex POME. The microbial fuel cell (MFC) showed maximum power density of 3004 mW m(-2) after continuous feeding with artificial wastewater containing acetate substrate. Subsequent replacement of the acetate substrate with complex substrate of POME recorded maximum power density of 622 mW m(-2). Based on 16S rDNA analyses, relatively higher abundance of Deltaproteobacteria (88.5%) was detected in the MFCs fed with acetate artificial wastewater as compared to POME. Meanwhile, members of Gammaproteobacteria, Epsilonproteobacteria and Betaproteobacteria codominated the microbial consortium of the MFC fed with POME with 21, 20 and 18.5% abundances, respectively. CONCLUSIONS: Enriched electrochemically active bacteria originated from POME demonstrated potential to generate bioenergy from both acetate and complex POME substrates. Further improvements including the development of MFC systems that are able to utilize both fermentative and nonfermentative substrates in POME are needed to maximize the bioenergy generation. SIGNIFICANCE AND IMPACT OF THE STUDY: A better understanding of microbial structure is critical for bioenergy generation from POME using MFC. Data obtained in this study improve our understanding of microbial community structure in conversion of POME to electricity.

The quantitative significance of Syntrophaceae and syntrophic partnerships in methanogenic degradation of crude oil alkanes.

Libraries of 16S rRNA genes cloned from methanogenic oil degrading microcosms amended with North Sea crude oil and inoculated with estuarine sediment indicated that bacteria from the genera Smithella (Deltaproteobacteria, Syntrophaceace) and Marinobacter sp. (Gammaproteobacteria) were enriched during degradation. Growth yields and doubling times (36 days for both Smithella and Marinobacter) were determined using qPCR and quantitative data on alkanes, which were the predominant hydrocarbons degraded. The growth yield of the Smithella sp. [0.020 g(cell-C)/g(alkane-C)], assuming it utilized all alkanes removed was consistent with yields of bacteria that degrade hydrocarbons and other organic compounds in methanogenic consortia. Over 450 days of incubation predominance and exponential growth of Smithella was coincident with alkane removal and exponential accumulation of methane. This growth is consistent with Smithella's occurrence in near surface anoxic hydrocarbon degrading systems and their complete oxidation of crude oil alkanes to acetate and/or hydrogen in syntrophic partnership with methanogens in such systems. The calculated growth yield of the Marinobacter sp., assuming it grew on alkanes, was [0.0005 g(cell-C)/g(alkane-C)] suggesting that it played a minor role in alkane degradation. The dominant methanogens were hydrogenotrophs (Methanocalculus spp. from the Methanomicrobiales). Enrichment of hydrogen-oxidizing methanogens relative to acetoclastic methanogens was consistent with syntrophic acetate oxidation measured in methanogenic crude oil degrading enrichment cultures. qPCR of the Methanomicrobiales indicated growth characteristics consistent with measured rates of methane production and growth in partnership with Smithella.

Anaerobic biodegradation of longer-chain n-alkanes coupled to methane production in oil sands tailings.

Extraction of bitumen from mined oil sands ores produces enormous volumes of tailings that are stored in settling basins (current inventory ≥ 840 million m(3)). Our previous studies revealed that certain hydrocarbons (short-chain n-alkanes [C(6)-C(10)] and monoaromatics [toluene, o-xylene, m-xylene]) in residual naphtha entrained in the tailings are biodegraded to CH(4) by a consortium of microorganisms. Here we show that higher molecular weight n-alkanes (C(14), C(16), and C(18)) are also degraded under methanogenic conditions in oil sands tailings, albeit after a lengthy lag (~180 d) before the onset of methanogenesis. Gas chromatographic analyses showed that the longer-chain n-alkanes each added at ~400 mg L(-1) were completely degraded by the resident microorganisms within ~440 d at ~20 °C. 16S rRNA gene sequence analysis of clone libraries implied that the predominant pathway of longer-chain n-alkane metabolism in tailings is through syntrophic oxidation of n-alkanes coupled with CO(2) reduction to CH(4). These studies demonstrating methanogenic biodegradation of longer-chain n-alkanes by microbes native to oil sands tailings may be important for effective management of tailings and greenhouse gas emissions from tailings ponds.

Response of sulfate-reducing bacteria to an artificial oil-spill in a coastal marine sediment.

