Evolution of N-balance with qualitative expert evaluation approach.

Pollution of rivers by nitrate is a major issue. Many land use units are considered as net nitrate producers when the input dominates the uptake (e.g. agricultural areas), or in the opposite, net consumers (e.g. wetlands), but the role of their spatial organization and temporal dynamics together across the watershed is unclear. Here, we used a Nitrate-related Ecological Functions (NEF) concept, together with an expert-based analysis in a Geographical Information System, to investigate the role of two opposite landscape types in the nitrate regulation across the Garonne river watershed (France). At any point in a watershed, there is nitrate production (NP) and nitrate removal (NR). The nitrate net balance (NNB) between NP and NR functions can be neutral (NB, Neutral Balance) when nitrate fluxes balance over space and time. The first landscape type, called Actual, was obtained using a set of 7 actual environmental variables, as land cover types, soil organic matter content and wetlands presence. The second landscape type, called Natural, described a non-anthropized landscape, using the same layer types as the Actual landscape. Potentials in NP and NR for each class in each map layer were rated by a set of experts according to their scientific knowledge. NP, NR and by difference, NNB maps were obtained, overlaid and compared to provide an evaluation of the potential for each landscape. In both landscapes, NNB were largely balanced (Actual = 48% and Natural = 67%). In the Actual landscape, NNB were secondly dominated by an imbalance toward NP (43%) and in the Natural landscape secondly imbalanced toward NR (32%). We constructed 'maps of disagreement' between both landscapes to provide a spatially explicit assessment of NNB evolution caused by changing land cover. We found that 67% of the agricultural areas and 60% of the artificial areas of the watershed had been subjected to a loss of nitrate ecological functions from Natural to Actual landscapes. Some management practices able to modify these factors may improve ecological functions and diminish the NEF disagreement of the watershed.

Anaerobic Digestion Alters Copper and Zinc Speciation.

Anaerobic digestion is a widely used organic waste treatment process. However, little is known on how it could alter the speciation of contaminants in organic waste. This study was focused on determining the influence of anaerobic digestion on the speciation of copper and zinc, two metals that generally occur at high concentration in organic waste. Copper and zinc speciation was investigated by X-ray absorption spectroscopy in four different raw organic wastes (predigestion) and their digested counterparts (postdigestion, i.e., digestates). The results highlighted an increase in the digestates of the proportion of amorphous or nanostructured copper sulfides as well as amorphous or nanostructured zinc sulfides and zinc phosphate as compared to raw waste. We therefore suggest that the environmental fate of these elements would be different when spreading either digestates or raw waste on cropland.

Soil genotoxicity assessment--results of an interlaboratory study on the Vicia micronucleus assay in the context of ISO standardization.

The Vicia micronucleus assay was standardized in an international protocol, ISO 29200, "Assessment of genotoxic effects on higher plants-Vicia faba micronucleus test," for soil or soil materials (e.g., compost, sludge, sediment, waste, and fertilizing materials). The aim of this interlaboratory study on the Vicia micronucleus assay was to investigate the robustness of this in vivo assay in terms of its applicability in different countries where each participant were asked to use their own seeds and reference soil, in agreement with the ISO 29200 standard. The ISO 29200 standard protocol was adopted for this study, and seven laboratories from three countries (France, Italy, and Brazil) participated in the study. Negative and positive controls were correctly evaluated by 100 % of the participants. In the solid-phase test, the micronucleus frequency (number of micronuclei/1,000 cells) varied from 0.0 to 1.8 for the negative control (i.e., Hoagland's solution) and from 5.8 to 85.7 for the positive control (i.e., maleic hydrazide), while these values varied from 0.0 to 1.7 for the negative control and from 14.3 to 97.7 for the positive control in the case of liquid-phase test. The variability in the data obtained does not adversely affect the robustness of the protocol assessed, on the condition that the methodology described in the standard ISO 29200 is strictly respected. Thus, the Vicia micronucleus test (ISO 29200) is appropriate for complementing prokaryotic or in vitro tests cited in legislation related to risk assessment of genotoxicity potential.

New direct contact approach to evaluate soil genotoxicity using the Vicia faba micronucleus test.

