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A bacterium (strain G5G6) that grows anaerobically with toluene was isolated from a polluted aquifer (Banisveld, the Netherlands). The bacterium uses Fe(III), Mn(IV) and nitrate as terminal electron acceptors for growth on aromatic compounds. The bacterium does not grow on sugars, lactate or acetate. Phylogenetic analysis of the 16S rRNA gene sequence indicated that strain G5G6 belonged to the Betaproteobacteria. Its closest, but only distantly related, cultured relative is Sterolibacterium denitrificans Chol-1S(T) (94.6% similarity of the 16S rRNA genes), a cholesterol-oxidizing, denitrifying bacterium. Strain G5G6 possesses the benzylsuccinate synthase A (bssA) gene encoding the alpha-subunit of Bss, which catalyzes the first step in anaerobic toluene degradation. The deduced BssA amino acid sequence is closely related to those of Azoarcus and Thauera species, which also belong to the Betaproteobacteria. Strain G5G6 is the first toluene-degrading, iron-reducing bacterium that does not belong to the Geobacteraceae within the Deltaproteobacteria. Based on phylogenetic and physiological comparison, strain G5G6 could not be assigned to a described species. Therefore, strain G5G6 (DSMZ 19032(T)=JCM 14632(T)) is a novel taxon of the Betaproteobacteria. We propose the name Georgfuchsia toluolica gen. nov., sp. nov.

The performance of a full-scale (500 m3) sulfidogenic synthesis gas fed gas-lift reactor treating metal- and sulfate-rich wastewater was investigated over a period of 128 weeks. After startup, the reactor had a high methanogenic activity of 46 Nm3.h(-1). Lowering the carbon dioxide feed rate during the first 6 weeks gradually lowered the methane production rate. Between weeks 8 and 93, less than 1% of the hydrogen supplied was used for methanogenesis. Denaturing gradient gel electrophoresis analysis of polymerase chain reaction-amplified 16S rRNA gene fragments showed that the archaeal community decreased in diversity but did not disappear completely. After the carbon dioxide feed rate increased in week 88, the methane production rate also increased, confirming that methane production was carbon dioxide limited. Even though lowering the carbon dioxide feed appeared to affect part of the sulfate-reducing community, it did not prevent achieving the desired rates of sulfate reduction. The average sulfate conversion rate was 181 kg.h(-1) for the first 92 weeks. After 92 weeks, the sulfate input rate was increased and from week 94 to 128, the average weekly sulfate conversion rate was 295 kg.h(-1) (SD+/-87). Even higher sulfate conversion rates of up to 400 kg.h(-1) could be sustained for weeks 120-128. The long-term performance and stability together with the ability to control methanogenesis demonstrates that synthesis gas fed reactor can be used successfully at full scale to treat metal and sulfate-rich wastewater.

A thermophilic spore-forming bacterium (strain AMP) was isolated from a thermophilic methanogenic bioreactor that was fed with cobalt-deprived synthetic medium containing methanol as substrate. 16S rRNA gene analysis revealed that strain AMP was closely related to the acetogenic bacterium Moorella thermoacetica DSM 521(T) (98.3% sequence similarity). DNA-DNA hybridization showed 75.2 +/- 4.7% similarity to M. thermoacetica DSM 521(T), suggesting that strain AMP is a M. thermoacetica strain. Strain AMP has a unique one-carbon metabolism compared to other Moorella species. In media without cobalt growth of strain AMP on methanol was only sustained in coculture with a hydrogen-consuming methanogen, while in media with cobalt it grew acetogenically in the absence of the methanogen. Addition of thiosulfate led to sulfide formation and less acetate formation. Growth of strain AMP with CO resulted in the formation of hydrogen as the main product, while other CO-utilizing Moorella strains produce acetate as product. Formate supported growth only in the presence of thiosulfate or in coculture with the methanogen. Strain AMP did not grow with H(2)/CO(2), unlike M. thermoacetica (DSM 521(T)). The lack of growth with H(2)/CO(2) likely is due to the absence of cytochrome b in strain AMP.

A hydrogenotrophic, sulfate-reducing bacterium, designated strain SB1(T), was isolated from sulfidogenic sludge of a full-scale synthesis-gas-fed bioreactor used to remediate wastewater from a zinc smelter. Strain SB1(T) was found to be an abundant micro-organism in the sludge at the time of isolation. Hydrogen, formate, pyruvate, lactate, malate, fumarate, succinate, ethanol and glycerol served as electron donors for sulfate reduction. Organic substrates were incompletely oxidized to acetate. 16S rRNA gene sequence analysis showed that the closest recognized relative to strain SB1(T) was Desulfovibrio gigas DSM 1382(T) (97.5 % similarity). The G+C content of the genomic DNA of strain SB1(T) was 62.2 mol%, comparable with that of Desulfovibrio gigas DSM 1382(T) (60.2 mol%). However, the level of DNA-DNA relatedness between strain SB1(T) and Desulfovibrio gigas DSM 1382(T) was only 56.0 %, indicating that the two strains are not related at the species level. Strain SB1(T) could also be differentiated from Desulfovibrio gigas based on phenotypic characteristics, such as major cellular fatty acid composition (anteiso-C(15 : 0), iso-C(14 : 0) and C(18 : 1) cis 9) and substrate utilization. Strain SB1(T) is therefore considered to represent a novel species of the genus Desulfovibrio, for which the name Desulfovibrio paquesii sp. nov. is proposed. The type strain is SB1(T) (=DSM 16681(T)=JCM 14635(T)).

