Vesicular arbuscular mycorrhizal mediation of grass response to acidic and heavy metal depositions

Summary The perennial bunchgrassEhrharta calycina was grown with and without V.A.M. fungal infection (Glomus fasciculatum) in a sandy loam exposed to a range of acidic and heavy metal depositions. Heavy metals (Cu, Ni, Pb, Zn, Fe, and Co) were applied in simulated rain (pH 3.0, 4.0, and 5.6) at deposition rates approximating those observed to result from smelter efluents. Metal concentrations in the roots and shoots of mycorrhizal plants were greater than those of non-mycorrhizal plants. Mycorrhizal enhancement of plant metal uptake increased with greater acidity and higher heavy metal content of treatment. The growth of mycorrhizal plants was reduced compared to non-mycorrhizal plants when metal depostion was combined with simulated acid rain. We propose that mycorrhizal enhancement of heavy metal uptake caused reduced growth in plants exposed to acidic and heavy metal depositions.     

Characterization of microbial community and kinetics for spent sulfidic caustic applied autotrophic denitrification

Spent sulfidic caustic was applied to sulfur utilizing autotrophic denitrification as the simultaneous source of electron donor and alkalinity. The two experiment set-up of upflow anoxic hybrid growth reactor (UAHGR) and upflow anoxic suspended growth reactor (UASGR) was adopted and nitrate removals were similar in both reactors. Approximately 90% of the initial nitrate was denitrified at nitrate loading rate of 0.15∼0.40 kgNO₃ ⁻/m³·d. The experimental stoichiometric ratio of sulfate production to nitrate removal was ranged from 1.5 to 2.1 mgSO₄ ²⁻/mgNO₃ ⁻. During the operation period, denaturing gradient gel electrophoresis (DGGE) analysis of polymerase chain reaction (PCR)-amplified 16S rDNA fragments for the sludge sample of both reactors showed the change of microbial communities. Thiobacillus denitrificans-like microorganism occupied 28.5% (18 clones) of the 63 clones by cloning the PCR products from the sludge sample of UAHGR. Acidovorax avenae, which can reduce nitrate to nitrogen gas while oxidizing phenol (heterotrophic denitrifier), was also found in 7 clones (11.1%). Although an organic carbon source was not added to the medium, a microorganism (Kaistella koreensis) capable of oxidizing organic compounds was found in 7 clones (11.1%). Therefore, the microbial community of spent sulfidic caustic applied autotrophic denitrification process well corresponds to the substrate components of spent sulfidic caustic. Through the batch cultivation of microorganisms in UAHGR, the microbial kinetic coefficients of spent sulfidic caustic applied autotrophic denitrification were estimated to be μ _max = 0.097 h⁻¹, k _d = 0.0021 h⁻¹, K _s = 200 mgNO₃ ⁻/L, and Y = 0.31 mgMLVSS/mgNO₃ ⁻.     

Early reproduction and increased reproductive allocation in metal-adapted populations of the terrestrial isopod Porcellio scaber

Organisms inhabiting metal-contaminated areas can be stressed by metal exposure and are possibly subject to selection, resulting in increased metal tolerance and changes in growth and/or reproduction characteristics. In a previous study it was found that in the terrestrial isopod Porcellio scaber, sampled from the vicinity of a zine smelter, the body size was small and the brood size was large compared to isopods from a reference area. To assess whether these differences were due to genetic differentiation between strains, isopods collected from a reference wood, a zinc smelter area and a lead mine were cultured on non-polluted food, while growth, reproduction and metal concentrations were studied in first and second laboratory generations. The isopods from the three populations differed in age and weight at first reproduction, although there were hardly any differences in growth. The females of the mine and the smelter population started to reproduce earlier, at a lower weight, which resulted in fewer young per female. However, reproductive allocation (=wight of young relative to the weight of the mother) was higher in mine and smelter isopods. We conclude that the isopods at the metal-contaminated sites have been selected for early reproduction and increased reproductive allocation. The results indicate that populations inhabiting metal-polluted sites have probably undergone evolutionary changes. This study showed that growth and reproduction characteristics of different populations under laboratory conditions may provide information on selection processes in the field.     

