Copper toxicity on Eisenia fetida in a vineyard soil: a combined study with standard tests, genotoxicity assessment and gut metagenomic analysis.

Copper (Cu) toxicity is a pressing concern for several soils, especially in organic viticulture. The objective of this work was to assess Cu toxicity on the non-target organism Eisenia fetida, employing both traditional and novel tools for early identification of Cu-induced damages. In addition to traditional tests like avoidance and reproductive toxicity experiments, other tests such as the single cell gel electrophoresis (SCGE) and gut microbiome analysis were evaluated to identify early and more sensitive pollution biomarkers. Four sub-lethal Cu concentrations were studied, and the results showed strong dose-dependent responses by the earthworm avoidance test and the exceeding of habitat threshold limit at the higher Cu doses. An inverse proportionality was observed between reproductive output and soil Cu concentration. Bioaccumulation was not detected in earthworms; soil concentrations of potentially bioavailable Cu were not affected by E. fetida presence or by time. On the contrary, the SCGE test revealed dose-dependent genotoxicity for the 'tail length' parameter already at the second day of Cu exposition. Gut microbiome analysis a modulation of microbial composition, with the most aboundant families being Pectobateriaceae, Comamonadaceae and Microscillaceae. Bacillaceae increased over time and showed adaptability to copper up to 165 mg/kg, while at the highest dose even the sensitive Acetobacteriaceae family was affected. The research provided new insights into the ecotoxicity of Cu sub-lethal doses highlighting both alterations at earthworms' cellular level and changes in their gut microbiota.

Copper Monitoring in Vineyard Soils of Central Italy Subjected to Three Antifungal Treatments, and Effects of Sub-Lethal Copper Doses on the Earthworm Eisenia fetida.

The extensive employment of copper-based fungicides has increased copper concentration in vineyard soils. The present study's objectives were to monitor copper concentration in two vineyard soils during two cropping seasons and study the ecotoxicological effects on the earthworm Eisenia fetida. Total, soluble, and bioavailable copper fractions were measured at the end of two cropping seasons and different depths in two vineyards of central Italy, characterised by three anticryptogamic control methods: copper compounds, chitosan, and combined treatments of them. A laboratory experiment to assess the effects on Eisenia fetida was conducted with soil samples collected in the vineyards with a mean copper concentration of 60 mg/kg and two higher concentrations of 90 and 150 mg/kg. Results showed low levels of total copper concentration in the first 20 cm of soils, regardless of antifungal treatment, highlighting prudent management of the vineyards under study, but the soluble fractions showed a significant increase in all samples during the two cropping seasons. At the dose of 150 mg/kg, earthworms suffer during the first two days, showing weight loss and DNA damage, but they are able to recover until day 28, showing no permanent harm at this copper concentration in soil.

Sub-Lethal Effects of Pesticides on the DNA of Soil Organisms as Early Ecotoxicological Biomarkers.

This review describes the researches performed in the last years to assess the impact of pesticide sub-lethal doses on soil microorganisms and non-target organisms in agricultural soil ecosystems. The overview was developed through the careful description and a critical analysis of three methodologies based on culture-independent approaches involving DNA extraction and sequencing (denaturing gradient gel electrophoresis, DGGE; next-generation sequencing, NGS) to characterize the microbial population and DNA damage assessment (comet assay) to determine the effect on soil invertebrates. The examination of the related published articles showed a continuous improvement of the possibility to detect the detrimental effect of the pesticides on soil microorganisms and non-target organisms at sub-lethal doses, i.e., doses which have no lethal effect on the organisms. Considering the overall critical discussion on microbial soil monitoring in the function of pesticide treatments, we can confirm the usefulness of PCR-DGGE as a screening technique to assess the genetic diversity of microbial communities. Nowadays, DGGE remains a preliminary technique to highlight rapidly the main differences in microbial community composition, which is able to give further information if coupled with culture-dependent microbiological approaches, while thorough assessments must be gained by high-throughput techniques such as NGS. The comet assay represents an elective technique for assessing genotoxicity in environmental biomonitoring, being mature after decades of implementation and widely used worldwide for its direct, simple, and affordable implementation. Nonetheless, in order to promote the consistency and reliability of results, regulatory bodies should provide guidelines on the optimal use of this tool, strongly indicating the most reliable indicators of DNA damage. This review may help the European Regulation Authority in deriving new ecotoxicological endpoints to be included in the Registration Procedure of new pesticides.

