The influence of mycorrhizal symbiosis and fertilizer amendments on establishment of vegetation in heavy metal mine spoil

Biomass production of Andropogon gerardii and Festuca arundinacea was assessed in mine tailings (chat), a material containing high levels of zinc. The effects of organic and inorganic fertilizer amendments, the addition of an expanded clay material, and mycorrhizal fungi on the revegetation of chat were assessed. Plant growth in chat was best with mycorrhizal inoculation combined with nitrogen (either organic or inorganic) and phosphorus fertilization. Plant growth was also achieved if the chat material was amended with expanded clay and N and P fertilizer. However, the biomass produced in contaminated soil did not equal that of similarly fertilized uncontaminated soil. Regression analysis and canonical discriminate analysis revealed significant differences in the responses of the two plant species to the various chat amendments. Although F. arundinacea generally produced greater biomass than A. gerardii, it tended to accumulate more zinc in the shoots than A. gerardii. Therefore, if plant cover is to be used as forage or if wind-blown plant dry matter is of concern, A. gerardii may be more appropriate than F. arundinacea for revegetation of these toxic sites. Alternatively, if maximum plant cover is of primary importance to reduce wind or water erosion from contaminated soils, F. arundinacea may be the species of choice.     

Effects of mycorrhizae and fertilizer amendments on zinc tolerance of plants

In soils containing elevated levels of zinc, plant growth may be impaired because of Zn interference with P uptake by plants and because of detrimental effects of Zn toxicity itself. Because mycorrhizal fungi are known to improve uptake of plant P, the beneficial effects of mycorrhizal symbiosis on Zn tolerance of Andropogon gerardii Vitm. were assessed in soil amended with various levels of Zn and P. In the absence of P amendment, mycorrhizal fungi stimulated plant growth, but the degree of benefit depended on the inoculum source and the soil Zn level. Mycorrhizal fungi from a Zn contaminated site were more effective in increasing plant biomass at higher levels of Zn in the soil, whereas plant growth at lower levels of soil Zn was greater with mycorrhizal fungi from a non-contaminated site. Mycorrhizal fungus inoculation had no effect on shoot Zn concentration; however, inoculation significantly improved the plant P nutrition and therefore resulted in a high shoot P/Zn concentration ratio at all the soil Zn levels. To a certain extent, addition of P to the soil alleviated the Zn toxicity that had inhibited plant growth, but plant biomass tended to decrease with increasing soil Zn levels. Although P amendment improved P uptake, it also resulted in increased shoot Zn uptake.     

Interactions between arbuscular mycorrhizae and heavy metals under sand culture experiment

A sand culture experiment was established to determine interactions between arbuscular mycorrhizae and heavy metals. Mycorrhizal infection rates, spore densities, maize root and shoot weights, and heavy metal contents in maize were as indexes of responses of arbuscular mycorrhizal fungi (Acaulospora laevis, Glomus caledonium and Glomus manihotis) to heavy metals (Cu and Cd). The mycorrhizal infection rates of G. caledonium were the highest among these three mycorrhizal fungi, but the sporulating ability of G. caledonium was the poorest in the heavy metal treatments. The shoot and root weights of non-mycorrhizal plants were usually greater than those of mycorrhizal plants when the Cu concentrations in solutions are less than 3 mg l(-1) or Cd concentrations less than 1 mg l(-1). When Cd concentrations were 0.5 and 1 mg(-1), the root and shoot weights of plants inoculated with A. laevis were significantly (p < 0.05) lower than those of other treatments. Copper concentrations in shoots of mycorrhizal plants were higher than those of non-mycorrhizal ones at all Cu concentrations in solution, especially at low Cu concentrations. As to A. laevis, Cu concentrations in roots and shoots of the host were higher than those of non-mycorrhizal plants in these treatments. Thus A. laevis was sensitive to Cu and Cd, especially Cd, and G. caledonium was more tolerant to these two heavy metals. It is suggested that G. caledonium might be a promising mycorrhizal fungus for bioremediation of heavy metal contaminated soil.     