In situ mesocosm experiments using a calcareous sand flat from a coastal area of the island of Mallorca in the Mediterranean Sea were performed in order to study the response of sulfate-reducing bacteria (SRB) to controlled crude oil contamination, or heavy contamination with naphthalene. Changes in the microbial community caused by the contamination were monitored by a combination of comparative sequence analysis of 16S rRNA genes, fluorescence in situ hybridization, cultivation approaches and metabolic activity rates. Our results showed that crude oil and naphthalene negatively influenced the total microbial community as the natural increase in cell numbers due to the seasonal dynamics was attenuated. However, both contaminants enhanced the sulfate reduction rates, as well as the culturability of SRB. Our results suggested the presence of autochthonous deltaproteobacterial SRBs that were able to degrade crude oil or polycyclic aromatic hydrocarbons such as naphthalene in anaerobic sediment layers.

Transcription of fdh and hyd in Syntrophobacter spp. and Methanospirillum spp. as a diagnostic tool for monitoring anaerobic sludge deprived of molybdenum, tungsten and selenium.

Formate dehydrogenases and hydrogenases contain molybdenum or tungsten and/or selenium. These enzymes are crucial for interspecies formate and hydrogen transfer between propionate degrading Syntrophobacter spp. and methanogenic Methanospirillum spp. Here we used reverse transcription of total RNA followed by quantitative PCR (RT-qPCR) with specific primers to get insight into interspecies formate and hydrogen transfer. Transcriptional regulation of formate dehydrogenases and hydrogenases in Syntrophobacter and Methanospirillum spp. in a propionate-fed up-flow anaerobic sludge bed (UASB) reactor was examined. In both microorganisms formate dehydrogenase and hydrogenase coding genes (fdh and hyd respectively) were transcribed simultaneously. During 249 days in which molybdenum, tungsten and selenium were not supplied to the reactor feed, the microbial activity and transcription of fdh and hyd in Syntrophobacter spp. decreased. Transcription of fdh and hyd in Methanospirillum spp. did not decrease, but transcription of fdh increased when after 249 days molybdenum, tungsten and selenium were supplied to the reactor feed. The developed RT-qPCR is a technique that can give rapid information about active processes in methanogenic granular sludge and may contribute to predict metal limitation and failure in UASB reactors.

Evolutionary divergence and biogeography of sympatric niche-differentiated bacterial populations.

Using multiple lines of evidence from denaturing gradient gel electrophoresis, environmental sequences and TaqMan quantitative PCR assays targeting a functional gene for sulfate respiration (dsr) affiliated with the geochemically important genus Desulfobulbus, we revealed strongly restricted distributions of specific genotypes and populations correlated with sampling position along an estuarine gradient free of dispersal barriers. Evidence of evolutionary divergence of populations was provided by three complementary analyses. First, analysis of molecular variance rejected the null hypothesis that genetic diversity within each sampling site was not significantly different than that of all sites pooled together (P<0.0001). Second, UniFrac and Parsimony tests showed phylogenetic clustering of sampling sites was highly significant (P<0.001). Third, pairwise F(ST) statistics showed significant evolutionary divergence of populations based on the location in the estuary. To test the hypothesis that environmental niche-driven evolutionary divergence can create and maintain microbial biogeography, we used both statistical inference and an experimental manipulation to assess the independent effects of environment and geography. Significant effects of each on genotype distributions and population divergence supported the hypothesis. Our data are consistent with both sympatric and parapatric models of speciation, and suggest niche partitioning can contribute to evolutionary divergence and observable biogeographic patterns in microbial communities even among closely related taxa at limited spatial scales without significant barriers to dispersal.

Composition and dynamics of sulfate-reducing bacteria during the waterflooding process in the oil field application.

The composition and dynamics of sulfate-reducing bacteria (SRB) during the waterflooding process of Daqing Oilfield were investigated in this study. PCR-DGGE analysis indicated that the microbial communities were significantly different in each treatment unit, and the dominant members were mainly close to Clostridium sp., Thauera sp., Hydrogenophaga sp., Pseudomonas sp., Eubacterium sp. and Arcobacter sp. However, the members belonging to SRB were relatively few and mainly consisted of Desulfovibrio sp. and Desulfovibrio profundus. According to APS (adenosine-5'-phosphosulfate reductase) gene clone library and sequence analysis, it was found that SRB mainly belonged to Proteobacteria and Deltaproteobacteria. In the library, the genus Desulfovibrio of Deltaproteobacteria was about 65.2% while genus Desulfomonas, Desulfomicrobium, Desulfohalobium, Desulfonatronum, Desulfbulbus, Desulforhopalus, Desulfnema and Desulforhopalus were occupied nearly 34.8%. It was also proved that most of SRB in the oil field were enriched in the ground treatment unit and they are not in the liquid of oil well.

