Isoprene Production by Sphagnum Moss Is Balanced by Microbial Uptake, as Revealed by Selective Inhibitors

Abstract

Northern peatlands, ecosystems which store enormous amounts of carbon, and yet are major sources of methane and plant-derived volatiles including isoprene, are predicted to be greatly affected by climate change. Isoprene, the major volatile secondary metabolite released by plants, can support the carbon and energy needs of a variety of bacteria. Here we show that Sphagnum moss from an acidic bog harboured highly active isoprene degraders which consumed the vast majority of the plant-produced isoprene, preventing its release to the atmosphere. We quantified the potential for microbial isoprene uptake in the moss and, using alkyne inhibitors specific to either isoprene monooxygenase of bona fide isoprene degraders, or to the enzymes of other microbes capable of its fortuitous co-oxidation, we show that methane utilizers, for example, did not oxidise significant isoprene in incubations. Our technique enabled the separate quantification of plant isoprene production and microbial uptake, revealing that although atmospheric isoprene concentrations are typically low, the microbes contained in, or in close association with the moss were capable of isoprene uptake at the plant-generated isoprene concentration. Analysis of the bacterial community suggested that the isoprene degraders in this environment belonged to novel groups distinct from extant strains with this capability.

Keywords: Sphagnum; biogeochemical cycles; isoprene; methanotrophs; monooxygenase; peatlands; volatiles.

FIGURE 1

Consumption of (A) methane or (B) isoprene, by microcosms containing Sphagnum moss, peat and bog water, in the presence of the inhibitors acetylene or 1‐octyne, in comparison with no inhibitor (Second Incubations). Error bars show the SEM, n = 6 (live samples), n = 3 (killed controls).

FIGURE 2

(A) Isoprene uptake or production by moss microcosms incubated under controlled lighting conditions, with or without the addition of isoprene, or without isoprene but with the inhibitor of microbial isoprene uptake, 1‐octyne, exposed to, or protected from light (Third Incubations). (B) the rates of isoprene production or consumption calculated from the data shown in panel A. Different upper‐case letters (A, B, C) indicate statistically significant differences between treatments (p < 0.001). The data show the mean ± SEM (n = 3).

FIGURE 3

Microbial phylogeny based on 16S rRNA gene sequencing of Sphagnum moss shoots as sampled (Moss) and following enrichment with isoprene for 39 days (Enriched moss). Taxa increased in relative abundance over twofold between the two timepoints are shown in bold.

FIGURE 4

Phylogenetic tree showing the relationship between isoA sequences obtained from Dersingham Bog (in bold) with those of known isoprene degraders. The tree, constructed using the maximum likelihood method in Mega 7 (Kumar et al. 2016), is based on nucleotide sequences of the aligned proteins. Bootstrap values (500 replications) are shown at the nodes. The scale bar shows substitutions per site. The relative abundance of ASVs obtained in this study from amplicon sequencing of isoprene‐enriched samples is shown in parentheses, and two sequences from long read metagenomic sequencing are identified by their read numbers.

FIGURE 5

Gene sequences assumed to be responsible for isoprene degradation retrieved from DNA extracted from Dersingham moss. The iso gene clusters are aligned with those of reference strain Variovorax sp. WS11, a well‐characterised isoprene‐degrading isolate (Dawson et al. ; Larke‐Mejía et al. 2019).