Variations in the extraction and clean-up efficiency of DNA during anaerobic digestion of wastewater sludge at mesophilic and thermophilic temperatures

Abstract

The effect of anaerobic sludge digestion on the isolation of DNA and clean-up efficiency of obtained DNA isolates was studied. Influent and digestate of a continuous full-scale reactor were sampled twice. Also, batch digesters were operated at 35, 45 and 55 °C, namely, AnD.35, AnD.45 and AnD.55. The batch reactors were sampled at 3-to-4-day time intervals during the digestion. A sharp decline in the extraction yield was observed over time for the batch reactors. In AnD.55, the extraction yield declined by 96 % in 12 days. The purity of the DNA isolates was assessed by purity ratios, which included absorption at 260 nm to that at 280 nm (Abs260/280) and to that at 230 nm (Abs260/230). Abs260/230 before clean-up declined as a result of digestion in all reactors, the most severe case being AnD.55 decreasing from 1.91 to 0.47 in nine days. The efficiency of clean-up was evaluated by analysing DNA recovery and changes in the purity ratios. AnD.35 and the full-scale reactor had the highest recoveries (≥ 80 %). Furthermore, a 4.1-fold increase was observed in Abs260/230. Additionally, non-linear correlations were found between purity parameters and the DNA concentration (R2 > 0.7). The observations of the current investigation strongly indicate that the change in DNA extraction yield was at least partly due to the change in DNA extraction efficiency and not just the change in the actual DNA content. This is an important consideration for the quantification of biomass, specific genes, and microbial populations, where DNA isolation plays a key role.