Dual augmentation for aerobic bioremediation of MTBE and TCE pollution in heavy metal-contaminated soil

In this work we isolated from soil and characterized several bacterial strains capable of either resisting high concentrations of heavy metals (Cd²⁺ or Hg²⁺ or Pb²⁺) or degrading the common soil and groundwater pollutants MTBE (methyl-tert-butyl ether) or TCE (trichloroethylene). We then used soil microcosms exposed to MTBE (50 mg/l) or TCE (50 mg/l) in the presence of one heavy metal (Cd 10 ppm or Hg 5 ppm or Pb 50 or 100 ppm) and two bacterial isolates at a time, a degrader plus a metal-resistant strain. Some of these two-membered consortia showed degradation efficiencies well higher (49–182% higher) than those expected under the conditions employed, demonstrating the occurrence of a synergetic relationship between the strains used. Our results show the efficacy of the dual augmentation strategy for MTBE and TCE bioremediation in the presence of heavy metals.     

Phytosociological and geographical survey of Mesohromion communities in Western Europe

A survey is given of the floristic composition and geographical distribution of limestone grasslands assigned to the Mesobromion ereeti, in Western Europe.This survey is based upon 800 vegetation relevés from various investigators throughout the area. Relevant tables have been summarized into a synoptic table with 30 columns. Each column represents a community described as an association or a subassociation.The synoptic table shows a division into a NW European group (Ireland, England, Denmark, and South Sweden) and a Central-W European group (Germany, The Netherlands, Belgium, France, and Switzerland). Associations belonging to the former have, as a rule, a smaller distribution area than the latter.In the latter group only two associations can be distinguished: Mesobrometum erecti and Antherico-Brometum ass. nov. The Mesobrometum is widespread and includes many subassociations. The Antherico-Brometum shows a transition towards the Xerobromion erecti.According to Barkman et al., (1976) the often used name Gentiano-Koelerietum has to be seen as a not-correct synonym for the Mesobrometum erecti.The Mesobromion grasslands can, in general, not be divided based upon their agricultural management system.An attempt is made to interpret the gradual transition in floristic composition of the limestone grasslands from NW to SE, with climatological data.Plant nomenclature follows Heukels & Van Ooststroom (1975) or, else, Clapham et al. (1968) for phanerogams and Vander Wijk et al. (1969) for bryophytes.I am much indebted to Prof. Dr. J. J. Barkman, who encouraged me to tackle the present study and stimulated me throughout its duration.     

Characterization of a heavy metal-tolerant endomycorrhizal fungus from the surroundings of a zinc refinery

An arbuscular mycorrhizal fungus was isolated from the rhizosphere of Agrostis capillaris growing in the contaminated surroundings of a zinc refinery in The Netherlands. After examination of the infection pattern and the spores, it was characterized as Scutellospora dipurpurescens, which was first isolated from a reclaimed coal mine area in West Virginia.     

Pattern of species dominance and factors affecting community composition in areas deforested due to air pollution

Vegetation of mountain areas affected by SO₂ pollution (Kruné hory Mts., Czech Republic) was analysed using multivariate methods. Communities with prevailing species Calamagrostis villosa, a rhizomatous grass expanding into deforested sites, were sampled by Braun-Blanquet relevé method. Canonical correspondence analysis was used to assess the effect of environmental variables (soil removal, deforestation, and shading). To test the effects of light, moisture, soil acidity and nitrogen, mean sample indicator values were correlated a posteriori with sample axes on ordination scores. Light, soil acidity, moisture, and site history (in terms of past deforestation and soil removal applied in reclamation procedures) were found to be the main factors responsible for the community composition. Nitrogen level had not a significant effect on the community composition. When analyzing the whole data set, i.e. including also remnants of natural spruce forests, light was the factor affecting at most the composition of communities. Within the bare spot vegetation, if treated separately, the highest variation was found along the soil acidity/moisture gradient. The effect of soil removal was only obvious at early successional stages. Species diversity increased with moisture and decreased with soil acidity. Species exhibiting S- and/or R-strategy are successful on extremely acid soils whereas forbs present in bare spots appear to be supported by disturbances.     

