Bacterial CS2 Hydrolases from Acidithiobacillus thiooxidans Strains Are Homologous to the Archaeal Catenane CS2 Hydrolase

Carbon disulfide (CS₂) and carbonyl sulfide (COS) are important in the global sulfur cycle, and CS₂ is used as a solvent in the viscose industry. These compounds can be converted by sulfur-oxidizing bacteria, such as Acidithiobacillus thiooxidans species, to carbon dioxide (CO₂) and hydrogen sulfide (H₂S), a property used in industrial biofiltration of CS₂-polluted airstreams. We report on the mechanism of bacterial CS₂ conversion in the extremely acidophilic A. thiooxidans strains S1p and G8. The bacterial CS₂ hydrolases were highly abundant. They were purified and found to be homologous to the only other described (archaeal) CS₂ hydrolase from Acidianus strain A1-3, which forms a catenane of two interlocked rings. The enzymes cluster in a group of β-carbonic anhydrase (β-CA) homologues that may comprise a subclass of CS₂ hydrolases within the β-CA family. Unlike CAs, the CS₂ hydrolases did not hydrate CO₂ but converted CS₂ and COS with H₂O to H₂S and CO₂. The CS₂ hydrolases of A. thiooxidans strains G8, 2Bp, Sts 4-3, and BBW1, like the CS₂ hydrolase of Acidianus strain A1-3, exist as both octamers and hexadecamers in solution. The CS₂ hydrolase of A. thiooxidans strain S1p forms only octamers. Structure models of the A. thiooxidans CS₂ hydrolases based on the structure of Acidianus strain A1-3 CS₂ hydrolase suggest that the A. thiooxidans strain G8 CS₂ hydrolase may also form a catenane. In the A. thiooxidans strain S1p enzyme, two insertions (positions 26 and 27 [PD] and positions 56 to 61 [TPAGGG]) and a nine-amino-acid-longer C-terminal tail may prevent catenane formation.     

The effect of inoculation and the type of carrier material used on the biofiltration of methyl sulphides

 Low elimination capacities (less than 10 g m^-3 day^-1) were observed for the odorant dimethyl sulphide (Me₂S) when either wood bark or compost was used as the carrier material in a laboratory-scale biofilter. Enrichment experiments were set up by incubation of garden soil samples during 4 weeks with 100 ppm (v/v) headspace concentrations of both Me₂S and dimethyl disulphide (Me₂S₂). After transfer to a mineral medium, Me₂S- and Me₂S₂-degrading enrichment cultures were obtained for all five soil samples tested, both compounds being converted stoichiometrically to sulphuric acid. Upon inoculation of the laboratory-scale biofilter with one of these enrichment cultures (±120 g cell dry weight m^-3 reactor), the elimination capacity for Me₂S increased in a 3-week period to 35 g m^-3 day^-1 and 680 g m^-3 day^-1 when wood bark and compost were used as the respective carrier materials. Both inoculated biofilters were able to degrade Me₂S₂, however the elimination capacities obtained for Me₂S₂ were lower (e.g. 24 g m^-3 day^-1 for the wood bark filter) compared to those for Me₂S. For both inoculated biofilters, a gradual decrease of the elimination capacity for the methyl sulphides was observed as a result of acidification of the carrier material, suggesting that pH regulation is necessary if long-term biofiltration experiments are to be performed. Received: 6 June 1995/Received revision: 10 August 1995/Accepted: 22 August 1995     

Role of Thiobacillus thioparus in the biodegradation of carbon disulfide in a biofilter packed with a recycled organic pelletized material

