13C incorporation into DNA as a means of identifying the active components of ammonia-oxidizer populations


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Abstract

Aims:To identify active CO2-assimilating species of ammonia-oxidizing bacteria in fresh water sediment.Methods and Results:Enrichment cultures were incubated in the presence of 13C labelled CO2, and 13C-DNA successfully resolved from 12C-DNA by caesium chloride density gradient ultracentrifugation of DNA extracts. Ammonia-oxidizer DNA recovered from these gradients was amplified and characterised by Temporal Temperature Gradient Gel Electrophoresis (TTGE), with confirmatory sequence analysis to identify the metabolically active components of the population.Conclusions:The 12C-DNA fraction was dominated by nitrosospiras, in contrast to the 13C-DNA fraction which was largely nitrosomonad DNA, in support of the hypothesis that nitrosomonads out-compete nitrosospiras in laboratory culture.Significance and Impact of the Study:The use of stable isotype incorporation into ammonia-oxidizer DNA could therefore circumvent the problems associated with RNA detection to identify metabolically active species in situ.

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