Structure and function of the soil microbial community in microhabitats of a heavy metal polluted soil

 Particle-size fractionation of a heavy metal polluted soil was performed to study the influence of environmental pollution on microbial community structure, microbial biomass, microbial residues and enzyme activities in microhabitats of a Calcaric Phaeocem. In 1987, the soil was experimentally contaminated with four heavy metal loads: (1) uncontaminated controls; (2) light (300 ppm Zn, 100 ppm Cu, 50 ppm Ni, 50 ppm V and 3 ppm Cd); (3) medium; and (4) heavy pollution (two- and threefold the light load, respectively). After 10 years of exposure, the highest concentrations of microbial ninhydrin-reactive nitrogen were found in the clay (2–0.1 μm) and silt fractions (63–2 μm), and the lowest were found in the coarse sand fraction (2,000–250 μm). The phospholipid fatty acid analyses (PLFA) and denaturing gradient gel electrophoresis (DGGE) separation of 16S rRNA gene fragments revealed that the microbial biomass within the clay fraction was predominantly due to soil bacteria. In contrast, a high percentage of fungal-derived PLFA 18 : 2ω6 was found in the coarse sand fraction. Bacterial residues such as muramic acid accumulated in the finer fractions in relation to fungal residues. The fractions also differed with respect to substrate utilization: Urease was located mainly in the <2 μm fraction, alkaline phosphatase and arylsulfatase in the 2–63 μm fraction, and xylanase activity was equally distributed in all fractions. Heavy metal pollution significantly decreased the concentration of ninhydrin-reactive nitrogen of soil microorganisms in the silt and clay fraction and thus in the bulk soil. Soil enzyme activity was reduced significantly in all fractions subjected to heavy metal pollution in the order arylsulfatase >phosphatase >urease >xylanase. Heavy metal pollution did not markedly change the similarity pattern of the DGGE profiles and amino sugar concentrations. Therefore, microbial biomass and enzyme activities seem to be more sensitive than 16S rRNA gene fragments and microbial amino-sugar-N to heavy metal treatment. Received: 21 January 2000     

重金属污染矿区复垦土壤微生物生物量及酶活性的研究

对铜矿废弃地复垦土壤微生物生物量及酶活性研究结果表明 ,与对照土壤相比 ,矿区复垦土壤微生物生物量C、N和P均有所降低 ,微生物商Cmic Corg可作为矿区重金属污染土壤微生物学敏感指标之一 ;酶活性变化与此相似 ,脲酶、脱氢酶和酸性磷酸酶与对照差异显著 ,其他则与对照差异明显 ,一定程度削弱了矿区土壤中C、N营养元素周转速率和能量循环     

Influence of heavy metals on the functional diversity of soil microbial communities

Three soil types-Calcaric Phaeozem, Eutric Cambisol and Dystric Lithosol-in large container pots were experimentally contaminated with heavy metals at four different levels (light pollution: 300 ppm Zn, 100 ppm Cu, 50 ppm Ni, 50 ppm V and 3 ppm Cd; medium pollution: twofold concentrations; heavy pollution: threefold concentrations; uncontaminated control). We investigated the prognostic potential of 16 soil microbial properties (microbial biomass, respiration, N-mineralization, 13 soil enzymes involved in cycling of C, N, P and S) with regard to their ability to differentiate the four contamination levels. Microbial biomass and enzyme activities decreased with increasing heavy metal pollution, but the amount of decrease differed among the enzymes. Enzymes involved in the C-cycling were least affected, whereas vartous enzyme activities related to the cycling of N, P and S showed a considerable decrease in activity. In particular, arylsulfatase and phosphatase activities were dramatically affected. Their activity decreased to a level of a few percent of their activities in the corresponding unpolluted controls. The data suggest that aside from the loss of rare biochemical capabilities-such as the growth of organisms at the expense of aromatics (Reber 1992)-heavy metal contaminated soils lose very common biochemical propertities which are necessary for the functioning of the ecosystem. Cluster analysis as well as discriminant analysis underline the similarity of the enzyme activity pattern among the controls and among the polluted soils. The trend toward a significant functional diversity loss becomes obvious already at the lowest pollution level. This implies that concentrations of heavy metals in soils near the current EC limits will most probably lead to a considerable reduction in decomposition and nutrient cycling rates. We conclude that heavy metal pollution severely decreases the functional diversity of the soil microbial community and impairs specific pathways of nutrient cycling.Dedicated to Professor J. C. G. Ottow on the occasion of his 60th birthday     