Fifty years vegetation development of a xerothermic calcareous grassland in Central Europe after heavy disturbance

Fifty years ago on the south-facing slope of a limestone hill 4 km NE of Göttingen (Germany) a small plot of 4 m² size was chosen in a xerothermic calcareous grassland (Mesobromion alliance) for investigation. The formerly grazed slope was abandoned a few years prior to the study, but in 1987 intermittent grazing started again. The mean annual temperature in Göttingen (1953–2003) was 8.8 °C on the average; the annual mean increased by 1.1 °C during the study period. Mean annual precipitation amounted to 642 mm (393–1093 mm). The mean monthly precipitation during summer (April–August) varied in the range between 29 and 101 mm. In the years 1953–2003 the presence of species was recorded annually in 400 subplots (100 cm² each). In part of this time also the amount of bare soil within the plot was recorded, and standing crop and the nitrogen contents in leaves of Bromus erectus were measured in the immediate surrounding of the plot.In fall 1953 the vegetation was removed completely in half of the plot, in the other half the two dominant grasses Bromus erectus and Brachypodium pinnatum were left. The differences between the two halves vanished already after 2 years (starting phase 1953–1955). Thus, only results concerning the plot as a whole are reported here. During the investigation three periods could be distinguished. After the starting phase in period I (1955–1986) total presence, presence of the most important species, species richness, number of endangered species, diversity and evenness increased. In this way the grassland regenerated after the heavy disturbance, and a dense, species-rich vegetation formed. Only a few species showed maximum values already within the first 10 years and faded out later.Period II (1987–1997) spanned 10 years of grazing. During this time total presence, presence of many important species, species richness, number of endangered species, diversity and evenness decreased, whereas areas with bare soil increased. Only two species showed maximum values during this phase, and just one new species invaded (Lolium perenne). During this period a more open type of grassland was formed. Period III (1998–2003) spanned 5 years without grazing and only one grazing year (2002). Here, a partial regeneration of a more closed grassland took place with traits similar to period I.Several phenomena occurred continuously through all three periods. Bromus erectus always was the dominating species. Nevertheless, proportions of graminoids (in % total presence) were lowered, and the proportion of herbs increased. The ratio Bromus erectus/Brachypodium pinnatum was determined mainly by Brachypodium. Seedlings of woody plants were always present, but in most cases survived only one summer. Shrubs did not settle inside the plot, although small bushes developed in the immediate surrounding. In summary the 50-year process can be described as a slow recovery of a calcareous grassland after very heavy disturbance, modified by intermittent grazing, without formation of shrub vegetation.     

Uptake of heavy metals by <Emphasis Type="Italic">Stylonychia mytilus</Emphasis> and its possible use in decontamination of industrial wastewater

The heavy metal resistant ciliate, Stylonychia mytilus, isolated from industrial wastewater has been shown to be potential bioremediator of contaminated wastewater. The ciliate showed tolerance against Zn²⁺ (30 μg/mL), Hg²⁺ (16 μg/mL) and Ni²⁺ (16 μg/mL). The metal ions slowed down the growth of the ciliate as compared with the culture grown without metal stress. The reduction in cell population was 46% for Cd²⁺, 38% for Hg²⁺, 23% for Zn²⁺, 39% for Cu²⁺ and 51% for Ni²⁺ after 8 days of metal stress. S. mytilus reduced 91% of Cd²⁺, 90% of Hg²⁺ and 98% of Zn²⁺ from the medium after 96 h of incubation in a culture medium containing 10 μg/mL of the respective metal ions. Besides this, the ciliate could also remove 88% of Cu²⁺ and 73% Ni²⁺ from the medium containing 5 μg/mL of each metal after 96 h. The ability of Stylonychia to take up variety of heavy metals from the medium could be exploited for metal detoxification and environmental clean-up operations.     

Cloning and expression analysis of SKn-type dehydrin gene from bean in response to heavy metals

A heavy metal responsive gene PvSR3 (GenBank accession number U54703) encoding an acid dehydrin was isolated from a mercuric chloride-treated bean (Phaseolus vulgaris L.) leaf cDNA library by differential screening using cDNAs derived from treated and untreated plants. The PvSR3 cDNA is 981-bp long and has a 606-bp open-reading frame with a 202-residue-deduced amino acid sequence. The PvSR3 sequence contains two conserved repeats of the characteristic lysine-rich K segment (EKKGIMDKIKEKLPG) preceded by an 8-serine residue stretch, whereas the Y segment (DEYGNP) conserved motif is absent. The deduced protein has a calculated molecular weight of 23 kDa and an isoelectric point of 5.2. Sequence similarity and comparative analysis showed that PvSR3 shares 70 and 73% similarity with the dehydrin of poplar and pepper, respectively. Southern hybridizations indicated that PvSR3 was a low copy-number gene. Northern blot analysis revealed that PvSR3 mRNA was weakly detected in seedling leaves. However, the gene expression was strongly stimulated by heavy metals, such as mercury, cadmium, arsenic, and coppper, whereas virus infection and salt had little effect on it. In contrast, PvSR3 was not responsive to drought or abscisic acid (ABA), and was downregulated by UV radiation. Furthermore, PvSR3 was upregulated by the exogenous signaling molecules, including salicylic acid (SA) and hydrogen peroxide (H₂O₂). It is suggested that PvSR3 is extremely related to heavy metal stress, and might play an important role in metal detoxification and resistance to the damage caused by heavy metals.     

