Retromobilization of heavy metal resistance genes in unpolluted and heavy metal polluted soil

Abstract: Retromobilization of the nonconjugative (Tra⁻Mob⁺) IncQ vector, pMOL155, and the non-mobilizable (Tra⁻Mob⁻) vector, pMOL149, by means of the IncP plasmids RP4 and pULB113 (RP4::Mu3A), was studied in plate matings and in soil microcosms, and compared with direct and triparental mobilization. Both vectors harbour the czc genes, originating from Alcaligenes eutrophus , which code for resistance to Co, Zn, and Cd. The donor of the czc genes was Escherichia coli which did not express these genes. The recipient, Alcaligenes eutrophus , expressed the czc genes very well. Retromobilization, direct and triparental mobilization of pMOL155 was observed in sterile soil. Both the addition of nutrients and heavy metals significantly enhanced the number of (retro)transconjugants. Retromobilization was also detected in nutrient amended nonsterile soil, but the presence of the autochthonous soil biota strongly reduced the number of retrotransconjugants and also prevented their increase upon application of heavy metals to the soil. Retromobilization of the czc genes, cloned in pMOL149, by using pULB113 was also observed, yet only in sterile, nutrient amended, heavy metal polluted soil.     

Arousing sleeping genes: shifts in secondary metabolism of metal tolerant actinobacteria under conditions of heavy metal stress

Numerous microbial habitats are strongly influenced by elevated levels of heavy metals. This type of habitat has developed either due to ore mining and metal processing or by pedogenesis above metal-rich base rocks. Most actinobacteria are soil-borne microbes with a remarkable capability for the synthesis of a broad variety of biologically active secondary metabolites. One major obstacle in identifying secondary metabolites, however, is the known phenomenon of sleeping gene clusters which are present, but silent under standard screening conditions. Here, we proceed to show that sleeping gene clusters can be awakened by the induction in heavy metal stress. Both, a chemical and a biological screening with extracts of supernatant and biomass of 10 strains derived from metal contaminated and non-contaminated environments was carried out to assay the influence of heavy metals on secondary metabolite patterns of metal tolerant actinobacteria. Metabolite patterns of cultures grown in complex and minimal media were compared to nickel (or cadmium) spiked parallels. Extracts of some strains grown in the presence of a metal salt displayed intense antibiosis against Escherichia coli, Mycobacterium smegmatis, Staphylococcus aureus and Candida albicans. Contrarily to the widely held opinion of metals as hindrance in secondary metabolism, metals thus can induce or enhance synthesis of possibly potent and medically relevant metabolites in metal tolerant strains. Hence, re-screening of existing strain libraries as well as identification of new strains from contaminated areas are valid strategies for the detection of new antibiotics in the future.     

Local adaptation of microbial communities to heavy metal stress in polluted sediments of Lake Erie

Microbial communities must balance the assimilation of biologically necessary metals with resistance to toxic metal concentrations. To investigate the impact of heavy metal contaminants on microbial communities, we performed two experiments measuring extracellular enzyme activities (EEA) in polluted and unpolluted sediments of Lake Erie. In the first experiment, inoculations with moderate concentrations of copper and zinc appreciably diminished EEA from uncontaminated sites, whereas EEA from contaminated sediments increased or were only negligibly affected. In the second experiment, we compared the effects of three separate metals (i.e. copper, arsenic, and cadmium) on microbial community metabolism in polluted and unpolluted locations. Although copper and arsenic elicited differential effects by inhibiting EEA only in unpolluted sediments, cadmium inhibited EEA in both polluted and unpolluted sediments. Multivariate analyses of EEA from polluted sediments revealed direct associations among hydrolytic enzymes and inverse or absent associations between hydrolases and oxidases; these associations demonstrated resilience to heavy metal stress. In contrast, addition of heavy metals to unpolluted sediments appeared to have a higher impact on the multivariate pattern of EEA associations as revealed by an increase in the number of associations, more inverse relationships, and potential enzymatic trade-offs. The results of this study suggest community-level adaptations through the development of resistance mechanisms to the types and local levels of heavy metals in the environment.     