Phytoremediation Potential of Crop Plants in Countering Nickel Contamination in Carbonation Lime Coming from the Sugar Industry.

The phytoremediation potential of four crop species cultivated on carbonation lime coming from the sugar industry with water-soluble nickel (Ni) exceeding the Italian legal limit of 10 µg L-1 was assessed. Two autumn-winter species (spinach and canola) were tested with and without the addition of bentonite in a greenhouse experiment in order to overcome prolonged unfavourable weather conditions. Two spring-summer species (sunflower and sorghum) were grown in outdoor boxes. Plant species were selected among crops of interest for phytoremediation and their rotation throughout the year enable to maintain a permanent vegetation cover. Nickel concentration in different plant tissues and the concentrations of soluble and bioavailable Ni in lime were measured. In the greenhouse study, soluble Ni decreased below the legal limit in all the tests, and the combined effect of bentonite and plants reduced Ni in lime mainly in the bioavailable fraction. Spinach and sunflower emerged to be more suitable for phytoextraction than canola and sorghum, because of the higher concentration of the metal in the epigeal portions. The results from the outdoor experiment highlighted that sorghum has a good phytostabilisation potential since its ability to accumulate Ni mainly at the root level and to attract a significant amount of bioavailable Ni in the rhizosphere. This study arose from a real scenario of environmental contamination and investigated the potential of different approaches on the bioremediation of a specific industrial waste product.

Zeolite and bentonite as nickel sequestrants in carbonation lime coming from the sugar industry.

A laboratory trial was performed to test the sequestration capacity of two minerals (bentonite and zeolite) at three initial concentrations (2.5, 5 and 10%) in order to counter water-soluble nickel (Ni) exceeding the Italian legal limit (10 µg L-1) in carbonation lime disposed of in a field and sampled for an 85-day lab study. The results show a noticeable reduction in water-soluble and bioavailable Ni in lime after the addition of sequestrants, especially at the dose of 5% bentonite or zeolite, thereby indicating a "ceiling effect" of the sequestrant, i.e., an increasing dose could reduce the adsorption capacity and be less effective. The alkaline pH and the presence of organic matter could be the main factors affecting the good performance of sequestrant addition, causing an increase in the negative charge of the organic and mineral colloids and the formation of unavailable Ni precipitates. The 85-day experiment seems to be sufficient to reach an adsorption equilibrium for water-soluble nickel, while for the bioavailable form a longer period appears to be necessary.

Iron Speciation in Organic Matter Fractions Isolated from Soils Amended with Biochar and Organic Fertilizers.

The role and distribution of iron (Fe) species in physical soil fractions have received remarkably little attention in field-scale systems. Here, we identify and quantify the Fe phases into two fractions (fine sand, FSa, and fine silt and clay, FSi + Cl), isolated from an agricultural soil unamended and amended with different organic materials, by Fe K-edge extended X-ray absorption fine structure (EXAFS) spectroscopy. The linear combination fitting and wavelet transform of EXAFS data revealed noticeable differences between unamended FSa and FSi + Cl fractions. Specifically, the FSi + Cl fraction was mainly characterized by ferrihydrite (48%) and Fe(III)-soil organic matter (SOM) complexes (37%), whereas in the FSa fraction, ferrihydrite still represented a major phase (44%), with a lower contribution from Fe(III)-SOM (18%). In the FSa fraction, the addition of the organic amendments resulted in an increase of Fe-SOM complexes (31-35%) and a decrease of ferrihydrite (28-29%). By contrast, in the amended FSi + Cl fractions, the added organic matter led to negligible changes in percent ferrihydrite. Therefore, regardless of the amendment type, the addition of organic matter to soil increased the capability of the coarse fraction (FSa) to stabilize organic carbon, thus pointing out that the role of FSa in carbon sequestration in agricultural soils at a global scale may be overlooked.

Evaluation of gene expression of different molecular biomarkers of stress response as an effect of copper exposure on the earthworm EIsenia Andrei.

