13CO2 pulse labelling of plants in tandem with stable isotope probing:: methodological considerations for examining microbial function in the rhizosphere

Recently developed (CO2)-C-13 pulse labelling and stable isotope probing (SIP) methods offer the potential to track C-13-labelled plant photosynthate into phylogenetic groups of microbial taxa in the rhizosphere, permitting an examination of the link between soil microbial diversity and carbon flow in situ. We tested the feasibility of this approach to detect functional differences in microbial communities utilising recently fixed plant photosynthate in moisture perturbed grassland turfs. Specifically, we addressed two questions: (1) How does moisture perturbation (three treatments; continual wetting, drying, and drying followed by rewetting) affect the assimilation of C-13-labelled exudates carbon into the soil microbial community?; (2) Can C-13 deposited in soil from pulse-labelled plants be used to identify microbes utilising plant exudates using SIP methodologies? Net CO2 fluxes showed that prior to (CO2)-C-13 pulse labelling, all treatments were photosynthetically active, but differences were observed in night time respiration, indicating moisture treatments had impacted on net CO2 efflux. Measurements of pulse-derived C-13 incorporated into soil RNA over 2 months showed that there was only evidence of C-13 enrichment in the continuously wetted treatments. However, isotopic values represented only a 0.1-0.2 C-13 at.% increase over natural abundance levels and were found to be insufficient for the application of RNA-SIP. These findings reveal that in this experimental system, the microbial uptake of labelled carbon from plant exudates is low, and further optimisation of methodologies may be required for application of SIP to natural plant-soil systems where C-13 tracer dilution is a consideration. (C) 2004 Elsevier B.V. All rights reserved.