Effects of heavy metal contamination and remediation on soil microbial communities in the vicinity of a zinc smelter as indicated by analysis of microbial community phospholipid fatty acid profiles

Heavy metal contamination in an area immediately surrounding a zinc smelter has resulted in destruction of over 485 hectares of forest. The elevated levels of heavy metals in these soils have had significant impacts on the population size and overall activity of the soil microbial communities. Remediation of these soils has resulted in increases in indicators of biological activity and viable population size, which suggest recovery of the microbial populations. Questions remain as to how the metal contamination and subsequent remediation at this site have impacted the population structure of the soil microbial communities. In the current study, microbial communities from this site were analyzed by the phospholipid fatty acid (PLFA) procedure. Principal component analysis of the PLFA profiles indicated that there were differences in the profiles for soils with different levels of metal contamination, and that soils with higher levels of metal contamination showed decreases in indicator PLFAs for mycorrhizal fungi, Gram-positive bacteria, fungi, and actinomycetes. PLFA profiles for remediated sites indicated that remediated soils showed increases in indicator PLFAs for fungi, actinomycetes, and Gram-positive bacteria, compared to unremediated metal contaminated soils. These data suggest a change in the population structure of the soil microbial communities resulting from metal contamination and a recovery of several microbial populations resulting from remediation.     

Microbial activity and community structure of a soil after heavy metal contamination in a model forest ecosystem

We assessed the effects of chronic heavy metal (HM) contamination on soil microbial communities in a newly established forest ecosystem. We hypothesized that HM would affect community function and alter the microbial community structure over time and that the effects are more pronounced in combination with acid rain (AR). These hypotheses were tested in a model forest ecosystem consisting of several tree species (Norway spruce, birch, willow, and poplar) maintained in open top chambers. HMs were added to the topsoil as filter dust from a secondary metal smelter and two types of irrigation water acidity (ambient rain vs. acidified rain) were applied during four vegetation periods. HM contamination strongly impacted the microbial biomass (measured with both fumigation-extraction and quantitative lipid biomarker analyses) and community function (measured as basal respiration and soil hydrolase activities) of the soil microbial communities. The most drastic effect was found in the combined treatment of HM and AR, although soil pH and bioavailable HM contents were comparable to those of treatments with HM alone. Analyses of phospholipid fatty acids (PLFAs) and terminal restriction fragment length polymorphisms (T-RFLPs) of PCR-amplified 16S ribosomal DNA showed that HM treatment affected the structure of bacterial communities during the 4-year experimental period. Very likely, this is due to the still large bioavailable HM contents in the HM contaminated topsoils at the end of the experiment.     

Near-infrared spectroscopy for analysis of chemical and microbiological properties of forest soil organic horizons in a heavy-metal-polluted area

