Cryptic diversity in the well-studied terrestrial worm Cognettia sphagnetorum (Clitellata: Enchytraeidae)

The terrestrial worm Cognettia sphagnetorum has been used as a model in several studies focusing on research areas such as climate change as well as forest and soil ecology; it has also been shown to play a key role in the decomposition of organic matter and nutrient cycling. Cognettia is an enchytraeid genus commonly found in acidic terrestrial habitats, such as coniferous forests and bogs. In this study, the diversity of the genus, with particular focus on the morphospecies C. sphagnetorum in northern Europe, is assessed using four molecular markers, the mitochondrial COI (cytochrome c oxidase subunit I) and16S (16S ribosomal RNA), and the nuclear H3 (Histone 3) and ITS (Internal Transcribed Spacer). The datasets were first delimited into Molecular Operational Units (MOTUs) and the existence of global barcoding-gaps was tested. Single gene-trees were then estimated for all genes using Bayesian Inference, and a species tree was estimated with all markers combined using the multi-species coalescence. The results show that in northern Europe the genus consists of at least eight MOTUs supported by all genes except H3. Four of these MOTUs were within the morphotaxon C. sphagnetorum and two within Cognettia glandulosa. C. sphagnetorum s.l. was found to be non-monophyletic in all gene-trees, as well as in the species tree. As the MOTUs were well separated and non-monophyly was observed within C. sphagnetorum s.l., we conclude that the MOTUs are best treated as separate species. Given that cryptic diversity was found in this genus, we recommend that material of Cognettia used in future studies should be identified using molecular barcodes.     

Why does temperature affect relative uptake rates of nitrate, ammonium and glycine: A test with Eucalyptus pauciflora

Few studies have examined how temperature affects uptake of nitrate, ammonium and amino acids from soil. This study tests the hypothesis that cool temperatures favour uptake of the amino acid glycine while warm temperatures favour uptake of inorganic forms of N such as nitrate. We used glasshouse-grown ectomycorrhizal seedlings of the sub-alpine tree species Eucalyptus pauciflora Sieber ex Spreng. Seedlings were grown in soil (humic umbrosol, from species' habitat) that was dominated by amino acids and ammonium with only small amounts of nitrate. To examine if root physiology affects temperature responses of N uptake, we measured uptake from ¹⁵N-labelled hydrosolutions containing equimolar 100 μmol L⁻¹ mixtures of ammonium, nitrate and glycine at temperatures from 5 to 35 °C. We also examined if the effect of temperature on uptake of N forms was due to plant-microbe competition by following the fate of equimolar amounts of labelled ammonium, nitrate and glycine injected into the soil at temperatures of 5 °C and 25 °C. Hydrosolution experiments showed that uptake of glycine was favoured by warm temperatures and inorganic N by cool temperatures. In contrast, when ¹⁵N was injected into soil the uptake of glycine was favoured by low temperatures and nitrate by warm temperatures. At 25 °C, glycine was 17% of the N taken up from soil and nitrate was 51%; whereas at 5 °C glycine was 30% of the N taken up from soil and nitrate was 23%. Microbes were better competitors than seedlings for all forms of N, but temperature did not affect microbial preference for the different N forms. Hence, while microbes limit N available for plant uptake, they do not seem to be the cause of the greater plant uptake of glycine at cool temperatures and nitrate at warm temperatures. Intact uptake of glycine by plants was suggested by the positive relationship between uptake of ¹³C and ¹⁵N and detection by GC-MS of intact , ¹⁵N glycine molecules in roots. In conclusion, uptake of glycine is favoured by cool temperatures and nitrate by warm temperatures, but this is apparently not a function of root physiology or competition with soil microbes.     

Earthworms (Millsonia anomala, Megascolecidae) do not increase rice growth through enhanced nitrogen mineralization

Earthworms have been shown to increase plant growth in 75% of the experiments that have compared plant growth in their presence and absence. However, the relative importance of the different mechanisms advanced to explain such a stimulatory effect has never been tested. In a laboratory experiment, we observed increased growth of rice plants in the presence of earthworms (Millsonia anomala, Megascolecidae) and demonstrated that enhanced nitrogen release (generally considered as the principal mechanism involved in earthworm positive effect on plants) was not responsible for this result: earthworms had the same stimulatory effect on plant growth (+20%) irrespective of whether the soil (provided with different amounts of mineral-N fertilizer) was either N-limited or N-saturated. We discuss alternative explanations for the observed variations in rice production.     

