Novel phages of Pseudomonas syringae unveil numerous potential auxiliary metabolic genes.

Relatively few phages that infect plant pathogens have been isolated and investigated. The Pseudomonas syringae species complex is present in various environments, including plants. It can cause major crop diseases, such as bacterial canker on apricot trees. This study presents a collection of 25 unique phage genomes that infect P. syringae. These phages were isolated from apricot orchards with bacterial canker symptoms after enrichment with 21 strains of P. syringae. This collection comprises mostly virulent phages, with only three being temperate. They belong to 14 genera, 11 of which are newly discovered, and 18 new species, revealing great genetic diversity within this collection. Novel DNA packaging systems have been identified bioinformatically in one of the new phage species, but experimental confirmation is required to define the precise mechanism. Additionally, many phage genomes contain numerous potential auxiliary metabolic genes with diversified putative functions. At least three phages encode genes involved in bacterial tellurite resistance, a toxic metalloid. This suggests that viruses could play a role in bacterial stress tolerance. This research emphasizes the significance of continuing the search for new phages in the agricultural ecosystem to unravel novel ecological diversity and new gene functions. This work contributes to the foundation for future fundamental and applied research on phages infecting phytopathogenic bacteria.

The use of copper as plant protection product contributes to environmental contamination and resulting impacts on terrestrial and aquatic biodiversity and ecosystem functions.

Copper-based plant protection products (PPPs) are widely used in both conventional and organic farming, and to a lesser extent for non-agricultural maintenance of gardens, greenspaces, and infrastructures. The use of copper PPPs adds to environmental contamination by this trace element. This paper aims to review the contribution of these PPPs to the contamination of soils and waters by copper in the context of France (which can be extrapolated to most of the European countries), and the resulting impacts on terrestrial and aquatic biodiversity, as well as on ecosystem functions. It was produced in the framework of a collective scientific assessment on the impacts of PPPs on biodiversity and ecosystem services in France. Current science shows that copper, which persists in soils, can partially transfer to adjacent aquatic environments (surface water and sediment) and ultimately to the marine environment. This widespread contamination impacts biodiversity and ecosystem functions, chiefly through its effects on phototrophic and heterotrophic microbial communities, and terrestrial and aquatic invertebrates. Its effects on other biological groups and biotic interactions remain relatively under-documented.

Improved assessment of the impacts of plant protection products on certain soil ecosystem services requires better consideration of terrestrial microalgae and cyanobacteria.

There is growing scientific and societal consciousness that the environmental risks and impacts of plant protection products (PPPs) cannot be properly assessed without considering ecosystem services. However, the science on this issue remains incomplete and fragmented, as recently illustrated in a collective scientific assessment that pointed out the limited knowledge on the risks and impacts of PPPs on soil ecosystem services, which are clearly overlooked. Beside soil ecosystem services, certain key players involved in these services are largely overlooked in the scientific literature on the risks and impacts of PPPs, namely soil microbial photosynthetic communities. Here, we followed the principles of evidence-based logic chain approaches to show the importance of considering these microorganisms when studying the impacts of PPPs on certain services provided by soil ecosystems, with a focus on regulating and maintenance services that play a role in the regulation of baseline flows and extreme events. Terrestrial microalgae and cyanobacteria are ubiquitous photosynthetic microorganisms that, together with other soil micro- and macro-organisms, play key roles in the ecosystem functions that underpin these ecosystem services. There is an extensive literature on the ecotoxicological effects of PPPs on different organisms including soil microorganisms, but studies concerning soil microbial photosynthetic communities are very scarce. However, there is scientific evidence that herbicides can have both direct and indirect impacts on these microbial photosynthetic communities. Given that they play key functional roles, we argue that soil microbial photosynthetic communities warrant greater attention in efforts to assess the environmental risks and impacts of PPPs and, ultimately, help preserve or restore the regulating and maintenance services provided by soil ecosystems.

Linking ecotoxicological effects on biodiversity and ecosystem functions to impairment of ecosystem services is a challenge: an illustration with the case of plant protection products.