The arbuscular mycorrhizal fungus Glomus mosseae induces growth and metal accumulation changes in Cannabis sativa L

The effect of arbuscular mycorrhiza on heavy metal uptake and translocation was investigated in Cannabis sativa. Hemp was grown in the presence and absence of 100 microg g-1 Cd and Ni and 300 microg g-1 Cr(VI), and inoculated or not with the arbuscular mycorrhizal fungus Glomus mosseae. In our experimental condition, hemp growth was reduced in inoculated plants and the reduction was related to the degree of mycorrhization. The percentage of mycorrhizal colonisation was 42% and 9% in plants grown in non-contaminated and contaminated soil, suggesting a significant negative effect of high metal concentrations on plant infection by G. mosseae. Soil pH, metal bioavailability and plant metal uptake were not influenced by mycorrhization. The organ metal concentrations were not statistically different between inoculated and non-inoculated plants, apart from Ni which concentration was significantly higher in stem and leaf of inoculated plants grown in contaminated soil. The distribution of absorbed metals inside plant was related to the soil heavy metal concentrations: in plant grown in non-contaminated soil the greater part of absorbed Cr and Ni was found in shoots and no significant difference was determined between inoculated and non-inoculated plants. On the contrary, plants grown in artificially contaminated soil accumulated most metal in root organ. In this soil, mycorrhization significantly enhanced the translocation of all the three metals from root to shoot. The possibility to increase metal accumulation in shoot is very interesting for phytoextraction purpose, since most high producing biomass plants, such as non-mycorrhized hemp, retain most heavy metals in roots, limiting their application.     

Survival strategies of plants associated with arbuscular mycorrhizal fungi on toxic mine tailings

A field survey of metal concentrations and arbuscular mycorrhizal (AM) components of plants growing on five mining sites was conducted in Chenzhou City, Hunan Province, Southern China and a control site in Hong Kong. Significant differences were observed in the average concentrations of total heavy metals (Pb, Zn, Cu, Cd) and one metalloid (As) in contaminated soils compared with the control site. Gramineae and Compositae were the dominant plant families growing on mine tailings, with Chrysanthemum moritolium (common chrysanthemum), Cynodon dactylon (Bermuda grass), Miscanthus florodulus (Sword grass) and Pteris vittata (Ladder brake fern) commonly found at all sites. AM fungal colonization was detected in most of the plants. Comparing the four common plant species, three components of mycorrhizal colonization (arbuscules, vesicles and coiled hyphae) were found in the roots of C. dactylon and P. vittata growing at Do Shun Long (DSL) mine site. Concentrations of As in fronds were 24-fold higher than in roots of P. vittata with the highest mycorrhizal colonization rate (73%) among all sampling sites. Extensive mycorrhizal colonization (85%) was also recorded in the roots of C. dactylon with As accumulation 57 times higher than in shoots. The four common plants found in metal contaminated sites had developed different strategies for survival in the contaminated sites with the aid of indigenous AM fungi.     

The effects of multiple metal contamination on ectomycorrhizal Scots pine (Pinus sylvestris) seedlings

Experiments were conducted to investigate the effects of single and multiple metal contamination (Cd, Pb, Zn, Sb, Cu) on Scots pine seedlings colonised by ectomycorrhizal (ECM) fungi from natural soil inoculum. Seedlings were grown in either contaminated field soil from the site of a chemical accident, soils amended with five metals contaminating the site, or in soil from an uncontaminated control site. Although contaminated and metal-amended soil significantly inhibited root and shoot growth of the Scots pine seedlings, total root tip density was not affected. Of the five metals tested in amended soils, Cd was the most toxic to ECM Scots pine. Field-contaminated soil had a toxic effect on ECM fungi associated with Scots pine seedlings and caused shifts in ECM species composition on ECM seedlings. When compared to soils amended with only one metal, soils amended with a combination of all five metals tested had lower relative toxicity and less accumulation of Pb, Zn and Sb into seedlings. This would indicate that the toxicity of multiple metal contamination cannot be predicted from the individual toxicity of the metals investigated.     

