This study examines the comparative analysis of Avian Pathogenic Escherichia coli (APEC) isolates through various bioinformatics tools, including phylogenetic analysis, 3D structure prediction, ADMET evaluation, molecular docking and molecular dynamics simulation. APEC strains significantly impact poultry health, leading to colibacillosis and subsequent economic losses in the poultry industry. The study underscores the importance of in silico methodologies in vaccine development, aiding in identifying antigens and optimizing vaccine design. We isolated E. coli strains from table and hatchable eggs using Tryptic Soy Agar, MacConkey Agar and Eosin Methylene Blue Agar media. The genomic analysis revealed approximately 4,288 protein-coding genes, highlighting the organism's complex structure and diverse antigenic types contributing to its virulence. The classification of E. coli based on serogroups and phylogroups enables a deeper understanding of its pathogenicity. By employing advanced bioinformatics techniques, we aim to elucidate the virulence factors associated with APEC strains and leverage this knowledge to enhance disease prevention strategies in poultry. This comprehensive analysis could pave the way for more effective vaccine formulations and improved management practices to mitigate the impact of APEC-related diseases in avian populations.