Identification and characterisation of isoprene-degrading bacteria in an estuarine environment.

Approximately one-third of volatile organic compounds (VOCs) emitted to the atmosphere consists of isoprene, originating from the terrestrial and marine biosphere, with a profound effect on atmospheric chemistry. However, isoprene provides an abundant and largely unexplored source of carbon and energy for microbes. The potential for isoprene degradation in marine and estuarine samples from the Colne Estuary, UK, was investigated using DNA-Stable Isotope Probing (DNA-SIP). Analysis at two timepoints showed the development of communities dominated by Actinobacteria including members of the genera Mycobacterium, Rhodococcus, Microbacterium and Gordonia. Representative isolates, capable of growth on isoprene as sole carbon and energy source, were obtained from marine and estuarine locations, and isoprene-degrading strains of Gordonia and Mycobacterium were characterised physiologically and their genomes were sequenced. Genes predicted to be required for isoprene metabolism, including four-component isoprene monooxygenases (IsoMO), were identified and compared with previously characterised examples. Transcriptional and activity assays of strains growing on isoprene or alternative carbon sources showed that growth on isoprene is an inducible trait requiring a specific IsoMO. This study is the first to identify active isoprene degraders in estuarine and marine environments using DNA-SIP and to characterise marine isoprene-degrading bacteria at the physiological and molecular level.

Isolation of isoprene degrading bacteria from soils, development of isoA gene probes and identification of the active isoprene-degrading soil community using DNA-stable isotope probing.

Emissions of biogenic volatile organic compounds (bVOCs), are an important element in the global carbon cycle, accounting for a significant proportion of fixed carbon. They contribute directly and indirectly to global warming and climate change and have a major effect on atmospheric chemistry. Plants emit isoprene to the atmosphere in similar quantities to emissions of methane from all sources and each accounts for approximately one third of total VOCs. Although methanotrophs, capable of growth on methane, have been intensively studied, we know little of isoprene biodegradation. Here, we report the isolation of two isoprene-degrading strains from the terrestrial environment and describe the design and testing of polymerase chain reaction (PCR) primers targeting isoA, the gene encoding the active-site component of the conserved isoprene monooxygenase, which are capable of retrieving isoA sequences from isoprene-enriched environmental samples. Stable isotope probing experiments, using biosynthesized (13) C-labelled isoprene, identified the active isoprene-degrading bacteria in soil. This study identifies novel isoprene-degrading strains using both culture-dependent and, for the first time, culture-independent methods and provides the tools and foundations for continued investigation of the biogeography and molecular ecology of isoprene-degrading bacteria.

Regulation of plasmid-encoded isoprene metabolism in Rhodococcus, a representative of an important link in the global isoprene cycle.

Emissions of biogenic volatile organic compounds (VOCs) form an important part of the global carbon cycle, comprising a significant proportion of net ecosystem productivity. They impact atmospheric chemistry and contribute directly and indirectly to greenhouse gases. Isoprene, emitted largely from plants, comprises one third of total VOCs, yet in contrast to methane, which is released in similar quantities, we know little of its biodegradation. Here, we report the genome of an isoprene degrading isolate, Rhodococcus sp. AD45, and, using mutagenesis shows that a plasmid-encoded soluble di-iron centre isoprene monooxygenase (IsoMO) is essential for isoprene metabolism. Using RNA sequencing (RNAseq) to analyse cells exposed to isoprene or epoxyisoprene in a substrate-switch time-course experiment, we show that transcripts from 22 contiguous genes, including those encoding IsoMO, were highly upregulated, becoming among the most abundant in the cell and comprising over 25% of the entire transcriptome. Analysis of gene transcription in the wild type and an IsoMO-disrupted mutant strain showed that epoxyisoprene, or a subsequent product of isoprene metabolism, rather than isoprene itself, was the inducing molecule. We provide a foundation of molecular data for future research on the environmental biological consumption of this important, climate-active compound.