While transitions (C <-> T, G <-> A) are the most prevalent form of substitution reported in bacteria [18], transitions from C to T and G to A were found to be equally likely with transversions from C to A and G to C for both species ( Figure 8 ). As these rates have not been corrected for the G+C% of the parent organisms (both with <50% G+C), the proportion of substitutions which transition/transverse in higher G+C% organisms is likely larger than observed here. The novelty of this observation, in light of accepted substitution mechanisms, suggests that these substitutions are a consequence of the PGM sequencer, which is further reinforced by the prevalence of low confidence flow-calls for these substitutions. emPCR is an unlikely source of these errors, as transitions are the dominant substitution type for 454 GS FLX Titanium sequencing [16]. Considering that the PGM introduces substitution errors at a rate of between 0.04%–0.1%, rare variants (single nucleotide polymorphisms) which occur at a frequency greater than 0.1% may be detected using the PGM platform.