A method to assess micronucleus (MN) induction in Vicia faba roots by direct contact exposure to a solid matrix was developed. The procedure comprised a 5-d germination period, as in the well-known method using aqueous extracts. However, the seeds were here sown directly into the test soil whereas a culture period is necessary before exposing seedlings to a liquid medium. One soil under forest and two contaminated soils from areas affected by industrial installations and a coke works were used. Three durations of direct exposure were tested: 2, 5 and 7 d. The optimal duration was evaluated at 2 d to observe maximal MN induction without observing toxicity symptoms. The methodology using aqueous extracts was applied to the same three soils: MN frequency was higher than in the direct contact assay but the ratios of MN frequencies from tested soils in comparison to the negative control were lower. However, for each soil, both the direct contact method and the aqueous extract exposure led to the same risk assessment diagnosis. The evaluation of a concentration range of a polycyclic aromatic hydrocarbons (PAH)-contaminated soil showed a dose-dependent MN frequency when the seeds were allowed to germinate before sowing in the soil: the soil genotoxicity was the highest at intermediate doses. The direct contact method was found to be rapid, sensitive and well suited to the evaluation of soil quality.

Bioavailability of Cu and Zn in raw and anaerobically digested pig slurry.

The impact of anaerobic digestion on the bioavailability of copper and zinc from pig slurry was assessed. Both chemical and biological approaches were used independently on raw slurry (RS) and anaerobically digested pig slurry (DS). This work, using ultracentrifugation pellets from the same pig slurry before and after an anaerobic treatment, confirmed that Cu and Zn behave differently in terms of bioavailability, and contrasting results were obtained by chemical and biological assessments. A chemical approach combined a preliminary study of the pH effect on particulate/dissolved metal partitioning, sequential extraction, and biochemical fractionation. This approach tended to show a lower mobility of metals from digested slurry (DS). A biological approach was carried out with Zea mays and Vicia faba to study Cu and Zn uptake in soil amended with RS or DS. This assay could not differentiate the two slurries.

Potential role of NADPH-oxidase in early steps of lead-induced oxidative burst in Vicia faba roots.

The mechanism of oxidative burst induced by lead in Vicia faba excised roots was investigated by luminol-dependent chemiluminescence. Results showed that lead triggered a rapid and dose-dependent increase in chemiluminescence production. In this study, specific inhibitors of putative reactive oxygen species (ROS) sources were used to determine the mechanism of lead-induced ROS generation. This generation was sensitive to dephenylene iodonium (DPI), quinacrine and imidazole, some inhibitors of the NADPH-oxidase and not inhibited by other putative ROS sources inhibitors. Data reported in this work clearly demonstrated the pivotal role of NADPH-oxidase-like enzyme in early steps of lead-induced oxidative burst. To investigate the respective implication of calmodulin and protein kinase (PK) in lead-induced NADPH-oxidase activation, excised roots were treated with the calmodulin inhibitor W7 or with the PK inhibitor staurosporine. The chemiluminescence generation inhibition by these inhibitors illustrated the role of PK in lead-induced NADPH-oxidase activation and revealed a calmodulin-dependent step. Using the calcium entry blocker La(3+) or different concentrations of calcium in the extra-cellular medium, our data highlighted the implication of Ca(2+) channel in lead-induced oxidative burst.

Particle size and metal distributions in anaerobically digested pig slurry.

Particle size distribution and trace element patterns were studied in a full-scale anaerobic digestion plant treating pig slurry. Mass balance was established for major (N, P, K, Ca, Fe, Mg and S) and minor (Al, Cu, Mn and Zn) elements. Most of the elements were conserved through the process but part of the P, Ca, Mg and Mn was deposited as crystals lining the digester. In the dry matter of the slurry, Cu and Zn occurred at between 170 and 2600 mg kg(-1) due to pig diet supplements. Analyses of particle size distributions in raw and digested slurries showed a general shift in distribution towards larger sizes due to degradation of small and easily degradable particles as well as formation of large microbial filaments. Graded sieving of digested slurry showed metals to be mainly present on 3-25 microm particles. Less than 2% Cu and Zn was removed by passage through a 250 microm rotary screen.