Mars simulated exposure and the characteristic Raman biosignatures of amino acids and halophilic microbes

Though Raman bands of α-amino acids (AA) are well documented, often only the strongest intensity bands are quoted as identifiers (e.g. Jenkins et al., 2005; De Gelder et al., 2007; Zhu et al., 2011). Unknown regolith mixtures on Mars-sampling missions could obscure these bands. Here the case is made for determining, via a statistical method, sets of characteristic bands to be used as identifiers, independent of band intensity or number of bands (Rolfe et al., 2016). AA have upwards of 25 potentially identifying bands and this method defines sets of 10–19 bands per AA. Examination of AA-doped Mars-like basalt resulted in a maximum of eight bands being identified, as some characteristic bands were obscured by mineral bands, including the strongest intensity band in some cases. This proved the need for characteristic bands to be defined, enabling successful identification of AA. The ESA ExoMars Rover mission will crush and then pass the sample to the Raman Laser Spectrometer. We crushed a Mars-like basalt to a similar grain size expected to be created by the rover. Our samples were doped with 1 % (by weight) AA samples, resulting in no detection of AA, because of loss of original spatial context and spaces between the grains. We recommend that Raman spectroscopy on future missions should be conducted before the sample is crushed. Halite-entombed halophilic microbes, known to survive being entombed, were exposed to Mars-like surface (including temperature, pressure, atmospheric composition and UV) and freeze-thaw cycle (plus pressure and atmospheric composition) conditions. This test on the survival of the microbes showed that survival rates quickly deteriorated in surface conditions, but freeze-thaw cycle samples had well preserved Raman biosignatures, indicating that similar signatures could be detectable on Mars if similar life persists in evaporitic material or brines today

Microbial metabolism of isoprene: a much-neglected climate-active gas

The climate-active gas isoprene is the major volatile produced by a variety of trees and is released into the atmosphere in enormous quantities, on a par with global emissions of methane. While isoprene production in plants and its effect on atmospheric chemistry have received considerable attention, research into the biological isoprene sink has been neglected until recently. Here, we review current knowledge on the sources and sinks of isoprene and outline its environmental effects. Focusing on degradation by microbes, many of which are able to use isoprene as the sole source of carbon and energy, we review recent studies characterizing novel isoprene degraders isolated from soils, marine sediments and in association with plants. We describe the development and use of molecular methods to identify, quantify and genetically characterize isoprene-degrading strains in environmental samples. Finally, this review identifies research imperatives for the further study of the environmental impact, ecology, regulation and biochemistry of this interesting group of microbes

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

Sphingopyxis sp. Strain OPL5, an Isoprene-Degrading Bacterium from the Sphingomonadaceae Family Isolated from Oil Palm Leaves

The volatile secondary metabolite, isoprene, is released by trees to the atmosphere in enormous quantities, where it has important effects on air quality and climate. Oil palm trees, one of the highest isoprene emitters, are increasingly dominating agroforestry over large areas of Asia, with associated uncertainties over their effects on climate. Microbes capable of using isoprene as a source of carbon for growth have been identified in soils and in the tree phyllosphere, and most are members of the Actinobacteria. Here, we used DNA stable isotope probing to identify the isoprene-degrading bacteria associated with oil palm leaves and inhabiting the surrounding soil. Among the most abundant isoprene degraders of the leaf-associated community were members of the Sphingomonadales, although no representatives of this order were previously known to degrade isoprene. Informed by these data, we obtained representatives of the most abundant isoprene degraders in enrichments, including Sphingopyxis strain OPL5 (Sphingomonadales), able to grow on isoprene as the sole source of carbon and energy. Sequencing of the genome of strain OPL5, as well as a novel Gordonia strain, confirmed their pathways of isoprene degradation and broadened our knowledge of the genetic and taxonomic diversity of this important bacterial trait

Poplar phyllosphere harbors disparate isoprene-degrading bacteria

The climate-active gas isoprene (2-methyl-1,3-butadiene) is released to the atmosphere in huge quantities, almost equaling that of methane, yet we know little about the biological cycling of isoprene in the environment. Although bacteria capable of growth on isoprene as the sole source of carbon and energy have previously been isolated from soils and sediments, no microbiological studies have targeted the major source of isoprene and examined the phyllosphere of isoprene-emitting trees for the presence of degraders of this abundant carbon source. Here, we identified isoprene-degrading bacteria in poplar tree-derived microcosms by DNA stable isotope probing. The genomes of isoprene-degrading taxa were reconstructed, putative isoprene metabolic genes were identified, and isoprene-related gene transcription was analyzed by shotgun metagenomics and metatranscriptomics. Gram-positive bacteria of the genus Rhodococcus proved to be the dominant isoprene degraders, as previously found in soil. However, a wider diversity of isoprene utilizers was also revealed, notably Variovorax, a genus not previously associated with this trait. This finding was confirmed by expression of the isoprene monooxygenase from Variovorax in a heterologous host. A Variovorax strain that could grow on isoprene as the sole carbon and energy source was isolated. Analysis of its genome confirmed that it contained isoprene metabolic genes with an identical layout and high similarity to those identified by DNA-stable isotope probing and metagenomics. This study provides evidence of a wide diversity of isoprene-degrading bacteria in the isoprene-emitting tree phyllosphere and greatly enhances our understanding of the biodegradation of this important metabolite and climate-active gas