There is growing interest in using the ecosystem services framework for environmental risk assessments of chemicals, including plant protection products (PPPs). Although this topic is increasingly discussed in the recent scientific literature, there is still a substantial gap between most ecotoxicological studies and a solid evaluation of potential ecotoxicological consequences on ecosystem services. This was recently highlighted by a collective scientific assessment (CSA) performed by 46 scientific experts who analyzed the international science on the impacts of PPPs on biodiversity, ecosystem functions, and ecosystem services. Here, we first point out the main obstacles to better linking knowledge on the ecotoxicological effects of PPPs on biodiversity and ecological processes with ecosystem functions and services. Then, we go on to propose and discuss possible pathways for related improvements. We describe the main processes governing the relationships between biodiversity, ecological processes, and ecosystem functions in response to effects of PPP, and we define categories of ecosystem functions that could be directly linked with the ecological processes used as functional endpoints in investigations on the ecotoxicology of PPPs. We then explore perceptions on the possible links between these categories of ecosystem functions and ecosystem services among a sub-panel of the scientific experts from various fields of environmental science. We find that these direct and indirect linkages still need clarification. This paper, which reflects the difficulties faced by the multidisciplinary group of researchers involved in the CSA, suggests that the current gap between most ecotoxicological studies and a solid potential evaluation of ecotoxicological consequences on ecosystem services could be partially addressed if concepts and definitions related to ecological processes, ecosystem functions, and ecosystem services were more widely accepted and shared within the ecotoxicology community. Narrowing this gap would help harmonize and extend the science that informs decision-making and policy-making, and ultimately help to better address the trade-off between social benefits and environmental losses caused by the use of PPPs.

Main conclusions and perspectives from the collective scientific assessment of the effects of plant protection products on biodiversity and ecosystem services along the land-sea continuum in France and French overseas territories.

Preservation of biodiversity and ecosystem services is critical for sustainable development and human well-being. However, an unprecedented erosion of biodiversity is observed and the use of plant protection products (PPP) has been identified as one of its main causes. In this context, at the request of the French Ministries responsible for the Environment, for Agriculture and for Research, a panel of 46 scientific experts ran a nearly 2-year-long (2020-2022) collective scientific assessment (CSA) of international scientific knowledge relating to the impacts of PPP on biodiversity and ecosystem services. The scope of this CSA covered the terrestrial, atmospheric, freshwater, and marine environments (with the exception of groundwater) in their continuity from the site of PPP application to the ocean, in France and French overseas territories, based on international knowledge produced on or transposable to this type of context (climate, PPP used, biodiversity present, etc.). Here, we provide a brief summary of the CSA's main conclusions, which were drawn from about 4500 international publications. Our analysis finds that PPP contaminate all environmental matrices, including biota, and cause direct and indirect ecotoxicological effects that unequivocally contribute to the decline of certain biological groups and alter certain ecosystem functions and services. Levers for action to limit PPP-driven pollution and effects on environmental compartments include local measures from plot to landscape scales and regulatory improvements. However, there are still significant gaps in knowledge regarding environmental contamination by PPPs and its effect on biodiversity and ecosystem functions and services. Perspectives and research needs are proposed to address these gaps.

Soil Photosynthetic Microbial Communities Mediate Aggregate Stability: Influence of Cropping Systems and Herbicide Use in an Agricultural Soil.

Edaphic cyanobacteria and algae have been extensively studied in dryland soils because they play key roles in the formation of biological soil crusts and the stabilization of soil surfaces. Yet, in temperate agricultural crop soils, little is understood about the functional significance of indigenous photosynthetic microbial communities for various soil processes. This study investigated how indigenous soil algae and cyanobacteria affected topsoil aggregate stability in cereal cropping systems. Topsoil aggregates from conventional and organic cropping systems were incubated in microcosms under dark or photoperiodic conditions with or without a treatment with an herbicide (isoproturon). Physicochemical parameters (bound exopolysaccharides, organic carbon) and microbial parameters (esterase activity, chlorophyll a biomass, and pigment profiles) were measured for incubated aggregates. Aggregate stability were analyzed on the basis of aggregate size distribution and the mean weight diameter (MWD) index, resulting from disaggregation tests. Soil photosynthetic microbial biomass (chl a) was strongly and positively correlated with aggregate stability indicators. The development of microalgae crusts in photoperiodic conditions induced a strong increase of the largest aggregates (>2 mm), as compared to dark conditions (up to 10.6 fold and 27.1 fold, in soil from organic and conventional cropping systems, respectively). Concomitantly, the MWD significantly increased by 2.4 fold and 4.2 fold, for soil from organic and conventional cropping systems. Soil microalgae may have operated directly via biochemical mechanisms, by producing exopolymeric matrices surrounding soil aggregates (bound exopolysaccharides: 0.39-0.45 µg C g-1 soil), and via biophysical mechanisms, where filamentous living microbiota enmeshed soil aggregates. In addition, they may have acted indirectly by stimulating heterotrophic microbial communities, as revealed by the positive effect of microalgal growth on total microbial activity. The herbicide treatment negatively impacted soil microalgal community, resulting in significant decreases of the MWD of the conventional soil aggregates (up to -42% of the value in light treatment). This study underscores that indigenous edaphic algae and cyanobacteria can promote aggregate formation, by forming photosynthetic microbiotic crusts, thus improving the structural stability of topsoil, in temperate croplands. However, the herbicide uses can impair the functional abilities of algal and cyanobacterial communities in agricultural soils. ORIGINALITY/SIGNIFICANCE: Edaphic algal and cyanobacterial communities are known to form photosynthetic microbial crusts in arid soils, where they drive key ecosystem functions. Although less well characterized, such communities are also transiently abundant in temperate and mesic cropped soils. This microcosm study investigated the communities' functional significance in topsoil aggregate formation and stabilization in two temperate cropping systems. Overall, our results showed that the development of indigenous microalgal communities under our experimental conditions drove higher structural stability in topsoil aggregates in temperate cropland soils. Also, herbicide use affected photosynthetic microbial communities and consequently impaired soil aggregation.

