Phylogenetic relationships and divergence time estimates
In the phylogenetic tree, we observed that the haplotypes were clearly
divided into two clades, i.e., clade 1 and clade 2, and the S morph was
derived from the ancestral L morph (Figure 2). Clade 1 included only
Korean L morph haplotypes, while clade 2 included haplotypes from Korea,
Japan, and the USA, as well as both L and S morph haplotypes. The Korean
and Japanese S morph haplotypes were monophyletic, implying that the
wing transformation event took place only once in the history of the
species. The USA haplotype (Hap 36) diverged earlier and was not
monophyletic with the S morph haplotypes. Based on molecular dating, the
two clades diverged approximately 2.7078 MYA (95% HPD: 0.0053–9.278
MYA), and the divergence of the S morph from the L morph occurred around
0.2 MYA (95% HPD: 0.0003–0.7164 MYA) (Figure 2).
Wing morphology and Wolbachia infection
Unlike all of the L morph individuals, which proved to be infected withWolbachia , Wolbachia infection was polymorphic in the S
morph individuals. None of the Korean S morph populations (Hap 7)
harbored Wolbachia , but the Japanese S morph populations
collected from the mid-northern part of Japan, i.e., Ishikawa and Toyama
(Hap 22), were completely infected whereas populations from Tokyo (Hap
25, 26) and Gifu (Hap 24) were free of Wolbachia (Figure 2). The
wing development pattern correlated strongly with Wolbachiainfection status in this ant species (n = 142, Pearson
χ2 = 100.339, df = 1, P <
0.001). These results also suggest that Wolbachia is not involved
in clonal reproduction in the ant species because clonal reproduction
occurs in both wing morphs (Ohkawara et al., 2006).