This study reports the biodegradation of carbon disulfide (CS₂) in air biofilters packed with a pelletized mixture of composted manure and sawdust. Experiments were carried out in two lab-scale (1.2 L) biofiltration units. Biofilter B was seeded with activated sludge enriched previously on CS₂-degrading biomass under batch conditions, while biofilter A was left as a negative inoculation control. This inoculum was characterized by an acidic pH and sulfate accumulation, and contained Achromobacter xylosoxidans as the main putative CS₂ biodegrading bacterium. Biofilter operation start-up was unsuccessfully attempted under xerophilic conditions and significant CS₂ elimination was only achieved in biofilter A upon the implementation of an intermittent irrigation regime. Sustained removal efficiencies of 90–100 % at an inlet load of up to 12 g CS₂ m^?3 h^?1 were reached. The CS₂ removal in this biofilter was linked to the presence of the chemolithoautotrophic bacterium Thiobacillus thioparus, known among the relatively small number of species with a reported capacity of growing on CS₂ as the sole energy source. DGGE molecular profiles confirmed that this microbe had become dominant in biofilter A while it was not detected in samples from biofilter B. Conventional biofilters packed with inexpensive organic materials are suited for the treatment of low-strength CS₂ polluted gases (IL <12 g CS₂ m^?3 h^?1), provided that the development of the adequate microorganisms is favored, either upon enrichment or by inoculation. The importance of applying culture-independent techniques for microbial community analysis as a diagnostic tool in the biofiltration of recalcitrant compounds has been highlighted.     

Carbon disulfide oxidation by a microbial consortium from a trickling filter

Biological oxidation rates of CS₂ with a mixed microbial culture obtained from a trickling filter were optimal with 3 mM CS₂, pH 7, 30°C and SO₄ ^2– below 25 g l^–1. Degradation rates were 3.4 mg CS₂/g_proteinmin and 13.8 mg H₂S/g_proteinmin. The concentrations of intermediates (H₂S, COS and S°) and the product (SO₄ ^2–) of CS₂ oxidation were measured. The biological oxidation was due principally to Gram negative bacteria.     

An evaluation of carbon disulphide as a sulphur fertilizer and as a nitrification inhibitor

There was little release of extractable SO₄-S during four weeks from CS₂ applied by injecting into two S-deficient soils. In this incubation experiment, the rate of CS₂ was 30 μg S g⁻, placement was injection at 9 cm depth, soil temperature was 20°C, and soil moisture tension was 33 kPa. The yield of barley forage after seven weeks in the greenhouse showed only small increases from 10 or 30 μg S g⁻¹ of CS₂ as compared to Na₂SO₄, on the two soils. While CS₂ supplied little plant available S in the short term, it was an effective inhibitor of nitrification. In the laboratory, or in the field, the injection of CS₂ (with N fertilizers) at a point 9 cm into the soils either stopped or reduced nitrification. In one laboratory experiment, 35 μg of CS₂ g⁻¹ of soil with urea reduced nitrification for at least four weeks; and in another experiment 20 μg of CS₂ g⁻¹ of soil with aqua NH₃ nearly or completely inhibited nitrification at 20 days. In two field experiments, 3 and 12 μg of CS₂ g⁻¹ of soil (or 6 and 24 kg ha⁻¹) with aqua NH₃ inhibited nitrification from October to the subsequent May. In addition, CS₂ reduced the amount of ammonium produced from the soil N, both in these two field experiments and in the laboratory experiments. That is to say, CS₂ injected at a point, inhibited both nitrification and ammonification. In other field experiments, CS₂ at a rate of 10 kg ha⁻¹ was injected in bands 9 cm deep with urea in October, and by May there was still reduced nitrification. Less than half of the fall-applied urea alone was recovered as mineral N, but with the application of CS₂ the recovery was increased to three-quarters. The yield and N uptake of barley grain was increased where fall-applied banded urea or aqua NH₃ received banded CS₂, (NH₄)₂CS₃, or K₂CS₃. The average increase in yield from fall-applied fertilizer, from inhibitor with fall-applied fertilizer, and from spring-applied fertilizer was 800, 1370, and 1900 kg ha⁻¹, respectively. In the same order, the apparent % recovery of fertilizer N in grain was 24, 42, and 60.     

