The selection pressures acting on the sporulation pathwayare highlighted by the sheer number of mutations discovered within the entire data set that occur in proteins known to play roles in sporulation. Nine of the 38 mutations (23%) found in all lineages were in genes that directly or indirectly regulate either entry into stationary phase or sporulation; this number exceeds the number that would be expected if mutations were to occur by chance, since less than 5% of B. subtilis genes are dedicated to regulatory processes of any kind [50, 51] . In addition to the mutations found within the ''military'' lineage, the two variants of ATCC 49822 shown in Figure 2 differ by mutations in rpoB (Table S5) which also plays a role in entry into sporulation [52] . Null mutations in spo0F resulting in asporogenous phenotypes contribute to colony morphology variation in B. anthracis, B. thuringiensis and B. subtilis [53, 54, 55] . Enhanced in vitro ''fitness'' is also a likely driver behind the recovery of asporogenic B. anthracis mutants that were discovered during the investigation into the B. anthracis attacks of 2001 [56] . Because the process of sporulation is highly energy-intensive and irreversible once commenced, mutants that delay sporulation (or fail to sporulate altogether) to take advantage of remaining nutrients would out-compete wild-type cells during repeated passage in vitro in the absence of other selection pressures, as has been demonstrated in extended in vitro evolution studies with B. subtilis under relaxed sporulation conditions [57] . This may not be universally the case, since gain-of-function mutations in sporulation such as those observed in this studymay compete favorably with wild-type cells if cannibalism of vegetative cells by sporulating bacteria is the dominant selective pressure [58] . Finally, horizontally transferred genetic elements can have dramatic effects on sporulation: for example, recent studies of phage lysogeny in B. anthracis have revealed the ability of several integrated phages to positively affect the kinetics of sporulation upon lysogeny of commonly used B. anthracis strains [59] .