Philip Ineson

Elevated CO2, litter chemistry, and decomposition: a synthesis

The results of published and unpublished experiments investigating the impacts of elevated [CO2] on the chemistry of leaf litter and decomposition of plant tissues are summarized. The data do not support the hypothesis that changes in leaf litter chemistry often associated with growing plants under elevated [CO2] have an impact on decomposition processes. A meta-analysis of data from naturally senesced leaves in field experiments showed that the nitrogen (N) concentration in leaf litter was 7.1% lower in elevated [CO2] compared to that in ambient [CO2]. This statistically significant difference was: (1) usually not significant in individual experiments, (2) much less than that often observed in green leaves, and (3) less in leaves with an N concentration indicative of complete N resorption. Under ideal conditions, the efficiency with which N is resorbed during leaf senescence was found not to be altered by CO2 enrichment, but other environmental influences on resorption inevitably increase the variability in litter N concentration. Nevertheless, the small but consistent decline in leaf litter N concentration in many experiments, coupled with a 6.5% increase in lignin concentration, would be predicted to result in a slower decomposition rate in CO2-enriched litter. However, across the assembled data base, neither mass loss nor respiration rates from litter produced in elevated [CO2] showed any consistent pattern or differences from litter grown in ambient [CO2]. The effects of [CO2] on litter chemistry or decomposition were usually smallest under experimental conditions similar to natural field conditions, including open-field exposure, plants free-rooted in the ground, and complete senescence. It is concluded that any changes in decomposition rates resulting from exposure of plants to elevated [CO2] are small when compared to other potential impacts of elevated [CO2] on carbon and N cycling. Reasons for experimental differences are considered, and recommendations for the design and execution of decomposition experiments using materials from CO2-enrichment experiments are outlined.

Identification of active methylotroph populations in an acidic forest soil by stable- isotope probing

Stable-isotope probing (SIP) is a culture-independent technique that enables the isolation of DNA from micro-organisms that are actively involved in a specific metabolic process. In this study, SIP was used to characterize the active methylotroph populations in forest soil (pH 3.5) microcosms that were exposed to (13)CH(3)OH or (13)CH(4). Distinct (13)C-labelled DNA ((13)C-DNA) fractions were resolved from total community DNA by CsCl density-gradient centrifugation. Analysis of 16S rDNA sequences amplified from the (13)C-DNA revealed that bacteria related to the genera Methylocella, Methylocapsa, Methylocystis and Rhodoblastus had assimilated the (13)C-labelled substrates, which suggested that moderately acidophilic methylotroph populations were active in the microcosms. Enrichments targeted towards the active proteobacterial CH(3)OH utilizers were successful, although none of these bacteria were isolated into pure culture. A parallel analysis of genes encoding the key enzymes methanol dehydrogenase and particulate methane monooxygenase reflected the 16S rDNA analysis, but unexpectedly revealed sequences related to the ammonia monooxygenase of ammonia-oxidizing bacteria (AOB) from the beta-subclass of the PROTEOBACTERIA: Analysis of AOB-selective 16S rDNA amplification products identified Nitrosomonas and Nitrosospira sequences in the (13)C-DNA fractions, suggesting certain AOB assimilated a significant proportion of (13)CO(2), possibly through a close physical and/or nutritional association with the active methylotrophs. Other sequences retrieved from the (13)C-DNA were related to the 16S rDNA sequences of members of the Acidobacterium division, the beta-Proteobacteria and the order Cytophagales, which implicated these bacteria in the assimilation of reduced one-carbon compounds or in the assimilation of the by-products of methylotrophic carbon metabolism. Results from the (13)CH(3)OH and (13)CH(4) SIP experiments thus provide a rational basis for further investigations into the ecology of methylotroph populations in situ.

Detection and classification of atmospheric methane oxidizing bacteria in soil.

Well-drained non-agricultural soils mediate the oxidation of methane directly from the atmosphere, contributing 5 to 10% towards the global methane sink. Studies of methane oxidation kinetics in soil infer the activity of two methanotrophic populations: one that is only active at high methane concentrations (low affinity) and another that tolerates atmospheric levels of methane (high affinity). The activity of the latter has not been demonstrated by cultured laboratory strains of methanotrophs, leaving the microbiology of methane oxidation at atmospheric concentrations unclear. Here we describe a new pulse-chase experiment using long-term enrichment with 12CH4 followed by short-term exposure to 13CH4 to isotopically label methanotrophs in a soil from a temperate forest. Analysis of labelled phospholipid fatty acids (PLFAs) provided unambiguous evidence of methane assimilation at true atmospheric concentrations (1.8–3.6 p.p.m.v.). High proportions of 13C-labelled C18 fatty acids and the co-occurrence of a labelled, branched C17 fatty acid indicated that a new methanotroph, similar at the PLFA level to known type II methanotrophs, was the predominant soil micro-organism responsible for atmospheric methane oxidation.

