Assessing the ability of Tigecycline and Meropenem to clear intra-macrophage Klebsiella pneumoniae

The emergence of hypervirulent Klebsiella pneumoniae of serotype K1 and K2 are a major cause of life-threatening, community-acquired infections. Recent studies demonstrated that Kp can persist for long periods of time within the spleen and liver and survive within macrophages. We aimed to explore whether two clinically relevant antimicrobials differed in their capacity to clear within-macrophage Kp. The mouse monocyte cell line J774a were used to model Kp macrophage infection (cultured in RPMI, 10% Fetal-bovine-serum, 37oC, 5%CO2). Cells were harvested and seeded in a 96-well plate at 2x106 cells/mL and incubated overnight. The following day, cells were infected with hypervirulent, K1 Kp (NTUH-K2044) for one hour at an MOI of 10. A 1-hour treatment with gentamicin and polymyxin-Bwas used to kill extracellular Kp. Cells were then washed, and incubated with serial 2-fold dilutions of meropenem and tigecycline for 6 hours. In parallel, a killing assay was performed with antibiotic, and Kp in cell culture media alone to compare the intracellular and extracellular activity of each antibiotic. We show that whilst the majority of the inoculum was resistant to phagocytosis, a small fraction of Kp were able to adhere to macrophages, enter the cell, and persist for up to 6h. Furthermore, we demonstrate that lower concentrations of tigecycline were required to inhibit intracellular Kp, compared with meropenem, which required concentrations in excess of the planktonic MIC to clear the intracellular niche. These data indicate that there is reason to re-examine the antimicrobial treatment regimens for hypervirulent Kp infection with a focus on intracellularly active drugs.