Sex‐determining mechanisms change repeatedly throughout evolution, and it is difficult to track this continual process. The Japanese soil‐frog Glandirana rugosa is a remarkable evolutionary witness to the ongoing process of the evolution of sex‐ determining modes. The two geographic groups, designated XY and Neo‐ZW, have homologous sex chromosomes, yet display opposite types of sex chromosomes, XX‐ XY and ZZ‐ZW, respectively. These two groups are sympatric at the edges of their respective ranges in Central Japan. In this study, we discovered molecular evidence that the eastern part of the Neo‐ZW group (Neo‐ZW2 subgroup), which is found near the sympatric area, shares mitochondrial haplotypes with the XY group. By analysing single nucleotide polymorphism (SNP) loci, we have also discovered that the representative nuclear genome of the Neo‐ZW2 subgroup shares allele clusters with both the XY group and another part of the Neo‐ZW group (Neo‐ZW1 subgroup), indicating a hybrid origin of the Neo‐ZW2. Further analysis of sex‐linked SNP loci revealed that the alleles on the W chromosomes of the Neo‐ZW2 were derived mostly from X chromosomes, while alleles on the Z chromosomes originated from the Z chromosomes of the Neo‐ZW1 subgroup and partly from the Y chromosomes of the XY group. Our study revealed that admixture of the two opposite sex‐ chromosome systems reconstructed a female heterogametic system by recycling the X chromosomes into new W chromosomes. This work offers an illustrative example of how de novo sex‐chromosome systems can arise by recycling material from ancestral sex chromosomes
Background: The extent to which sex reversal is associated with transitions in sex determining systems (XX-XY, ZZ-ZW, etc.) or abnormal sexual differentiation is predominantly unexplored in amphibians. This is in large part because most amphibian taxa have homomorphic sex chromosomes, which has traditionally made it challenging to identify discordance between phenotypic and genetic sex in amphibians, despite all amphibians having a genetic component to sex determination. Recent advances in molecular techniques such as genome complexity reduction and high throughput sequencing present a valuable avenue for furthering our understanding of sex determination in amphibians and other taxa with homomorphic sex chromosomes like many fish and reptiles. Results: We use DArTseq as a novel approach to identify sex-linked markers in the North American green frog (Rana clamitans melanota) using lab-reared tadpoles as well as wild-caught adults from seven ponds either in undeveloped, forested habitats or suburban ponds known to be subject to contamination by anthropogenic chemicals. The DArTseq methodology identified 13 sex-linked SNP loci and eight presence-absence loci associated with males, indicating an XX-XY system. Both alleles from a single locus show partial high sequence homology to Dmrt1, a gene linked to sex determination and differentiation throughout Metazoa. Two other loci have sequence similarities to regions of the chimpanzee and human X-chromosome as well as the chicken Z-chromosome. Several loci also show geographic variation in sex-linkage, possibly indicating sex chromosome recombination. While all loci are statistically sex-linked, they show varying degrees of female heterozygosity and male homozygosity, providing further evidence that some markers are on regions of the sex chromosomes undergoing higher rates of recombination and therefore further apart from the putative sex determining locus. Conclusion: The ease of the DArTseq platform provides a useful avenue for future research on sex reversal and sex chromosome evolution in vertebrates, particularly for non-model species with homomorphic or cryptic or nascent sex chromosomes.