Nodulation and growth of common bean (Phaseolus vulgaris) under water deficiency

Three experiments were conducted in order to investigate the effect of water deficiency on nodulation, rhizobial diversity and growth of common bean. In the first experiment, the effect of water deficiency was studied on two soil samples under glasshouse conditions. A significant decrease in nodulation and shoot dry weight production was observed. The molecular characterization of the root nodule isolates by PCR-RFLP of 16S rRNA and nodC genes showed that the nodulation by Rhizobium etli was severely inhibited. The in vitro analysis of salt tolerance indicated that drought stress favoured nodulation by salt-tolerant strains. In the second experiment, the effect of water deficiency was studied on sterilized sand using Rhizobium tropici CIAT899^T and Ensifer meliloti bv. mediterranense 4H41 as inoculants. The results showed that strain 4H41, which is the more salt tolerant, was more competitive and more effective under water deficiency than strain CIAT899^T. In the third experiment, the strain 4H41 was used to inoculate four fields. A significant increase in nodule number, shoot dry weight and grain yield was observed even in the non-irrigated soils. This work constitutes the first report of a strain enhancing the growth and the grain yield of common bean under water deficiency.     

Temporal patterns of nutrient availability around nests of leaf-cutting ants (Atta colombica) in secondary moist tropical forest

Leaf-cutting ants consume up to 10% of canopy leaves in the foraging area of their colony and therefore represent a key perturbation in the nutrient cycle of tropical forests. We used a chronosequence of nest sites on Barro Colorado Island, Panama, to assess the influence of leaf-cutting ants (Atta colombica) on nutrient availability in a neotropical rainforest. Twelve nest sites were sampled, including active nests, recently abandoned nests (<1 year) and long-abandoned nests (>1 year). Waste material discarded by the ants down-slope from the nests contained large concentrations of nitrogen and phosphorus in both total and soluble forms, but decomposed within one year after the nests were abandoned. Despite this, soil under the waste material contained high concentrations of nitrate and ammonium that persisted after the disappearance of the waste, although soluble phosphate returned to background concentrations within one year of nest abandonment. Fine roots were more abundant in soil under waste than control soils up to one year after nest abandonment, but were not significantly different for older sites. In contrast to the waste dumps, soil above the underground nest chambers consistently contained lower nutrient concentrations than control soils, although this was not statistically significant. We conclude that the ‘islands of fertility’ created by leaf-cutting ants provide a nutritional benefit to nearby plants for less than one year after nest abandonment in the moist tropical environment of Barro Colorado Island.     

The HFC/HCFC breakdown product trifluoroacetic acid (TFA) and its effects on the symbiosis between Bradyrhizobium japonicum and soybean (Glycine max)

The environmental impact of hydrofluorocarbon (HFC) and hydrochlorofluorocarbon (HCFC) are under intense study due to the resistance of the breakdown product, trifluoroacetic acid (TFA), to further degradation. TFA has come under scrutiny due to its rapid and complete partitioning into aqueous phases of the environment, eventually allowing for deposition of TFA into soil via precipitation. Evidence exists that TFA may be toxic to soil microbes and plants, with little or no degradation occurring in soils. Uptake by plants and microorganisms and its similarity to acetate, implies its potential to effect the symbiotic nitrogen-fixing partners Bradyrhizobium japonicum and soybean (Glycine max). A preliminary study was performed in accordance with the Alternative Fluorocarbon Environmental Acceptability Study (AFEAS). Those results are presented here in addition to the findings of further experimentation on the initial interaction of B. japonicum with soybean. We used three levels of TFA (0.67, 6.74 and 67.40 μl TFA kg⁻¹ soil; 0.003, 0.031 and 0.314 μl TFA l⁻¹) for soil and hydroponics conditions and three levels (10, 100 μM and 1 mM) in bacterial culture. The results demonstrate that TFA affects growth of B. japonicum significantly, but does not affect PHB accumulation. Also no F⁻ was found in cultures grown on TFA. Attachment of B. japonicum to soybean roots was enhanced with the lowest level of acetate or TFA and was significantly reduced with 1 mM acetate or TFA. Cultures grown on acetate or acetate with TFA do not attach well, with those grown with 1 mM TFA the least. Both effects may be attributed to pH. Soybean seedlings had significantly retarded development with levels of TFA at or above 6.74 μl TFA kg⁻¹ soil and 0.031 μl TFA l⁻¹ nutrient solution. No nodules formed on those plants treated with these levels of TFA except in the hydroponics trials. Nodule location was not affected regardless of the TFA level. At the lowest level used we found no effects on soybean or symbiotic nitrogen fixation. In some cases, nodulation was enhanced, but nodule weight reduced. Anaerobically isolated bacteroids had normal levels of acetylene reduction activity regardless of the level of TFA used. In summary, soybean is much more sensitive to low levels of TFA than its symbiotic counterpart B. japonicum. No detrimental effects on symbiotic nitrogen fixation in soybean should be expected unless large bioaccumulation of TFA occurs in agricultural areas.     