Invasion of alien macroalgae in different Mediterranean habitats

The study evaluated different macroalgal invasions in the main Mediterranean coastal habitats on hard bottom. Biodiversity, species composition and structure of macroalgal assemblages were compared among non-invaded areas and areas invaded by the Chlorophyta Caulerpa racemosa var. cylindracea and by the turf-forming Rhodophyta Womersleyella setacea in three different habitats: shallow rocky bottom, deep rocky bottom and dead matte of the seagrass Posidonia oceanica. Results showed that alien macroalgae constituted a relevant component of benthic assemblages in invaded areas of the Mediterranean Sea. Assemblages invaded by Womersleyella setacea and Caulerpa racemosa showed lower values of diversity and large differences in the structure and species composition related to non-nvaded assemblages. The species that mostly suffered from invasion were erect species reproducing sexually; moreover, the dominance of W. setacea led to low abundance of native filamentous algae, while C. racemosa colonization seemed particularly threatening for encrusting algae. All the studied habitats appeared highly invasible by alien macroalgae, even if W. setacea appeared more invasive in deeper habitats, while colonization of C. racemosa seemed more serious in shallower habitats; the dead matte of P. oceanica represented a suitable substrate for the spread of both species. Differences among assemblages in different habitats were reduced in invaded areas.     

Life on the edge: hydrogen sulfide and the fish communities of a Mexican cave and surrounding waters

Most eucaryotic organisms classified as living in an extreme habitat are invertebrates. Here we report of a fish living in a Mexican cave (Cueva del Azufre) that is rich in highly toxic H₂S. We compared the water chemistry and fish communities of the cave and several nearby surface streams. Our study revealed high concentrations of H₂S in the cave and its outflow (El Azufre). The concentrations of H₂S reach more than 300 μM inside the cave, which are acutely toxic for most fishes. In both sulfidic habitats, the diversity of fishes was heavily reduced, and Poecilia mexicana was the dominant species indicating that the presence of H₂S has an all-or-none effect, permitting only few species to survive in sulfidic habitats. Compared to habitats without H₂S, P. mexicana from the cave and the outflow have a significantly lower body condition. Although there are microhabitats with varying concentrations of H₂S within the cave, we could not find a higher fish density in areas with lower concentrations of H₂S. We discuss that P. mexicana is one of the few extremophile vertebrates. Our study supports the idea that extreme habitats lead to an impoverished species diversity.     

Seasonality of VAM infection in three populations of Agrostis capillaris (Gramineae) on soil with or without heavy metal enrichment

Three populations of the perennial grass Agrostis capillaris, growing on limestone derived clay with and without natural enrichment of the heavy metals cadmium, lead, and zinc, and on a sandy soil polluted by a metal smelter have been investigated with regard to the percentage and seasonality of infection with vesicular-arbuscular mycorrhizal (VAM) fungi and its impact on mineral nutrition.In all populations VAM infection was lowest during winter, and highest during late summer and autumn. The population at the metal smelter site was less infected by VAM fungi than both other populations. The concentration of mineral nutrients for the three populations was clearly related to the soil concentration, but hardly modified by the degree of VAM infection.     

