Abstract
<title>Abstract</title> <p> <italic>Klebsiella pneumoniae</italic> is a Gram-negative opportunistic pathogen responsible for a wide range of healthcare-associated infections and is recognized for its remarkable genomic plasticity. While comparative genomic studies have predominantly focused on protein-coding genes associated with antimicrobial resistance and virulence, intergenic regions (IGRs), which contain promoters, transcription factor-binding sites, untranslated regions, and other regulatory elements, remain poorly characterized at the population level. In this study, we performed a comparative genomic analysis of 451 <italic>K. pneumoniae</italic> genomes to investigate the diversity and evolutionary patterns of both coding sequences and IGRs. Pangenome analysis revealed an open pangenome. Phylogenomic and average nucleotide identity analyses demonstrated high genomic similarity while identifying distinct evolutionary lineages. Comparative analysis of IGRs revealed substantial sequence diversity and variable conservation, indicating that non-coding regions contribute to genome evolution alongside protein-coding genes. Gene presence–absence analysis revealed extensive variation in accessory gene content, while IGR-based comparisons demonstrated substantial diversity and variable conservation of non-coding regions across the analyzed genomes. These findings provide a comprehensive overview of both coding and non-coding genomic diversity in <italic>K. pneumoniae</italic> and highlight the importance of incorporating intergenic regions into comparative genomic studies to improve our understanding of bacterial evolution, adaptation, and genome organization. </p>