Release and consumption of nitrogen by snail feces in Negev Desert soils

Snail grazing and feces production have been shown to be major components of the nitrogen (N) budget of Negev Desert ecosystems. However, the movement of N from feces into soil N cycling processes has not been studied. In this study, we measured immediate N release from different types of snail feces following wetting of dry desert soils, and characterized potential net N mineralization and nitrification and soil respiration over a 12-day incubation under laboratory conditions. The dynamics of morganic N exhibited two distinct phases during the 12-day incubation: (1) immediate release of inorganic N following wetting of the soil and (2) decline of inorganic N from day 1 today 12 of the incubation. The immediate pulse of N release from this one wetting event (6–25 mg N m^-2) was larger than annual atmospheric inputs of N to Negev Desert ecosystems (<2 mg N m^-2); however, from 50 to 80% of the N released upon wetting was consumed by the end of the incubation. There were differences in inorganic N release and respiration from feces from different kinds of snails, and from feces from the same species of snail fed different plants. The results suggest that while snail feces contribute significant amounts of plant available N to Negev ecosystems, plants must compete with other sinks for this N.     

Rapid intrinsic rates of amino acid biodegradation in soils are unaffected by agricultural management strategy

Amino acids represent one of the largest inputs of dissolved organic nitrogen to soil and consequently they constitute a major component of the organic N cycle. The effect of agricultural management on the rate of amino acid turnover in soil, however, remains largely unknown. The aim of this study was to evaluate in long-term field experiments the effect of fertilizer addition (N, P and K), grazing, pH manipulation (lime addition), vegetation cover and shifts (grassland versus arable) and drainage on the mineralization of ¹⁴C-labelled amino acids in agricultural topsoils. Our results showed that the intrinsic rate of amino acid mineralization was rapid for all management regimes, irrespective of the tested soil type. The average (±SEM) half-life of the amino acids in all soils (n=155) was calculated to be 2.3±0.5 h. The relative amount of amino acid-C partitioned into respiration (25% of total C) versus biomass production (75% of total C) was also unaffected by management strategy. The rate of amino acid mineralization was shown to be slightly sensitive to soil pH, peaking at around pH_(2CaCl) 5.0 with an approximate twofold reduction at the pH extremes (pH 3.8 and 6.4). We conclude that management regime has little effect on the intrinsic rate of amino acid mineralization in agricultural soils. We propose therefore that total microbial activity rather than microbial diversity or community structure is likely to be the key determinant governing amino acid turnover in agricultural soils.     

The effects of Glycine max and Helianthus annuus on nutrient availability in two soils

The effects of two plant species—soybean (Glycine max), and sunflower (Helianthus annuus) on nutrient availability in two soils (an organically farmed soil, OF, and a native grassland soil, GS, both Alfisols)—were measured with anion exchange membranes (Plant Root Simulator, PRS^TM probes) in a greenhouse study. Vegetation (especially sunflower) in the OF soil caused significant reductions in soil N and K availability (which was interpreted as due to uptake), and significant increases in P, S, Cu, Fe, Mn, and Zn availability. The increases in the latter case were consistent with the results of a previous study showing rhizosphere-enhanced mineralization of native soil organic matter in this soil. Vegetation had no significant effects on Ca, Mg, B, or Al availability in the OF soil and no significant effects on any measured nutrient in the GS soil. Collectively, these results show that the presence of plants can have either a negative or a positive effect on soil nutrient availability, and that plant uptake and soil nutrient availability are interdependent.     