Ferrous Iron and Sulfur Oxidation and Ferric Iron Reduction Activities of Thiobacillus ferrooxidans Are Affected by Growth on Ferrous Iron, Sulfur, or a Sulfide Ore

Eight strains of Thiobacillus ferrooxidans (laboratory strains Tf-1 [= ATCC 13661] and Tf-2 [= ATCC 19859] and mine isolates SM-1, SM-2, SM-3, SM-4, SM-5, and SM-8) and three strains of Thiobacillus thiooxidans (laboratory strain Tt [= ATCC 8085] and mine isolates SM-6 and SM-7) were grown on ferrous iron (Fe²⁺), elemental sulfur (S⁰), or sulfide ore (Fe, Cu, and Zn). The cells were studied for their aerobic Fe²⁺ - and S⁰-oxidizing activities (O₂ consumption) and anaerobic S⁰-oxidizing activity with ferric iron (Fe³⁺) (Fe²⁺ formation). Fe²⁺-grown T. ferrooxidans cells oxidized S⁰ aerobically at a rate of 2 to 4% of the Fe²⁺ oxidation rate. The rate of anaerobic S⁰ oxidation with Fe³⁺ was equal to the aerobic oxidation rate in SM-1, SM-3, SM-4, and SM-5, but was only one-half or less that in Tf-1, Tf-2, SM-2, and SM-8. Transition from growth on Fe²⁺ to that on S⁰ produced cells with relatively undiminished Fe²⁺ oxidation activities and increased S⁰ oxidation (both aerobic and anaerobic) activities in Tf-2, SM-4, and SM-5, whereas it produced cells with dramatically reduced Fe²⁺ oxidation and anaerobic S⁰ oxidation activities in Tf-1, SM-1, SM-2, SM-3, and SM-8. Growth on ore 1 of metal-leaching Fe²⁺-grown strains and on ore 2 of all Fe²⁺-grown strains resulted in very high yields of cells with high Fe²⁺ and S⁰ oxidation (both aerobic and anaerobic) activities with similar ratios of various activities. Sulfur-grown Tf-2, SM-1, SM-4, SM-6, SM-7, and SM-8 cultures leached metals from ore 3, and Tf-2 and SM-4 cells recovered showed activity ratios similar to those of other ore-grown cells. It is concluded that all the T. ferrooxidans strains studied have the ability to produce cells with Fe²⁺ and S⁰ oxidation and Fe³⁺ reduction activities, but their levels are influenced by growth substrates and strain differences.     

Novel eubacteria able to grow on carbon disulfide

Four eubacterial strains able to grow on carbon disulfide (CS₂) as sole energy substrate were isolated from soil and leaves of the CS₂-producing tree Quercus lobata. Three of the isolates (strains KS1, KS2, and KL1) were gram-negative, facultatively methylotrophic, and heterotrophic, and capable of growth on a wide range of inorganic and organic sulfur compounds. Biochemical and physiological properties differed slightly among the three strains, but all are proposed to be novel thiobacillus species. Growth yields on CS₂ in batch and chemostat culture ranged from 3.3 g dry wt/mol CS₂ (batch) to a maximum growth yield (Y_max) of 11.1 g dry wt/mol (chemostat). Chemostat data for two of the strains growing, autotrophically on thiosulfate gave Y_max values of 7.4 and 7.1 g dry wt/mol, which fall within the range observed with thiobacilli. The three new Thiobacillus strains had DNA containing 39.8 (KS2), 47.8 (KS1), and 50.5 (KL1) mol% G+C. All three were unusual in being able to grow not only on thiosulfate (aerobically or with denitrification), but also on CS₂, carbonyl sulfide and methylated sulfides as sole energy substrates, and one was unique in being able to grow also on substituted thiophenes. They are the first organisms described to be capable, of anaerobic growth with denitrification on CS₂. The fourth isolate (strain KL2) was gram-positive non-motile and nonspore-forming, with 39.0 mol% G+C. It had a restricted range of sulfur-containing growth substrates, could not grow methylotrophically or on autotrophic substrates other than CS₂, and is not yet classifiable These organisms extend the range of eubacteria known to be capable of CS₂ breakdown and demonstrate that several types of facultatively chemolithotrophic bacteria, able to grow exclusively on CS₂, are associated with a CS₂-producing plant.     