Autotrophic denitrification and inhibitory effect caused by the injection of spent sulfidic caustic in a modified Ludzack-Ettinger process

As spent sulfidic caustic (SSC) from petroleum plants contains a high concentration of alkalinity and sulfur compounds, SSC can be applied in sewage treatment system as an electron donor for autotrophic denitrification. In our previous study, the reuse of SSC in the biological nitrogen process was successful, and some neutralization may be required for stable treatment performance. In this study, the pH of SSC was neutralized to 12.0 from 13.3, and the modified Ludzack-Ettinger process was conducted for 90 days with the municipal wastewater. Some toxic effects of SSC on microorganisms were tested via a specific oxygen uptake rate (SOUR) assay. According to the SOUR assay, as compared with no SSC injection condition, SOUR was reduced by approximately 5.4% when 4 mL SSC/L was injected and the effective concentration of a toxicant causing 50% inhibition of the microorganism’s activity (EC₅₀) was 22.6 mL/L. During the days of operation, the COD removal and nitrification efficiency were over 53.0 and 98.2%, respectively. The TN removal efficiency was 56.6% and the nitrogen removal rate (NRR) was 0.15 kg/m³·d when the hydraulic retention time (HRT) in the anoxic tank was 3 h. The ratio of nitrifying bacteria was unaffected by the HRT, and Nitrobacter spp. and Nitrospira genus existed at similar ratios. The ratio of T. denitrificans increased after the injection of SSC and was approximately 6.5%.     

Fate of lead in the soil and its effects onPinus halepensis

Summary Chemical analysis of samples of needles of Aleppo pine (Pinus halepensis), humus and mineral soil collected in the vicinity of the Pb-smelter in Lavrion, Attica, showed that an area of about 2 km around the smelter is heavily polluted primarily with Pb and also with Zn, S and F^–. Most of the Pb was found accumulated in the humus and the top layer of the mineral soil.About 50% of the Pb found in the samples of Aleppo pine needles could be removed by washing them in a dilute detergent solution. One and 15% of the total Pb from Pb-polluted soil and humus samples, respectively, was extracted with normal ammonium acetate solution. No Pb or a very small amount was extracted with distilled H₂O from humus and soil samples treated with concentrations of Pb as high as 3000 ppm and incubated in the laboratory for 30 days.Humus samples treated with up to 1500 ppm Pb or Pb-polluted humus samples collected from points leeward of the smelter had no adverse effects on the germination of Aleppo pine seeds. When the germination experiment was conducted on Pb-polluted humus samples from points windward of the smelter, where a temporary pine needle death had been observed, the percentage seed germination, for two years, was found reduced and negatively correlated to the distance of the humus sampling point from the Pb-smelter.In nursery experiments Pb concentrations of up to 2000 ppm, added to a calcareous soil, had no adverse effect on height increase of Aleppo pine seedlings. Very little Pb was transported from the roots to the needles after seven months growth of the seedlings on either Pb-amended soil or on Pb-polluted soil from Lavrion. Concentrations of Pb in the roots of the seedlings, however, were found high in both cases and almost proportional to concentrations of Pb in the soil.     

Sperm production in an extremophile fish,the cave molly (<Emphasis Type="Italic">Poecilia mexicana</Emphasis>, Poeciliidae,Teleostei)

A prominent trade-off in life history theory and evolution balances the costs of reproduction with those of basic somatic needs. Hence, reproductive efforts may be reduced in environments where additional energy is required for somatic maintenance. Here, we investigated male sperm stores in Atlantic mollies (Poecilia mexicana) from a sulfidic cave and several sulfidic and non-sulfidic surface habitats. We found significant differences among populations in the number of sperm stripped per male, which was also correlated with differences in gonad weights. The largest sperm stores were detected in males from non-sulfidic surface creeks, while males from a partially sulfidic surface system had lower sperm counts, and males from completely sulfidic systems, surface as well as subterranean, had even fewer available sperm. We conclude that the extreme environmental conditions in sulfidic habitats appear to constrain male sperm production, since hydrogen sulfide as a naturally occurring toxin requires energy-demanding adaptations. Furthermore, we examined sperm counts of lab-reared cave and surface mollies in response to energy limitation. Males from stock populations were placed under high and low food treatments for a 2-week period and then stripped of sperm. Sperm counts of surface mollies tended to be reduced by low food availability, whereas sperm counts of cave mollies did not significantly vary between food treatments, which likely points towards a higher starvation resistance in cave mollies.     

