Stable carbon isotope fractionation as tracer of carbon cycling in anoxic soil ecosystems

doi:10.1016/j.copbio.2016.07.001

Current Opinion in Biotechnology

Volume 41, October 2016, Pages 122-129

https://doi.org/10.1016/j.copbio.2016.07.001 Get rights and content

Highlights

•
Quantification of the relative contribution of different methanogenic pathways.
•
Quantification of the relative contribution of different acetogenic pathways.
•
Quantification of the relative contribution of different substrates (e.g., C₃ vs. C₄ plants) to the degradation process.
•
Importance of the knowledge of fractionation factors.

While the structure of microbial communities can nowadays be determined by applying molecular analytical tools to soil samples, microbial function can usually only be determined by physiological experiments requiring incubation of samples. However, analysis of stable isotope fractionation might be able to analyse microbial function without incubation in soil samples. We describe the limitations of diagnosing and quantifying carbon flux pathways in soil by using the determination of stable carbon isotope composition in soil compounds and emphasize the importance of determining stable isotope fractionation factors for defined biochemical pathways. Fractionation factors are sufficiently different for some central biochemical pathways in anaerobic degradation of organic carbon. Thus, it is possible to quantify the relative contribution of CH₄ production by hydrogenotrophic or aceticlastic methanogenic pathways, and of acetate formation by chemolithotrophic (acetyl-CoA synthase) or heterotrophic (fermentation) pathways. In addition, stable isotope analysis may allow the differentiation between different organic substrates used for degradation, for example, the relative contribution of root exudation versus soil organic matter degradation, provided the different substrates are sufficiently distinct in their isotopic compositions (e.g., mixture of C₃ and C₄ plants) and the carbon conversion pathways display only small fractionation factors or are identical for the different substrates.

Graphical abstract

Introduction

Carbon cycling in soil ecosystems is mainly the result of input of carbon substrates by plants and degradation by soil microorganisms. The input by plants is mostly in the form of litter, dead plant material or root exudates. The degradation, however, is achieved by a complex soil microbial community comprising many different biochemical pathways, which depend on the chemical nature of the organic substrate and the environmental conditions (Figure 1). In general, the anaerobic degradation of soil organic matter can be separated in the hydrolysis and fermentation of polymeric substrates to short-chain fatty acids, alcohols, CO₂ and H₂. Further fermentation results in degradation to acetate, CO₂ and H₂, which are the dominant precursors of methanogenesis. In anaerobic systems, CH₄ and CO₂ are stable end products. Oxidation of CH₄ usually requires the presence of O₂ and methane oxidizing bacteria. Understanding the carbon cycling in soil ecosystems therefore requires knowledge of both the structure and the function of the microbial communities. There has recently been much progress in elucidating the structure of microbial communities by applying molecular analytical tools, in particular sequencing of microbial genes and gene products [1]. These analyses just require sampling of soil and — the sometimes technically demanding — extraction of nucleic acids or proteins. While some information on microbial functioning can be obtained by combining genomic and metaproteomic approaches [2, 3, 4] or using stable isotope probing techniques [5, 6], the in situ functions of the microbial communities usually can only be analyzed by incubation and measurement of the temporal change of analytically accessible variables. However, analysis of stable isotope signatures in soil samples might overcome this problem, since the isotopic signatures partially reflect the microbial functioning [7]. Our review intends to explore the limits of using stable carbon isotopic signatures for elucidating the microbial functional pathways of organic carbon degradation in soil, anaerobic degradation of small organic molecules originating from hydrolysis and fermentation, and methane production in particular.

