Distribution and diversity of isoprene degrading bacteria in the terrestrial environment

Isoprene is the most abundantly produced biogenic volatile organic compound (BVOC) and is an important climate active gas that plays a complex role in atmospheric chemistry. Through a combination of cultivation, and cultivation-independent techniques this project set out to investigate the diversity and distribution of isoprene degrading bacteria, and to investigate the factors that influence their abundance in complex microbial communities in a range of environments. DNA-Stable isotope probing (DNA-SIP) was combined with amplicon sequencing techniques and metagenomics, investigating the isoprene degrading community harboured by a high isoprene emitting willow (Salix fragilis) in the UK, and one of the highest known emitters, the oil palm tree (Elaeis guineensis) in Malaysia. Willow leaves harboured a diverse community of isoprene degrading bacteria and represented the first phyllosphere environment to contain communities of Methylobacterium and Mycobacterium that could metabolise isotopically-labelled isoprene during a DNA-SIP experiment. A Mycobacterium metagenome-assembled genome (MAG) was recovered, containing the first known instance of two complete isoprene degradation metabolic gene clusters in a single genome. Malaysian oil palm soil and phyllosphere studies showed that the soil environment harboured a novel and much more diverse community of isoprene degrading bacteria than the associated phyllosphere. Amplicon sequencing of the isoA gene (encoding the α-subunit of the isoprene monooxygenase) showed that genomes from phyllosphere and soil communities contained a novel diversity of isoA which is essential for bacterial degradation of isoprene. The abundance of isoprene degrading bacteria in a number of environments was also investigated with a combination of metagenomic gene mining and qPCR. qPCR methods were carried out with newly-designed gene probes, validated in this study, that target the isoA gene. Results showed that isoA containing bacteria are found in environments that do not emit isoprene, and make up 0.02% - 1% of the whole microbial community in a given environment