Identification of Sex-Specific Genetic Markers of Mosquitofish

The livebearing eastern (Gambusia holbrooki) and western (Gambusia affinis) mosquitofish are two highly invasive species that have been released for mosquito control in many tropical countries. The appearance of both species is quite similar, except for a small morphological difference of the male intromittent organ (gonopodium). In addition, in some G. holbrooki populations a few males display a black spotting pattern that sons inherit from the father. Surprisingly, these two sister species differ in their sex determination mechanism: while G. holbrooki has an XX/XY sex determination system in which males are the heterogametic sex, in G. affinis sex is determined by a WZ/ZZ system with a much larger female W-chromosome. So far, nothing is known about the mechanisms underlying sex determination and pigmentation in Gambusia. In order to identify sex-specific genetic markers in both species, we generated RAD-seq data from a minimum of 26 males and females of each species. In addition, we sequenced the genome of a spotted G. holbrooki male and a G. affinis female with Illumina 250 bp paired-end technology. After analyzing the RAD-seq data with STACKS and assembling the genomes with DISCOVAR, sex-specific RAD-tags were mapped to the Gambusia genomes and the platyfish (Xiphophorus maculatus) reference genome. We could identify sex-specific markers in both species, of which some are linked to genes. Further analysis by qRT-PCR, RNA-seq, and in situ hybridization will help to understand the roles that these genes play in mosquitofish sex determination and pigmentation.