Evaluating commercial DNA extraction kits for bacterial community profiling in waterlogged archaeological wood

Understanding bacterial communities responsible for the slow biodeterioration of waterlogged archaeological wood is critical for reconstructing decay processes, predicting preservation trajectories, and informing conservation strategies. The extraction of sufficient and amplifiable bacterial DNA from waterlogged archaeological wood is essential for robust sequencing and accurate bacterial community profiling. However, DNA yield and inferred community composition can vary depending on the extraction protocol used. In this study, we evaluated the performance of three commercially available DNA extraction kits - Bead-Beat Micro AX Gravity, DNeasy PowerSoil Pro and MagAttract PowerSoil Pro DNA – on samples from a single waterlogged Neolithic wooden pole recovered from a submerged archaeological site in Denmark and preserved under anoxic conditions. Quantitative analysis showed that the Bead-beat Micro AX Gravity kit yielded the highest DNA concentrations by Qubit fluorometry and lowest PCR cycle number in quantitative PCR. Elevated DNA signals were observed in some extraction blanks during Qubit quantification; however, these blanks did not produce sequencing reads, indicating that the signal likely represented non-amplifiable material. Comparative analysis of bacterial community composition, based on 16S rRNA gene amplicon sequencing (V4 region, primers 515F/806R), revealed minimal variation between extraction methods, but distinct taxonomic profiles were observed across different sampling areas within a single archaeological wooden pole. These results indicate that while extraction protocol has limited influence on bacterial community composition in this context, spatial heterogeneity within samples can affect results. Protocols yielding higher DNA concentrations may therefore be advantageous when working with degraded archaeological materials or DNA-intensive downstream applications. However, extraction methods optimized for the recovery of highly fragmented DNA may offer additional advantages for ancient heavily degraded substrates.