Application des outils biomoléculaires 16S, MALDI-TOF et métagénomique pour la détermination des variantes taxonomiques bactériennes telluriques

Abstract

The traditional bacterial culture techniques are not sufficient to study the terrestrial bacterial populations. By against, molecular biology techniques have a power of high discrimination and they are the current methods for the identification of different bacterial taxonomic variants, allowing a finer approach in the diversity studies. The full exploration of bacterial diversity has always been a challenge; this is due to the complexity, to heterogeneity of the soil matrix and the existence of populations numerically small and rarely detected by conventional techniques. In this thesis, we used three molecular techniques (MALDI-TOF MS, the 16S and metagenomics) to achieve the following objectives: (i) to demonstrate the applicability of the MALDI-TOF MS in the field of soil microbiology, (ii) measure the bacterial diversity of the region ""El-Meridj"" (city of Constantine, Algeria) through metagenomics and (iii) study the effect of physicochemical characteristics on this biodiversity. In the first part, we confirmed the susceptibility of proteomics technology MALDI-TOF MS to identify all our bacterial isolates (9 strains) in species level with a score greater than 2,000 and in record time compared to phenotypic techniques and genotypic techniques (amplification and sequencing of the 16S rDNA). In the second part, the 454 pyrosequencing of metagenomes isolated from three soil samples (lake soil, forest soil and cultivated soil) reveals the presence of 11 phyla, 20 families and 13 genera. Thus, the calculation of diversity indices (Shannon-Weaver index, Inverse Simpson index and Chao1 index) showed that the soil in the area ""El-Meridj"" contains significant heterogeneity of bacterial population and a fair distribution of species richness which can be correlated to the alkaline pH of this region and the high organic content. Our work clearly shows that it is possible to improve access to different telluric bacterial taxonomic variations by the application of MALDI-TOF MS as routine identification system to soil microbiology laboratories and metagenomics for diversity studies