The paper reports the results of a laboratory test on the bioaccumulation and toxicological effects of sub-lethal soil concentration of copper, a widely used fungicide in organic farming, on DNA damage, a critical marker increasingly used in ecotoxicology in the earthworm Eisenia andrei. In the same experimental setting we evaluated gene expression of classical biomarker of stress induced by xenobiotic. [Heat Shock Protein 70 (HSP70) and Metallothionein (MET)], as well as genes coding for enzymes involved in detoxification of reactive oxygen species [Superoxide dismutase (SOD) and catalase (CAT)]. Additionally, expression of genes involved in the immune response were investigated: a Toll-like receptor (TLR), a receptor with cytolytic activity named Cytolytic Factor (CCF) and two antimicrobial peptides, fetidin (FET) and lysenin (LYS). Results showed significant time-dependent bioaccumulation of Cu and DNA damage at concentrations remarkably lower than those found in most agricultural soils worldwide. MET was increased as was FET and TLR. The present work gives new insights into the mechanisms of sub-lethal toxicity of copper as an environmental pollutant and in the identification of novel sub-lethal biomarkers of cellular response to the stressor such as immune response genes.

Density-based fractionation of soil organic matter: effects of heavy liquid and heavy fraction washing.

Physical fractionation methods used in soil organic matter (SOM) research commonly include density-based procedures with heavy liquids to separate SOM pools with varying turnover rates and functions. Once separated, the heavy SOM pools are often thoroughly rinsed with water to wash off any residues of the heavy liquids. Using four soils with contrasting properties, we investigated the effects of using either sodium polytungstate (SPT) or sodium iodide (NaI), two of the most commonly used heavy liquids, on the distribution of organic carbon (C) and total nitrogen (N) in free light, intra-aggregate light, and mineral-associated heavy SOM pools isolated by a common fractionation scheme. We also determined the effects of washing the mineral-associated heavy SOM fractions on the recovery of organic C and total N after separation. Because of its smaller viscosity compared to that of NaI, SPT consistently yielded greater intra-aggregate and smaller mineral-associated soil organic C contents. We also confirm that some commercial SPT products, such as the one used here, can contaminate organo-mineral heavy pools with N during density-based fractionation procedures. We do not recommend the repeated washing of heavy fractions separated with Na-based heavy liquids, as this can mobilize SOM.

The role of Fe(III) in soil organic matter stabilization in two size fractions having opposite features.

Soil organic matter (SOM) protection, stability and long-term accumulation are controlled by several factors, including sorption onto mineral surfaces. Iron (Fe) has been suggested as a key regulator of SOM stability, both in acidic conditions, where Fe(III) is soluble, and in near-neutral pH environments, where it precipitates as Fe(III) (hydr)oxides. The present study aimed to probe, by sorption/desorption experiments in which Fe was added to the system, the mechanisms controlling Fe(III)-mediated organic carbon (C) stabilization; fine silt and clay (FSi + Cl) and fine sand (FSa) SOM fractions of three soils under different land uses were tested. Fe(III) addition caused a decrease in the organic C remaining in solution after reaction, indicating an Fe-mediated organic C stabilization effect. This effect was two times larger for FSa than for FSi + Cl, the former fraction being characterized by both low specific surface area and high organic C content. The organic C retained in the solid phase after Fe-mediated stabilization has relatively low sensitivity to desorption. Moreover, attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy indicated that Fe-mediated organic C stabilization can be mainly ascribed to the formation of complexes between carbohydrate OH functional groups and Fe oxides. These results demonstrate that the binding of labile SOM compounds to Fe(III) contributes to its preservation, and that the mechanisms involved (flocculation vs. coating) depend on the size fractions.

DNA damage in different Eisenia andrei coelomocytes sub-populations after in vitro exposure to hydrogen peroxide.

Earthworms play an essential role in providing soil fertility and may represent an important soil contamination bio-indicator. They are able to ingest soil particles, adsorb substances throughout the intestinal epithelium into the coelomic cavity, where chemicals can come in direct contact with coelomic fluid. Earthworm coelomic fluid shelters leucocytes (coelomocytes) that differ significantly both structurally and functionally. Cellular variability could lead to different susceptibility towards contaminants possibly present in soil ecosystem. In order to define population specific dose response to chemicals and to identify a homogeneous cell population to be used as a relevant biomarker, we investigated different coelomocytes subpopulation, obtained by Percoll density gradient centrifugation (5-35 %), exposed ex vivo to H2O2 in the range of concentration 15-120 µM. DNA damage levels were assessed by the comet assay on unseparated coelomocytes and on three enriched cellular fractions (light, medium and heavy density subpopulations). All tested samples showed a dose-response genotoxic effect following H2O2 exposure. Moreover, light density sub-population appeared more susceptible to oxidative insult highlighted by a significant increase in DNA damage indexes at lower concentrations of H2O2. Present data suggested that in these experimental condition coelomocytes light fraction may represent a more sensitive biomarker of genotoxic insult.