Section snippets

Isotopic signatures, isotope effects, and fractionation factors

The ¹³C isotopic signature of a particular carbon compound is given by its ratio R = ¹³C/¹²C and is usually denoted relative to a standard (st) as δ¹³C = 10³ (R/R_st − 1) [8, 9]. The reactions in a biochemical pathway, especially those involving the cleavage of carbon bonds, often display characteristic fractionation factors (α) or enrichment factors (ɛ = 10³[1 − α]), which define the extent to which the carbon atoms have been fractionated during the conversion of a substrate to a product. For a reaction

Determination of fractionation factors for microbial pathways

The compound specific stable isotope analysis (CSIA) of δ¹³C usually requires an isotope ratio mass spectrometer (IRMS), which measures the δ¹³C in CO₂. If a particular carbon compound can be isolated (e.g., by chromatography) and converted to CO₂ (e.g., by combustion), the average δ¹³C of all its C atoms can be determined usually using a GC–IRMS or HPLC–IRMS system. For some simple compounds (e.g., CO₂, CH₄), the δ¹³C can also be determined using laser-based absorption spectrometry, for

Diagnosis and quantification of carbon flux pathways in soil

The measurement of δ¹³C of individual soil carbon compounds may allow the diagnosis of particular biochemical pathways that operated until the time of sampling [60, 61, 62]. The usual approach is defining theoretically possible C flux paths that can be differentiated by their fractionation factors and testing them by δ¹³C analysis of their substrates and products. The crucial point is that the pathways to be discriminated must display sufficiently different fractionation factors, reflected in

Conclusions and future perspectives

Stable isotope fractionation can be useful for distinguishing different carbon flux pathways provided the ranges covered by the fractionation factors associated with the individual pathways do not overlap. This is nicely the case for anaerobic processes involved in the degradation of organic matter, thus allowing the differentiation and even quantification of methanogenesis by hydrogenotrophic or aceticlastic pathways, and of acetogenesis by chemolithotrophic (acetyl-CoA synthase) or

References and recommended reading

Papers of particular interest, published within the period of review, have been highlighted as:

• of special interest
•• of outstanding interest

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Stable carbon isotope fractionation as tracer of carbon cycling in anoxic soil ecosystems

Highlights

Graphical abstract

Introduction

Section snippets

Isotopic signatures, isotope effects, and fractionation factors

Determination of fractionation factors for microbial pathways

Diagnosis and quantification of carbon flux pathways in soil

Conclusions and future perspectives

References and recommended reading

Soil Biol Biochem

Biotechnol Adv

Org Geochem

Geochim Cosmochim Acta

Marine Geol

Org Geochem

Curr Opin Biotechnol

Biochim Biophy Acta-Proteins Proteomics

Phytochemistry

Chem Geol

Org Geochem

Plant Soil

Org Geochem

Appl Environ Microbiol

Geochim Cosmochim Acta

Global Change Biol

Appl Environ Microbiol

Org Geochem

Appl Environ Microbiol

Environ Sci Technol

Sci Total Environ

Science

Science

Geochim Cosmochim Acta

Appl Environ Microbiol

Soil Biol Biochem

PLoS One

Appl Environ Microbiol

Org Geochem

Org Geochem

Geochim Cosmochim Acta

Frontiers Microbiol

Soil Biol Biochem

PLoS One

Org Geochem

J Mol Microbiol Biotechnol

Curr Opin Biotechnol

Global Biogeochem Cycles

Back to the future of soil metagenomics

Front Microbiol

Community proteomics of a natural microbial biofilm

Science

Metagenomics as a tool for the investigation of uncultured microorganisms

Genetika

Metagenomic analyses: past and future trends

Appl Environ Microbiol

How well do we know VPDB? Variability of delta C-13 and delta O-18 in CO2 generated from NBS19-calcite

Rapid Commun Mass Spectrom

Stable isotope fractionation of tetrachloroethene during reductive dechlorination by Sulfurospirillum multivorans and Desulfitobacterium sp strain PCE-S and abiotic reactions with cyanocobalamin

Appl Environ Microbiol

Influence of mass transfer on stable isotope fractionation

Appl Microbiol Biotechnol

How well do we know VPDB? Variability of delta C-13 and delta O-18 in CO₂ generated from NBS19-calcite