In industrial areas, heavy metals may accumulate in forest soil organic horizons, affecting soil microorganisms and causing changes in the chemical composition of the accumulated organic matter. The objectives of this study were to test the ability of near-infrared spectroscopy (NIRS) to detect heavy metal effects on the chemical composition of forest soil O horizons and to test whether NIRS may be used to quantitatively determine total and exchangeable concentrations of Zn and Pb (Zn_t, Pb_t, Zn_ex, Pb_ex) and other chemical and microbial properties in forest soil O horizons polluted with heavy metals. The samples of O horizons (n = 79) were analyzed for organic C (C_org), total N and S (N_t, S_t), Zn_t, Pb_t, Zn_ex, Pb_ex, basal respiration (BR), microbial biomass (C_mic) and C_mic-to-C_org ratio. Spectra of the samples were recorded in the Vis-NIR range (400–2,500 nm). To detect heavy-metal-induced changes in the chemical composition of O horizons principal components (PC1–PC7) based on the spectral data were regressed against Zn_t + Pb_t values. A modified partial least squares method was used to develop calibration models for prediction of various chemical and microbial properties of the samples from their spectra. Regression analysis revealed a significant relationship between PC3 and PC5 (r = −0.27 and −0.34, respectively) and Zn_t + Pb_t values, indicating an effect of heavy metal pollution on the spectral properties of the O horizons and thus on their chemical composition. For quantitative estimations, the best calibration model was obtained for C_org-to-N_t ratio (r = 0.98). The models for C_org, N_t, and microbial properties were satisfactory but less accurate. NIRS failed to accurately predict S_t, C_org-to-S_t, Zn_t, Pb_t, Zn_ex, and Pb_ex.     

Effects of heavy metal accumulation in apple orchard soils on microbial biomass and microbial activities

Abstract

The effects of heavy metals (Cu, Pb, and As) accumulated in apple orchard surface soils on the microbial biomass, dehydrogenase activity, and soil respiration were investigated. The largest concentrations of total Cu, Pb, and As found in the soils used were 1,010, 926, and 166 mg kg^?1 soil, respectively. The amounts of microbial biomass C and N, expressed on a soil organic C and soil total N basis, respectively, were each negatively correlated with the amounts of total, 0.1 M HCI-extractable, and 0.1 M CaCl₂-extractable Cu as logarithmic functions, the correlation coefficient being lowest for the 0.1 M CaCl₂extractable Cu. Nevertheless, they were not correlated with the soil pH which was controlling the solubility of Cu in 0.1 M CaCl₂. The dehydrogenase activity expressed per unit of soil organic C was also negatively correlated with the amounts of total, 0.1 M HCI-extractable Cu, and 0.1 M CaCl₂-extractable Cu as logarithmic functions. However, the correlation coefficient was highest for the 0.1 M CaCl₂-extractable Cu. Although the soil respiration per unit of soil total organic C did not show any significant correlations with the total concentrations of heavy metals, it showed negative significant correlations with the amount of 0.1 M HCI-extractable Cu, and to a greater extent, with the amount of 0.1 M CaCl₂-extractable Cu. Both the dehydrogenase activity and respiration per unit of soil total organic C increased significantly with increasing soil pH. These results suggested that in apple orchard soils with heavy metal accumulation the microbial biomass was adversely affected by the slightly soluble Cu, whereas the microbial activities by the readily soluble Cu whose amount depended on the soil pH. The respiration per unit of microbial biomass C showed a positive significant correlation with the logarithmic concentration of total Cu. Furthermore, the contribution of fungi to substrate-induced respiration increased with increasing total Cu content in the soils.     

重金属污染土壤间作修复的研究进展

种植单一的超富集植物修复重金属污染土壤,不但中断农业生产导致经济收益降低,而且因生物量较低、修复周期长等诸多弊端导致修复效果不甚理想。间作作为一种传统的农艺管理方式,利用生态位和生物多样性原理等能提高农作物对资源的有效利用,对共植的农作物种类增量提质。在中、轻度污染土壤修复中利用间作体系,通过调控超富集植物与农作物的生长发育,促进超富集植物根系低分子量有机酸(LMWOAs)的分泌,降低其根际土壤p H,增加重金属活性,从而增加超富集植物对重金属的吸收,同时抑制农作物根系LMWOAs的分泌,以减少农作物对重金属的吸收,提高其产量和品质,实现"边生产边修复",提高土地利用率,并增加经济效益。本文根据近几年来国内外相关文献,综述了间作条件下超富集植物和农作物生物量、生理生化响应、重金属吸收、转运、富集等方面的变化,以及间作对土壤环境质量的影响,并对间作修复重金属污染土壤领域的发展趋势,如超富集植物和农作物间作的信号转导和分子生物学机制、间作体系下两类植物根际微生物类群的差异及其功能机制,以及构建高效间作体系提高重金属污染土壤的修复效率等方面进行了展望。     

Is the soil microbial community related to the basal area of trees in a Scots pine stand?