重金属污染土壤接种丛枝菌根真菌对蚕豆毒性的影响

采用盆栽实验的方法,研究了重金属(包括Cu、Zn、Pb和Cd)复合污染和接种丛枝菌根真菌(arbuscular mycorrhizal fungi,AMF)Glomus mosseae对蚕豆(Vicia faba)生长及DNA损伤的影响.结果表明,虽然接种菌根真菌对蚕豆生物量的影响并不显著,但是却显著影响植物对重金属的吸收,接种菌根真菌对蚕豆吸收4种重金属元素的作用有差异.采用单细胞凝胶电泳(single cell gel electrophoresis,SCGE)法研究接种菌根真菌对蚕豆叶片的DNA损伤的影响,与重金属吸收的结果相吻合.结果表明,接种处理可显著增加蚕豆叶片的DNA损伤程度,这与接种处理可提高植物的重金属吸收相一致.     

重金属污染土壤的植物修复技术与其他技术联用的进展与前景

随着工业和经济的发展,土壤重金属污染越来越严重,因此重金属污染土壤的修复迫在眉睫。植物修复技术因其经济、环保等优点已经成为国内外研究的热点,但存在修复植物稀少的问题。将其他技术与植物修复技术联用可能是突破植物修复这一局限性的关键举措之一。介绍了基因工程技术与植物修复技术联用的研究进展;总结了微生物技术与植物修复技术联用中植物根际促生细菌、菌根真菌的作用;概述了材料化学技术联合植物修复的应用现状,着重介绍了土壤改良剂和螯合剂的实际应用。最后对基因工程、微生物、材料化学等技术联合植物修复技术在研究和应用中的未来方向进行了前景展望,旨在为植物修复技术发展提供更多新的思路和参考。     

Metallophyte status of violets of the section Melanium

Violets from metal-enriched soils have controversially been described as both heavy-metal accumulators and excluders in the literature. The present study solves the issue for violets of the section Melanium (zinc violets, Viola lutea ssp. calaminaria and V. lutea ssp. westfalica; hartsease or wild pansy, Viola tricolor; and mountain pansy, V. lutea). The aims were to determine the concentrations of heavy metals in the soil and in the roots and shoots of field-collected plants, to evaluate the potential impact of colonisation by arbuscular mycorrhizal fungi on heavy-metal concentrations in the plant tissues, and to quantitatively define the localisation of the elements in root cross-sections. When these violets grow in low-metal soils, higher concentrations of the heavy metals were found in the roots and shoots than in the soil, whereas the opposite was seen in samples from high-metal soils. Under all field conditions examined, the roots of all of these species were colonised by arbuscular mycorrhizal fungi. However, V. tricolor was marginally colonised when the concentrations of Zn and P were higher in the soil. Determination of the spatial distribution of the elements in root cross-sections of these violets indicates tissue-specific deposition of elements within the vascular tissue, the cortex, and the rhizodermis. These data indicate that violets of the section Melanium are heavy-metal excluders.     

Metal tolerating methylotrophic bacteria reduces nickel and cadmium toxicity and promotes plant growth of tomato (Lycopersicon esculentum L)

Inoculation of plants with microorganisms may reduce the toxicity of heavy metals to plants in contaminated soils. In this study, we have shown that the plant growth promoting bacteria Methylobacterium oryzae strain CBMB20 and Burkholderia sp. strain CBMB40 from rice reduce the toxicity of Ni and Cd in tomato and promote plant growth under gnotobiotic and pot culture experiments. The bacterial strains bound considerable amounts of Ni(II) and Cd(II) in their growing and resting cells and showed growth in the presence of NiCl2 and CdCl2. In gnotobiotic assay, inoculation with the bacterial strains reduced the ethylene emission and increased the tolerance index of the seedlings against different concentrations of NiCl2/CdCl2. In pot experiments carried out with non-polluted, Ni and Cd supplemented Wonjo-Mix bed soil, the results clearly demonstrated reduction in the accumulations of Ni(II) and Cd(II) in roots and shoots, with significant increase in the plant growth attributes with bacterial inoculations compared to untreated control. Strain CBMB20 performed better than CBMB40 in reducing the heavy metal accumulations in plants. Our results suggest conclusively, that protection against the heavy metals toxicity is rendered by these bacterial strains by reducing their uptake and further translocation to shoots in plants and promote the plant growth by other PGP characteristics.     