GENETIC DIFFERENTIATION AND SELECTION AGAINST MIGRANTS IN EVOLUTIONARILY REPLICATED EXTREME ENVIRONMENTS

We investigated mechanisms of reproductive isolation in livebearing fishes (genus Poecilia) inhabiting sulfidic and nonsulfidic habitats in three replicate river drainages. Although sulfide spring fish convergently evolved divergent phenotypes, it was unclear if mechanisms of reproductive isolation also evolved convergently. Using microsatellites, we found strongly reduced gene flow between adjacent populations from different habitat types, suggesting that local adaptation to sulfidic habitats repeatedly caused the emergence of reproductive isolation. Reciprocal translocation experiments indicate strong selection against immigrants into sulfidic waters, but also variation among drainages in the strength of selection against immigrants into nonsulfidic waters. Mate choice experiments revealed the evolution of assortative mating preferences in females from nonsulfidic but not from sulfidic habitats. The inferred strength of sexual selection against immigrants (RI_s) was negatively correlated with the strength of natural selection (RI_m), a pattern that could be attributed to reinforcement, whereby natural selection strengthens behavioral isolation due to reduced hybrid fitness. Overall, reproductive isolation and genetic differentiation appear to be replicated and direct consequences of local adaptation to sulfide spring environments, but the relative contributions of different mechanisms of reproductive isolation vary across these evolutionarily independent replicates, highlighting both convergent and nonconvergent evolutionary trajectories of populations in each drainage.     

Avoidance of willows from moderately polluted area by leaf beetle,Melasoma lapponica: effects of emission or induced resistance?

We studied preference for willows along a pollution gradient on the Kola Peninsula, Russia, by the leaf beetle, Melasoma lapponica. Multiple tests with leaf disks demonstrated low preference for Salix borealis, S. caprea and S. phylicifolia from the plot situated 14 km from the smelter, in comparison with conspecific plants from plots situated at 1 and 29 km distances. This pattern was observed when testing beetles orginating from any plot both in 1993 and 1994, using both young and mature leaves of S. borealis. Although fumigation of S. borealis with realistic SO₂ concentration (100 g/m³) increased plant palatability, preference for plants from our study plots did not correlate with plot-specific mean SO₂ concentrations. Furthermore, no correlation with foliar concentrations of the main metal pollutants (Ni and Cu) was found. Palatability of plants was negatively correlated with population density of M. lapponica, which peaked in the moderately polluted plot 14 km from the smelter. Within this plot, beetles clearly preferred non-damaged bushes of S. borealis to previously damaged bushes. We therefore conclude that low preference of S. borealis from the moderately polluted area was caused by plant resistance induced by severe damage from M. lapponica in previous years rather than by pollution impact. However, S. caprea and S. phylicifolia had little damage from M. lapponica, and low palatability of these species in the moderately polluted plot suggests changes in plant quality similar to changes in heavily damaged bushes of S. borealis.     

Changes in antioxidant enzyme activity in the fine roots of black poplar (Populus nigra L.) and cottonwood (Populus deltoides Bartr. ex Marsch) in a heavy-metal-polluted environment

The effect of heavy metal deposition onto soil from a copper smelter on lipid peroxidation and antioxidant enzyme activity in the fine roots of two poplars (Populus nigra L. and Populus deltoides Bartr. ex Marsch) was analyzed. The subjects were mature trees growing in real environments. In both analyzed species, heavy metals stimulated the overproduction of free radicals in fine roots (measured as malondialdehyde level), which was directly proportional to advancing senescence. In young fine roots, heavy metals caused a decrease in guaiacol peroxidase activity and presumably disturbed the lignification process. Catalase was highly sensitive to the presence of heavy metals in the soil. In contrast, ascorbate peroxidase and glutathione reductase activities were unaffected by heavy metals. In the case of superoxide dismutase, a clear increase in enzyme activity was observed only in P. nigra under drought conditions, whereas it was inhibited in polluted stands.     