Soil C and N statuses determine the effect of maize inoculation by plant growth-promoting rhizobacteria on nitrifying and denitrifying communities.

Maize inoculation by Azospirillum stimulates root growth, along with soil nitrogen (N) uptake and root carbon (C) exudation, thus increasing N use efficiency. However, inoculation effects on soil N-cycling microbial communities have been overlooked. We hypothesized that inoculation would (i) increase roots-nitrifiers competition for ammonium, and thus decrease nitrifier abundance; and (ii) increase roots-denitrifiers competition for nitrate and C supply to denitrifiers by root exudation, and thus limit or benefit denitrifiers depending on the resource (N or C) mostly limiting these microorganisms. We quantified (de)nitrifiers abundance and activity in the rhizosphere of inoculated and non-inoculated maize on 4 sites over 2 years, and ancillary soil variables. Inoculation effects on nitrification and nitrifiers (AOA, AOB) were not consistent between the three sampling dates. Inoculation influenced denitrifiers abundance (nirK, nirS) differently among sites. In sites with high C limitation for denitrifiers (i.e. limitation of denitrification by C > 66%), inoculation increased nirS-denitrifier abundance (up to 56%) and gross N2O production (up to 84%), likely due to increased root C exudation. Conversely, in sites with low C limitation (<47%), inoculation decreased nirS-denitrifier abundance (down to -23%) and gross N2O production (down to -18%) likely due to an increased roots-denitrifiers competition for nitrate.

Agrifood systems and the microbial safety of fresh produce: Trade-offs in the wake of increased sustainability.

Fresh produce has been a growing cause of food borne outbreaks world-wide prompting the need for safer production practices. Yet fresh produce agrifood systems are diverse and under constraints for more sustainability. We analyze how measures taken to guarantee safety interact with other objectives for sustainability, in light of the diversity of fresh produce agrifood systems. The review is based on the publications at the interface between fresh produce safety and sustainability, with sustainability defined by low environmental impacts, food and nutrition security and healthy life. The paths for more sustainable fresh produce are diverse. They include an increased use of ecosystem services to e.g. favor predators of pests, or to reduce impact of floods, to reduce soil erosion, or to purify run-off waters. In contrast, they also include production systems isolated from the environment. From a socio-economical view, sustainability may imply maintaining small tenures with a higher risk of pathogen contamination. We analyzed the consequences for produce safety by focusing on risks of contamination by water, soil, environment and live stocks. Climate change may increase the constraints and recent knowledge on interactions between produce and human pathogens may bring new solutions. Existing technologies may suffice to resolve some conflicts between ensuring safety of fresh produce and moving towards more sustainability. However, socio-economic constraints of some agri-food systems may prevent their implementation. In addition, current strategies to preserve produce safety are not adapted to systems relying on ecological principles and knowledge is lacking to develop the new risk management approaches that would be needed.

Pollution-induced community tolerance (PICT) as a tool for monitoring Lake Geneva long-term in situ ecotoxic restoration from herbicide contamination.