Microbial cellulose decomposition in soils from a rifle range contaminated with heavy metals

The objective of this study was to assess the effects of heavy metals on microbial decomposition of cellulose in heavy metal-contaminated soils using a cotton strip assay. The assay is a measure of the potential of soil microorganisms to decompose the plant polymer, cellulose. Cellulolytic activity in soil was assessed by determining the reduction in tensile strength of the buried cotton strips over a 25- and 45-day period. Soils were obtained from a rifle range that contain high levels of lead, copper and zinc. The site has been used for approximately 50 years, resulting in metal levels of up to 30,000 mg/kg of lead, 4000 mg/kg of copper and 600 mg/kg of zinc in the most contaminated soils. All the metal-contaminated soils had lower degradation rates than the uncontaminated soils tested. Among the contaminated soils, however, the heavy metal concentration was not the major factor in determining the loss in tensile strength of the cotton strips, where cellulose decomposition was governed by other soil physicochemical properties. Soil with a higher cation exchange capacity, readily oxidisable material and volatile solids content had the greatest loss in tensile strength of cotton strips. Microbial adaptation to the presence of high concentrations of soil heavy metals and reduced bioavailability of metals is the likely explanation for this phenomenon.     

Growth response and phytoextraction of copper at different levels in soils by Elsholtzia splendens

Phytoremediation is a promising approach for cleaning up soils contaminated with heavy metals. Information is needed to understand growth response and uptake mechanisms of heavy metals by some plant species with exceptional capability in absorbing and superaccumulating metals from soils. Greenhouse study, field trial, and old mined area survey were conducted to evaluate growth response and Cu phytoextraction of Elsholtzia splendens in contaminated soils, which has been recently identified to be tolerant to high Cu concentration and have great potential in remediating contaminated soils. The results from this study indicate that the plant exhibited high tolerance to Cu toxicity in the soils, and normal growth was attained up to 80 mg kg(-1) available soil Cu (the NH4OAc extractable Cu) or 1000 mg kg(-1) total Cu. Under the field conditions, a biomass yield of 9 ton ha(-1) was recorded at the soil available Cu level of 77 mg kg(-1), as estimated by the NH4OAc extraction method. Concentration-dependent uptake of Cu by the plant occurred mainly at the early growth stage, and at the late stage, there is no difference in shoot Cu concentrations grown at different extractable soil Cu levels. The extractability of Cu from the highly polluted soil is much greater by the roots than that by the shoots. The NH4OAc extractable Cu level in the polluted soil was reduced from 78 to 55 mg kg(-1) in the soil after phytoextraction and removal of Cu by the plant species for one growth season. The depletion of extractable Cu level in the rhizosphere was noted grown in the mined area, even at high Cu levels, the NH4OAc extractable Cu in the rhizosphere was 30% lower than that in the bulk soil. These results indicate that phytoextraction of E. splendens can effectively reduce the plant-available Cu level in the polluted soils.     

Potential contribution of arbuscular mycorrhiza to cadmium immobilisation in soil

The contribution of arbuscular mycorrhiza (AM) to immobilisation of Cd in substrate was studied in two experiments. In the first experiment, substrates prepared by cultivating tobacco, either non-mycorrhizal or inoculated with the AM fungus Glomus intraradices were enriched with a range of Cd concentrations, and Cd toxicity in the substrates was assessed using root growth tests with lettuce as a test plant. The tests revealed lower Cd toxicity in the mycorrhizal than in the non-mycorrhizal substrate, and the difference increased with increasing total Cd concentration in the substrates. In the second experiment, extraradical mycelium (ERM) of G. intraradices exposed in vivo to Cd was collected and analysed on Cd concentration. The ERM accumulated 10–20 times more Cd per unit of biomass than tobacco roots. While Cd concentrations were lower in the biomass of mycorrhizal plants than of non-mycorrhizal plants, Cd immobilisation by ERM did not affect the total Cd content in mycorrhizal tobacco.

It is concluded that mycorrhiza may decrease Cd toxicity to plants by enhancing Cd immobilisation in soil. The results therefore suggest a potential role of AM symbiosis in the phytostabilisation of contaminated soils, where high Cd availability inhibits plant growth.     

Plant accumulation of potentially toxic elements in sewage sludge as affected by soil organic matter level and mycorrhizal fungi.

