3.1 Sex ratio and geographic distribution of sexual and parthenogenetic populations
Results of sex check by sex-check PCR and crossing experiments were concordant, except for some samples from p24, p27, and Argentina which had both female and male markers in sex-check PCR (Figure 1; Table 1; Table S1). In crossing experiments, samples from p27 released gametes fused with male gametes of p22 (Table S1) and did not function as male gametes. Therefore, the samples having both markers were considered female (at least, phenotypically).
Populations were considered sexual when at least one male was identified, or parthenogenic when no males could be identified in samples of ten or more individuals (Table 1). For p1, although the sex of only two individuals were examined, this population was considered parthenogenetic (Table 1), since sexual reproduction was not observed in this population in a previous study (Kogame et al., 2005). Along the Sea of Japan coast, sexual populations were distributed from Kyushu to Hokkaido and overlapped with parthenogenetic populations on the west coast of Hokkaido (Figure 1). Along the Pacific coast, sexual populations were limited to the South of Choshi, Chiba Prefecture (the sampling locality of p24 and p25) and parthenogenetic populations were found North of Choshi (Figure 1). Sexual and parthenogenetic populations were parapatric in two localities: Esashi, the west coast of Hokkaido (p4 and p5) and Choshi (p24 and p25). At these localities, the two types of populations were 1 km and 0.3 km apart along the coast, respectively (Figure S1). In both localities, parthenogenetic populations were found in more upper part of intertidal zone and more wave-exposed areas, compared with sexual populations. No males were found among European and Argentinean samples (four and 17 samples, respectively; Table S1).