The dissipation of three fungicides in a biobed organic substrate and their impact on the structure and activity of the microbial community.

Biopurification systems (BPS) have been introduced to minimise the risk for point source contamination of natural water resources by pesticides. Their depuration efficiency relies mostly on the high biodegradation of their packing substrate (biomixture). Despite that, little is known regarding the interactions between biomixture microflora and pesticides, especially fungicides which are expected to have a higher impact on the microbial community. This study reports the dissipation of the fungicides azoxystrobin (AZX), fludioxonil (FL) and penconazole (PC), commonly used in vineyards, in a biomixture composed of pruning residues and straw used in vineyard BPS. The impact of fungicides on the microbial community was also studied via microbial biomass carbon, basal respiration and phospholipid fatty acid analysis. AZX dissipated faster (t1/2 = 30.1 days) than PC (t1/2 = 99.0 days) and FL (t1/2 = 115.5 days). Fungicides differently affected the microbial community. PC showed the highest adverse effect on both the size and the activity of the biomixture microflora. A significant change in the structure of the microbial community was noted for PC and FL, and it was attributed to a rapid inhibition of the fungal fraction while bacteria showed a delayed response which was attributed to indirect effects by the late proliferation of fungi. All effects observed were transitory and a full recovery of microbial indices was observed 60 days post-application. Overall, no clear link between pesticide persistence and microbial responses was observed stressing the complex nature of interactions between pesticides in microflora in BPS.

Losses and dissipation of penconazole in vineyard soil as affected by mid-row management system.

A field experiment was performed with the aim to assess the amount of penconazole losses during field application and the spatial variability of penconazole concentration in a vineyard soil, under two different management techniques (tilled and grass covered). The field dissipation of penconazole under the two techniques was also followed for 114 days, highlighting the effects of spatial variability of the initial concentration and of the grass covering in terms of the different soil metabolic activity. Data found show that a high percentage of penconazole, from 42.5 % to 67.43 %, can reach the soil during the treatments, despite the fact that penconazole is applied to the foliage. The high values of the coefficient of variation for data within rows show considerable variability in all applications, ranging from 30 to 65 in the first application, from 35 to 79 in the second and from 36 to 75 in the third. Since the applications of penconazole occurred under almost same climatic conditions in terms of wind speed and direction, the high variability of concentration of penconazole found within mid-rows was attributed to the uneven slope of the vineyard causing an irregular speed of the sprayer equipment. However, least significant differences of the mean values did not end in a significant difference of penconazole concentration among mid-rows for all applications, indicating that the variability between rows does not contribute to the overall variability. The calculated half-life values for penconazole in tilled soil were 62.4 days for tilled and 33.0 for grassed soil, highlighting the tendency of penconazole to faster dissipate in grassed than in tilled soil. Grassed soil was characterised by a higher metabolic activity in terms of microbial biomass carbon content, basal respiration and total hydrolytic activity, than tilled soil. A significant correlation between penconazole dissipation and total hydrolytic activity, was found. The faster dissipation of penconazole in grassed soil was attributed to the higher amount of the total hydrolytic activity which, in grassed soil, was 1.5 fold-higher than in tilled soil.

Fungicides degradation in an organic biomixture: impact on microbial diversity.

Biological systems are being developed all over EU countries to protect water-bodies from pesticide contamination at farm level. A laboratory experiment was carried out to test the efficiency of a mixture of compost and straw in bio-degrading different mixtures of fungicides usually applied in vineyards. At the same time the effects of fungicide applications on microbial community of biomixture were also evaluated. Results showed that the biomixture had a good capability of degrading pesticides. Indeed, at the end of the experiment (112 days), the concentration of most of the pesticides was close to complete degradation. Denaturing gradient gel electrophoresis (DGGE) analysis showed an evident modification of microbial diversity after the addition of fungicides. However, at the end of degradation process, no significant changes in the composition of microbial community were seen. In this specific substrate used in the biomixture, yeast flora and ascomycete filamentous fungi seemed to be involved in the degradation activity.

Degradation of isoproturon and bentazone in peat- and compost-based biomixtures.