Active root-inhabiting microbes identified by rapid incorporation of plant-derived carbon into RNA

Plant roots harbor a large diversity of microorganisms that have an essential role in ecosystem functioning. To better understand the level of intimacy of root-inhabiting microbes such as arbuscular mycorrhizal fungi and bacteria, we provided 13CO2 to plants at atmospheric concentration during a 5-h pulse. We expected microbes dependent on a carbon flux from their host plant to become rapidly labeled. We showed that a wide variety of microbes occurred in roots, mostly previously unknown. Strikingly, the greatest part of this unsuspected diversity corresponded to active primary consumers. We found 17 bacterial phylotypes co-occurring within roots of a single plant, including five potentially new phylotypes. Fourteen phylotypes were heavily labeled with the 13C. Eight were phylogenetically close to Burkholderiales, which encompass known symbionts; the others were potentially new bacterial root symbionts. By analyzing unlabeled and 13C-enriched RNAs, we demonstrated differential activity in C consumption among these root-inhabiting microbes. Arbuscular mycorrhizal fungal RNAs were heavily labeled, confirming the high carbon flux from the plant to the fungal compartment, but some of the fungi present appeared to be much more active than others. The results presented here reveal the possibility of uncharacterized root symbioses.

PAHs associated with the leaves of three deciduous tree species. I — Concentrations and profiles

Results for the concentrations of total polycyclic aromatic hydrocarbons (Sigma PAH) and the PAH profile in leaves from three deciduous tree species from the same woodland are presented, and discussed with reference to environmental and leaf-related variables. There were significant differences between oak, ash and hazel leaves in their Sigma PAH concentrations (sum of 23 PAHs), and in the relative contribution of individual PAHs to the sum. Leaves exhibiting pubescence (hairiness) were found to have significantly higher Sigma PAH concentrations than hairless leaves, regardless of their position in the vegetation strata of the wood. Hazel leaves from the understorey had a PAH profile consisting of a greater proportion of the 4-, 5- and 6-ring PAHs than oak or ash from the canopy. This was concluded to be the result of the filtering effect of the main canopy on the air passing over and through it, with subsequent transfer of particles and attendant PAHs to the understorey below. The proportion of Sigma PAH contributed by the 6-ring PAH in hazel leaves was negatively correlated with distance from the southern edge of the canopy. It is proposed that the predominantly windward edges of the woodland, where atmospheric turbulence is likely to be greatest, favoured the deposition of particle-bound PAHs to leaves.

Carbon assimilation and turnover in grassland vegetation using anin situ13CO2 pulse labelling system

A mobile laboratory was developed to administer a controlled flow of (13)C labelled CO(2) at ambient concentrations ( approximately 350 ppm) in the field. The stable isotope delivery (SID) system consists of an isotope-mixing unit with flow control to a series of 12 independent labelling chambers. In-line CPU controlled infrared gas analysers allow automated measurement of chamber CO(2) concentrations and gas flow management. A preliminary experiment was established on an upland pasture located at the NERC Soil Biodiversity experimental site, Sourhope, UK, in August 1999. The objective of this investigation was to determine the magnitude of pulse-derived C incorporation into a typical upland plant community. To achieve this, the SID system was deployed to pulse-label vegetation with CO(2) enriched with (13)C (50 atom %) at ambient concentrations ( approximately 350 ppm) on two consecutive days in August 1999. Samples of headspace CO(2), shoot and root were taken on four occasions over a period of 28 days after (13)C labelling. These materials were then prepared for (13)C/(12)C ratio determination by continuous-flow/combustion/isotope ratio mass spectrometry (CF-C-IRMS). Results showed that pulse derived CO(2)-C was assimilated at a rate of 128 +/- 32 microg g shoot-C hour(-1). Dynamic samplings showed that pulse-derived (13)C concentrations in the labelled plant tissues declined by 77.4 +/- 6% after 48 hours. The rapid decline in (13)C concentrations in plant matter was the result of C loss from the plant in the form of respired CO(2) and root exudates, and dilution by subsequent unlabelled C assimilates. This novel system offers considerable potential for in situ tracer investigations.