The main energy sources of soil microorganisms are litter fall, root litter and exudation. The amount on these carbon inputs vary according to basal area of the forest stand. We hypothesized that soil microbes utilizing these soil carbon sources relate to the basal area of trees. We measured the amount of soil microbial biomass, soil respiration and microbial community structure as determined by phospholipid fatty acid (PLFA) profiles in the humus layer (FH) of an even-aged stand of Scots pine (Pinus sylvestris L.) with four different basal area levels ranging from 19.9 m² ha⁻¹ in the study plot Kasper 1 to 35.7 m² ha⁻¹ in Kasper 4. Increasing trend in basal respiration, total PLFAs and fungal-to-bacterial ratio was observed from Kasper 1 to Kasper 3 (basal area 29.2 m² ha⁻¹). The soil microbial community structure in Kasper 3 differed from that of the other study plots.     

Experimentally induced effects of heavy metal on microbial activity and community structure of forest mor layers

This study compared the toxic effects of adding chromium (Cr), zinc (Zn), lead (Pb), molybdenum (Mo), nickel (Ni), and cadmium (Cd) at three dose levels to mor layer samples in laboratory experiments. Microbial activity in the form of soil respiration was monitored for 64 days. At the end of the experimental period, the composition of the soil microbial community structure was analysed by phospholipid fatty acid (PLFA) analysis. The metals added induced changes in the microbial community structure and affected respiration negatively, indicating toxicity. The microbial community structure (principal component analysis of the PLFA pattern) for all metals was significantly related to microbial activity (cumulative respiration), indicating intimate links between microbial community structure and activity. The most striking result in this study was that the shift in the microbial community because of metal stress was similar for all metals. Thus, the PLFA i16:0 increased most in relative abundance in metal-polluted soils, followed by other PLFAs indicative of Gram-positive bacteria (10Me16:0, 10Me17:0, 10Me18:0, a17:0 and br18:0). The PLFA 16:1ω5 was consistently negatively affected by metal stress, as were the PLFAs 18:1, 18:1ω7 and 19:1a. However, a significant separation between Cr- and Cd-polluted soils was observed in the response of the PLFA cy19:0, which decreased in abundance with Cr stress, and increased with Cd stress. Furthermore, the PLFA 18:2w6, indicating fungi, only increased with Cr and Zn stress. The effective doses of the metals, ranked with regard to background metal concentrations, decreased in the order: Zn > Cr > Pb > Mo > Ni > Cd. We concluded that interpretation of results of microbial activity from experiments of metal toxicity should include microbial structural patterns and background metal concentrations.     

Temporal response of soil biochemical properties in a pastoral soil after cultivation following high application rates of undigested sewage sludge

Soil biochemical properties were measured annually between 1995 and 1999 in soil from an 8-ha site that had received over 1,000 wet tonnes ha^–1 undigested sewage sludge, 1–4 years earlier. Basal respiration generally declined with time and was usually greatest in the untreated control area. This trend was attributed to a similar trend in soil moisture content. In contrast, microbial biomass C increased with time and also generally increased with sludge treatment age. Microbial biomass C, and to a lesser extent sulphatase activity, accurately predicted the order of sludge application to the site. This was perceived as a function of time since tillage and pasture establishment, with activities increasing in parallel to the build up of C residues in the soil, and not an effect of sludge or its composition. Except immediately after sludge application, there was no effect on N mineralisation and nitrification. None of the biochemical properties was strongly correlated with heavy metal concentrations. Our results suggest that there was little effect on soil biochemical properties, either adverse or beneficial, of adding raw sewage sludge to this site. Although a companion study showed considerable mobility and plant uptake of heavy metals, this difference could mainly be attributed to a different sampling strategy and the effects of intensive liming of the site.     

