Fig. 1. Evolutionary divergence of eggshell morphology and rho expression pattern between D. melanogaster and D. virilis. (A) The egg of D. virilis has four DAs, an ancestral trait, as found in the Hirtodrosophila outgroup of the genus Drosophila. The two DAs found in D. melanogaster is a more recently evolved trait. D. virilis and D. melanogaster diverged 40-60 million years ago (mya). (B,C) The eggshells of D. melanogaster (B) and D. virilis (C) are shown with anterior to the left (dorsal view). (D,E) Schematic comparison of the rho expression patterns between D. melanogaster and D. virilis during oogenesis. Dorsal lateral views of stage 10A- to 12-egg chambers are shown with rho expression (blue). Anterior is to the left. (D) D. melanogaster rho was expressed in a saddle-shaped zone at the dorsal anterior at stage 10A. This expression was refined into an L-shaped stripe on either side of the dorsal midline at stage 10B. Expression was then restricted to the anterior row lacking the midline region at stage 11, as it continued until stage 12. (E) D. virilis rho was expressed in two domains at the dorsal-lateral sides at stage 10A. No expression occurred in the midline, unlike in D. melanogaster. The expression was repressed at the posterior region and refined into a Vshaped stripe missing its apex at stage 10B. Expression was upregulated at the front and rear of each stripe, and expression in the rear moved anteriorly at stage 11. Subsequently, the expression was divided into four domains at stage 12, corresponding to the positions of the four DAs.