Species associations and metal contents of algae from rivers polluted by heavy metals

SUMMARY. The algal flora of the Rivers Hayle and Gannel. whieh drain copper and lead mining regions of Cornwall, are described and compared. Although fluctuations in the rarer members of the algal communities were observed, the dominant filamentous algae at most sites did not change over the year of study. Both the total algal abundance and the number of species were depressed at high metal sites. Associations of species that were evident in field samples were confirmed and correlated with water metal levels by principal component analysis. The close similarity between the flora of similar sites on the copper-polluted River Hayle and the lead-polluted River Gannei implies that the degree of metal pollution, rather than the polluting metal per se, determines the species present. All mine sites were characterized by a Microspora -community whereas a Zygnemales community of Spirogyra and Mougeotia species was typical of low metal pollution. Moderately polluted sites downsteam of the mines had an intermediate flora of Zygnemaies, Microsporales. Ulotrichales and gelatinous Volvocales and Tetrasporales species. No species could be said to invariably indicate metal pollution; the most abundant species at highly contaminated sites were also those with the widest distributions. Field samples of filamentous algae (mainly Spirogyra, Zygogonium, Mougeotia and Microspora species) contained metal concentrations several orders of magnitude greater than ambient levels. For copper and iron, both algal metal contents and concentration ratios (μg g-1 algae/μg ml-1 water) were positive functions of water metal levels, although algal iron appeared to plateau at water concentrations of about 1 mg ml-1. Algal lead concentrations, but not the concentration ratios, also were positively correlated with water lead levels. Thus, for these three metals, the algal metal contents were indicative of ambient conditions. In contrast, algal zinc concentrations were nearly constant and consequently the concentration ratios for this metal were inversely related to water levels. This result suggests that unlike the uptake of other metals, uptake of zinc by these aigae may be strictly regulated.     

治理重金属污染河流底泥的生物淋滤技术

治理重金属污染河流底泥的生物技术是指利用生物体来消除或降低重金属毒性的方法,包括微生物修复、植物修复和植物.微生物联合修复,具有成本低、去除效率高、脱毒后污泥脱水性能好等优点,近年来在国际上备受关注.本文介绍了近10年来生物技术在治理重金属污染河流底泥方面的研究成果,着重介绍了日益受关注的生物淋滤技术,从微生物学、分子生物学和生物表面活性剂等方面的发展作了详细描述,展望了生物修复在河流底泥重金属污染应用中的前景.     

Genetic structure of Suillus luteus populations in heavy metal polluted and nonpolluted habitats

The genetic structure of populations of the ectomycorrhizal basidiomycete Suillus luteus in heavy metal polluted and nonpolluted areas was studied. Sporocarps were collected at nine different locations and genotyped at four microsatellite loci. Six of the sampling sites were severely contaminated with heavy metals and were dominated by heavy metal-tolerant individuals. Considerable genetic diversity was found within the geographical subpopulations, but no reduction of the genetic diversity, current or historic, was observed in subpopulations inhabiting polluted soils. The genetic differentiation between the geographical subpopulations was low, and no evidence for clustering of subpopulations from polluted soils vs. subpopulations from nonpolluted soils was found. These results indicate that heavy metal pollution has a limited effect on the genetic structure of S. luteus populations, and this may be due to the high frequency of sexual reproduction and extensive gene flow in S. luteus, which allows rapid evolution of the tolerance trait while maintaining high levels of genetic diversity.     