Chemical monitoring revealed a regular decrease in herbicide concentration in Lake Geneva since last decades that may be linked to an ecotoxic restoration of nontarget phytoplanktonic communities. The Pollution-induced community tolerance (PICT) approach was tested as a tool to monitor the ecotoxic restoration of Lake Geneva for herbicides from 1999 to 2011. We conducted monthly assessments in 1999 and in 2011 for the tolerance of the phytoplankton communities to two herbicides (atrazine and copper), using PICT bioassays. The taxonomical composition of the communities was determined on the same collecting dates. The herbicide concentration decrease during the 12 years significantly influenced the composition of communities. The PICT monitoring indicated that a significant tolerance decrease in the community to both herbicides accompanied the herbicide concentration decrease. PICT measurements for atrazine and copper also changed at the intra-annual level. These variations were mainly due to community composition shifts linked to seasonal phosphorus and temperature changes. PICT monitoring on a seasonal basis is required to monitor the mean tolerance of communities. PICT appeared to be a powerful tool that reflected the toxic effects on environmental communities and to monitor ecotoxic ecosystem restoration.

Soil microbial respiration and PICT responses to an industrial and historic lead pollution: a field study.

We performed a field investigation to study the long-term impacts of Pb soil contamination on soil microbial communities and their catabolic structure in the context of an industrial site consisting of a plot of land surrounding a secondary lead smelter. Microbial biomass, catabolic profiles, and ecotoxicological responses (PICT) were monitored on soils sampled at selected locations along 110-m transects established on the site. We confirmed the high toxicity of Pb on respirations and microbial and fungal biomasses by measuring positive correlations with distance from the wall factory and negative correlation with total Pb concentrations. Pb contamination also induced changes in microbial and fungal catabolic structure (from carbohydrates to amino acids through carboxylic malic acid). Moreover, PICT measurement allowed to establish causal linkages between lead and its effect on biological communities taking into account the contamination history of the ecosystem at community level. The positive correlation between qCO2 (based on respiration and substrate use) and PICT suggested that the Pb stress-induced acquisition of tolerance came at a greater energy cost for microbial communities in order to cope with the toxicity of the metal. In this industrial context of long-term polymetallic contamination dominated by Pb in a field experiment, we confirmed impacts of this metal on soil functioning through microbial communities, as previously observed for earthworm communities.

The Water Cycle, a Potential Source of the Bacterial Pathogen Bacillus cereus.

The behaviour of the sporulating soil-dwelling Bacillus cereus sensu lato (B. cereus sl) which includes foodborne pathogenic strains has been extensively studied in relation to its various animal hosts. The aim of this environmental study was to investigate the water compartments (rain and soil water, as well as groundwater) closely linked to the primary B. cereus sl reservoir, for which available data are limited. B. cereus sl was present, primarily as spores, in all of the tested compartments of an agricultural site, including water from rain to groundwater through soil. During rain events, leachates collected after transfer through the soil eventually reached the groundwater and were loaded with B. cereus sl. In groundwater samples, newly introduced spores of a B. cereus model strain were able to germinate, and vegetative cells arising from this event were detected for up to 50 days. This first B. cereus sl investigation in the various types of interrelated environments suggests that the consideration of the aquatic compartment linked to soil and to climatic events should provide a better understanding of B. cereus sl ecology and thus be relevant for a more accurate risk assessment of food poisoning caused by B. cereus sl pathogenic strains.

Enhanced co-tolerance and co-sensitivity from long-term metal exposures of heterotrophic and autotrophic components of fluvial biofilms.

Understanding the interactive effects of multiple stressors on ecosystems has started to become a major concern. The aim of our study was therefore to evaluate the consequences of a long-term exposure to environmental concentrations of Cu, Zn and As on the pollution-induced community tolerance (PICT) of lotic biofilm communities in artificial indoor channels. Moreover, the specificity of the PICT was assessed by evaluating the positive and negative co-tolerance between these metals. Photosynthetic efficiency and substrate-induced respiration (SIR), targeting the autotrophic and heterotrophic communities respectively were used in short-term inhibition bioassays with Cu, Zn and As to assess sensitivities of pre-exposed biofilms to the metals tested. Diversity profiles of a phototrophic, eukaryotic and prokaryotic community in biofilms following the different treatments were determined and analyzed with principal component analysis. The results demonstrated that pre-exposure to metals induced structural shifts in the community and led to tolerance enhancements in the phototrophic and heterotrophic communities. On the other hand, whatever the functional parameter used (i.e. photosynthesis and SIR), communities exposed to Cu were more tolerant to Zn and vice versa. Furthermore, only phototrophic communities pre-exposed to As developed tolerance to Cu but not to Zn, whereas no co-tolerance between Cu and As was observed in the heterotrophic communities. Finally, phototrophic and heterotrophic communities exposed to Cu and Zn became more sensitive to As, reflecting a negative co-tolerance between these metals. Overall, our findings support the fact that although the mode of action of the different metals is an important driver for the structure and thus the tolerance of the communities, it appears that the detoxification modes are the most important factors for the occurrence of positive or negative co-tolerance.