Extreme habitats as refuge from parasite infections? Evidence from an extremophile fish

Living in extreme habitats typically requires costly adaptations of any organism tolerating these conditions, but very little is known about potential benefits that trade off these costs. We suggest that extreme habitats may function as refuge from parasite infections, since parasites can become locally extinct either directly, through selection by an extreme environmental parameter on free-living parasite stages, or indirectly, through selection on other host species involved in its life cycle. We tested this hypothesis in a small freshwater fish, the Atlantic molly (Poecilia mexicana) that inhabits normal freshwaters as well as extreme habitats containing high concentrations of toxic hydrogen sulfide. Populations from such extreme habitats are significantly less parasitized by the trematode Uvulifer sp. than a population from a non-sulfidic habitat. We suggest that reduced parasite prevalence may be a benefit of living in sulfidic habitats.     

Ant predation ofEriocrania miners in a polluted area

We tested the hypothesis that air pollution may affect population densities of birch-feeding leafminers via changes in ant predation. Foraging activity of three ant species (Formica rufa, F. fusca andF. lemani), predation rates and population densities of both solitary and gregariousEriocrania (Lepidoptera: Eriocraniidae) miners were investigated at 13 sites around the Harjavalta copper-nickel smelter, SW Finland. Ant species differed in their distribution patterns relative to pollution. However, the total percentage of birch trees foraged by ants (all species combined) showed no correlation with the distance from the factory complex. As a result, no clear trends in predation rates were apparent in relation to the distance from the pollution source for either solitary or gregariousEriocrania species. Densities of the solitaryEriocrania species tended to increase with the distance from the pollution source whereas densities of the gregariousE. haworthi peaked close to the factory complex. No corresponding differences in predation rates between solitary and gregarious miners were found. Ant predation, thus, did not explain density patterns ofEriocrania miners in the polluted area.     

Heavy metal pollution monitoring using the brown seaweed Padina durvillaei in the coastal zone of the Santa Rosalía mining region, Baja California Peninsula, Mexico

The coastal marine sediments near Santa Rosalía on the eastern coast of the Baja California Peninsula (Mexico) are heavily polluted by metals due to copper mining and smelting over the past century (up to 1984). The present study compares the accumulation of metals in the brown seaweed Padina durvillaei from the central segment of the coast of Santa Rosalía (polluted by Co, Cu, Mn and Zn) and from two segments north and south of the known “hot spot”. The seaweed samples were collected in May and August 2004 and June 2005 at 13 stations located along the Santa Rosalía mining region. Heavy metal concentrations were measured by instrumental neutron activation analysis (Co and Fe) directly in dried homogenized subsamples or by flame atomic absorption spectrophotometry (Cd, Cu, Mn, Ni, Pb and Zn) after complete strong acid digestion of sub-samples. The means and standard deviations of the concentrations in dry tissues of Padina durvillaei specimens for all the studied metals and stations lie in the following sequence: Cd (3.6 ± 1.6 mg kg⁻¹) < Co (6.5 ± 6.1 mg kg⁻¹) < Pb (7.8 ± 6.2) < Ni (9.96 ± 5.28 mg kg⁻¹) < Cu (53 ± 38 mg kg⁻¹) < Zn (63 ± 43 mg kg⁻¹) < Mn (295 ± 269 mg kg⁻¹) < Fe (2243 ± 2325 mg kg⁻¹). This increase of the average concentrations was statistically significant. Principal Component Analysis showed that Factor 1 (36.46%) displays high positive loadings for Co, Cu, Mn and Zn, reflecting the influence of local anthropogenic pollution on the seaweed composition, while Factor 2 (26.91%) is important for Co, Fe and Ni and probably corresponds to the adsorption or accumulation of terrigenous elements of the regional origin, while Factor 3, with a high positive loading for Pb and a high negative loading for Cd, is probably controlled by local and regional anthropogenic input of Pb and episodic supply of Cd by local upwellings. The results of ANOVA for each element do not show any significant differences between the average concentrations for Cd, Fe, Ni and Pb in the seaweed of the three segments, or between the central and southern segments for Cu, Mn and Zn. Cobalt contents in the seaweed from the northern and central segments are, however, significantly different from the southern segment. This indicates that element concentrations in Padina durvillaei generally do not follow the drastic increases and gradients of Cu, Co, Mn and Zn contents detected in surface sediments. The apparent contradiction could be explained by a low geochemical mobility of these metals in the polluted sediments, composed mainly of stable residues of smelter wastes, with very low content of organic matter usually driving diagenetic mobilization of some metals into interstitial waters and then to the overlying water.     