Dark septate endophyte (DSE) fungi isolated from metal polluted soils: Their taxonomic position, tolerance, and accumulation of heavy metals In Vitro

To understand the possible role of the plant root associated fungi on metal tolerance, their role in the uptake of heavy metals and the potential transfer of these metal ions to the plant, three strains of dark septate endophytic (DSE) fungi were isolated from a waste smelter site in southwest China, and one strain was isolated from a non-contaminated site. According to molecular phylogenetic analysis of the ITS 1-5.8S rDNA-ITS 2 gene regions and morphological characteristics, one is identified as Exophiala pisciphila, and the other three are non-sporulating fungi under the experiment condition with the nearest phylogenetic affinities to the Thysanorea papuana strain EU041814. Tolerance and accumulation abilities of the three DSE strains for metals were investigated in liquid culture. Minimum inhibitory concentrations (MIC) of Pb, Zn, and Cd were determined. It was demonstrated that the tolerance of the DSE strains varied between metal species and strains. The E. pisciphila strain is able to accumulate lead and cadmium over 20% and 5% of dry weight of biomass, respectively. Partial of the sequestrated metals can be washed with CaCh. Morphological and enzyme activity changes taking place in the presence of excessive Pb, Cd, and/or Zn also indicate that the mechanism of heavy metal tolerance and accumulation of the DSE strains would be a complex process. The findings indicated promising tolerance and accumulation of the DSE strains with potential values in metal cycling and restoration of soil and water system.     

Activity and heavy metal resistance of non-specific esterases in leaf beetle Chrysomela lapponica from polluted and unpolluted habitats

We compared the general activity and heavy metal resistance of non-specific esterases in two populations of the leaf beetle Chrysomela lapponica from habitats severely contaminated by heavy metals (mostly Ni and Cu) and two populations from unpolluted habitats. Concentrations of Ni and Cu in adult beetles from the most polluted site were 7.7 and 3.6 times higher that in beetles from unpolluted habitats. Larval esterases showed higher activity and lower susceptibility to heavy metals than esterases of adults. Larval esterase activity did not differ between populations from polluted and unpolluted sites, but adult beetles from polluted localities had lower esterase activity than beetles from unpolluted habitats. Both Cu and Ni sulfates in millimolar concentrations in vitro suppressed esterase activity of larvae from unpolluted habitats, but caused no negative effect on esterases of larvae from polluted sites. Similarly, inhibition of adult esterase activity by Ni was stronger in beetles from unpolluted localities than in beetles from polluted localities. This indicates that resistance of non-specific esterases to heavy metals is higher in leaf beetle populations from contaminated environment.     

AM真菌对重金属污染土壤生物修复的应用与机理

土壤重金属污染威胁人类健康和整个生态系统,而高效、低耗、安全的生物修复技术显示出了极大的应用潜力,特别是利用植物-微生物共生体增强生物修复效应的应用.丛枝菌根(Arbuscular Mycorrhizae,AM)真菌是一类广泛分布于土壤生态系统中的有益微生物,能与90％以上的陆生高等植物形成共生体.研究发现,AM真菌能够增强宿主植物对土壤中重金属胁迫的耐受性.当前,利用AM真菌开展重金属污染土壤的生物修复已经引起环境学家和生态学家的广泛关注.基于此,围绕AM真菌在重金属污染土壤生物修复作用中的最新研究进展,从物理性防御体系的形成、对植物生理代谢的调控、生化拮抗物质的产生、基因表达的调控等角度探究AM真菌在重金属污染土壤生物修复中的作用机理,以期为利用AM真菌开展重金属污染的生物修复提供理论依据,并对本领域未来的发展和应用前景进行了展望.     

