If the in situ growth rate of filamentous bacteria in activated sludge can be quantified, researchers can more accurately assess the effect of operating conditions on the growth of filaments and improve the mathematical modeling of filamentous bulking. We developed a method to quantify the in situ specific growth rate of Sphaerotilus natans (a model filament) in activated sludge using the species-specific 16S rRNA:rDNA ratio. Primers targeting the 16S rRNA of S. natans were designed, and real-time PCR and RT-PCR were used to quantify DNA and RNA levels of S. natans, respectively. A positive linear relationship was found between the rRNA:rDNA ratio (from 440 to 4500) and the specific growth rate of S. natans (from 0.036 to 0.172 h) using chemostat experiments. The in situ growth rates of S. natans in activated sludge samples from three water reclamation facilities were quantified, illustrating how the approach can be applied in a complex environment such as activated sludge.
The goal of this project was to develop a species-specific method to quantify the in-situ growth rate of one filament and one floc-former in activated sludge. Chemostats were used to study pure cultures of Sphaerotilus natans, as the representative filament, and Arthrobacter globiformis, as the representative floc-former, to determine the relationship between growth rate and RNA:DNA ratio for each species. Real-time qPCR and reverse transcription real-time qPCR were used to measure DNA and RNA levels respectively. The relationship between RNA:DNA ratio and growth rate was found to be positive and linear, with R 2 of 0.58 and 0.98 for S. natans and A. globiformis respectively. This relationship was used to determine in-situ growth rate of S. natans in activated sludge from the aeration basin of a full-scale WWTP and in samples collected from a previous study on the effect of different substrates on bulking. A. globiformis could not be detected in any of the WWTP samples. This study represents the first time qPCR and RT-qPCR have been used to quantify in-situ microbial growth rates in activated sludge.
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