BACTERIAL SOIL DIVERSITY IN HERBICIDE RESISTANT TRANSGENIC SUGARCANE FIELD

Francisco Dini Andreote (ESALQ/USP); Fernando Dini Andreote (EMBRAPA); Armando Cavalcante Franco Dias (CENA/USP); Welington Luiz de Araújo (NIB - UMC)

The advanced of molecular approaches, based on culture-independent methods, have been largely contributing to understanding the structure and dynamics of microbial communities in environmental samples. In this context, this work aimed to properly sampling and measuring the total bacterial community in sugarcane bulk soil (variety SP80-1842) and its transgenic isoline (IMI-1) resistant to imazapyr herbicide, in three different cultivation conditions: i) conventional plants (SP80-1842) with manual weeds control (CV); ii) transgenic plants with manual weeds control (TC) and iii) transgenic plants with herbicide imazapyr application (TH). The bulk soil samples were collected in three different stages of sugarcane development and analyzed firstly by 16S rRNA-based PCR-denaturing gradient gel electrophoresis (PCR-DGGE), using primers for total bacterial community and for specific bacterial groups (Actinobacteria, Alphaproteobacteria and Betaproteobacteria). The results revealed that the structure of these assemblages kept constant among the different cultivation conditions and over time sampling. Additionally, 16S rRNA clone libraries were constructed for the three different cultivations conditions. A total of 61, 57 and 67 sequences from each of the CV, TC and TH libraries, respectively, were obtained. The phylogenetic affiliation of the sequences was achieved by using RDPQuery, where a range of groups, considering each OTU at 97% of similarity, were described. Clone sequences affiliated with the Proteobacteria, Actinobacteria, Firmicutes and Acidobacteria were, respectively, most abundant. Some other groups affiliated with Nitrospira, Chloroflexi, Thermomicrobia, Bacteroidetes, Gemmatimonadetes and Verrucomicrobia were described and clones affiliated with candidate division TM7 and OP10 were also encompassed. Furthermore, a rarefaction curve and diversity and richness indexes were assigned using DOTUR, and a statistic test for multiple comparisons among libraries was conduced by S-Libshuff. These results show a small shift in the TC library when compared to CV and TH. It evidences the low coverage of libraries CV and TH in some evolutionary distances when compared to TC, indicating the possible presence of a more rich community associated with transgenic plants, and also remaking the importance in using polyphasic approaches to address a microbial ecology issue.