The escalating global population, diminishing agricultural lands, and the overarching global climate crisis are significantly impacting pepper cultivation. These challenges exacerbate the vulnerability of pepper plants to various biotic and abiotic factors, particularly viral diseases, resulting in diminished yield and quality. Cucumber mosaic virus (CMV), a notable concern for peppers in the Solanaceae family, is causing substantial quality and yield losses, with no effective chemical control methods currently available. This study focuses on exploring the genetic structure of CMV isolates obtained from pepper production areas in Kilis province and comparing these regions through the High-Resolution Melting (HRM) method. CMV isolates, cultivated in tobacco plants, have their partial coat protein sequences compared with those of other CMV isolates registered in the gene bank. The nucleotide sequences of identified CMV isolates are phylogenetically grouped and compared using an HRM graph. The HRM graph effectively distinguishes Kilis 3 and Kilis 4 isolates, clustered similarly to sequence patterns, from other isolates. The study highlights the utility of HRM analyses in identifying differences between isolates before determining sequence patterns. In the gene bank comparison, Kilis CMV isolates distinguished from others. Similarities were observed with isolates from Iran's Balsam (Impatiens balsamina - LC066478), Türkiye's Radish (Raphanus sativus - LC0665051), and Wild Turnip (Rapistrum rugosum - LC066514, LC066511, LC066517). The study found that Kilis 7 CMV isolate, transferred mechanically to 24 different pepper genotypes (C. annuum) from the local population, revealed susceptibility to CMV in the 24 lines developed from the Elbeyli population in Kilis province.