To investigate the origins and stages of vertebrate adaptive radiation, we reconstructed 17 the spatial and temporal histories of genetic variants underlying major phenotypic axes of 18 diversification from the genomes of 202 Caribbean pupfishes. Ancient standing variation from 19 disparate spatial sources was reassembled into new combinations which are under strong 20 selection for adaptation to novel trophic niches on only a single island throughout the Caribbean. 21 This occurred in three stages: first, standing variation associated with feeding behavior swept, 22 103 7 which scale-eater candidate adaptive variants were significantly enriched, with relevant terms 104 corresponding to major axes of divergence in this radiation highlighted in bold (FDR < 0.01; full 105 list of terms with FDR < 0.05 in Table S4). 106 107 Even though both trophic specialists are endemic to SSI, we also found nearly all their 108 candidate adaptive variants to occur as standing genetic variation across the Caribbean 109 (molluscivore: 100%; scale-eater: 98%; Fig. 2A). Furthermore, nearly half these variants were 110 ancient and also found in Cualac or Megupsilon outgroups to Cyprinodon (41% and 55% of 111 scale-eater and molluscivore adaptive variants, respectively), which diverged over 5 Mya (22). 112 However, most adaptive variants did not show any evidence of selection in five other focal high- 113 coverage Caribbean generalist populations (only 2% and 6% of scale-eater and molluscivore 114 variants, respectively; Fig. S3) and strong linkage disequilibrium among adaptive variants in SSI 115 trophic specialists was not observed in these focal populations (Fig. 2B). Thus, novel trophic 116 specialists within a microendemic adaptive radiation were almost entirely assembled from 117 ancient standing genetic variation through strong selection for new adaptive combinations of 118 alleles. 119 Multiple lines of evidence suggest that more hybridization and adaptive introgression 120 took place in SSI populations than other Caribbean island populations, consistent with the 121 hypothesis that hybridization triggered adaptive radiation. First, the strongest signal of 122 introgression across the Caribbean was into the root node of the SSI radiation (Fig. 3B). Second, 123 trophic specialists on San Salvador experienced at least twice as much adaptive introgression as 124 other generalist populations across the Caribbean (P < 0.006; Fig. 3C). Third, based on 125 129 9 130 Fig. 3. A history of hybridization across the Caribbean. A) A summary map of adaptive 131 introgression into SSI trophic specialists from focal generalist populations across the Caribbean, 132 with thickness of arrows proportional to the number of outlier regions of the fd statistic. B) 133 149 (mediated through foraging behavior: scale-eating or snail-eating), trophic morphology, and 150 sexual communication signals. Based on both GO annotations of genes near candidate adaptive 151 variants and genome-wide association mapping (GWAS) in Caribbean pupfishes for ...