The degradation of methanethiol (MT) at 30 degrees C under saline-alkaline (pH 8-10, 0.5M Na(+)) conditions was studied in a lab-scale Upflow Anaerobic Sludge Blanket (UASB) reactor inoculated with estuarine sediment from the Wadden Sea (The Netherlands). At a sodium concentration of 0.5M and a pH between 8 and 9 complete MT degradation to sulfide, methane and carbon dioxide was possible at a maximum loading rate of 22mmolMTL(-1)day(-1) and a hydraulic retention time of 6h. The presence of yeast extract (100mg/L) in the medium was essential for complete MT degradation. 16S rRNA based DGGE and sequence analysis revealed that species related to the genera Methanolobus and Methanosarcina dominated the archaeal community in the reactor sludge. Their relative abundance fluctuated in time, possibly as a result of the changing operational conditions in the reactor. The most dominant MT-degrading archaeon was enriched from the reactor and obtained in pure culture. This strain WR1, which was most closely related to Methanolobus taylorii, degraded MT, dimethyl sulfide (DMS), methanol and trimethylamine. Its optimal growth conditions were 0.2M NaCl, 30 degrees C and pH 8.4. In batch and reactor experiments operated at pH 10, MT was not degraded.

A bacterium, strain BC, was isolated from a benzene-degrading chlorate-reducing enrichment culture. Strain BC degrades benzene in conjunction with chlorate reduction. Cells of strain BC are short rods that are 0.6 microm wide and 1 to 2 microm long, are motile, and stain gram negative. Strain BC grows on benzene and some other aromatic compounds with oxygen or in the absence of oxygen with chlorate as the electron acceptor. Strain BC is a denitrifying bacterium, but it is not able to grow on benzene with nitrate. The closest cultured relative is Alicycliphilus denitrificans type strain K601, a cyclohexanol-degrading nitrate-reducing betaproteobacterium. Chlorate reductase (0.4 U/mg protein) and chlorite dismutase (5.7 U/mg protein) activities in cell extracts of strain BC were determined. Gene sequences encoding a known chlorite dismutase (cld) were not detected in strain BC by using the PCR primers described in previous studies. As physiological and biochemical data indicated that there was oxygenation of benzene during growth with chlorate, a strategy was developed to detect genes encoding monooxygenase and dioxygenase enzymes potentially involved in benzene degradation in strain BC. Using primer sets designed to amplify members of distinct evolutionary branches in the catabolic families involved in benzene biodegradation, two oxygenase genes putatively encoding the enzymes performing the initial successive monooxygenations (BC-BMOa) and the cleavage of catechol (BC-C23O) were detected. Our findings suggest that oxygen formed by dismutation of chlorite can be used to attack organic molecules by means of oxygenases, as exemplified with benzene. Thus, aerobic pathways can be employed under conditions in which no external oxygen is supplied.

A stable anoxic enrichment culture was obtained that degraded benzene with chlorate as an electron acceptor. The benzene degradation rate was 1.65 mM benzene per day, which is similar to reported aerobic benzene degradation rates but 20-1650 times higher than reported for anaerobic benzene degradation. Denaturing gradient gel electrophoresis of part of the 16S rRNA gene, cloning and sequencing showed that the culture had a stable composition after the seventh transfer. Five bacterial clones were further analyzed. Two clones corresponded to bacteria closely related to Alicycliphilus denitrificans K601. The three other clones corresponded to bacteria closely related to Zoogloea resiniphila PIV-3A2w, Mesorhizobium sp. WG and Stenotrophomonas acidaminiphila. DGGE analysis of cultures grown with different electron donors and acceptors indicated that the bacterium related to Alicycliphilus denitrificans K601 is able to degrade benzene coupled to chlorate reduction. The role of the other bacteria could not be conclusively determined. The bacterium related to Mesorhizobium sp. WG can be enriched with benzene and oxygen, but not with acetate and chlorate, while the bacterium related to Stenotrophomonas acidaminophila grows with acetate and chlorate, but not with benzene and oxygen. As oxygen is produced during chlorate reduction, an aerobic pathway of benzene degradation is most likely.

