Phenotypic Divergence Among Genomically Closely Related Klebsiella pneumoniae Clinical Isolates

Klebsiella pneumoniae can cause a wide array of infections and exhibit substantial phenotypic heterogeneity depending on the infection site. However, less is known about phenotypic heterogeneity among bacteria isolated from the same infection site. We screened K. pneumoniae clinical isolates taken from various infection sites and found that 4 of 29 isolates produced two colony morphologies on antibiotic-supplemented chromogenic agar, which we classified as solid and gelatinous based on their appearance. The paired colony phenotypes exhibited similar growth in rich and chemically defined media and comparable carbon-source utilization, but differed in antimicrobial susceptibility and virulence-associated traits. Gelatinous colonies tended to produce more capsular polysaccharide, and displayed greater colony spreading, whereas solid colonies produced more biofilm. To determine whether the divergent phenotypes could occur in a closely related genomic background, we performed Oxford Nanopore whole-genome sequencing of the solid and gelatinous colonies from MCFM21. Mapping of solid-colony reads to the gelatinous-colony assembly demonstrated 99.86% primary-read mapping with genome-wide coverage across the 5.29 Mb chromosome and 198 kb circular contig. Haploid variant calling identified one strongly supported single-nucleotide variant and 30 candidate indels. Together, these findings demonstrate that differences in K. pneumoniae colony morphology, capsule, biofilm, colony spreading, and antimicrobial susceptibility can occur between closely related populations recovered from the same clinical specimen.