Active microbial RNA turnover in a grassland soil estimated using a 13CO2 spike

Rhizosphere microbes are critical to the initial transfer and transformation of root carbon inputs to the soil but our understanding of the activity of these organisms remains constrained by their limited culturability. In this study we combined isotopic 13C tracer and molecular approaches to measure the incorporation of recently assimilated plant C into soil microbial RNA and DNA pools as a means to determine the turnover of the ‘active’ rhizosphere community. This required the development of a method for the extraction, purification and preparation of small-sample soil DNA and RNA (<5 μg C) for isotope analysis. Soil, plant and respired CO2 samples were collected from a 13CO2 pulse-chase experiment at intervals for 20 days post-labelling. The peak of 13C release in soil/root respired CO2 came between 5 and 48 h after 13CO2 pulse-labelling and was followed by a secondary peak of soil heterotroph 13C respiration after 136 h. Results showed that both soil DNA and RNA rapidly incorporated recent photosynthate with greatest 13C found in the ‘active’ microbial RNA fraction reflecting higher rates of microbial RNA turnover. The dilution rate of the pulse derived 13C in RNA-C was used to estimate a microbial RNA turnover of approximately 20% day−1 with a 15–20 day residence time for photosynthate derived 13C in the RNA pool. The findings of this work confirm the rapid transfer of photosynthate C inputs through soil microorganisms to the atmosphere as CO2 and the potential of the biomolecular-isotope tracer approach in soil C research.

Relationships between enchytraeid worms (Oligochaeta), climate change, and the release of dissolved organic carbon from blanket peat in northern England

Over a period of 1 yr, we examined the dynamics of a community of enchytraeid worms in a blanket peat soil in relation to the prevailing climate and the release of dissolved organic carbon (DOC). Our objectives were two-fold: first, we aimed to establish whether there is a relationship between climatic variables and the density of enchytraeid populations; and also assess whether changes in the density of enchytraeid populations, caused by climate, were related to the release of DOC. Our second aim was to predict the consequences of atmospheric warming for the enchytraeid community at the site, and, in turn, the biological contribution to DOC release. Positive linear relationships between the abundance of enchytraeids and soil temperature were found, as well as with concentrations of DOC in the soil solution: We calculate that at mean field density, enchytraeids currently account for up to 26% of the DOC produced in the surface (upper 10 cm) blanket peat. These relationships suggest that future soil warming caused by climate change could increase enchytraeid abundance by 43% and C release by 11% in these peatlands, based on an increase in mean monthly air temperatures of 2.5 °C.

Enchytraeid worm (Oligochaeta) influences on microbial community structure, nutrient dynamics and plant growth in blanket peat subjected to warming

Our aim was to determine whether the response of below-ground feedback processes to atmospheric warming affects nutrient dynamics and primary production in a model peatland ecosystem. Specifically, we examined the interactions between a dominant soil animal of a blanket peat ecosystem (Enchytraeidae, Oligochaeta) and microbes in response to soil warming (to 6°C above current mean summer temperatures), and the consequences of these interactions for nutrient mineralisation and the growth of the graminoid Festuca ovina L. Enchytraeids reduced soil microbial biomass (total PLFA) by 23%, but did not affect soil nutrient availability or plant nutrient content. Enchytraeids did, however, increase C mineralisation by 8%, measured as dissolved organic carbon (DOC) release in the soil solution. Atmospheric warming increased plant nutrient uptake (increasing shoot N and P contents by 12 and 11%, respectively), but reduced the function of enchytraeids with respect to their role in DOC release (by 16%). These findings suggest that in the short term, independent of the effects of enchytraeids, warming may have reduced the ability of the soil microbial biomass to immobilise nutrients and may have relaxed the competition for nutrients between plants and microbes in these nutrient poor soils, increasing the ability of plants to act as a nutrient sink. The results suggest that although soil warming may disrupt biological interactions that are presently operating in these ecosystems, this may not necessarily result in detrimental effects on ecosystem function, which we determined by the growth of F. ovina.

Decomposition of Pinus sylvestris litter in litter bags: Influence of underlying native litter layer

Abstract Pinus sylvestris (L.) litter confined in 1-mm mesh litter bags was incubated on layers of either P. sylvestris or Picea abies (Dietra.) litter of different origins (native litters) under standard conditions in the laboratory. The decomposition rate of the confined litter was measured both as evolution of CO 2 and as mass loss. The native litter played an important part in governing the rate of decomposition of the confined litter, with the native litters richest in N and Ca resulting in greater decomposition rates (up to 15% more) of the confined litter than the nutrient poor native litters. Both inter- and intra-species effects were observed and these are discussed in terms of chemical and physical interactions between the confined and native litters.