不同分布区胀果甘草原生境土壤微生物群落结构特征及其影响因素

胀果甘草(Glycyrrhiza inflata)主要分布于新疆、甘肃的荒漠区,是耐盐性最强的药用甘草,在改良盐碱地土壤中发挥着重要作用,其原生境土壤微生物群落结构特征是揭示种群分布影响因素及盐碱地修复机制的重要依据。从胀果甘草5个主产区采集原生境土壤,测定土壤理化指标,并采用高通量测序技术,结合Spearman、dbRDA等方法开展微生物群落组成及多样性特征研究,揭示不同分布区的优势微生物群落特征和影响因子。结果表明：真菌群落中曲霉属(Aspergillus)、地丝霉属(Geomyces)、镰刀菌属(Fusarium)和细菌群落中的寡养单胞菌属(Stenotrophomonas)、Marinimicrobium、Idiomarina是野生胀果甘草原生境土壤中的优势微生物类群。不同分布区的土壤真菌多样性和丰富度具有显著差异,但土壤细菌多样性和丰富度差异不显著;部分分布区土壤中的真菌和细菌种类差异较大。土壤理化因子中,土壤含水量和总含盐量对真菌和细菌的群落分布、丰富度有显著影响。原生境含水量与曲霉属(Aspergillus)、镰刀菌属(Fusarium)、链格孢属(Alternaria)...     

Morphological changes in an acidophilic bacterium induced by heavy metals

The Acidiphilium strains inhabit acidic mine regions where they are subjected to occasional environmental stresses such as high and low temperatures, exposure to various heavy metals, etc. Change in morphology is one of the strategies that bacteria adopt to cope with environmental stresses; however, no study on this aspect has been reported in the case of Acidiphilium sp. This work is an attempt using the acidophilic heterotrophic bacterium Acidiphilium symbioticum H8. It was observed that the maximum alterations in size occurred when the bacterium was exposed to sub-inhibitory concentrations of Cu and Cd. Loosely packed coccobacillus-type normal cells formed characteristic chains of coccoidal lenticular shape with constrictions at the junctions between them in the presence of Cd; Cu induced transformation of cells to become round shaped; Ni caused the cells to aggregate, but Zn showed no effect. Respective metal depositions on the cell surface were confirmed by scanning electron microscopy equipped with energy dispersive X-ray analysis. Cell bound Ca²⁺ ions were replaced by these metal ions and measured by inductively coupled plasma mass spectrometry from the culture filtrate. Cell shape changed only after the addition of sub-inhibitory concentrations of the metals, but in growth inhibitory concentrations it was similar to the normal cells.     

Length of burning rotation and community composition in high-level Calluna-Eriophorum bog in N England

Studies on a long-term experiment on Calluna-Eriophorum bog indicate that the length of time between management fires affects the post-fire development. A short rotation (every 10 years) results in increased dominance by Eriophorum spp. while a long rotation (every 20 years) results in greater abundance of Calluna vulgaris after fire. Variations in post-fires species abundances are related to pre-fire stand composition coupled with the patchy distribution of regeneration centers of rhizomatous species. The usefulness of burning this vegetation type in management for herbivore grazing is questioned.Nomenclature follows Tutin et al. (1964 80) for vascular plants. Smith (1978) for mosses, Watson (1981) for liverworts and Hawksworth, James & Coppins (1980) for lichens. Following common practice. Calluna vulgaris is referred to simply as Calluna. Acknowledgements. I thank M. Raves and the Nature Conservancy Council for permission to work on Moor House National Nature Reserve and for the use of data from Reserve Records, V. J. Hobbs for field assistance, and Prof. C. H. Gimingham for help and advice during the study. I also thank all who worked at Moor House for creating a pleasant working environment. The long-term experiment was planned by Dr R. J. Elliot and analyzed initially by Dr G. I. Forrest.     