Microbial biomass,activity, and organic matter accumulation in soils contaminated with heavy metals

Chemical characteristics and some parameters related to biological components were determined in 16 soils from a fairly homogeneous area in the north of Italy, contaminated with different levels of heavy metals. Correlation analysis of the parameters studied showed close positive relationships among the metals and with the organic C content in the soils studied. Negative relationships were observed among the heavy metals, soil respiration, and the ratio between evolved CO₂–C and microbial biomass C per unit time (specific respiratory activity). This was ascribed to an adverse heavy metal effect on the soil microflora, which appeared to increase the accumulation of organic matter as the heavy metal content increased, probably because the biomass was less effective in mineralising soil organic matter under these conditions.     

Influence of inorganic and organic fertilization on soil microbial biomass, metabolic quotient and heavy metal bioavailability

 We studied the long-term effects (12 years) of municipal refuse compost addition on the total organic carbon (TOC), the amount and activity of the microbial biomass (soil microbial biomass C, B_C and metabolic quotient qCO₂) and heavy metal bioavaiability in soils as compared to manuring with mineral fertilizers (NPK) and farmyard manure (FYM). In addition, we studied the relationships between among the available fraction [Diethylenetriaminopentacetic acid (DTPA) extractable] of heavy metals and their total content, TOC and B_C. After 12 years of repeated treatments, the TOC and B_C of control and mineral fertilized plots did not differ. Soils treated with FYM and composts showed a significant increase in TOC and B_C in response to the increasing amounts of organic C added. Values of the B_C/TOC ratio ranged from 1.4 to 2, without any significative differences among soil treatments. The qCO₂ increased in the organic-amended soil and may have indicated microbial stress. The total amounts of metals in treated soils were lower than the levels permitted by the European Union in agricultural soils. DTPA-extractable metals increased in amended soils in response to organic C. A multiple regression analysis with stepwise selection of variables was carried out in order to discriminate between the influence exerted on DTPA-extractable metals by their total content, TOC and B_C. Results showed that each metal behaved quite differently, suggesting that different mechanisms might be involved in metal bioavailability Received: 31 October 1997     

Soil microbial effects of a Cu-Ni smelter in southwestern Finland

Summary The influence of a Cu-Ni smelter on the soil microbial parameters: physiological groups of bacteria, soil respiration, fungal hyphal length, and green-needle litter decomposition, were investigated. The microbial parameters were reduced and this was significantly explained (P<0.01) by the supplied environmental variables: exchangeable Ca, Mg, K, Mn, Cu, Ni, Cd, Zn, soil moisture, pH, and organic C as loss on ignition (Canoco, RDA-analysis). The importance of measuring exchangeable cations for major and trace elements appeared to be a relevant factor that must be considered when establishing relationships between microbial populations, their activity and the effect of heavy metals.     

Soil C balance in a long-term field experiment in relation to the size of the microbial biomass

Soil C balances were calculated in a field experiment started in 1956. Treatments include a fallow and soils receiving different N fertilizers or organic amendments. By assuming the absence of a priming effect, the degree of mineralization of crop residues and organic amendments was calculated. Crop residue mineralization was not affected by a more than 50% decrease in the size of the microbial biomass in soil fertilized with (NH₄)₂SO₄, which had caused the pH of this soil to drop from 6.6 to 4.4. More C had accumulated per unit C input in peat-and sewage sludge-amended soils than in any of the other soils, suggesting that peat and sewage sludge were more resistant to microbial attack. Recalcitrance of substrate C was an adequate explanation for the low ratio of biomass C to soil C in the peat-amended soils, but not in the sewage sludge-amended soil. There was a close linear relationship (r=0.94) between the content of microbial biomass C in the soil measured in 1990 and cumulative C losses from the soil since 1956. Compared to the relationship between soil biomass C and soil organic C concentrations, the linear relationship between microbial C and cumulative C losses suggested that the significantly reduced biomass in the sewage sludge-amended soil was at least partially due to the presence of toxic substances (presumably elevated heavy metal concentrations) in this soil and was probably not affected by the somewhat low pH (5.3) in this soil.     