BACKGROUND: The composition and properties of a biomixture used in a biobed are decisive for pesticide sorption and degradation. This study was performed to investigate the capability of compost-based substrates in mixtures with citrus peel and vine branch straw and peat-based substrates in mixtures with soil and vine branch straw at different levels in order to degrade isoproturon and bentazone. RESULTS: Dissipation and mineralisation rates of both pesticides were determined, and metabolic activity was followed as respiration. Compost-based substrates showed faster pesticide dissipation in the presence of lignocellulosic materials, as in garden compost and vine branch straw. The increasing content of vine branch straw in peat-based substrates does not seem to affect dissipation of the parent compounds. Low mineralisation rate was observed in all treatments. CONCLUSION: Higher pesticide degradation was observed in the lignocellulosic substrates, probably because of the development of lignin-degrading microorganisms which have shown to be robust and are able to degrade recalcitrant pesticides. Copyright © 2010 Society of Chemical Industry.

Effect of pesticide inoculation, duration of composting, and degradation time on the content of compost fatty acids, quantified using two methods.

Compost biobeds can promote biodegradation of pesticides. The microbial community structure changes during the composting process, and simple methods can potentially be used to follow these changes. In this study the microbial identification (MIDI) and ester-linked (EL) procedures were used to determine the composition of fatty acid methyl esters (FAMEs) in composts aged 3 and 12 months, inoculated with 3 recalcitrant pesticides (azoxystrobin, chlorotoluron, and epoxyconazole and a coapplication of all three) after 0, 56, and 125 days of degradation. Pesticide persistence was high, and after 125 days the residue was 22 to 70% of the applied amount depending mostly on the composting age. Seventy-one FAMEs belonging to nine groups were detected. The EL method provided three times as many detections as did the MIDI method and was more sensitive for all FAME groups except alcohol. Thirty-six and five FAMEs were unique to the EL and MIDI methods, respectively. The extraction method was of importance. The EL method provided a higher number of detections for 57 FAMEs, and the MIDI method provided a higher number for 9 FAMEs, while the two methods were equal for 5 FAMEs; thus, the EL method provided a more uniform overall FAME profile. Effects of the other factors were not always clear. Inoculation with pesticide did not influence the FAME profile with the MIDI method, while it influenced cyclopropane and monounsaturated content with the EL method. Composting age and degradation time had an effect on some groups of FAMEs, and this effect was greater with the EL method. The use of some FAMEs as biomarkers to follow microbial community succession was likely influenced by the type of compost and other factors.

Retention capacity of an organic bio-mixture against different mixtures of fungicides used in vineyards.

A laboratory experiment was carried out to test the efficiency of a bio-mixture made up of pruning residues at two (PR2) and five (PR5) years of composting and wheat straw (STW) in the biological cleaning of water contaminated by different mixtures of fungicides usually employed in vineyards. The experiment was conducted and reproduced at a scale of 1:100 of operating field conditions. Commercial formulates of penconazole (PC), (RS)-1-[2-(2,4-dichlorophenyl)pentyl]-1H-1,2,4-triazole), dimetomorph (DM), (EZ)-4-[3-(4-chlorophenyl)-3-(3,4-dimethoxyphenyl)acryloyl]morpholine), azoxystrobin (AZ), (methyl (E)-2-{2-[6-(2-cyanophenoxy)pyrimidin-4-yloxy]phenyl}-3-methoxyacrylate), iprovalicarb (IP), (isopropyl 2-methyl-1-[(RS)-1-p-tolylethyl]carbamoyl-(S)-propylcarbamate), metalaxyl (MX), (methyl N-(methoxyacetyl)-N-(2,6-xylyl)-DL-alaninate), fludioxonil (FL), (4-(2,2-difluoro-1,3-benzodioxol-4-yl)-1H-pyrrole-3-carbonitrile) and cyprodinil (CY), (4-cyclopropyl-6-methyl-N-phenylpyrimidin-2-amine) were mixed in water and discharged into the bio-mixture following the time schedule of the treatments carried out in the grapevine in real field conditions. At each treatment, contaminated water with fungicides was circulated repeatedly through the bio-mixture to enhance the sorption of fungicides. In fact, it retained them between 98-100% with the exception of MX of which it was able to retain only 90.5%. The adsorption/desorption experiment showed that repeated circulation of water, instead of enhancing MX retention, can easily remove about 30% of MX already adsorbed by the bio-mixture. This finding suggests that water contaminated by very mobile pesticides should be discharged at the end of field treatments without re-circulating the water in order to avoid the release of pesticides weakly adsorbed on the bio-mixture.