The microbiology of isoprene cycling in aquatic ecosystems

Isoprene (2-methyl-1,3-butadiene) is emitted in vast quantities (>500 Tg C yr-1). Most isoprene is emitted by trees, but there is still incomplete understanding of the diversity of isoprene sources. The reactivity of isoprene in the atmosphere has potential implications for both global warming and global cooling, with human health implications also arising from isopreneinduced ozone formation in urban areas. Isoprene emissions from terrestrial environments have been studied for many years, but our understanding of aquatic isoprene emissions is less complete. Several abundant phytoplankton taxa produced isoprene in the laboratory, and the relationship between chlorophyll a and isoprene production has been used to estimate emissions from marine environments. The aims of this review are to highlight the role of aquatic environments in the biological cycling of isoprene and to stimulate further study of isoprene metabolism in marine and freshwater environments. From a microbial ecology perspective, the isoprene metabolic gene cluster, first identified in Rhodococcus sp. AD45 (isoGHIJABCDEF) and subsequently found in every genome-sequenced isoprene-degrader, provides the ideal basis for molecular studies on the distribution and diversity of isoprene-degrading communities. Further investigations of isopreneemitting microbes, such as the influence of environmental factors and geographical location, must also be considered when attempting to constrain estimates of the flux of isoprene in aquatic ecosystems. We also report isoprene emission by the scleractinian coral Acropora horrida and the degradation of isoprene by the same coral holobiont, which highlights the importance of better understanding the cycling of isoprene in marine environments

Spatial and temporal variability of biogenic isoprene emissions from a temperate estuary

[1] Isoprene is important for its atmospheric impacts and the ecophysiological benefits it affords to emitting organisms; however, isoprene emissions from marine systems remain vastly understudied compared to terrestrial systems. This study investigates for the first time drivers of isoprene production in a temperate estuary, and the role this production may play in enabling organisms to tolerate the inherently wide range of environmental conditions. Intertidal sediment cores as well as high and low tide water samples were collected from four sites along the Colne Estuary, UK, every six weeks over a year. Isoprene concentrations in the water were significantly higher at low than high tide, and decreased toward the mouth of the estuary; sediment production showed no spatial variability. Diel isoprene concentration increased with light availability and decreased with tidal height; nighttime production was 79% lower than daytime production. Seasonal isoprene production and water concentrations were highest for the warmest months, with production strongly correlated with light (r2 = 0.800) and temperature (r2 = 0.752). Intertidal microphytobenthic communities were found to be the primary source of isoprene, with tidal action acting as a concentrating factor for isoprene entering the water column. Using these data we estimated an annual production rate for this estuary of 681 μmol m−2 y−1. This value falls at the upper end of other marine estimates and highlights the potentially significant role of estuaries as isoprene sources. The control of estuarine isoprene production by environmental processes identified here further suggests that such emissions may be altered by future environmental change

Genome characterisation of an isoprene-degrading Alcaligenes sp. isolated from a tropical restored forest

Isoprene is a climate-active biogenic volatile organic compound (BVOC), emitted into the atmosphere in abundance, mainly from terrestrial plants. Soil is an important sink for isoprene due to its consumption by microbes. In this study, we report the ability of a soil bacterium to degrade isoprene. Strain 13f was isolated from soil beneath wild Himalayan cherry trees in a tropical restored forest. Based on phylogenomic analysis and an Average Nucleotide Identity score of >95%, it most probably belongs to the species Alcaligenes faecalis. Isoprene degradation by Alcaligenes sp. strain 13f was measured by using gas chromatography. When isoprene was supplied as the sole carbon and energy source at the concentration of 7.2 × 105 ppbv and 7.2 × 106 ppbv, 32.6% and 19.6% of isoprene was consumed after 18 days, respectively. Genome analysis of Alcaligenes sp. strain 13f revealed that the genes that are typically found as part of the isoprene monooxygenase gene cluster in other isoprene-degrading bacteria were absent. This discovery suggests that there may be alternative pathways for isoprene metabolism