Reverse-flow strategy in biofilters treating CS2 emissions

The bacteriostatic properties of carbon disulphide (CS₂) hamper its biodegradation in conventional biofilters. The response of four biofilters operating in downflow mode and reverse-flow mode was compared in a laboratory-scale plant treating CS₂ under sudden short-term changes in operating conditions. A process shutdown for 24 h, an inlet concentration increase and an interruption of the inlet air humidification for 48 h at an empty bed residence time (EBRT) of 240 s did not impact significantly on biodegradation performance, regardless of flow mode. Nevertheless, a reduction in the EBRT to 60 s resulted in a significant decrease in removal efficiency in all the biofilters. The CS₂ degradation profile showed that the reverse-flow mode strategy rendered a more homogenous distribution of biomass along the bed height. The benefits of the reverse-flow mode were demonstrated even when the unidirectional flow mode was re-established.     

Conserved Gating Elements in TRPC4 and TRPC5 Channels

TRPC4 and TRPC5 proteins share 65% amino acid sequence identity and form Ca²⁺-permeable nonselective cation channels. They are activated by stimulation of receptors coupled to the phosphoinositide signaling cascade. Replacing a conserved glycine residue within the cytosolic S4–S5 linker of both proteins by a serine residue forces the channels into an open conformation. Expression of the TRPC4_G503S and TRPC5_G504S mutants causes cell death, which could be prevented by buffering the Ca²⁺ of the culture medium. Current-voltage relationships of the TRPC4_G503S and TRPC5_G504S mutant ion channels resemble that of fully activated TRPC4 and TRPC5 wild-type channels, respectively. Modeling the structure of the transmembrane domains and the pore region (S4-S6) of TRPC4 predicts a conserved serine residue within the C-terminal sequence of the predicted S6 helix as a potential interaction site. Introduction of a second mutation (S623A) into TRPC4_G503S suppressed the constitutive activation and partially rescued its function. These results indicate that the S4–S5 linker is a critical constituent of TRPC4/C5 channel gating and that disturbance of its sequence allows channel opening independent of any sensor domain.     

Isolation of a carbon disulfide utilizing <Emphasis Type="Italic">Thiomonas</Emphasis> sp. and its application in a biotrickling filter

The carbon disulfide (CS₂)-oxidizing bacterium Thiomonas sp. WZW was enriched and isolated using activated sewage sludge as inoculum. Growth of Thiomonas sp. WZW was observed on CS₂, thiosulfate, dimethylsulfide (DMS), dimethyldisulfide (DMDS), and H₂S. No growth occurred on dimethylsulfoxide, methanol, acetate, and on complex media with glucose, yeast extract, or tryptone. DMDS-grown cells respired CS₂, DMS, and DMDS, while thiosulfate-grown cells did not respire CS₂. Chemostat cultures growing on thiosulfate could be rapidly adapted to growth on CS₂. Growth was observed between pH 6 and 8. The K _s values for CS₂, thiosulfate, and sulfide of CS₂-grown cells were between 5 and 10 μM. CS₂ was inhibitory above 0.3 mM. A lab-scale biotrickling filter with lava stone as carrier material for treatment of CS₂-polluted air was inoculated with Thiomonas sp. WZW. A rapid start up (95% removal in 1 week) was obtained at an inlet CS₂ concentration of 2 cmol l⁻¹ and an initial space velocity (SV) of 54 h⁻¹. Subsequent thiosulfate addition for a week during start up increased the removal to 99%. The step-wise increase of SV to 130 h⁻¹ and a CS₂ concentration to 3 μmol l⁻¹ resulted in a stable performance with a removal efficiency of 95%. Feeding mixtures of volatile sulfur compounds showed simultaneous conversion of H₂S, CS₂, dimethyldisulfide (DMDS), and DMS, with a preference in this order.     

