Friday, 14 July 2006

Relations between Biodegradation, Microbial Genetic Potential and Global Structure of Bacterial Communities in a Silty Cultivated Soil: Case of 2,4-D.

Laure Vieublé-Gonod, UMR INRA - INA PG - Environnement & Grandes Cultures, Bldg EGER, Thiverval Grignon, 78850, France, Fabrice Martin laurent, UMR Microbiologie et Géochimie des Sols, INRA-Université de Bourgogne, 17 Rue Sully, BP 86510, Dijon, 21065, France, and Claire Chenu, UMR Biogeochimie des Milieux Continentaux, INRA-CNRS-UPMC, Bldg EGER, Thiverval-Grignon, 78850, France.

Major soil processes such as pesticides fate can not only be explained with abiotic factors. Knowing the key role of telluric microorganisms in the degradation of these compounds, an understanding of soil functioning requires the description of the structure, the composition, the density and the biodiversity of soil microbial communities. The possible relations between the biodiversity of soil microbial communities and their functions still remain poorly documented, mainly because of a lack of adequate methods. For the last years, the development of molecular techniques offer new insights in soil functioning. Only a few studies reported the quantification of indigenous microbial populations such as atrazine degraders and denitrifiers. Consequently, kinetics of the genetic potential of microbial functions such as pesticide degradation is poorly described. If microorganisms influence the fate of pesticides, pesticide application may reciprocally affect soil microbial communities according to two principal ways: (i) through the toxic effect of the molecule or (ii) by favoring the growth of specific degraders able to use this molecule as a carbon source and thereby modifying the equilibrium between microbial populations in soil. In order to search for possible links between the functioning of specific soil microbial communities, their genetic potential and their genetic structure, we studied 2,4-D biodegradation. 2,4-D has been one of the most heavily used herbicides in the world for agricultural and turf grass applications. The 2,4-D degradation pathway, leading to 2-chloromaleylacetate production, involves six enzymes coded by the tfdA, B, C, D, E and F genes. 2-chloromaleylacetate is then entirely mineralized. In this context, the objectives of our work were to estimate (i) the impact of 2,4-D application on the genetic structure of bacterial communities, (ii) 2,4-D mineralization and (iii) 2,4-D-degrading genetic potential, by quantifying tfdA sequences, coding an enzyme specifically involved in the first step of 2,4-D mineralization leading to 2,4-dichlorophenol accumulation. To respond to these objectives and particularly to establish links between the presence of specific microorganisms and the processes they catalyse, we combined isotope measurements with molecular analyses. The genetic structure of soil bacterial communities, the 14C-2,4-D mineralization and the 2,4-D degrading genetic potential estimated by real time PCR targeted on tfdA sequences were depicted in soil microcosms treated or not with 2,4-D. The impact of 2,4-D on the structure of bacterial populations was followed with a a Ribosomal Intergenic Spacer Analysis (RISA) at different dates of incubation. 2,4-D was preferentially used by soil microorganisms to produce energy rather than to synthesize new cells. The 2,4-D degrading genetic potential increased rapidly following 2,4-D application. This increase was correlated with the one of 14C microbial biomass. These results suggested that in this soil 2,4-D degrading microbial communities may preferentially use the tfd pathway to degrade 2,4-D. Furthermore, the maximum of tfdA sequences corresponded to the maximum rate of 2,4-D mineralization. RISA revealed that the genetic structure of bacterial communities was significantly modified in response to 2,4-D application. This impact was only observed during the intense phase of 2,4-D biodegradation. Seven days after 2,4-D application the genetic structure of bacterial recovered its initial state.

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