In situ spatio-temporal changes in pollution-induced community tolerance to zinc in autotrophic and heterotrophic biofilm communities.

Pollution-induced community tolerance (PICT) uses increased tolerance in populations at contaminated sites as an indicator of contaminant effects. However, given the broad structural and functional complexity that characterizes biological communities, the acquisition of PICT could vary with (i) target community, (ii) intensity of toxicant exposure, (iii) the species succession stage, and (iv) the physicochemical characteristics of the studied site. To assess the spatio-temporal changes of zinc-induced tolerance in fluvial biofilm communities, we conducted an in situ study in Osor River (North-East Catalonia, Spain), which has zinc contamination. Biofilms were developed for 5 weeks in a non-metal-polluted site, and were then transferred to different sites in Osor River with different levels of zinc contamination. The spatio-temporal changes of biofilm PICT to zinc was determined using photosynthetic activity bioassays and respiration-induced aerobic bioassays at T(0), and at 1, 3 and 5 weeks of exposure. We also performed physicochemical characterization of the sites, taxonomic analysis of diatoms, bacterial and fungal diversity and profiled pigments of phototrophic communities. We used multivariate ordination to analyze results. In addition to natural species succession, the intensity of metal pollution exerted structural pressure by selecting the most metal-tolerant species, but differently depending on the type of biofilm. Zn-tolerance values indicated that exposure to high levels of zinc had effects that were similar to a longer exposure to lower levels of zinc.

Impact of chronic and acute pesticide exposures on periphyton communities.

Aquatic ecosystems face variable exposure to pesticides, especially during floodings which are associated with short bursts of high contaminant concentrations that influence biological systems. A study was undertaken to highlight the impact of the herbicide diuron applied in mixture with the fungicide tebuconazole on natural periphyton during flooding events. Periphyton were grown in two series of two lotic outdoor mesocosms: one series was non-contaminated while the other was exposed to chronic contamination. After 4weeks, one channel of each series was exposed to three successive pulses, with each pulse followed by one week of recovery. Impacts on periphyton were assessed by using Denaturing Gel Gradient Electrophoresis to characterize eukaryotic community structure. At a functional scale, photosynthetic efficiency was quantified during each pulse, and the induced tolerance to diuron was estimated by performing short-term inhibition tests based on photosynthetic efficiency. Moreover, pesticide concentrations in the water column and periphyton matrix were measured. Diuron was adsorbed in the periphyton during each pulse and desorbed 13h after pulse end. The different pulses affected the eukaryotic community structures of the control biofilms, but not of the chronically exposed ones. During the first pulse, photosynthetic efficiency was correlated with pesticide concentration in the water phase, and there was no difference between periphyton from chronically contaminated channels and control channels. However, during the second and third pulses, the photosynthetic efficiency of periphyton chronically exposed to pesticides appeared to be less impacted by the acute pulsed exposure of pesticide. These changes were consistent with the acquisition of induced tolerance to diuron since only after the third pulse that periphyton from chronic channel became tolerant to diuron. Our experimental study indicates that the effects of pulsed acute exposures to pesticides on periphyton depended on whether the communities had previously been exposed to the same stressors or not.

Use of the MicroResp™ method to assess pollution-induced community tolerance to metals for lotic biofilms.

Understanding the ecological status of aquatic ecosystems and the impact of anthropogenic contamination requires correlating exposure to toxicants with impact on biological communities. Several tools exist for assessing the ecotoxicity of substances, but there is still a need for new tools that are ecologically relevant and easy to use. We have developed a protocol based on the substrate-induced respiration of a river biofilm community, using the MicroResp™ technique, in a pollution-induced community tolerance approach. The results show that MicroResp™ can be used in bioassays to assess the toxicity toward biofilm communities of a wide range of metals (Cu, Zn, Cd, Ag, Ni, Fe, Co, Al and As). Moreover, a community-level physiological profile based on the mineralization of different carbon substrates was established. Finally, the utility of MicroResp™ was confirmed in an in-situ study showing gradient of tolerance to copper correlated to a contamination gradient of this metal in a small river.

