<title>Abstract</title> Peppers (<italic>Capsicum</italic> spp.) rank among the most widely consumed spices globally. Fruit color, serving as a determinant for use in food colorants and cosmeceuticals and an indicator of nutritional content, significantly influences market quality and price. Cultivated <italic>Capsicum</italic> species display extensive phenotypic diversity, especially in fruit coloration. Our study leveraged the genetic variance within four <italic>Capsicum</italic> species (<italic>Capsicum baccatum, Capsicum chinense, Capsicum frutescens</italic>, and <italic>Capsicum annuum</italic>) to elucidate the genetic mechanisms driving color variation in peppers and related Solanaceae species. We analyzed color metrics and chromatic attributes (Red, Green, Blue, L*, a*, b*, Luminosity, Hue, and Chroma) on samples cultivated over six years (2015–2021). We resolved genomic regions associated with fruit color diversity through SNPs obtained from Genotyping by Sequencing (GBS) and genome-wide association study (GWAS) with a Multi-Locus Mixed Linear Model (MLMM). Significant SNPs with FDR correction were identified, within the Cytochrome P450, MYB-related genes, Pentatricopeptide repeat proteins, and ABC transporter family were the most common among the four species, indicating comparative evolution of fruit colors. We further validated the role of a pentatricopeptide repeat-containing protein (Chr01:31205460) and a cytochrome P450 enzyme (Chr08:45351919) via competitive allele-specific PCR (KASP) genotyping. Our findings advance the understanding of the genetic underpinnings of <italic>Capsicum</italic> fruit coloration, with developed KASP assays holding potential for applications in crop breeding and aligning with consumer preferences. This study provides a cornerstone for future research into exploiting <italic>Capsicum'</italic>s diverse fruit color variation.

Genetic Tapestry of Capsicum Fruit Colors: A Comparative Analysis of Four Cultivated Species