Open AccessUse of Field-Based Stable Isotope Probing To Identify Adapted Populations and Track Carbon Flow through a Phenol-Degrading Soil Microbial Community
Author(s) -
Christopher M. DeRito,
Graham M. Pumphrey,
Eugene L. Madsen
Publication year - 2005
Publication title -
applied and environmental microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.552
H-Index - 324
eISSN - 1070-6291
pISSN - 0099-2240
DOI - 10.1128/aem.71.12.7858-7865.2005
Subject(s) - stable isotope probing , microbial population biology , phenol , environmental chemistry , chemistry , soil microbiology , microorganism , microbial biodegradation , isotopes of carbon , substrate (aquarium) , bacteria , biology , soil water , total organic carbon , organic chemistry , ecology , genetics
The goal of this field study was to provide insight into three distinct populations of microorganisms involved in in situ metabolism of phenol. Our approach measured13 CO2 respired from [13 C]phenol and stable isotope probing (SIP) of soil DNA at an agricultural field site. Traditionally, SIP-based investigations have been subject to the uncertainties posed by carbon cross-feeding. By altering our field-based, substrate-dosing methodologies, experiments were designed to look beyond primary degraders to detect trophically related populations in the food chain. Using gas chromatography-mass spectrometry (GC/MS), it was shown that13 C-labeled biomass, derived from primary phenol degraders in soil, was a suitable growth substrate for other members of the soil microbial community. Next, three dosing regimes were designed to examine active members of the microbial community involved in phenol metabolism in situ: (i) 1 dose of [13 C]phenol, (ii) 11 daily doses of unlabeled phenol followed by 1 dose of [13 C]phenol, and (iii) 12 daily doses of [13 C]phenol. GC/MS analysis demonstrated that prior exposure to phenol boosted13 CO2 evolution by a factor of 10. Furthermore, imaging of13 C-treated soil using secondary ion mass spectrometry (SIMS) verified that individual bacteria incorporated13 C into their biomass. PCR amplification and 16S rRNA gene sequencing of13 C-labeled soil DNA from the 3 dosing regimes revealed three distinct clone libraries: (i) unenriched, primary phenol degraders were most diverse, consisting of α-, β-, and γ-proteobacteria and high-G+C-content gram-positive bacteria, (ii) enriched primary phenol degraders were dominated by members of the generaKocuria andStaphylococcus , and (iii) trophically related (carbon cross-feeders) were dominated by members of the genusPseudomonas . These data show that SIP has the potential to document population shifts caused by substrate preexposure and to follow the flow of carbon through terrestrial microbial food chains.