Spectroscopic studies of cobalt-substituted ferredoxin fromClostridium pasteurianum

Summary Ferredoxin fromClostridium pasteurianum substituted with two Co atoms did not give any cobalt EPR signal at 8 K as isolated, but upon reduction with sodium dithionite, a broad signal appeared withg values that indicate highspin (S=3/2) Co(II). These signals were distinct from Co(II)-dithiothreitol signals, and disappeared upon reoxidation with air. Under anaerobic incubation of apoferredoxin with Co(II), a green derivative showed a visible spectrum typical of tetrahedral Co(Il)-thiolate coordination, which shifted dramatically upon exposure to air. The¹H-NMR spectrum of the aerobically isolated protein is reported at 300 MHz; magnetic susceptibility measurements were indicative of a diamagnetic species. These spectroscopic studies indicate that Co(II)-substituted ferredoxin is oxidized to low-spin Co(III)-ferredoxin in the presence of sulfide and oxygen. The diamagnetic Co(III) state could reversibly be reduced to highspin Co(II) by sodium dithionite.     

Effect of inorganic N on the population,in vitro colonization and morphology of Acetobacter diazotrophicus (syn. Gluconacetobacter diazotrophicus)

We investigated whether Acetobacter diazotrophicus (syn.Gluconacetobacter diazotrophicus) could be recovered only from sugarcane plants either with low or no application of fertiliser N. We report here the enrichment and enumeration of A. diazotrophicus from high N-fertilised samples where high heterotrophic populations reduce the numbers of A. diazotrophicus ultimately diminshing its isolation frequency as reported earlier. The growth medium of micropropagated sugarcane seedlings of the varieties Co 8021, Co 86249, Co 86010, Co 86032, and Co 87025 was amended with potassium nitrate, ammonium nitrate, ammonium chloride and urea. The colonisation and AR activity of A. diazotrophicus were affected in the presence of high levels (25 mM) of ammonium chloride and ammonium nitrate but remained unaffected in low levels of N (i.e 1/10th of MS liquid medium) and with high levels of potassium nitrate (25 mM) and urea (500 ppm). A. diazotrophicus was detected in the inoculated plants both at low and high levels of N based on the amplification of a specific 16S rRNA gene fragment using PCR based method targeting a stretch of 445 bp with primers AC and DI. High levels of N in the growth medium induced morphological changes on A. diazotrophicus cells resulting in long pleomorphic cells. The percentage of pleomorphic cells was in the decending order from NH₄NO₃, NH₄Cl, KNO₃, and urea. These changes were more prominent in ammonium chloride and ammonium nitrate than potassium nitrate, urea and N free medium. The morphological changes and the increased heterotrophic populations may play a role on the survival ofA. diazotrophicus in high N-fertilised samples/environments.     