Phenotypic Characteristics of Thiobacillus ferrooxidansStrains

Phenotypicpolymorphism of Thiobacillus ferrooxidansstrains isolated from various ecological niches was studied. The strains differed both in rates of growth and oxidation of Fe²⁺, S⁰, FeS₂, and sulfide minerals contained in concentrate. Each strain, irrespective of its original environment, required a period of adaptation to a new substrate. Strains TFN-d, TFBk, TFO, and TFL-2, isolated from ores and concentrates rich in oxidizable substrates, showed an equal adaptation rate (five culture transfers) but differed in their adaptation efficiency. Strain TFV-1, isolated from low-grade ore and showing the lowest rates of growth and oxidation of all the substrates, required five culture transfers to adapt to S⁰and FeS₂and seven culture transfers to adapt to the concentrate. It is concluded that the phenotypic properties of the strains correlate with their genotypic polymorphism and the environmental conditions under which their microevolution took place.     

Localisation régionale des gènes humains IDHS,MDHS, PGK, α-GAL,G6PD par l'hybridation cellulaire interspécifique

Summary 22 independent man-hamster (HGPRT^–) hybrids using male human cells with balanced reciprocal translocation t(X;2)(p22;q32) were analysed for human genes localized on chromosome 2 (IDH_S, MDH_S), on chromosome X (PGK, GAL, G6PD) and for the different chromosomes in relation with the balanced reciprocal translocation (chr.2, chr.2q^–, chr.Xp⁺).The following results were obtained:The chromosomes 2 and 2q^– are absent in the 22 hybrids.In 9 hybrids, the absence of MDH_S in spite of the presence of the chromosome Xp⁺ indicates that the gene for MDH_S is not localized on this chromosome (or that the gene for MDH_S is not on the segment 2q32-2qter translocated on X).In 14 hybrids, the three markers of X (PGK, GAL, G6PD) and IDH_S are expressed in the presence of the chromosome Xp⁺. This result indicates that the genes for these markers are on Xp⁺ or that the genes PGK, GAL, G6PD are on X without the Xp22-Xter segment, translocated on the chr.2, and that the gene for IDH_S is on the 2q32-2qter segment translocated on X.In 8 hybrids, in the absence of the intact chromosome Xp⁺, the higher percentage of the presence of G6PD (7 hybrids) and the lower percentage of the presence of IDH_S (3 hybrids) are explained by the fact that these hybrids selected in HAT medium had to retain a segment of Xp⁺ bearing the human gene HGPRT. G6PD appeared very close to HGPRT and IDH_S very distant from HGPRT.The study of the different correlations between the presence and the absence of these four markers on Xp⁺ in the different hybrids indicates the following order on the chromosome Xp⁺ from p to q: IDH_s — PGK — GAL — G6PD.

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Groupe INSERM: Directeur J. Frézal

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Groupe CNRS, ER, 149: Directeur J. de Grouchy     

The Ca-dependent interaction of calpastatin domain L with the C-terminal tail of the Cav1.2 channel

To demonstrate the interaction of calpastatin (CS) domain L (CS_L) with Cav1.2 channel, we investigated the binding of CS_L with various C-terminus-derived peptides at ≈ free, 100 nM, 10 μM, and 1 mM Ca²⁺ by using the GST pull-down assay method. Besides binding with the IQ motif, CS_L was also found to bind with the PreIQ motif. With increasing [Ca²⁺], the affinity of the CS_L–IQ interaction gradually decreased, and the affinity of the CS_L–PreIQ binding gradually increased. The results suggest that CS_L may bind with both the IQ and PreIQ motifs of the Cav1.2 channel in different Ca²⁺-dependent manners.     

