Characterization and evolution of natural aquatic biofilm communities exposed in vitro to herbicides.

River biofilms are assemblies of autotrophic and heterotrophic microorganisms that can be affected by pollutants such as those found in watersheds and wastewater treatment plants. In the laboratory, experimental biofilms were formed from river water, and their overall composition was investigated. Denaturing gradient gel electrophoresis (DGGE) and cytometry were used to assess the richness and diversity of these communities. The software Cytostack (available on request) was developed to treat and analyze the cytometric data. Measurements of chlorophyll-a and carotenoids were used to assess the global composition of the photoautotrophic community, whereas proteins, polysaccharides (PS) content, and esterase activities were used to assess overall changes in the mixed communities. We evaluated the effects that 3 weeks of treatment with the herbicides diuron and glyphosate (10 µg L(-1)) had on these biofilms. Exposed to diuron, bacterial communities adapted, changing their composition. Glyphosate inhibited growth of one autotrophic community but caused no chlorophyll deficit. As a whole, the biofilm acted as a micro-ecosystem, able to regulate and maintain a constant level of photosynthetic pigment through the structural adaptation of the autotrophic community. These results are one more proof that microbial diversity of aquatic biofilms is influenced by chemical stresses, potentially leading to disturbances within the ecosystems.

Response of spring and summer riverine microbial communities following glyphosate exposure.

Seasonal variation in the response of riverine microbial communities to an environmentally relevant exposure to glyphosate (about 10 microgl(-1)) was assessed on natural communities collected in spring and summer, using two 14-day microcosm studies. The two experiments showed no major effect of glyphosate on algal biomass (chlorophyll a concentrations), bacterial activity ([(3)H]thymidine incorporation), or bacterial community diversity (16S PCR-TTGE detection). Effects on algal community composition (genus-level taxonomic identification) and eukaryotic community diversity (18S PCR-DGGE on <100 microm organisms) were only detected on the samples collected in summer. This work demonstrates that even if the effects of a short pulse of glyphosate (10 microgl(-1)) on riverine microorganisms seem to be limited, the responses of natural microbial communities to glyphosate exposure (and probably to other pesticide exposures) can clearly vary between the experiments, and can be seasonally dependent.

Stimulation or inhibition: Leaf microbial decomposition in streams subjected to complex chemical contamination.

Leaf litter decomposition is a key mechanism in headwater streams, allowing the transfer of nutrients and energy into the entire food web. However, chemical contamination resulting from human activity may exert a high pressure on the process, possibly threatening the structure of heterotrophic microbial communities and their decomposition abilities. In this study, the rates of microbial Alnus glutinosa (Alnus) leaf decay were assessed in six French watersheds displaying different land use (agricultural, urbanized, forested) and over four seasons (spring, summer, autumn, winter). In addition, for each watershed at each sampling time, both upstream (less-contaminated) and downstream (more-contaminated) sections were monitored. Toxicities (estimated as toxic units) predicted separately for pesticides and pharmaceuticals as well as environmental parameters (including nutrient levels) were related to microbial decay rates corrected for temperature and a range of fungal and bacterial community endpoints, including biomass, structure, and activity (extracellular ligninolytic and cellulolytic enzymatic activities). Results showed that agricultural and urbanized watersheds were more contaminated for nutrients and xenobiotics (higher pesticides and pharmaceuticals predicted toxicity) than forested watersheds. However, Alnus decay rates were higher in agricultural and urbanized watersheds, suggesting compensatory effects of nutrients over xenobiotics. Conversely, fungal biomass in leaves was 2-fold and 1.4-fold smaller in urbanized and agricultural watersheds than in the forested watersheds, respectively, which was mostly related to pesticide toxicity. However, no clear pattern was observed for extracellular enzymatic activities except that ß-glucosidase activity positively correlated with Alnus decay rates. Together, these results highlight microbial communities being more efficient for leaf decomposition in polluted watersheds than in less contaminated ones, which is probably explained by changes in microbial community structure. Overall, our study showed that realistic chemical contamination in stream ecosystems may affect the biomass of Alnus-associated microbial communities but that these communities can adapt themselves to xenobiotics and maintain ecosystem functions.