Transplantation of microbenthic algal assemblages to assess structural and functional recovery after diuron exposure.

The potential of microbenthic algal assemblages to recover after diuron exposure was investigated. Microbenthic algal assemblages (periphyton) were grown on glass slides in correspondence to a diuron-polluted and adiuron-free sampling site of a river. After 5 weeks of colonization, the impacted periphyton was transferred by translocating the colonized glass slides to the unpolluted site. To monitor the changes in functional and structural parameters and to assess recovery, both the transferred and the local reference periphyton were sampled at the day of transfer (t0) and 1, 3, and 5 weeks after the transfer (t1, t3,t5). Structural transitions of eukaryotic communities were characterized by denaturing gradient gel electrophoresis (DGGE) and functional ones were characterized by short-term incubation toxicity tests with diuron. As shown by Bray­Curtis similarity values based on DGGE band patterns, almost total structural recovery of the transferred periphyton took place 5 weeks after transfer. For the transferred periphyton, previous diuron exposure at the contaminated site induced the development of diuron tolerance,which decreased after 1 week and became very similar to the tolerance of the nonimpacted community after 5 weeks. It is concluded that 5-week-old eukaryotic periphyton communities are capable of restoring their structural and functional attributes after 5 weeks within a noncontaminated environment.

In situ assessment of periphyton recovery in a river contaminated by pesticides.

Recovery of bacterial and eukaryotic communities in biofilms naturally grown on stones was studied for 9 weeks after transferring them from a pesticide polluted downstream site of the river Morcille (Beaujolais, France) to a non-contaminated upstream site. Site-specific periphyton present on stones at both the down- and the upstream sampling site were collected to analyze the site-specific colonization. Throughout the experiment, structural and functional parameters were analyzed for the periphyton transferred and for the site-specific up- and downstream periphyton. Comparison between these three communities allowed quantifying recovery of the transferred one. Denaturing Gradient Gel Electrophoresis of PCR-amplified 16S and 18S rRNA gene fragments were used to assess prokaryotic and eukaryotic microbial community composition, respectively. Microscopy counts allowed characterizing the diatom taxa abundances. The sensitivity of the microalgal communities towards diuron and copper was investigated at the laboratory by short-term photosynthesis inhibition assays. The functional reaction of the bacterial communities towards copper was assessed by short-term respiration inhibition assays. The structure of transferred eukaryotic, bacterial and diatom communities was more similar to the structure of the downstream communities than to upstream ones even after 9 weeks acclimatization in particular for the bacterial community. In the same way, the community tolerance towards diuron and copper, as estimated by the EC50 values, was intermediate for the transferred biofilms compared to the local up- or downstream biofilm, even after 9 weeks of acclimatization. These results strongly suggest slow recovery, likely to be linked to long lasting exposure of pesticides and in particular copper adsorbed to the biofilm matrices and to the toughness for pioneer microorganisms to invade mature biofilms.

PO43- dependence of the tolerance of autotrophic and heterotrophic biofilm communities to copper and diuron.

Pollution-induced community tolerance (PICT) concept is based on the assumption that the toxicant exerts selection pressure on the biological communities when exposure reaches a critical level for a sufficient period of time and therefore sensitive species are eliminated. However, induced tolerance of microbial biofilm communities cannot be attributed solely to the presence of toxicants in rivers but also to various environmental factors, such as amount of nutrients. An experimental study was undertaken to highlight the potential impact of a phosphorus gradient on the sensitivity of periphytic microbial community to Cu and diuron. Biofilms were exposed to real-world levels of chronic environmental contamination of toxicants with a phosphorus gradient. Biofilm sensitivity to Cu and diuron was assessed by performing short-term inhibition tests based on photosynthetic efficiency to target photoautotrophs, extracellular enzyme activity (beta-glucosidase and leucine-aminopeptidase) and substrate-induced respiration activity to target heterotrophs. The impact of P-gradient associated to pollution was evaluated by measuring pesticide concentrations in biofilms, biomass parameters (chla, AFDW), bacterial cell density, photosynthetic efficiency and community structure (using 18S and 16S rDNA gene analysis to target eukaryotes and DGGE and HPLC pigment analysis to target bacteria and photoautotrophs). The obtained results show that depending on the studied toxicant and the used structural or functional parameter, the effect of the phosphorus gradient was variable. This highlights the importance of using a range of parameters that target all the biological communities in the biofilm. The PICT method can be regarded as a good tool for assessing anthropogenic environmental contamination, but it is necessary to dissociate the real impact of toxicants from environmental factors.