The effect of crop rotation on pesticide leaching in a regional pesticide risk assessment.

New modeling approaches that include the use of GIS are under development in order to allow a more realistic assessment of environmental contamination by pesticides. This paper reports a regional GIS-based risk assessment using a software tool able to simulate complex and real crop rotations at the regional scale. A single pesticide leaching assessment has been done. The mean annual pesticide concentration in leachate has been analyzed using both stochastic and deterministic approaches. The outputs of these simulations were mapped over the sampling locations of the regional pesticide monitoring program, demonstrating that GIS-based risk assessment can be used to establish new monitoring programs. A multiple pesticide leaching assessment for analyzing the risk related to pest control strategies in six different maize-based rotations has been carried out. Additive toxic units approach has been used. Crop rotation allows to mediate the risk related to pesticide use because forces the use of different compounds with different fate and toxicology properties.

The effect of initial concentration, co-application and repeated applications on pesticide degradation in a biobed mixture.

A 180 d laboratory experiment was conducted to investigate the degradation rates of chlorpyrifos (10 and 50 mg kg(-1)) and metalaxyl (100 mg kg(-1)) separately and co-applied in a biomix constituted by topsoil, vine-branches and urban-waste-garden compost. The effect of repeated application of metalaxyl was also investigated. Microbial biomass-C (MBC) content and metabolic quotient (qCO(2)) were measured to evaluate changes in microbial biomass size and activity induced by the presence of the two pesticides. Degradation rate decreased with increasing concentration of chlorpyrifos in all treatments. Metalaxyl half-life was significantly reduced in co-application with chlorpyrifos indicating a synergic interaction between the two pesticides in favour of enhanced degradation rate for metalaxyl but not for chlorpyrifos. Furthermore, repeated application resulted in a sharp reduction of metalaxyl half-life from 37 d after first application to 4 d after third application. MBC content was negatively influenced by the addition of pesticides but it started to recover immediately, in both separate and co-applied treatments, reaching the control value when pesticide residues were about 50% of the initial concentration. The qCO(2) reached a steady-state after about 20 d in separately applied and 40 d in co-applied treatments, indicating a tendency to arrive at a new metabolic equilibrium. In conclusion, the biomix tested has been shown to degrade pesticides relatively fast and to have a microbial community that is varied enough to allow selection of those microorganisms able to degrade metalaxyl and chlorpyrifos.

Measures to reduce pesticide spray drift in a small aquatic ecosystem in vineyard estate.

A field experiment is reported to ascertain the drift of two pesticides (chlorpyrifos and metalaxyl) in a vineyard in Italian climatic conditions and the effect of mitigation measures, such as buffer zones and tree rows, on pesticide drift contamination in a small aquatic system located inside the field. Results indicated that, in typical Italian agricultural conditions, spray drift in vineyards occurs at a distance of more than 24 m and adequate buffer zones are required to protect surface water bodies from direct contamination. The presence of tree rows in front of the water body inside the agricultural field, against the main wind direction, resulted in a very high reduction of the spray drift and of the ecotoxicological risk for aquatic ecosystem. In addition, a comparison between the data obtained in the experiment and the Drift Calculator procedure showed that the model failed when the procedure is used for short distances. However, concordance was found in terms of maximum drift distances.

Adaptation of the biobed composition for chlorpyrifos degradation to Southern Europe conditions.

Biobeds developed in Sweden bind and degrade pesticides from point sources. The objective of this work was to adapt the biobed to Italian operating conditions, for example, to identify organic materials as effective as those in the original Swedish composition. The capacity of urban and garden composts alone or mixed with citrus peel or straw to degrade chlorpyrifos and its metabolite TCP was compared to the typical Swedish biomix consisting of straw, peat, and soil. A tendency for higher 14C-chlorpyrifos mineralization and lower TCP levels was observed in the biomixes with garden compost alone or amended with straw. In a second trial, a high correlation of lower TCP with increasing levels of straw in typical Swedish biomixes was observed. Straw stimulates production of lignin-degrading enzymes such as manganese peroxidase (MnP), and further trials with pure MnP showed that this enzyme degrades TCP. Materials with an active lignin-degrading microflora are a prerequisite for effective dissipation of chlorpyrifos and non-accumulation of TCP. Thus, lignocellulosic materials as straw and garden composts should be present in biomixes to be used under Italian conditions.