Longitudinal changes in microbial planktonic communities of a French river in relation to pesticide and nutrient inputs.

To determine the effects of anthropic activities on river planktonic microbial populations, monthly water samples were collected for 11 months from two sampling sites characterized by differing nutrient and pesticide levels. The difference in trophic level between the two stations was particularly pronounced from May to November. Total pesticide concentrations were notably higher at the downstream station from April to October with a clear predominance of herbicide residues, especially the glyphosate metabolite aminomethylphosphonic acid (AMPA). From spring, algal biomass and density were favored by the high orthophosphate concentrations recorded at the downstream location. However, isolated drops in algal biomass were recorded at this sampling station, suggesting an adverse effect of herbicides on algal communities. No major difference was observed in bacterial heterotrophic production, density, or activity (CTC reduction) between the two sampling stations. No major variation was detected using the fluorescent in situ hybridization (FISH) method, but shifts in bacterial community composition were recorded by PCR-TTGE analysis at the downstream station following high nutrient and pesticide inputs. However, outside the main anthropic pollution period, the water's chemical properties and planktonic microbial communities were very similar at the two sampling sites, suggesting a high recovery potential for this lotic system.

Effects of the phenylurea herbicide diuron on natural riverine microbial communities in an experimental study.

The effects of the phenylurea herbicide diuron (10 microgl(-1)) on natural riverine microbial communities were investigated using a three-week laboratory microcosm study. During the first six days, a latency period was observed both in the algal and the bacterial communities despite favorable abiotic conditions and independently of diuron exposure. From the second week, an intense algal bloom (chlorophyll a concentrations and cell abundances) was observed in the uncontaminated microcosms but not in the treated microcosms. The bloom stimulated the bacterial community and led to an increase in heterotrophic bacterial production ([3H]thymidine incorporation), activity (CTC reduction) and cell abundance. In parallel, shifts in bacterial community composition were recorded by polymerase chain reaction (PCR)-temporal temperature gradient gel electrophoresis (TTGE) analysis, whereas no major variation was detected using the fluorescent in situ hybridization (FISH) method. In the treated microcosms, the diuron acted not by damaging the initial communities but by inhibiting the algal bloom and indirectly maintaining constant bacterial conditions throughout the experiment. These inhibitory effects, which were recorded in terms of abundance, activity and diversity, suggest that exposure to diuron can decrease the recovery capacities of microbial communities and delay the resumption of an efficient microbial food web despite favorable environmental conditions.

Comparative effects of the herbicides chlortoluron and mesotrione on freshwater microalgae.

Extensive use of herbicides in agriculture is accompanied by the risk of environmental contamination of aquatic ecosystems. The present study shows the effects of the herbicides chlortoluron and mesotrione on three microalgae species: two chlorophyceae (Pediastrum tetras, Ankistrodesmus fusiformis) and one diatom (Amphora coffeaeformis). The authors calculated the IC50 for one chlorophyceae and the diatom. The order of toxicity (median inhibitory concentration [IC50]) for mesotrione was A. coffeaeformis (13.1 mg/L) > A. fusiformis (56.1 mg/L) and A. fusiformis (0.05 mg/L) > A. coffeaeformis (0.08 mg/L) for chlortoluron. The impact of herbicides applied at 0.2 mg/L was then examined in Erlenmeyer flasks by monitoring for growth, pigment content, and metabolic activity. Algal responses varied widely according to species and herbicide. For example, chlortoluron showed a significant inhibitory effect on the growth of A. coffeaeformis, whereas mesotrione induced an increase in cellular density in A. fusiformis. Other cellular parameters, such as pigment content in P. tetras, were stimulated by both herbicides. The results obtained confirmed that microalgae cultures are clearly affected by acute and chronic exposition to herbicides. Further monitoring should be carried out in the field to assess the impact of sublethal levels of toxicity and the growth-enhancing effects of mesotrione and chlortoluron on natural algae communities.