南通沿海滩涂耐盐植物重金属抗性内生细菌的筛选及生物多样性

【目的】沿海滩涂耐盐植物重金属抗性内生细菌的筛选及其促生长潜在能力的研究有助于我们获得一些能够耐受并促进耐盐植物在被Cd2+、Pb2+、Hg2+、Cu2+,Zn2+等重金属离子污染的贫瘠的沿海滩涂上正常生长的菌株,达到既能够利用广袤的滩涂生物资源产生经济价值又能够净化生态环境的目的。【方法】以江苏南通沿海滩涂地区的4种耐盐植物为材料,采用稀释平板涂布法从中分离得到45株内生细菌,从中挑取23株代表性的菌株,对其进行抗重金属Cu2+、Pb2+、Cd2+、Zn2+,Hg2+的活性筛选;固氮、解磷、吲哚乙酸(IAA)的产生、1-氨基环丙烷-1-羧酸(ACC)脱氨酶活性等促生指标以及NaCl耐受能力的筛选。【结果】发现分离所得的大多数细菌能够耐受高浓度的Cu2+以及Pb2+,但是对Cd2+、Zn2+,Hg2+的耐受能力则较弱;26.1%的细菌具有固氮能力,21.7%的细菌具有解磷能力,60.9%的细菌能够产生IAA,39.1%的细菌含有ACC脱氨酶。对他们进行16S rRNA基因鉴定后发现,他们分属于芽胞杆菌属(Bacillus)、喜盐芽胞杆菌属(Halobacillus)、海洋芽胞杆菌属(Oceanobacillus)、微小杆菌属(Exiguobacterium)、沙雷氏菌属(Serratia)、短波单胞菌属(Brevundimonas)、弧菌属(Vibrio)、葡萄球菌属(Staphylococcus)共8个属,显示了丰富的多样性。其中菌株KLBMP 2432以及菌株KLBMP 2447为潜在的新种。【结论】沿海滩涂地区的耐盐植物内生细菌具有丰富多样的生物多样性以及促生长能力,且存在潜在的新种资源,并对重金属Cu2+,Pb2+具有较强的抗性。     

重金属胁迫下的微生物代谢组学研究进展

微生物的代谢活动易受到环境变化的影响,当环境中存在重金属污染时微生物会通过调节代谢降低自身所受的重金属的毒害。本文通过微生物代谢组学研究探讨重金属胁迫下微生物代谢活动的响应情况,介绍了微生物代谢组学的相关技术和方法,对其应用进行说明;基于重金属对微生物细胞的毒害作用,对重金属胁迫下微生物代谢组学的相关内容进行综述,发现在重金属胁迫下,微生物可以通过增加代谢活动进而产生更多的代谢物质来响应重金属的胁迫,其中微生物产生的胞外聚合物、草酸和柠檬酸等代谢物在微生物响应重金属胁迫中具有重要作用。微生物通过产生相应代谢物不仅使自身可以在重金属胁迫下生存,这些代谢物还可以使环境中重金属有所减少,这对于利用微生物资源修复重金属污染具有重要意义。     

Antioxidant enzyme responses of plants to heavy metal stress

Heavy metal pollutions caused by natural processes or anthropological activities such as metal industries, mining, mineral fertilizers, pesticides and others pose serious environmental problems in present days. Evidently there is an urgent need of efficient remediation techniques that can tackle problems of such extent, especially in polluted soil and water resources. Phytoremediation is one such approach that devices effective and affordable ways of engaging suitable plants to cleanse the nature. Excessive accumulation of metal in plant tissues are known to cause oxidative stress. These, in turn differentially affect other plant processes that lead to loss of cellular homeostasis resulting in adverse affects on their growth and development apart from others. Plants have limited mechanisms of stress avoidance and require flexible means of adaptation to changing. A common feature to combat stress factors is synchronized function of antioxidant enzymes that helps alleviating cellular damage by limiting reactive oxygen species (ROS). Although, ROS are inevitable byproducts from essential aerobic metabolisms, these are needed under sub-lethal levels for normal plant growth. Understanding the interplay between oxidative stress in plants and role of antioxidant enzymes can result in developing plants that can overcome oxidative stress with the expression of antioxidant enzymes. These mechanisms have been proving to have immense potential for remediating these metals through the process of phytoremediation. The aim of this review is to assemble our current understandings of role of antioxidant enzymes of plants subjected to heavy metal stress.     