A slow growing bacterial population able to utilize hexamethylelenetetramine (urotropine) as sole source of carbon, nitrogen and energy was isolated from soil. From this crude enrichment culture two bacteria were isolated and identified as Brevundimonas diminuta and a Phyllobacterium sp. by sequencing of 16S ribosomal DNA. These bacteria also grew on urotropine but at a lower rate than the enrichment culture. Addition of glucose to the latter resulted in growth of some yeasts that overgrew the bacteria. Assimilation of urotropine as sole nitrogen source is very common among yeasts, 46 out of 60 species tested showed this characteristic.

In multiple sclerosis intrathecal IgM synthesis correlates with an unfavourable disease course. Whether this reflects a pathogenic role of IgM, possibly in conjunction with complement, is a matter of debate. In a cross-sectional study we measured intrathecal synthesis of IgM and the complement component C3, and on cranial MRI lesion load and central brain atrophy in clinically active patients, 17 relapsing-remitting, 16 secondary progressive. Correlative analysis showed that in relapsing-remitting patients CSF IgM index correlated with cranial MRI T2 and T1 lesion load, and central brain atrophy; and the C3 index correlated with T2 lesion load. In secondary progressive patients CSF IgM index correlated with periventricular T2 lesion load. Our data are in favour of a pathogenic role of IgM in multiple sclerosis., (Copyright 2007 S. Karger AG, Basel.)

An anaerobic coculture was enriched from a hexachlorocyclohexane (HCH) polluted soil. The coculture reductively dechlorinates the beta-HCH isomer to benzene and chlorobenzene in a ratio of 0.5-2 depending on the amount of beta-HCH degraded. The culture grows with H(2) as electron donor and beta-HCH as electron acceptor, indicating that dechlorination is a respiratory process. Phylogenetic analysis indicated that the coculture consists of two bacteria that are both related to gram-positive bacteria with a low G + C content of the DNA. One bacterium was identified as a Dehalobacter sp. This bacterium is responsible for the dechlorination. The other bacterium was isolated and characterized as being a Sedimentibacter sp. This strain is not able to dechlorinate beta-HCH. The Dehalobacter sp. requires the presence of Sedimentibacter for growth and dechlorination, but the function of the latter bacterium is not clear. This is the first report on the metabolic dechlorination of beta-HCH by a defined anaerobic bacterial culture.

The biological anaerobic reductive dechlorination of beta-hexachlorocyclohexane under methanogenic conditions was tested in a number of contaminated soil samples from two locations in the Netherlands. Soils from a heavily polluted location showed rapid dechlorination of beta-hexachlorocyclohexane to benzene and chlorobenzene with lactate as electron donor. Soils from an adjacent slightly polluted location did not show substantial dechlorination of beta-hexachlorocyclohexane within 4 months. A heavily polluted sample was selected to optimise the dechlorination. All tested hexachlorocyclohexane isomers (alpha-, beta-, gamma-, and delta-), either added separately or simultaneously, were dechlorinated in this soil sample. The most rapid dechlorination was observed at a temperature of 30 degrees C. Dechlorination of beta-hexachlorocyclohexane was observed with acetate, propionate, lactate, methanol, H2, yeast extract and landfill leachate as electron donors. In a soil percolation column, packed with a selected heavily polluted soil sample, the presence of 10 mM sulphate in the influent led to simultaneous dechlorination of beta-hexachlorocyclohexane and sulphate reduction. When the column was fed with 10 mM nitrate instead of sulphate, dechlorination ceased immediately. After omitting nitrate from the influent, dechlorination activity recovered in about 1 month. Also in a separate column, the addition of nitrate from the start of the experiment did not result in dechlorination of beta-HCH. The significance of these experiments for in situ bioremediation of polluted soils is discussed.

Background & Aims: In several rat infection experiments, we have shown that dietary calcium inhibits intestinal colonization and translocation of invasive salmonella. The aim of the present study was to find out whether calcium is also protective against enterotoxigenic Escherichia coli (ETEC) infection. This was first tested in our rat model and subsequently verified in a human infection study., Methods: Rats were fed a purified diet with either a low or a high amount of calcium phosphate and orally infected with ETEC. In addition, a parallel, double-blind, placebo-controlled intervention study of 3 weeks was performed with 32 healthy men. Subjects largely maintained their habitual diet and consumed either regular milk products (calcium supply, 1100 mg/day) or placebo milk products (calcium supply, 60 mg/day). On day 10, subjects ingested a live but attenuated ETEC strain (strain E1392/75-2A), able to induce mild although short-lived symptoms. Primary outcomes studied were infection-induced diarrhea (total fecal output and relative fecal dry weight) and fecal mucin excretion., Results: In humans, ETEC induced diarrhea in both groups, in that total fecal output doubled and mean relative fecal dry weight dropped from 25% to 20%. Additionally, fecal mucin excretion was increased in both groups. All these fecal parameters were completely normalized in the calcium group on the second infection day, in contrast to the placebo group, which recovered on the third infection day. Likewise, supplemental calcium inhibited ETEC colonization and diarrhea in rats., Conclusions: Calcium in milk products improves human resistance to ETEC infection as it inhibits infectious diarrhea.