Geoalkalibacter subterraneus sp. nov., an anaerobic Fe(III)- and Mn(IV)-reducing bacterium from a petroleum reservoir, and emended descriptions of the family Desulfuromonadaceae and the genus Geoalkalibacter.

A strictly anaerobic Fe(III)-reducing bacterium, designated strain Red1(T), was isolated from the production water of the Redwash oilfield, USA. The cells were motile rods (1-5x0.5-0.6 microm) that stained Gram-negative and possessed polar flagella. Strain Red1(T) obtained energy from the reduction of Fe(III), Mn(IV), nitrate, elemental sulfur and trimethylamine N-oxide in the presence of a wide range of electron donors, including a variety of organic acids, alcohols, biological extracts and hydrogen. Strain Red1(T) was incapable of fermentative growth. The novel isolate grew optimally at 40 degrees C (temperature range for growth, 30-50 degrees C) and at pH 7 (pH range, 6-9) with 2 % (w/v) NaCl (NaCl range, 0.1-10 %, w/v). The DNA G+C content was 52.5 mol%. Phylogenetic analysis of the 16S rRNA gene sequence indicated that strain Red1(T) was a member of the order Desulfuromonadales within the class Deltaproteobacteria and most closely related to Geoalkalibacter ferrihydriticus Z-0531(T) (95.8 %), Desulfuromonas palmitatis SDBY1(T) (92.5 %) and 'Desulfuromonas michiganensis' BB1 (92.4 %). On the basis of phenotypic and phylogenetic differences, the novel strain is proposed to represent a novel species, Geoalkalibacter subterraneus sp. nov. (type strain Red1(T)=JCM 15104(T)=KCTC 5626(T)).

Field metabolomics and laboratory assessments of anaerobic intrinsic bioremediation of hydrocarbons at a petroleum-contaminated site.

Field metabolomics and laboratory assays were used to assess the in situ anaerobic attenuation of hydrocarbons in a contaminated aquifer underlying a former refinery. Benzene, ethylbenzene, 2-methylnaphthalene, 1,2,4- and 1,3,5-trimethylbenzene were targeted as contaminants of greatest regulatory concern (COC) whose intrinsic remediation has been previously reported. Metabolite profiles associated with anaerobic hydrocarbon decay revealed the microbial utilization of alkylbenzenes, including the trimethylbenzene COC, PAHs and several n-alkanes in the contaminated portions of the aquifer. Anaerobic biodegradation experiments designed to mimic in situ conditions showed no loss of exogenously amended COC; however, a substantive rate of endogenous electron acceptor reduction was measured (55 ± 8 µM SO(4) day(-1)). An assessment of hydrocarbon loss in laboratory experiments relative to a conserved internal marker revealed that non-COC hydrocarbons were being metabolized. Purge and trap analysis of laboratory assays showed a substantial loss of toluene, m- and o-xylene, as well as several alkanes (C(6)-C(12)). Multiple lines of evidence suggest that benzene is persistent under the prevailing site anaerobic conditions. We could find no in situ benzene intermediates (phenol or benzoate), the parent molecule proved recalcitrant in laboratory assays and low copy numbers of Desulfobacterium were found, a genus previously implicated in anaerobic benzene biodegradation. This study also showed that there was a reasonable correlation between field and laboratory findings, although with notable exception. Thus, while the intrinsic anaerobic bioremediation was clearly evident at the site, non-COC hydrocarbons were preferentially metabolized, even though there was ample literature precedence for the biodegradation of the target molecules.

Anaerobic degradation of p-xylene in sediment-free sulfate-reducing enrichment culture.

Anaerobic degradation of p-xylene was studied with sulfate-reducing enrichment culture. The enrichment culture was established with sediment-free sulfate-reducing consortium on crude oil. The crude oil-degrading consortium prepared with marine sediment revealed that toluene, and xylenes among the fraction of alkylbenzene in the crude oil were consumed during the incubation. The PCR-denaturing gradient gel electrophoresis (DGGE) analysis of 16S rRNA gene for the p-xylene degrading sulfate-reducing enrichment culture showed the presence of the single dominant DGGE band pXy-K-13 coupled with p-xylene consumption and sulfide production. Sequence analysis of the DGGE band revealed a close relationship between DGGE band pXy-K-13 and the previously described marine sulfate-reducing strain oXyS1 (similarity value, 99%), which grow anaerobically with o-xylene. These results suggest that microorganism corresponding to pXy-K-13 is an important sulfate-reducing bacterium to degrade p-xylene in the enrichment culture.

