Pulmonary Disease and NF-κB-Associated Responses in Mice Infected with Two Klebsiella pneumoniae Isolates

Hypervirulent Klebsiella pneumoniae (hvKP) causes invasive disease, but pulmonary macrophage responses remain incompletely defined. We compared strain 55055, used as a classical K. pneumoniae (cKP) comparator, with strain 46274, operationally classified as hvKP, using a pneumonia model, single-cell RNA sequencing of pooled lungs, and RAW 264.7 macrophages. At the same inoculum, strain 46274 had higher lung burdens and greater mortality than strain 55055; representative sections showed more extensive inflammatory-cell infiltration, whereas no pairwise difference in animal-level H&E scores remained significant after multiple-comparison adjustment. Because neither pulmonary burden nor disease stage was matched, immune-response differences cannot be separated from effects of unequal bacterial exposure or differences in disease progression. Descriptive pooled-library analysis showed differences in immune-cell composition and CellChat-inferred communication. In the single day-2 strain 46274 pooled library, SPP1+ macrophages comprised a larger fraction than in the corresponding strain 55055 pooled library and had high inflammatory-response, chemokine, and bacterial-recognition gene-set scores; this library-level observation was not independently validated in individual animals. Tissue-level staining showed numerical day-2 differences in CD86- and CD206-positive-cell abundance, but adjusted pairwise comparisons did not confirm strain-specific differences and the data do not demonstrate M1/M2 macrophage polarization. NF-κB-associated imaging findings remained qualitative. In vitro, strain 46274 showed lower adhesion to and phagocytosis by RAW 264.7 macrophages, but higher IL-6 and TNF-α readouts and stronger qualitative late p65 nuclear localization. These in vitro observations were obtained in RAW 264.7 cells within one experimental run and should not be extrapolated directly to primary pulmonary macrophages. Overall, in this strain-pair murine model, strain 46274 was associated with more severe pulmonary disease and NF-κB-associated inflammatory readouts.