Genomic analysis of Klebsiella pneumoniae causing community-acquired respiratory deaths among Zambian infants and children using targeted RNA-probe hybridization-capture metagenomics.

Klebsiella pneumoniae (Kp) is a leading cause of neonatal and infant deaths in sub-Saharan Africa and frequently associated with antimicrobial resistance. Previously, we identified Kp as a major cause of fatal community-associated lower respiratory infections among infants and children under five years in Lusaka, Zambia, using postmortem tissue sampling and pathogen specific multiplex qPCR. In this follow-up study, we employed a novel culture-independent RNA-probe hybridization-capture metagenomic sequencing approach, targeting Kp pan-genome core and accessory genes, to perform in-depth genomic analysis of Kp from eleven post-mortem lung biopsy samples from seven of these children. Analysis detected Kp in all cases except one, which identified Klebsiella quasipneumoniae subspecies similipneumoniae. Core-genome multi-locus sequence typing (cgMLST) revealed six clonal groups (CG607, CG1123, CG10072, CG280, CG3648, and CG10344) belonging to five sublineages (SL607, SL17, SL280, SL37, and SL10072), with perfect concordance between paired samples from the same case. Two infants sampled the same month harbored SL607 lineages sharing 621 out of 629 cgMLST alleles, suggesting clonal spread. Kp capsule (K) loci were detected in all but one case and included potential vaccine targets KL25, KL23, and KL122. Antimicrobial resistance genes were widespread among samples, particularly encoding resistance toward aminoglycosides, β-lactams, sulphonamides, tetracyclines, and trimethoprim. Extended spectrum β-lactamases were identified in four cases, three of which were bla CTX-M-15. The acquired Kp sideophore yersiniabactin (lineage ybt14) was identified in both cases associated with SL607, and the acquired siderophore aerobactin (lineage iuc5) was identified in one of these, suggesting possible convergence of antimicrobial resistance and hypervirulence. The detection of Kp with extensive antimicrobial resistance causing fatal community acquired pneumonia signals a deeply concerning epidemiologic shift from a largely nosocomial pathogen. This calls for urgent epidemiological investigations to better understand the burden, transmission dynamics, antimicrobial resistances, and potential vaccine targets for Kp in other community settings across sub-Saharan Africa.