The use of microbial parameters in monitoring soil pollution by heavy metals

Microbial parameters appear very useful in monitoring soil pollution by heavy metals, but no single microbial parameter can be used universally. Microbial activities such as respiration, C and N mineralization, biological N₂ fixation, and some soil enzymes can be measured, as can the total soil microbial biomass. Combining microbial activity and population measurements (e.g., biomass specific respiration) appears to provide more sensitive indications of soil pollution by heavy metals than either activity or population measurements alone. Parameters that have some form of internal control, e.g., biomass as a percentage of soil organic matter, are also advantageous. By using such approaches it might be possible to determine whether the natural ecosystem is being altered by pollutants without recource to expensive and long-running field experiments. However, more data are needed before this will be possible. Finally, new applications of molecular biology to soil pollution studies (e.g., genetic fingerprinting) which may also have value in the future are considered.     

Suitability of soil microbial parameters as indicators of heavy metal pollution

Several microbial parameters (microbial biomass, respiration, dehydrogenase, phosphatase, sulphatase, glucosidase, protease and urease activities) were measured in soils from five sites located in urban green areas close to roads differing in traffic density. Our aims were to evaluate the suitability of such parameters as field biomarkers of stress induced by heavy metal pollution, and to compare results obtained by single microbial parameters with results given by an index expressing the average microbial (AME) response of the microbial community. Data showed that all parameters were significantly reduced in the sites characterized by the highest load of metals in soil. Dehydrogenase, sulphatase, glucosidase activities and respiration, declined exponentially with increasing metal concentration, whereas phosphatase activity and AME decreased following a sigmoidal type relationship. In contrast, protease, urease and microbial biomass were not significantly correlated with soil metal concentration. Microbial parameters differed both in sensitivity to critical metal concentrations and in the rate of decline at increasing metal loads in soil. Due to the complex interplay of chemical, physical and biological factors which influence microbial activities and biomass, the proposed index (AME) appeared more suitable than single microbial parameters for a biomonitoring study of this type.     

Straw coverage alleviates seasonal variability of the topsoil microbial biomass and activity

Straw coverage on soil surface is a well-known practice for conserving soil and water. Seasonal variability of microbial biomass carbon (MBC) and basal respiration (BR) in surface soil (0–5 cm) was compared between conventional straw removal (SR) and straw coverage (SC) in a maize (Zea mays L.) field experiment, Northeast China. The straw coverage treatment significantly increased microbial energy, carbon and nutrient sources (soil C and N contents) and improved soil physical environment (moisture and porosity), and thus stimulated MBC and BR across the growth season of maize, as compared to the straw removal treatment. MBC and BR showed similar seasonal trends in soil temperature, with maximum values in summer. In both study years, the straw coverage treatment reduced seasonal variation of soil temperature, therefore it significantly moderated seasonal variability of MBC and BR. Our results demonstrate that straw coverage contributes to stabilizing soil microbial characteristics in season.     

Near-infrared characteristics of forest humus are correlated with soil respiration and microbial biomass in burnt soil

Near-infrared spectroscopy and soil physicochemical determinations (pH_H2O, organic matter content, total C content, NH _inf4 ^sup+ , total N content, cation-exchange capacity, and base saturation) were used to characterize fire-or wood ash-treated humus samples. The spectroscopic and the soil physicochemical analysis data from the humus samples were used separately to explain observed variations in soil respiration and microbial biomass C by partial least-square regression. The first regression component obtained from the physicochemical and spectroscopic characterization explained 10–12% and 60–80% of the biological variation, respectively. This suggests that information on organic material collected from near-infrared spectra is very useful for explaining biological variations in forest humus.     

The effect of tropical forest conversion on soil microbial biomass

We investigated the effects of converting forest to savanna and plough land on the microbial biomass in tropical soils of India. Conversion of the forest led to a significant reduction in soil organic C (40–46%), total N (47–53%), and microbial biomass C (52–58%) in the savanna and the plough land. Among forest, savanna, and plough land, basal soil respiration was maximum in the forest, but the microbial metabolic quotient (qCO₂ was estimated to be at a minimum in the forest and at a maximum in the plough land.