Fig 4

Fig 4

(A) Representative individuals imaged repeatedly during adult pattern formation. Images are aligned to show corresponding regions and rescaled to control for overall growth (frames and regions are selected from S1 Movie). Standard length (SL in mm) serves as a proxy for developmental stage as the relationship between development rate and days post fertilization (dpf) depends on rearing conditions [3]. By 7.0 mm SL, adult melanophores had started to develop in both species. Newly differentiating melanophores are marked by dashed squares, and the same cells are marked by dashed circles of the same color in subsequent images. Adult melanophores ultimately coalesced into dark pattern elements with movements by individual cells and rearrangements among cells particularly apparent in D. kyathit. As in D. rerio, melanophores were occasionally lost in both species (e.g., dashed triangle in D. kyathit) [25,99]. Densely arranged iridophores of the primary interstripe were evident by 7.0 mm SL in both species. Subsequently developing iridophores of stripes or spots (yellow arrowheads) appeared earlier in D. kyathit (7.0 mm SL) than D. quagga (9.2 mm SL), as did iridophores of secondary interstripes or “interspots” further ventrally (yellow arrowheads; 10.6 and 9.2 mm SL, respectively). To facilitate cell counting, fish were treated prior to imaging with epinephrine to contract melanin granules towards cell centers. Images shown are representative of 4 individuals of each species imaged throughout the stages of early adult pigment pattern development. (B–E) Pattern metrics during development of fish imaged repeatedly (splines over points), and in young adults and backcross progeny (most 22–25 mm SL, 4–5 months post-fertilization; see main text and S3 Fig). Individual points for melanophore nearest neighbor distances (C) represent median values calculated for all melanophores examined within each individual fish at time points with ten or more melanophores. *, P = 0.029 in B (F_1,21 = 5.52); ***, P<0.0002 in C (F_1,21 = 19.83), P<0.0001 in D (F_1,21 = 421.04), P<0.0001 in E (F_1,21 = 217.11). Scale bars, 100 μm.