Seasonal and diurnal CO2 efflux from red wood ant (Formica aquilonia) mounds in boreal coniferous forests

Organic mounds of the red wood ants (Formica rufa group; RWA) have been shown to be “hot spots” of carbon dioxide (CO₂) efflux from the European forest soils. However, little information is available on the variability of CO₂ effluxes from RWA mounds and on the factors regulating CO₂ efflux. We assessed the seasonal and diurnal changes in CO₂ effluxes, temperatures and volumetric water contents from mounds of Formica aquilona, the important RWA of the boreal forests in Finland. The daily average CO₂ efflux from RWA mounds ranged 1.1-6.9 g CO₂ m⁻² h⁻¹ during the active ant season (May-September), and from 0.2 to 1.1 g CO₂ m⁻² h⁻¹ during their dormant period (October-April). Mound CO₂ efflux from May to September was 3.6-6.0 times higher than from the surrounding forest floors, and most likely came from RWA respiration. Seasonal changes in mound CO₂ effluxes were significantly correlated with mound temperature, but not with volumetric water content (7% on average). The high CO₂ efflux associated with increased volumetric water content (up to 34%) after a RWA mound was abandoned indicated that these dry mound conditions restrict microbial decomposition of mound organic matter. CO₂ effluxes were highest at night and lowest during the day, which is likely due to an increased ant activity or numbers in the mound at night. Diurnal changes in mound CO₂ efflux were negatively correlated with air temperature, and positively correlated with the difference between the mound and air temperature. This suggests that thermal convection of warmer mound air to the colder outside air at night might be also a cause of the diurnal changes. We conclude that seasonal and diurnal variations in mound CO₂ effluxes are dependent on RWA activities and fluctuation in RWA mound and outside temperatures.     

长期施肥水稻土氮素剖面分布及温度对土壤氮素矿化特性的影响

利用太湖地区26a的水稻土长期定位试验,研究了长期施肥对水稻土剖面氮素分布和C/N的影响,分析了10℃和30℃下不同施肥处理土壤氮素矿化过程的差异,拟合了30℃下土壤累积矿化量与有效积温的方程式。结果表明:(1)长期施肥使表层土壤氮素累积量明显增加,0～20cm土层,施有机肥处理的含氮量普遍高于施化肥处理。20～30cm土层,化肥氮+磷处理(CNP)、化肥氮+磷+钾处理(CNPK)、秸秆+氮处理(CRN)及不施肥对照CK的含氮量高于有机肥+氮+磷处理(MNP)、有机肥+氮+磷+钾处理(MNPK)、秸秆+有机肥+氮处理(MRN)和仅施有机肥处理(M0),而40～50cm土层含氮量差异较小;(2)施有机肥或秸秆还田使表层土C/N值有降低趋势。0～10cm表层土壤的C/N,CNPK>MNPK,CK>M0,而施秸秆处理MRN的C/N最低。20～50cm土层,施有机肥处理的C/N普遍高于化肥处理,施化肥土壤下层的有机质分解程度较高。(3)在30℃下,土壤矿化过程达到稳定状态需要时间较短,且累积矿化量较高。在10℃下,培养后期的矿化速率高于30℃下的速率,但累积矿化量较低,低温条件引起了土壤氮素矿化率的降低。土壤累积矿化量与有效积温的关系符合有效积温方程式(p<0·01),K值较接近反应矿化初期各处理的矿化潜力,n值的差异反应MRN、MNP以及CNPK、CNP处理在矿化后期有更高潜力。     

Influence of Cognettia sphagnetorum (Oligochaeta: Enchytraeidae) on nitrogen mineralization in homogenized mor humus

Summary The influence of the enchytraeid species Cognettia sphagnetorum on N mineralization in homogenized mor humus was examined in a laboratory study. The mor humus was incubated in containers (150 ml) for 8 months at various temperatures and with different moisture levels. Two series were used, one with C. sphagnetorum and one without. The presence of enchytraeids in the cultures increased the level of NH₄ ⁺ and NO₃ ^- by about 18% compared with the cultures without enchytraeids. Almost 40% of this difference was explained by the decomposition of dead enchytraeids. Temperature and soil moisture were the most important factors controlling the mineralization rate. The optimum moisture for N mineralization was between pF 1.6 and 1.1.Dedicated to the late Prof. Dr. W. Kühnelt     