Plasmid Profiles of Acidithiobacillus ferrooxidans Strains Adapted to Different Oxidation Substrates

Plasmid profiles were studied in five Acidithiobacillus ferrooxidans strains of various origin cultivated on a medium with Fe²⁺, as well as adapted to such oxidation substrates as S⁰, FeS₂, and sulfide concentrate. The method used revealed plasmids in all A. ferrooxidans strains grown on a medium with Fe²⁺. One plasmid was found in strain TFL-2; two plasmids, in strains TFO, TFBk, and TFV-1; and three plasmids were detected in strain TFN-d. The adaptation of strain TFN-d to sulfide concentrate and the adaptation of strain TFV-1 to S⁰, FeS₂, or sulfide concentrate resulted in a change in the number of plasmids occurring in cells. In cells of strain TFN-d adapted to sulfide concentrate, the number of plasmids decreased from three to two. The number of plasmids in cells of strain TFV-1 adapted to different substrates varied from three to six depending on the energy source present in the medium: three plasmids were found after growth on FeS₂, four after growth on S⁰, and six after growth on sulfide concentrate. The possible role of plasmids in the adaptation of A. ferrooxidans to new energy substrates and in the regulation of the intensity of their oxidation is discussed.     

Bacterial Disproportionation of Elemental Sulfur Coupled to Chemical Reduction of Iron or Manganese

A new chemolithotrophic bacterial metabolism was discovered in anaerobic marine enrichment cultures. Cultures in defined medium with elemental sulfur (S⁰) and amorphous ferric hydroxide (FeOOH) as sole substrates showed intense formation of sulfate. Furthermore, precipitation of ferrous sulfide and pyrite was observed. The transformations were accompanied by growth of slightly curved, rod-shaped bacteria. The quantification of the products revealed that S⁰ was microbially disproportionated to sulfate and sulfide, as follows: 4S⁰ + 4H₂O → SO₄^2- + 3H₂S + 2H⁺. Subsequent chemical reactions between the formed sulfide and the added FeOOH led to the observed precipitation of iron sulfides. Sulfate and iron sulfides were also produced when FeOOH was replaced by FeCO₃. Further enrichment with manganese oxide, MnO₂, instead of FeOOH yielded stable cultures which formed sulfate during concomitant reduction of MnO₂ to Mn²⁺. Growth of small rod-shaped bacteria was observed. When incubated without MnO₂, the culture did not grow but produced small amounts of SO₄^2- and H₂S at a ratio of 1:3, indicating again a disproportionation of S⁰. The observed microbial disproportionation of S⁰ only proceeds significantly in the presence of sulfide-scavenging agents such as iron and manganese compounds. The population density of bacteria capable of S⁰ disproportionation in the presence of FeOOH or MnO₂ was high, > 10⁴ cm^-3 in coastal sediments. The metabolism offers an explanation for recent observations of anaerobic sulfide oxidation to sulfate in anoxic sediments.     

Chemolithoautotrophy in the marine,magnetotactic bacterial strains MV-1 and MV-2

Magnetite-producing magnetotactic bacteria collected from the oxic–anoxic transition zone of chemically stratified marine environments characterized by O₂/H₂S inverse double gradients, contained internal S-rich inclusions resembling elemental S globules, suggesting they oxidize reduced S compounds that could support autotrophy. Two strains of marine magnetotactic bacteria, MV-1 and MV-2, isolated from such sites grew in O₂-gradient media with H₂S or thiosulfate (S₂O₃^2–) as electron sources and O₂ as electron acceptor or anaerobically with S₂O₃^2– and N₂O as electron acceptor, with bicarbonate (HCO₃^–)/CO₂ as sole C source. Cells grown with H₂S contained S-rich inclusions. Cells oxidized S₂O₃^2– to sulfate (SO₄^2–). Both strains grew microaerobically with formate. Neither grew microaerobically with tetrathionate (S₄O₆^2–), methanol, or Fe²⁺ as FeS, or siderite (FeCO₃). Growth with S₂O₃^2– and radiolabeled ¹⁴C-HCO₃^– showed that cell C was derived from HCO₃^–/CO₂. Cell-free extracts showed ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) activity. Southern blot analyses indicated the presence of a form II RubisCO (cbbM) but no form I (cbbL) in both strains. cbbM and cbbQ, a putative post-translational activator of RubisCO, were identified in MV-1. MV-1 and MV-2 are thus chemolithoautotrophs that use the Calvin–Benson–Bassham pathway. cbbM was also identified in Magnetospirillum magnetotacticum. Thus, magnetotactic bacteria at the oxic–anoxic transition zone of chemically stratified aquatic environments are important in C cycling and primary productivity.     