Densities of microtime rodents along a pollution gradient from a copper-nickel smelter

Population densities of microtine rodents were studied along an air pollution gradient in the Kola Peninsula, Russia, by long-term and short-term trapping. The study area is affected by high sulphur dioxide and heavy metal emissions from the Severonikel copper-nickel smelter in Monchegorsk. The density of Clethrionomys rufocanus, the most abundant vole species in the area, was lowest close to the smelter and increased with distance up to the farthest, less polluted trapping sites. Clethrionomys glareolus, C. rutilus and Lemmus lemmus were absent from the most severely damaged area and were also scarce at the moderately polluted area 28 km south of the smelter. Although the population of C. glareolus has previously been cyclic at the moderately polluted area, we were unable to demonstrate any regular cycle. The most likely explanation for the low number of microtine rodents in the damaged and moderately polluted areas in a decrease in the quantity of important food plants: epiphytic lichens for C. glareolus and possibly C. rutilus, mosses for L. lemmus and seed plants, especially Vaccinium myrtillus, for C. rufocanus. Close to the smelter, direct toxic effects of heavy metals may also reduce population densities. The results show that pollutants may change the relative proportions of microtine species.     

Assessing the potential for biotic communities to recolonise freshwater wetlands affected by sulfidic sediments

1. The formation of sulfidic sediments in response to factors such as secondary salinisation and fertiliser usage is an emerging concern for the management of many freshwater wetlands. However, fundamental knowledge regarding the influence of sulfidic sediments on the aquatic biota is still lacking. 2. This study investigated the potential for biota to recolonise wetlands affected by sulfidic sediments, by assessing zooplankton hatching and aquatic plant germination following inundation with freshwater. Sediment samples were collected from 16 wetlands in the southern Murray‐Darling Basin, Australia, that ranged in condition from non‐impacted to possessing a known history of sulfidic sediments and/or acidification. 3. Principal Components Analysis indicated that the wetlands separated out into five different groups based on their sediment chemistry: non‐impacted, sulfidic, sulfidic and highly saline (sediment EC 46 800–209 000 μS cm^?1), sulfidic and potentially acidic (sediment pH 5.81–6.45 and ANC 0.07–0.31% CaCO₃), and sulfidic and acidic (sediment pH 4.37 and ANC 0.00% CaCO₃). 4. A viable dormant propagule bank was present in all wetlands, but the taxon richness of zooplankton and aquatic plants was significantly lower in wetlands affected by sulfidic sediments compared with those that were non‐affected. 5. This suggests that zooplankton and aquatic plants will be capable of recolonising wetlands that have accumulated sulfidic sediments via their propagule banks if the appropriate remediation measures are undertaken, although the communities developing are likely to be less diverse compared with those in non‐affected wetlands.     

Sulfur from benzothiophene and alkylbenzothiophenes supports growth of <Emphasis Type="Italic">Rhodococcus</Emphasis> sp. strain JVH1

Rhodococcus sp. strain JVH1 was previously reported to use a number of compounds with aliphatic sulfide bridges as sulfur sources for growth. We have shown that although JVH1 does not use the three-ring thiophenic sulfur compound dibenzothiophene, this strain can use the two-ring compound benzothiophene as its sole sulfur source, resulting in growth of the culture and loss of benzothiophene. Addition of inorganic sulfate to the medium reduced the conversion of benzothiophene, indicating that benzothiophene metabolism is repressed by sulfate and that benzothiophene is therefore used specifically as a sulfur source. JVH1 also used all six isomers of methylbenzothiophene and two dimethylbenzothiophene isomers as sulfur sources for growth. Metabolites identified from benzothiophene and some methylbenzothiophenes were consistent with published pathways for benzothiophene biodesulfurization. Products retaining the sulfur atom were sulfones and sultines, the sultines being formed from phenolic sulfinates under acidic extraction conditions. With 2-methylbenzothiophene, the final desulfurized product was 2-methylbenzofuran, formed by dehydration of 3-(o-hydroxyphenyl) propanone under acidic extraction conditions and indicating an oxygenative desulfination reaction. With 3-methylbenzothiophene, the final desulfurized product was 2-isopropenylphenol, indicating a hydrolytic desulfination reaction. JVH1 is the first microorganism reported to use all six isomers of methylbenzothiophene, as well as some dimethylbenzothiophene isomers, as sole sulfur sources. JVH1 therefore possesses broader sulfur extraction abilities than previously reported, including not only sulfidic compounds but also some thiophenic species.