Molecular techniques for understanding microbial abundance and activity in clay barriers used for geodisposal

Abstract Clays are commonly used in design concepts for geological disposal of nuclear waste. It is thus essential to identify and quantify microbial communities in clay-rich samples to study microbial processes during geological disposal. Although advances in culture-independent techniques have enabled detailed studies of microbial communities in diverse ecosystems, the efficiency and sensitivity of these molecular techniques depend on characteristics of the environment studied. Moreover, the outcome of nucleic acid-based approaches depends on the extraction method, primer specificity, PCR amplification, sequencing artifacts, and downstream bioinformatic analyses. Clays are recalcitrant to DNA extraction and are challenging for analysis by standard techniques using viability stains and measurement of metabolic activity. This chapter explores the impact of various sequencing and bioinformatic pipelines used for 16S rRNA gene profiling of microbial communities and compares the efficiency of different DNA extraction methods from clay. Moreover, we also discuss non-DNA-based techniques used to assess microbial activity and viability in clay samples.