ATP-dependent Pre-replicative Complex Assembly Is Facilitated by Adk1p in Budding Yeast

Pre-replicative complex (pre-RC) assembly is a critical part of the mechanism that controls the initiation of DNA replication, and ATP binding and hydrolysis by multiple pre-RC proteins are essential for pre-RC assembly and activation. Here, we demonstrate that Adk1p (adenylate kinase 1 protein) plays an important role in pre-RC assembly in Saccharomyces cerevisiae. Isolated from a genetic screen, adk1^G20S cells with a mutation within the nucleotide-binding site were defective in replication initiation. adk1Δ cells were viable at 25 °C but not at 37°C. Flow cytometry indicated that both the adk1-td (temperature-inducible degron) and adk1^G20S mutants were defective in S phase entry. Furthermore, Adk1p bound to chromatin throughout the cell cycle and physically interacted with Orc3p, whereas the Adk1^G20S protein had a reduced ability to bind chromatin and Orc3p without affecting the cellular ATP level. In addition, Adk1p associated with replication origins by ChIP assay. Finally, Adk1-td protein depletion prevented pre-RC assembly during the M-to-G₁ transition. We suggest that Adk1p regulates ATP metabolism on pre-RC proteins to promote pre-RC assembly and activation.     

Heterobimetallic platinum-bismuth aggregates derived from [Pt2(μ-S)2(PPh3)4]

The metalloligand [Pt₂(μ-S)₂(PPh₃)₄] reacts with Bi(S₂CNEt₂)₃ or Bi(S₂COEt)₃ in methanol to produce the orange cationic adducts [Pt₂(μ-S)₂(PPh₃)₄Bi(S₂CNEt₂)₂]⁺ and [Pt₂(μ-S)₂(PPh₃)₄Bi(S₂COEt)₂]⁺, respectively, isolated as their hexafluorophosphate salts. An X-ray structure determination on [Pt₂(μ-S)₂(PPh₃)₄Bi(S₂CNEt₂)₂]PF₆ reveals the presence of a six-coordinated bismuth centre with an approximately nido-pentagonal bipyramidal coordination geometry. Fragmentation pathways for both complexes have been probed using electrospray ionisation mass spectrometry; ions [Pt₂(μ-S)₂(PPh₃)₂Bi(S₂CXEt_n)₂]⁺ (X = O, n = 1, X = N, n = 2) are formed by selective loss of two PPh₃ ligands, and at higher cone voltages the species [(Ph₃P)PtS₂Bi]⁺ is observed. Ions formed by loss of CS₂ are also observed for the xanthate but not the dithiocarbamate ions.     