Responses of chronically contaminated biofilms to short pulses of diuron. An experimental study simulating flooding events in a small river.

An experimental study was undertaken to highlight the potential ecotoxicological impact of the herbicide diuron on biofilms during flooding events in a small river (Morcille) in the Beaujolais vineyard area (France). We investigated the responses of chronically contaminated biofilms exposed to short-term pulses (3 h) of diuron. Biofilms were grown in indoor microcosms that were either non-contaminated or exposed to low-level chronic contamination, and not exposed, or exposed to single or double pulses of two environmental concentrations (7 and 14 microg L(-1)) of diuron. Exposure to pollution and its impact on biofilms were assessed by measuring pesticide concentrations in biofilms, biomass parameters (chl a, AFDW), community structure (using 18S and 16S rDNA gene analysis by DGGE, and HPLC pigment analysis to target eukaryotes, bacteria and photoautotrophs, respectively) and by performing a physiological test. Control biofilms displayed very low diuron concentrations, whereas the herbicide was found in the contaminated biofilms. Nevertheless, diuron concentrations were not higher in the pulsed biofilms than in the non-pulsed ones. AFDW and chl ain vivo fluorescence increased in both microcosms during the experiment and biomass was higher in chronically exposed biofilms than in control ones. The impact on biomass was higher for the control double-pulsed biofilms than for the non-pulsed ones. Carbon incorporation by the chronically exposed biofilms was greater during the first 28 days of growth than during the first 28 days of growth in the control biofilms. Both single and double pulses inhibited carbon incorporation of all biofilm communities, especially of the control ones. Short-term inhibition of photosynthesis was never significantly different in exposed and non-exposed biofilms. Few differences in the pigment structure were found between chronically exposed and control biofilms, but pulses impacted on the pigment structure of all biofilm communities. Bacterial structural differences were observed between single-pulsed and non-pulsed biofilms, but not between double-pulsed and non-pulsed biofilms. The different pulses affected the eukaryotic community structures of the control biofilms, but not of the chronically exposed ones. Unlike the bacterial communities, the control eukaryotic communities were structurally different from the chronically exposed ones. This preliminary experimental study indicates that exposure to environmental concentrations of diuron and other agricultural contaminants and further exposure to diuron can have measurable effects on small river biofilm communities. The effects of a pulsed acute exposure to diuron on biofilms depended on whether the biofilms had previously been exposed to the same stressors or not.

Assessment of the effects of imidacloprid on the behavior of two earthworm species (Aporrectodea nocturna and Allolobophora icterica) using 2D terraria.

Earthworms are "ecosystem engineers" and changes in their main activities (creation of burrows and burial of organic matter) due to pollutants can have important effects on soil functions and indirectly on other components of the soil ecosystem. Using 2D terraria, we studied the behavior of two earthworm species (the endogeic Allolobophora icterica and the anecic Aporrectodea nocturna) exposed to sublethal concentrations (0.5 and 1 mg kg(-1) of dry soil) of imidacloprid, a widely used neonicotinoid insecticide. A second experiment was carried out to determine whether such imidacloprid concentrations were avoided by the two earthworm species we studied. In polluted soils, significant weight loss was observed in both species. No significant avoidance of imidacloprid was detected, whereas several modifications of the behavior were observed directly (burrowing dynamics, number of oscillations in the burrow system) or indirectly in the resulting burrow system architecture (area, topology, sinuosity maximal depth) according to the species and concentration tested. When exposed to imidacloprid (0.5 or 1 mg kg(-1)), A. icterica almost stopped burrowing after 24 h, whereas A. nocturna burrowed continuously but with a significantly lower rate compared to the control. Moreover, in both species, the burrow systems were always shallower when worms were exposed to imidacloprid. Since this last endpoint is easy to measure and is linked to water transfer in soils, it could be used in ecotoxicological studies provided it could be validated for a wide range of pollutants and concentrations.