Phytoremediation of heavy metal contaminated soil potential by woody plants on Tonglushan ancient copper spoil heap in China

Fast-growing metal-accumulating woody plants are considered potential candidates for phytoremediation of metals. Tonglushan mining, one of the biggest Cu production bases in China, presents an important source of the pollution of environment. The sample was collected at Tonglushan ancient copper spoil heap. The aims were to measure the content of heavy metal in the soil and woody plants and to elucidate the phytoremediation potential of the plants. The result showed that soil Cu, Cd and Pb were the main contamination, the mean contents of which were 3166.73 mg/kg, 3.66 mg/kg and 137.06 mg/kg respectively, which belonged to severe contamination. Fourteen species from 14 genera of 13 families were collected and investigated; except for Ligutrum lucidum, the other 13 woody plants species were newly recorded in this area. In addition, to assess the ability of metal accumulation of these trees, we proposed accumulation index. Data suggested that Platanus × acerilolia, Broussonetia papyrifera, Ligutrum lucidum, Viburnum awabuki, Firmiana simplex, Robina pseudoacacia, Melia azedarach and Osmanthus fragrans exhibited high accumulated capacity and strong tolerance to heavy metals. Therefore, Platanus × acerilolia and Broussonetia papyrifera can be planted in Pb contaminated areas; Viburnum awabuki, Firmiana simplex, Robina pseudoacacia and Melia azedarach are the suitable trees for Cd contaminated areas; Viburnum awabuki, Melia azedarach, Ligutrum lucidum, Firmiana simplex, Osmanthus fragrans and Robina pseudoacacia are appropriate to Cu, Pb and Cd multi-metal contaminated areas.     

Two naphthalene degrading bacteria belonging to the genera Paenibacillus and Pseudomonas isolated from a highly polluted lagoon perform different sensitivities to the organic and heavy metal contaminants

Two bacterial strains were isolated in the presence of naphthalene as the sole carbon and energy source from sediments of the Orbetello Lagoon, Italy, which is highly contaminated with both organic compounds and metals. 16S rRNA gene sequence analysis of the two isolates assigned the strains to the genera Paenibacillus and Pseudomonas. The effect of different contaminants on the growth behaviors of the two strains was investigated. Pseudomonas sp. ORNaP2 showed a higher tolerance to benzene, toluene, and ethylbenzene than Paenibacillus sp. ORNaP1. In addition, the toxicity of heavy metals potentially present as co-pollutants in the investigated site was tested. Here, strain Paenibacillus sp. ORNaP1 showed a higher tolerance towards arsenic, cadmium, and lead, whereas it was far more sensitive towards mercury than strain Pseudomonas sp. ORNaP2. These differences between the Gram-negative Pseudomonas and the Gram-positive Paenibacillus strain can be explained by different general adaptive response systems present in the two bacteria.     

Metal resistances of Chlorophyta from rivers polluted by heavy metals

SUMMARY. Two-hundred isolates, comprising 87 species of Chlorophyta, were obtained from sites along the Rivers Hayle and Gannel. which drain the ancient mining region of Cornwall. All isolates were tested for sensitivity to copper, lead, zinc and cadmium. In general, isolates were resistant to the metals normally present in their habitats. However, the distribution of metal sensitivities of the algae from a given site was broad; the effect of metal pollution was to shift the median response of a population toward higher metal resistance. Resistant algae of two general classes were identified: some normally sensitive species were metal-tolerant, presumably through genetic adaptation; other species were metal-resistant even when isolated from a non-polluted habitat. Many isolates of both types displayed multiple-resistances or co-tolerances. For example, copper tolerant isolates from high copper sites tended to be also lead resistant; however, algae from high lead sites were usually very copper sensitive. Zinc and cadmium resistances also were correlated among isolates from both zinc-polluted and non-polluted sites. General metal-insensitivity seemed to be common, particularly among gelatinous Chlamydomonas and Gloeococcus species. Thus, several evolutionary strategies appear to coexist among algae from metal polluted environments.