Genetic differentiation of Arthrobacter population from heavy metal-contaminated environment

Six samples containing extremely high concentration of Pb, Zn, and Cd were obtained from the layers of 5–10 cm and 25–30 cm three tailing piles, with ages of about 10, 20 and more than 80 years, respectively. Then, 48 bacterial strains were obtained from these samples, and subsequently their phylogenetic positions were determined by analysis on the partial sequence of 16S rRNA gene (fragment length ranging from 474 to 708 bp). These isolates were members of the Arthrobacter genus, phylogenetically close to A. keyseri and A. ureafaciens, with sequence ranging from 99.1% to 100%. Furthermore, genetic variation between subpopulations from different samples was revealed by analysis on their randomly amplified polymorphic DNA profile. Nei genetic distance showed that the greatest differentiation occurred between subpopulation A and C. Notably, either genetic distance between subpopulations from the layers of 5–10 cm and 25–30 cm of each tailing pile or between same layers of different tailing pile increased with the history of tailings. Moreover, correlation analysis showed that soluble Pb has a significantly negative relationship with Nei’ gene diversity of subpopulation. It was assumed that soluble Pb may be responsible for the reduced genetic diversity of the Arthrobacter population. Our data provided evidence that genetic differentiation of microbial populations was consistent with the changes of environmental factors, particularly heavy metals. Translated from Acta Ecologica Sinica, 2005, 25(10): 2569–2573 [译自: 生态学报]     

Effects of heavy metals on seed germination and early seedling growth of <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Seed is a developmental stage that is highly protective against external stresses in the plant life cycle. In this study, we analyzed toxicity of essential (Cu²⁺ and Zn²⁺) and non-essential heavy metals (Hg²⁺, Pb²⁺ and Cd²⁺) on seed germination and seedling growth in the model species Arabidopsis. Our results show that seedling growth is more sensitive to heavy metals (Hg²⁺, Pb²⁺, Cu²⁺ and Zn²⁺) in comparison to seed germination, while Cd²⁺ is the exception that inhibited both of these processes at similar concentrations. To examine if toxicity of heavy metals is altered developmentally during germination, we incubated seeds with Hg²⁺ or Cd²⁺ only for a restricted period during germination. Hg²⁺ displayed relatively strong toxicity at period II (12–24 h after imbibition), while Cd²⁺ was more effective to inhibit germination at period I (0–12 h after imbibition) rather than at period II. The observed differences are likely to be due in part to selective uptake of different ions by the intact seed, because isolated embryos (without seed coat and endosperm) are more sensitive to both Hg²⁺ and Cd²⁺ at period I. We assessed interactive toxicity between heavy metals and non-toxic cations, and found that Ca²⁺ was able to partially restore the inhibition of seedling growth by Pb²⁺ and Zn²⁺.     

Saccharomyces cerevisiae and Neurospora crassa contain heavy metal sequestering phytochelatin

In fungi, cellular resistance to heavy metal cytotoxicity is mediated either by binding of metal ions to proteins of the metallothionein type or by chelation to phytochelatin-peptides of the general formula (-Glu-Cys)_n-Gly. Hitherto, only one fungus, Candida glabrata has been shown to contain both metal inactivating systems. Here we show by unambiguous FAB-MS analysis that both a metallothionein-free mutant of Saccharomyces cerevisiae as well as a wildtype strain synthesize phytochelatin (PC₂) upon exposure to 250 M Cd²⁺ ions. The presence of Zn and/or Cu ions in the nutrient broth also induces PC₂ synthesis in this organism. By ¹⁰⁹Cd exchange and subsequent monobromobimane fluorescence HPLC, it could be shown that the presence of Cd²⁺ in the growth medium also induces phytochelatin synthesis in Neurospora crassa, which contains metallothioneins.     

Biosorption and bioaccumulation of lead by Penicillium sp. Psf-2 isolated from the deep sea sediment of the Pacific Ocean

A lead resistant fungus was isolated from the Pacific sediment. It was associated with Penicillium according to its partial sequences of 18S and ITS. The fungus could grow in the presence of 24 mM Pb(NO₃)₂ in a liquid medium, and no growth inhibition was observed at 4 mM and below. When growing in the presence of 4 mM Pb(NO₃)₂, the fungus accumulated a large amount of lead granules in the cell, as well as adsorbed on the outer layer of cell wall, as observed under a transmission electron microscope. The intracellular lead deposited either in the vicinity of the cytoplasm membrane or in the vacuoles, and also could aggregate into large particles in the cytoplasm. However, lead was not adsorbed on the thick inner wall of the fungus. Energy dispersive X-ray spectroscopy analysis showed that these granules or particles mainly consisted of lead, and other elements could hardly be detected. Selected area electron diffraction analysis showed that there were regular crystalline lattices in the lead precipitates, indicating that they were actually in the form of crystals to some extent. Therefore, both intracellular bioaccumulation and extracellular biosorption had contributed to the high resistance of this fungus to lead. These results suggest that this fungus can be used in biotreatment as a lead trapper.     