Leek (Allium ameloprasum) was grown in pot trials in two clay loams of contrasting organic contents, with and without indigenous mycorrhizal propagules. Sewage sludges containing varying levels of Cd, Cu and Zn were added. Extractable soil metals, plant growth, major nutrient content and accumulation of metals, and soil microbial indices were investigated. The aim was to establish whether soil organic content and mycorrhizal status affected plant and microbial exposure to these metals. Extractable metals were higher and responses to inputs more pronounced in the arable, lower organic matter soil, although only Cd showed a soil difference in the CaCl2 fraction. There were no metal toxic effects on plants and some evidence to suggest that they promoted growth. Uptake of each metal was higher in the larger plants of the grassland, higher organic matter soil. Inoculation with arbuscular mycorrhizal fungi increased root Cd and Zn concentrations. With the exception of Cd (roots) and Zn (shoots), higher inputs of sludge metals did not increase plant metals. Zn and Cu, but not Cd, concentrations were higher in roots than in shoots.     

Growth responses of crop and weed species to heavy metals in pot and field experiments

Greenhouse and field studies were performed to examine the growth responses and possible phytoremediation capacity towards heavy metals of several Brassicaceae (Brassica alba, Brassica carinata, Brassica napus and Brassica nigra) and Poaceae (durum wheat and barley). Soils used featured total concentrations of Cr, Cu, Pb and Zn largely exceeding the maximum levels permitted by the Italian laws. Different organic amendments were tested such as a compost and the plant growth-promoting rhizobacterium Bacillus licheniformis. In the greenhouse experiment, plant length, leaf area index and shoots dry matter were evaluated periodically for the Brassicaceae examined. Whereas plant length, grains production, weight of 1,000 seeds, ear fertility and tiller density were determined under field conditions at the end of the crop cycle for wheat and barley. In general, the species tested appeared to be tolerant to high heavy metal concentrations in soil, and slightly significant differences were found for all parameters considered. A marked growth increase was shown to occur for Brassicaceae cultivated on compost- and bacillus-amended contaminated soils, with respect to non-amended contaminated soils. With some exception, higher growth parameters were measured for wheat and barley plants cropped from contaminated soils in comparison to non-contaminated soils. Further, bacillus amendment enhanced the length of wheat and barley plants in both non-contaminated and contaminated soils, while different effects were observed for the other parameters evaluated.     

Bioavailability and plant accumulation of heavy metals and phosphorus in agricultural soils amended by long-term application of sewage sludge

Amendment of agricultural soils with municipal sewage sludges provides a valuable source of plant nutrients and organic matter. Nevertheless, addition of heavy metals and risks of eutrophication continue to be of concern. Metal behaviour in soils and plant uptake are dependent on the nature of the metal, sludge/soil physico-chemical properties and plant species. A pot experiment was carried out to evaluate plant production and heavy metal uptake, soil heavy metal pools and bioavailability, and soil P pools and possible leaching losses, in agricultural soils amended with sewage sludge for at least 10 years (F20) compared to non-amended soils (control). Sewage sludge application increased soil pH, N, Olsen-extractable-P, DOC and exchangeable Ca, Mg and K concentrations. Total and EDTA-extractable soil concentrations of Cu and Zn were also significantly greater in F20, and soil metal (Cu, Mn and Zn) and P fractionation altered. Compared to the control, in F20 relative amounts of acid-extractable (Mn, Zn), reducible (Mn, Zn) and oxidisable (Cu, Zn) metal fractions were greater, and a dominance of inorganic P forms was observed. Analyses of F20 soil solutions highlighted risks of PO4 and Cu leaching. However, despite the observed increases in metal bioavailability sewage sludge applications did not lead to an increase in plant shoot concentrations (in wild plants or crop species). On the contrary, depending on the plant species, Mn and Zn tissue concentrations were within the deficiency level for most plants.     

Arbuscular mycorrhiza enhanced arsenic resistance of both white clover (Trifolium repens Linn.) and ryegrass (Lolium perenne L.) plants in an arsenic-contaminated soil

In a compartmented cultivation system, white clover (Trifolium repens Linn.) and ryegrass (Lolium perenne L.), with their roots freely intermingled, or separated by 37 microm nylon mesh or plastic board, were grown together in an arsenic (As) contaminated soil. The influence of AM inoculation on plant growth, As uptake, phosphorus (P) nutrition, and plant competitions were investigated. Results showed that both plant species highly depended on mycorrhizas for surviving the As contamination. Mycorrhizal inoculation substantially improved plant P nutrition, and in contrast markedly decreased root to shoot As translocation and shoot As concentrations. It also showed that mycorrhizas affected the competition between the two co-existing plant species, preferentially benefiting the clover plants in term of nutrient acquisition and biomass production. Based on the present study, the role of AM fungi in plant adaptation to As contamination, and their potential use for ecological restoration of As contaminated soils are discussed.     