Survey of petroleum-degrading bacteria in coastal waters of Sunderban Biosphere Reserve

A survey of petroleum-degrading bacteria was carried out in the Indian part of deltaic Sunderbans to evaluate the distribution of the naturally occurring petroleum-degrading aerobic bacteria. Bacteriological analysis of surface water samples collected from five different locations in the Hooghly–Matla river mouth showed that, depending on the location, 0.08–2.0% of the heterotrophic bacteria culturable in marine agar medium could degrade crude petroleum hydrocarbons as the sole source of carbon. In the entire study area, the number of heterotrophic bacteria ranged from 1 × 10³ to 3.8 × 10⁵ c.f.u/ml, amongst which 2.7 × 10¹ to 6 × 10³ c.f.u/ml were petroleum degraders. There was a maximum number of petroleum-degrading bacteria in the waters of Haldia Port and its surrounding areas, where the water is highly polluted by hydrocarbon discharges from a nearby oil refinery and from the ships docking at the port. Among the isolates, identified on the basis of their Gram reaction, morphological and biochemical tests including the use of API20E strips, Pseudomonas, Mycobacterium, Klebsiella, Acinetobacter, Micrococcus, and Nocardia were the most common petroleum degraders. Other heterotrophic bacteria included several species of Escherichia, Klebsiella, non-oil-degrading Pseudomonas, Vibrio, Streptococcus, Staphylococcus and Bacillus. Following preliminary selection, five strains, showing best growth in medium with oil fraction as sole carbon source, were chosen for estimation of the efficiency of crude oil biodegradation. The selected strains belonged to Pseudomonas (two strains), Mycobacterium (two strains), and Nocardia (one strain). These strains degraded 47–78% of Arab-Mix crude oil over a period of 20 days. The best oil-degrading isolate, a strain of Pseudomonas aeruginosa, (BBW1), was found to degrade and multiply more rapidly in crude oil than the rest. BBW1 showed profuse growth in Bushnell Haas medium containing crude oil (as sole source of carbon) at high concentrations ranging from 0.2 to 20% (v/v), with optimum at 10%. As much as 75% of the oil was degraded within 72 h of incubation with the bacteria. Physicochemical analysis showed considerable decrease in initial boiling point and carbon residue of the degraded oil. The ability to degrade crude oil was found to be associated with a single 70-kb plasmid, pBN70. Resistance to the metals Mn²⁺ (50 mM), Mg²⁺ (200 mM), Zn²⁺ (6 mM), Ni²⁺ (10 mM) and antibiotics like ampicillin (10 g/ml), cephalexin (30 g/ml), nitrofurantoin (300 g/ml) and penicillin (10 U/ml) were plasmid-mediated.     

Effects of acute CS2 intoxication on liver protein and drug metabolism

Rats pretreated with phenobarbitone and their controls were exposed to 0.15% atmospheric CS₂ for 2 h. Liver protein metabolism and microsomal drug metabolizing enzyme activities were analyzed 1, 4 and 46 h later. In the pretreated rats the [¹⁴C]leucine uptake was at first inhibited, then liver RNA tended to increase. This increase was followed by a decline in the [¹⁴C]leucine uptake, while RNA content diminished to the control level. In the control rats (not pretreated with phenobarbital) the effects of CS₂ on liver protein metabolism were less; only at 4 h after the exposure was liver RNA increased and [¹⁴C]leucine uptake slightly stimulated. In the pretreated rats CS₂ had decreased microsomal P-450 by about 50% at 1 and 4 h, and the activity of 7-O-dealkylase of ethoxycoumarin had decreased even more. The measurable UDPglucuronosyltransferase of the liver microsomes of the pretreated rats had increased by 26% at 1 h and by 80% at 4 h after the CS₂ exposure. The in vivo activation of microsomal UDPglucuronosyltransferase may result from a stimulated lipid peroxidation of reticuloendothelial membranes by CS₂ metabolites in vivo, as suggested by the diene conjugation spectra. In control rats CS₂ depressed only the ethoxycoumarin deethylase activity. All the microsomal changes caused by CS₂ exposure were restored within 46 h.It is suggested that the different action of CS₂ on the liver protein metabolism of rats pretreated with phenobarbitone and controls results from the different stage of protein turnover in the two groups, i.e. in the barbituratetreated animals there is a phase of increased protein synthesis and accordingly the protein turnover is more sensitive to the action of CS₂.