The composition of pyrophosphate heavy metal detoxification granules in barnacles

A major detoxification mechanism involving the binding of excess metals to inorganic, insoluble metalliferous granules occurs in many invertebrates. X-ray microanalysis of granules extracted from the barnacles Elminius modestus Darwin and Semibalanus balanoides (L.) (Crustacea: Cirripedia) indicated that one granule type was predominant. Examples of this granule type usually contained Fe and P, or Zn and S, or a combination of all four elements as major constituents. The predominant form of P in the granules was pyrophosphate, P₂O⁴⁻₇, with oprthophosphate and other polyphosphates present in lower amounts. The S was present as organic S. Biochemical analysis showed high levels of Ca, Fe, Mg, K and Zn in the granules with Cu and Ag present in smaller quantities. Considerable variation was shown between granules.     

Heavy metal stress reduces the deposition of calcium oxalate crystals in leaves of Phaseolus vulgaris

Calcium oxalate (CaOx) crystals in plants may serve as a sink for the absorption of excess calcium, and they could play an important role in heavy metal detoxification. In this study, the effect of heavy metals and different calcium concentrations on the growth of calcium oxalate crystals in leaves of Phaseolus vulgaris was investigated. Different analytical techniques were used to determine the influence of exogenous lead and zinc on CaOx deposition and to detect a presence of these metals in CaOx crystals. We found a positive correlation between the calcium concentration in the nutrient medium and the production of calcium oxalate crystals in leaves of hydroponically grown plants. On the other hand, addition of the heavy metals to the nutrient medium decreased the number of crystals. Energy dispersive X-ray spectrometry did not detect the inclusion of heavy metals inside the CaOx crystals. Our investigation suggests that CaOx crystals do not play a major role in heavy metal detoxification in P. vulgaris but do play an important role in bulk calcium regulation.     

A taxometric analysis ofLimonium (Plumbaginaceae) in Western Europe

A taxometric analysis ofLimonium species in Western Europe has provided evidence for the subgeneric groupings proposed by other authors. A cluster analysis suggests that the origin of sexual and agamospermous species has taken place separately. Sexual and agamospermous species are usually clustered separately. The lack of a sexual process in many agamospermous species, as indicated by the distribution of incompatibility morphs and by very low pollen fertility, suggests that the evolution of sexual and agamospermous taxa has been by different processes. It is suggested that following the origin of agamospermous species or species groups by hybridisation, subsequent evolution in the agamospermous taxa has occurred by asexual means. Evidence from the distribution of pollen/stigma incompatibility morphs, especially the existence of self-compatible agamospermous species supports this hypothesis.     

Ecosystem respiration depends strongly on photosynthesis in a temperate heath

We measured net ecosystem CO₂ flux (F _n) and ecosystem respiration (R _E), and estimated gross ecosystem photosynthesis (P _g) by difference, for two years in a temperate heath ecosystem using a chamber method. The exchange rates of carbon were high and of similar magnitude as for productive forest ecosystems with a net ecosystem carbon gain during the second year of 293 ± 11 g C m⁻² year⁻¹ showing that the carbon sink strength of heather-dominated ecosystems may be considerable when C. vulgaris is in the building phase of its life cycle. The estimated gross ecosystem photosynthesis and ecosystem respiration from October to March was 22% and 30% of annual flux, respectively, suggesting that both cold-season carbon gain and loss were important in the annual carbon cycle of the ecosystem. Model fit of R _E of a classic, first-order exponential equation related to temperature (second year; R ² = 0.65) was improved when the P _g rate was incorporated into the model (second year; R ² = 0.79), suggesting that daytime R _E increased with increasing photosynthesis. Furthermore, the temperature sensitivity of R _E decreased from apparent Q ₁₀ values of 3.3 to 3.9 by the classic equation to a more realistic Q ₁₀ of 2.5 by the modified model. The model introduces R _photo, which describes the part of respiration being tightly coupled to the photosynthetic rate. It makes up 5% of the assimilated carbon dioxide flux at 0°C and 35% at 20°C implying a high sensitivity of respiration to photosynthesis during summer. The simple model provides an easily applied, non-intrusive tool for investigating seasonal trends in the relationship between ecosystem carbon sequestration and respiration.