Litter decomposition and nitrogen mineralization in newly formed gaps in a Danish beech (Fagus sylvatica) forest

Gap formation is suggested as an alternative forest management approach to avoid extreme changes in the N cycle of forest ecosystems caused by traditional management practises. The present study aimed to investigate the effect of gap formation on N availability in beech litter and mineral soil on sites, which experienced only little soil disturbance during tree harvest. N pools, litter decomposition, and N mineralization rates in mineral soil were studied in two gaps (17 and 30 m in diameter) in a 75-year-old managed European beech (Fagus sylvatica L.) forest in Denmark and related to soil temperature (5 cm depth) and soil moisture (15 cm depth). Investigations were carried out during the first 2 years after gap formation in measurement plots located along the north-south transect running through the centre of each gap and into the surrounding forest.An effect of gap size was found only for soil temperatures and litter mass loss: soil temperatures were significantly increased in the northern part of the large gap during the first year after gap formation, and litter mass loss was significantly higher in the smaller gap. All other parameters investigated revealed no effect of gap size. Nitrification, net mineralization, and soil N concentrations tended to be increased in the gaps. Cumulative rates of net mineralization were two fold higher in the gaps during the growing season (June-October), but a statistically significant increase was found only for soil NH₄-N concentrations during this period. Forest floor parameters (C:N ratios, mass loss, N release) were not significantly modified during the first year after gap formation, neither were the total C content nor the C:N ratio in mineral soil at 0-10 cm depth.     

The oribatid mite Scheloribates moestus (Acari: Oribatida) alters litter chemistry and nutrient cycling during decomposition

It is widely accepted that microarthropods influence decomposition dynamics but we know relatively little about their effects on litter chemistry, extracellular enzyme activities, and other finer-scale decomposition processes. Further, few studies have investigated the role of individual microarthropod species in litter decomposition. The oribatid mite Scheloribates moestus Banks (Acari: Oribatida) is abundant in many U.S. ecosystems. We examined the potential effects of S. moestus on litter decomposition dynamics and chemical transformations, and whether these effects are influenced by variation in initial litter quality. We collected corn and oak litter from habitats with large populations of S. moestus and in microcosms with and without mites measured respiration rates, nitrogen availability, enzyme activities, and molecular-scale changes in litter chemistry. Mites stimulated extracellular enzyme activities, enhanced microbial respiration rates by 19% in corn litter and 17% in oak litter over 62 days, and increased water-extractable organic C and N. Mites decreased the relative abundance of polysaccharides in decomposing corn litter but had no effect on oak litter chemistry, suggesting that the effects of S. moestus on litter chemistry are constrained by initial litter quality. We also compared the chemistry of mite feces to unprocessed corn litter and found that feces had a higher relative abundance of polysaccharides and phenols and a lower relative abundance of lignin. Our study establishes that S. moestus substantially changes litter chemistry during decomposition, but specific effects vary with initial litter quality. These chemical transformations, coupled with other observed changes in decomposition rates and nutrient cycling, indicate that S. moestus could play a key role in soil C cycling dynamics.     

稻田土壤氮素矿化的几种方法比较

结合盆栽实验,对稻田土壤氮素矿化的几种方法进行了比较,结果表明:KC I煮沸法和碱解氮法是很好的测试土壤供氮能力的方法,淹水培养法矿化氮量与植物全氮含量相关性不高;而采用淹水培养间歇淋洗法和淹水密闭连续培养法对稻田土壤进行土壤氮素矿化的淋溶试验,结果表明:淋洗有利于土壤氮素的矿化过程。     

Effects of Long-term Organic and Mineral N-fertilisation on Soil Microarthropods in Berlin (Germany), Madrid (Spain) and Tartu (Estonia)

Soil micro-arthropods play an important role in the nutrient cycle of agro-ecosystems and are considered to have a strong impact on microbial activity. The occurance of soil microarthropods was investigated in the International Organic Long-Term Field Experiment (IOSDV). The investigation took place in the experimental fields of Berlin/Germany, Madrid/Spain and Tartu/Estonia. Field plots "without organic fertisation", "farmyard manure" and "straw-beet-leaves-green manure" without mineral N-fertilisation and with 120kg/ha mineral N-fertilisation respectively have been investigated. Data shows that the effects of organic manure and mineral N-fertilisation are considerably influenced by specific site characteristics. The highest abundances were found in the experimental field of Madrid. In the two nothern sites soil microarthropods were found only in very low quantities. Data indicates that not only organic manure but also mineral N-fertilisation may influence the abundance of soil micro-arthropods positively.     

