<p>Currently, red rot caused by <i>Colletotrichum falcatum</i> Went is the major limiting factor in sugarcane production in India, as it frequently breaks down resistance in commercial varieties due to the emergence of new variants. From phylogenetic studies, it has been inferred that high phenotypic variation is mainly due to isolates adapting to newer cultivars, resulting in the breakdown of resistance. Hence, further studies were conducted to characterise pathogenicity-related genes/ proteins by utilizing phylogenetically differentiated virulent pathotype – <i>Cf</i>671 (CF06) and the least virulent pathotype – <i>Cf</i>92020. In the proteomic approach, mycelial proteins, secreted proteins and proteins from host–pathogen interaction of both the pathotypes were analyzed and unique proteins from the virulent pathotype were identified. Similarly, under the genomic approach, suppression subtractive hybridisation (SSH) was used to identify differentially expressed genes in the virulent pathotype. In both approaches, differentially expressed genes/ proteins were characterized from the virulent pathotype and common proteins expressed in mycelium, secretome and during host–pathogen interaction were identified. During host–pathogen interaction, Cytochrome P450, HSP20, and Redoxin were identified as potential targets for conventional management with inhibitors and fungicides. Based on SSH and interactive proteome results, Cytochrome P450 (CYP) was selected for further analysis, and its role in <i>C. falcatum</i> pathogenicity has been shown using azole fungicide as an inhibitor. Molecular analyses on the CYP gene revealed that there was a complete suppression of gene expression in the canes treated with azole fungicide. These results identified a potential fungicide target site of <i>C. falcatum</i> to manage the disease through efficient fungicides.</p>