TY - JOUR

T1 - Insights into the fate of a 13C labelled phenol pulse for stable isotope probing (SIP) experiments

AU - Manefield, Mike

AU - Griffiths, Rob

AU - McNamara, Niall P.

AU - Sleep, Darren

AU - Ostle, Nick

AU - Whiteley, Andrew

PY - 2007/5

Y1 - 2007/5

N2 - Stable isotope probing (SIP) using DNA or RNA as a biomarker has proven to be a useful method for attributing substrate utilisation to specific microbial taxa. In this study we followed the transfer of a 13C6-phenol pulse in an activated sludge micro-reactor to examine the resulting distribution of labelled carbon in the context of SIP. Most of the added phenol was metabolically converted within the first 100 min after 13C6-phenol addition, with 49% incorporated into microbial biomass and 6% respired as CO2. Less than 1% of the total 13C labelled carbon supplied was incorporated into microbial RNA and DNA, with RNA labelling 6.5 times faster than DNA. The remainder of the added 13C was adsorbed and/or complexed to suspended solids within the sludge. The 13C content of nucleic acids increased beyond the initial consumption of the 13C-phenol pulse. This study confirms that RNA labels more efficiently than DNA and reveals that only a small proportion of a pulse is incorporated into nucleic acids. Evidence of continued 13C incorporation into nucleic acids suggests that cross-feeding of the SIP substrate was rapid. This highlights both the benefits of using a biomarker that is rapidly labelled and the importance of sampling within appropriate timescales to avoid or capture the effects of cross-feeding, depending on the goal of the study.

AB - Stable isotope probing (SIP) using DNA or RNA as a biomarker has proven to be a useful method for attributing substrate utilisation to specific microbial taxa. In this study we followed the transfer of a 13C6-phenol pulse in an activated sludge micro-reactor to examine the resulting distribution of labelled carbon in the context of SIP. Most of the added phenol was metabolically converted within the first 100 min after 13C6-phenol addition, with 49% incorporated into microbial biomass and 6% respired as CO2. Less than 1% of the total 13C labelled carbon supplied was incorporated into microbial RNA and DNA, with RNA labelling 6.5 times faster than DNA. The remainder of the added 13C was adsorbed and/or complexed to suspended solids within the sludge. The 13C content of nucleic acids increased beyond the initial consumption of the 13C-phenol pulse. This study confirms that RNA labels more efficiently than DNA and reveals that only a small proportion of a pulse is incorporated into nucleic acids. Evidence of continued 13C incorporation into nucleic acids suggests that cross-feeding of the SIP substrate was rapid. This highlights both the benefits of using a biomarker that is rapidly labelled and the importance of sampling within appropriate timescales to avoid or capture the effects of cross-feeding, depending on the goal of the study.

KW - Microbial biomass

KW - DNA

KW - RNA

KW - Carbon

KW - Respiration

KW - CO

U2 - 10.1016/j.mimet.2007.01.019

DO - 10.1016/j.mimet.2007.01.019

M3 - Article

VL - 69

SP - 340

EP - 344

JO - Journal of Microbiological Methods

JF - Journal of Microbiological Methods

SN - 0167-7012

IS - 2

ER -