Disease caused by Pepper yellow leaf curl virus (PepYLCV) is one of the greatest threats to pepper (Capsicum spp.) cultivation in the tropics and subtropics. Resistance to PepYLCV was previously identified in a few Capsicum accessions, but no resistance QTLs have been mapped. This study aimed to elucidate the genetics of PepYLCV resistance in C. annuum L. Augmented inoculation by the viruliferous whitefly Bemisia tabaci was used to evaluate parental lines and an F2 segregating population derived from a cross between resistant C. annuum line LP97 and susceptible C. annuum line ECW30R. Final evaluation was performed six weeks after inoculation using a standardized 5-point scale (0 = no symptoms to 4 = very severe symptoms). A high-density linkage map was constructed using genotyping-by-sequencing (GBS) to identify single-nucleotide polymorphism (SNP) markers associated with PepYLCV resistance in the F2 population. QTL analysis revealed three QTLs, peplcv-1, peplcv-7, and peplcv-12, on chromosomes P1, P7, and P12, respectively. Candidate genes associated with PepYLCV resistance in the QTL regions were inferred. In addition, single markers Chr7-LCV-7 and Chr12-LCV-12 derived from the QTLs were developed and validated in another F2 population and in commercial varieties. This work thus provides not only information for mapping PepYLCV resistance loci in pepper but also forms the basis for future molecular analysis of genes involved in PepYLCV resistance.