Desulfotignum toluenicum sp. nov., a novel toluene-degrading, sulphate-reducing bacterium isolated from an oil-reservoir model column.

A Gram-negative, sulphate-reducing bacterium (strain H3(T)) was isolated from an oil-reservoir model column. The new isolate was able to oxidize toluene coupled to hydrogen sulphide production. For growth, the optimum salt concentration was 1.5 % (w/v), the optimum pH was 7.2 and the optimum temperature was 34 degrees C. The cells were straight to slightly curved rods, 0.6-1.0 microm in diameter and 1.4-2.5 microm in length. The predominant fatty acids were C(16 : 0), C(16 : 1)omega7c and C(17 : 0) cyclo, and the cells also contained dimethylacetals. Cloning and sequencing of a 1505 bp long fragment of the 16S rRNA gene showed that strain H3(T) is a member of the Deltaproteobacteria and is related closely to Desulfotignum balticum DSM 7044(T). The G+C content of the DNA was 52.0 mol% and the DNA-DNA similarity to D. balticum DSM 7044(T) was 56.1 %. Based on differences in DNA sequence and the unique property of toluene degradation, it is proposed that strain H3(T) should be designated a member of a novel species within the genus Desulfotignum, for which the name Desulfotignum toluenicum sp. nov. is proposed. The type strain is H3(T) (=DSM 18732(T)=ATCC BAA-1460(T)).

Desulfatiferula olefinivorans gen. nov., sp. nov., a long-chain n-alkene-degrading, sulfate-reducing bacterium.

A novel anaerobic, long-chain alkene-degrading, sulfate-reducing bacterium, strain LM2801T, was isolated from brackish sediment of a wastewater decantation facility of an oil refinery (Berre lagoon, France). Cells of strain LM2801T were Gram-negative, motile, slightly curved or vibrioid rods. Its optimum growth conditions were 30-36 degrees C, 6-10 g NaCl l(-1) and pH 7.5. Strain LM2801T incompletely oxidized long-chain alkenes (from C14 to C23) and fatty acids (C14 to C24). The DNA G+C content was 45.5 mol%. Sequence analyses of the 16S rRNA and dsrAB genes indicated that the strain was a member of the family Desulfobacteraceae within the Deltaproteobacteria. This novel isolate possesses phenotypic and phylogenetic traits that do not allow its classification as a member of any previously described genus. Therefore, strain LM2801T is described as a member of a new genus, Desulfatiferula gen. nov., of which Desulfatiferula olefinivorans sp. nov. is the type species. The type strain of Desulfatiferula olefinivorans is LM2801T (=DSM 18843T=JCM 14469T).

Pelobacter seleniigenes sp. nov., a selenate-respiring bacterium.

Strain KM(T) is a novel bacterium with the unique metabolic abilities of being able to respire selenate as the electron acceptor using acetate as the carbon substrate and possessing the ability to grow fermentatively on short-chain organic acids such as lactate, citrate and pyruvate. Strain KM(T) was isolated from a sediment enrichment culture of a highly impacted wetland system in New Jersey, USA. Strain KM(T) is able to reduce selenate as well as selenite to elemental selenium. The unique metabolic capabilities of strain KM(T) include the respiration of nitrate, poorly crystalline Fe(III) and anthraquinone disulfonate. Phylogenetic analysis of the 16S rRNA gene of the novel isolate indicates that strain KM(T) groups within the family Geobacteraceae in the class Deltaproteobacteria with approximately 96-97 % 16S rRNA gene sequence similarity to the closest known organisms Malonomonas rubra Gra Mal 1(T), Pelobacter acidigallici Ma Gal 2(T) and species of the genus Desulfuromusa. Recognized species of the genera Malonomonas and Pelobacter cannot use any inorganic electron acceptors, while strains of the genus Desulfuromusa do not ferment organic substrates. This contrasts with the ability of strain KM(T) to ferment organic compounds as well as to couple selenate reduction to acetate utilization. Based on 16S rRNA gene phylogeny and metabolic properties, strain KM(T) represents a novel species for which the name Pelobacter seleniigenes sp. nov. (type strain KM(T)=DSM 18267(T)=ATCC BAA-1388(T)) is proposed. Based on the phylogenetic grouping of species of the genus Pelobacter within the Desulfuromusa cluster, it is suggested that Malonomonas rubra Gra Mal 1(T) should also be included in this group.