Influence of arbuscular mycorrhizal fungi on the growth and nutrient status of bermudagrass grown in alkaline bauxite processing residue

A nursery experiment was conducted to evaluate the potential role of arbuscular mycorrhizal (AM) fungi in encouraging the vegetation cover on bauxite residue (red mud) sites. An alkali tolerant bermudagrass (Cynodon dactylon) adapted to local conditions were grown in red mud with different amendments with and without AM fungi to assess mycorrhizal effects on plant growth, mineral nutrition, metal uptake and neutralization of bauxite residue. Inoculation of AM fungi significantly increased the plant growth, nutrient uptake and reduced Fe, Al accumulation in plant tissue and also improved the soil physico-chemical and biochemical properties. Gypsum and sludge amended treatments inoculated with AM fungi had maximum biomass, nutrient uptake and reduced accumulation of metals. The neutralization of red mud was significant in presence of AM fungi than control. The experiment provided evidence for the potential use of bermudagrass in combination with AM fungi for ecological restoration of bauxite residue sites.     

Improvement of soil characteristics and growth of Dorycnium pentaphyllum by amendment with agrowastes and inoculation with AM fungi and/or the yeast Yarowia lipolytica

The effectiveness of two microbiologically treated agrowastes [dry olive cake (DOC) and/or sugar beet (SB)] on plant growth, soil enzymatic activities and other soil characteristics was determined in a natural soil from a desertified area. Dorycnium pentaphyllum, a legume plant adapted to stress situations, was the test plant to evaluate the effect of inoculation of native arbuscular mycorrhizal (AM) fungi and/or Yarowia lipolytica (a dry soil adapted yeast) on amended and non-amended soils. Plant growth and nutrition, symbiotic developments and soil enzymatic activities were limited in non-amended soil where microbial inoculations did not improve plant development. The lack of nodules formation and AM colonization can explain the limited plant growth in this natural soil. The effectiveness and performance of inocula applied was only evident in amended soils. AM colonization and spores number in natural soil were increased by amendments and the inoculation with Y. lipolytica promoted this value. The effect of the inoculations on plant N-acquisition was only important in AM-inoculated plants growing in SB medium. Enzymatic activities as urease and protease activities were particularly increased in DOC amended soil meanwhile dehydrogenase activity was greatest in treatments inoculated with Y. lipolytica in SB added soil. The biological activities in rhizosphere of agrowaste amended soil, used as indices of changes in soil properties and fertility, were affected not only by the nature of amendments but also by the inoculant applied. All these results show that the lignocellulosic agrowastes treated with a selected microorganism and its further interaction with beneficial microbial groups (native AM fungi and/or Y. lipolytica) is a useful tool to modify soil physico-chemical, biological and fertility properties that enhance the plant performance probably by making nutrients more available to plants.     

Soil contamination by crude oil: impact on the mycorrhizosphere and on the revegetation potential of forest trees

In vitro and greenhouse biotests were carried out to study the effects of various concentrations of crude oil on the mycorrhizosphere and the ability of ectomycorrhizal fungi to colonise Norway spruce and poplar seedlings grown on contaminated soil. Ectomycorrhizal fungi grown in pure cultures showed a variety of reactions to crude oil, ranging from growth stimulation to total inhibition of growth, depending on the species of fungi. Germination of poplar and spruce seeds was not significantly affected. The growth of spruce seedlings was not affected by crude oil, whereas that of poplar seedlings was significantly reduced at high concentrations. None of the concentrations had any effect on the degree of ectomycorrhizal and endomycorrhizal colonisation of poplar. With spruce, however, the ectomycorrhizal fungi showed species-specific reactions to increasing concentrations, in accordance with the results of the pure culture test. The length of time between soil contamination and seeding affects both seedling growth and the mycorrhizal infection potential of the soil. The results confirm the importance of mycorrhizal fungi in the bioremediation of soils contaminated by crude oil.