Abstract Xu and colleagues (Xu et al., 2021) recently suggested a new parameterisation of BWA-mem (Li, 2013) as an alternative to the current standard BWA-aln (Li and Durbin, 2009) to process ancient DNA sequencing data. The authors tested several combinations of the -k and -r parameters to optimise BWA-mem ’s performance with degraded and contaminated ancient DNA samples. They report that using BWA-mem with − k 19 − r 2.5 parameters results in a mapping efficiency comparable to BWA-aln with − I 1024 − n 0.03 (i.e. a derivation of the standard parameters used in ancient DNA studies; (Schubert et al., 2012)), while achieving significantly faster run times. We recently performed a systematic benchmark of four mapping software (i.e. BWA-aln , BWA-mem , NovoAlign ( http://www.novocraft.com/products/novoalign ), and Bowtie2 (Langmead and Salzberg, 2012) for ancient DNA sequencing data and quantified their precision, accuracy, specificity, and impact on reference bias (Oliva et al., 2021). Notably, while multiple parameterisations were tested for BWA-aln , NovoAlign , and Bowtie2 , we only tested BWA-mem with default parameters. Here, we use the alignment performance metrics from Oliva et al. to directly compare the recommended BWA-mem parameterisation reported in Xu et al. with the best performing alignment methods determined in the Oliva et al. benchmarks, and we make recommendations based on the results.

BWA-mem is not the best aligner for ancient DNA short reads