Sabine Stachowski-Haberkorn

Impact of Roundup on the marine microbial community, as shown by an in situ microcosm experiment

The effects of the herbicide Roundup (glyphosate) on natural marine microbial communities were assessed in a 7-day field experiment using microcosms. Bottles were maintained underwater at 6m depth, and 10% of their water content was changed every other day. The comparison of control microcosms and surrounding surface water showed that the microcosm system tested here can be considered as representative of the natural surrounding environment. A temporal temperature gradient gel electrophoresis (TTGE) was run on 16S and 18S rDNA-amplified extracts from the whole microbial community. Cluster analysis of the 16S gel showed differences between control and treatment fingerprints for Roundup at 1 microg L(-1) (ANOSIM, p=0.055; R=0.53), and 10 microg L(-1) (ANOSIM, p=0.086; R=0.40). Flow cytometry analysis revealed a significant increase in the prasinophyte-like population when Roundup concentration was increased to 10 microg L(-1). This study demonstrates that a disturbance was caused to the marine microbial community exposed to 1 microg L(-1) Roundup concentration, a value typical of those reported in coastal waters during a run-off event.

Multigenerational exposure of the microalga Tetraselmis suecica to diuron leads to spontaneous long-term strain adaptation

To investigate the ability of microalgae to develop stable, long-term resistance to herbicides, the marine microalga Tetraselmis suecica was exposed to the herbicide diuron (5 μg/L) for a 43-generation exposure period followed by a 12-generation depuration phase. During the first 25 generations, diuron-exposed cultures showed doubling times ranging from 1.95 to 2.6 days, which was 2 to 2.5-fold longer than control cultures. Between generations 25 and 38, during diuron exposure, two out of the three exposed cultures exhibited a spontaneous drop in doubling time. These results provided evidence of culture adaptation to diuron. To assess persistence of the diuron adaptation observed on growth performance, one of the adapted cultures (D3) was maintained for 12 months in unexposed conditions and then tested by a second, short-term exposure to diuron 5 μg/L, in parallel with a control culture (C1) for six generations. Flow cytometry analyses were used to monitor cell density, viability, morphology, relative chlorophyll content and intracellular reactive oxygen species (ROS) level. Under these conditions, diuron induced a strong increase of doubling time in exposed-C1 cultures (2.5-fold longer than unexposed-C1 cultures), but no significant increase occurred in exposed D3-cultures compared with unexposed D3- and unexposed C1-cultures, showing the persistence of adaptation in the previously-exposed strain D3. Intracellular ROS level showed the same trend. Significant differences were observed between these strains, with weaker effects of diuron on strain D3 compared with strain C1: forward scatter (FSC), representing relative cell size, decreased in exposed cultures (67.8% and 95% of the controls for C1 and D3, respectively), whereas FL3 as relative chlorophyll content increased in exposed cultures (115.6% and 108.6% of the controls for C1 and D3, respectively). Results of second exposure to diuron revealed that the adaptation of strain D3 had persisted after 12 months of depuration, as no growth impairment was observed. This study demonstrates the possible appearance of stable diuron resistance in microalgae in cases of strong, multigenerational chronic exposure to this herbicide in polluted environments.

The coming of age of microbial ecotoxicology: report on the first two meetings in France

Context and objectives Microorganisms are ubiquitous in soil, air, and water ecosystems, where they are key players of ecosystem services. Microbial ecotoxicology is an emerging interdisciplinary area of research which aims at investigating the impact of human activities on the diversity, abundance, and activity of microorganisms. In return, the results of such investigations hold the promise to provide novel ways of assessing in a sensitive way the impacts of diverse environmental disturbances and subsequent ecosystem responses. Thus and although the term itself is yet rarely encountered in the scientific literature, microbial ecotoxicology already addresses an increasing political and societal demand. In the French scientific landscape, which often mimics the famous (but sometimes indigestible) “mille-feuilles” pastry, microbial ecotoxicologists are scattered across many different research centers belonging to different research organizations and universities. This research field has thus lacked any visibility and remained unorganized until now. Formal organization of scientific activities may be considered a typical “froggies” concern (or ailment). Nevertheless, it is rather surprising that scientific journals and significant international conferences specifically devoted to microbial ecotoxicology have been missing so far, especially considering the plethoric range of journals and congresses devoted to microbial ecology and ecotoxicology. With these considerations in mind, the idea of organizing the French research community of microbial ecologists around concepts of ecotoxicology made its way, with the aim of sharing the necessity to overcome artificial boundaries that prevent progress in this promising field.

Evaluation of the partial renewal of in situ phytoplankton microcosms and application to the impact assessment of bentazon and dimethenamid

Microcosms, each consisting of 2L natural surface seawater maintained in 2.3-L glass bottles, were immersed at a depth of 6m. The renewal of 10% of microcosm volumes was carried out every other day. Phytoplankton-containing seawater was used for renewal (previously filtered through 25-, 50- or 200-μm cut-off). Phytoplankton community pigment analysis (by HPLC) and flow cytometry analysis were performed. After 13 days, data exhibited phytoplankton characteristics in microcosms in the same range as that of the natural surrounding sea water over the same period. Furthermore, in these microcosms, a negative correlation was observed between the filtration cut-off used for renewal water, and the total cell count. Herbicides were tested as commercial mixtures at 1, 10 and 100 μgL(-1) active substance. Both Frontier® (dimethenamid) and Basamais® (bentazon) induced significant modifications of the phytoplankton populations at every concentration tested. Such results suggest a possible disturbance in polluted coastal areas.

Combined effects of antifouling biocides on the growth of three marine microalgal species

The toxicity of the antifouling compounds diuron, irgarol, zinc pyrithione (ZnPT), copper pyrithione (CuPT) and copper was tested on the three marine microalgae Tisochrysis lutea, Skeletonema marinoi and Tetraselmis suecica. Toxicity tests based on the inhibition of growth rate after 96-h exposure were run using microplates. Chemical analyses were performed to validate the exposure concentrations and the stability of the compounds under test conditions. Single chemicals exhibited varying toxicity depending on the species, irgarol being the most toxic chemical and Cu the least toxic. Selected binary mixtures were tested and the resulting interactions were analyzed using two distinct concentration-response surface models: one using the concentration addition (CA) model as reference and two deviating isobole models implemented in R software; the other implementing concentration-response surface models in Excel®, using both CA and independent action (IA) models as reference and three deviating models. Most mixtures of chemicals sharing the same mode of action (MoA) were correctly predicted by the CA model. For mixtures of dissimilarly acting chemicals, neither of the reference models provided better predictions than the other. Mixture of ZnPT together with Cu induced a strong synergistic effect on T. suecica while strong antagonism was observed on the two other species. The synergy was due to the transchelation of ZnPT into CuPT in the presence of Cu, CuPT being 14-fold more toxic than ZnPT for this species. The two modelling approaches are compared and the differences observed among the interaction patterns resulting from the mixtures are discussed.