Decomposer animals (Lumbricidae, Collembola) and organic matter distribution affect the performance of Lolium perenne (Poaceae) and Trifolium repens (Fabaceae)

Decomposer animals stimulate plant growth by indirect effects such as increasing nutrient availability or by modifying microbial communities in the rhizosphere. In grasslands, the spatial distribution of organic matter (OM) rich in nutrients depends on agricultural practice and the bioturbation activities of large detritivores, such as earthworms. We hypothesized that plants of different functional groups with contrasting nutrient uptake and resource allocation strategies differentially benefit from sites in soil with OM accumulation and the presence of decomposer animals. In a greenhouse experiment we investigated effects of spatial distribution of ¹⁵N-labelled grass litter, earthworms and collembola on a simple grassland community consisting of Lolium perenne (grass) and Trifolium repens (legume). Litter aggregates (compared to homogeneous litter distribution) increased total shoot biomass, root biomass and ¹⁵N uptake by the plants. Earthworms and collembola did not affect total N uptake of T. repens; however, the presence of both increased ¹⁵N uptake by T. repens and L. perenne. Earthworms increased shoot biomass of T. repens 1.11-fold and that of L. perenne 2.50 fold. Biomass of L. perenne was at a maximum in the presence of earthworms, collembola and with litter concentrated in a single aggregate. Shoot biomass of T. repens increased in the presence of collembola, with L. perenne generally responding opposingly. The results indicate that the composition of the decomposer community and the distribution of OM in soil affect plant competition and therefore plant community composition.     

Decomposition processes under Bt (Bacillus thuringiensis) maize: Results of a multi-site experiment

The effects of maize expressing the Bacillus thuringiensis Cry1Ab protein (Bt maize) on decomposition processes under three different European climatic conditions were assessed in the field. Farming practices using Bt maize were compared with conventional farming practices using near-isogenic non-Bt maize lines under realistic agricultural practices. The litter-bag method was used to study litter decomposition and nitrogen mineralization dynamics of wheat straw. After 4 months incubation in the field, decomposition and mineralization were mainly influenced by climatic conditions with no negative effect of the Bt toxin on decomposition processes.     

Selective digestion of the proteinaceous component of humic substances by the geophagous earthworms Metaphire guillelmi and Amynthas corrugatus

Humic substances play a key role in the global carbon cycling and the sequestration of micropollutants in soil. The transformation of these substances by earthworms, the dominant soil macroinvertebrates of many terrestrial ecosystems, and the mechanisms involved are still obscure. We prepared two chemically identical humic model compounds that were specifically ¹⁴C-labeled either in the aromatic or the proteinaceous component, and added them to soil incubated with the geophagous earthworm species Metaphire guillelmi (anecic) and Amynthas corrugatus (endogeic). In the absence of the earthworms, both the aromatic and the proteinaceous components were mineralized at similarly low rates (5−8% after 9 days of incubation). In the presence of the earthworms, mineralization rate of the proteinaceous component was strongly stimulated (2-fold by M. guillelmi and 1.4-fold by A. corrugatus). The mineralization rate of the aromatic component was (slightly) stimulated (1.2-fold; P < 0.05) only by A. corrugatus. In all cases, the stimulated mineralization was accompanied by a transformation of radiolabeled humic acids to fulvic acids within the earthworm guts and by an incorporation of radiolabel into the earthworm tissues. Digestion of the proteinaceous component of humic acids by the earthworms was corroborated also by a decrease of extractable humic acids in fresh cast and a stimulated mineralization of soil nitrogen; in the case of M. guillelmi, the fresh cast contained sixfold more NH₄⁺ than the non-ingested soil. Our study provides direct evidence for the selective digestion of humic components by earthworms. Considering the ubiquity of geophagous earthworms and their large biomass, the alteration of the chemical structure of humic substances by the earthworms through their selective digestion of peptidic components may have significant impacts on the stability of humic substances and the bioavailability of micropollutants in soil.