Transferable AmpC hyperproduction in K. pneumoniae

Chromosomal and transferable AmpC ß-lactamases pose top resistance mechanisms in different Gram-negatives but knowledge regarding the latter, mostly about their regulation and virulence-related implications is far from being complete. To fill this gap we used Klebsiella pneumoniae (KP) and two different plasmid-encoded AmpCs [DHA-1, (AmpR regulator-linked, inducible) and CMY-2 (constitutive)] as models to perform a study in which we show that blockade of peptidoglycan recycling through AmpG permease inactivation abolished DHA-1 inducibility, but did not affect CMY-2 production, and neither altered KP pathogenic behavior. Moreover, whereas regular production of both AmpC-type enzymes did not attenuate KP virulence, when blaDHA-1 was expressed in an ampG-defective mutant, Galleria mellonella killing was significantly (but not drastically) attenuated. Spontaneous DHA-1 hyperproducer mutants were readily obtained in vitro, showing slight or insignificant virulence attenuations and high-level resistance to ß-lactams only mildly affected by basal production (e.g. ceftazidime, ceftolozane-tazobactam). By analyzing diverse DHA-1-harboring clinical KP strains we demonstrate that the natural selection of these hyperproducers is not exceptional (>10% of the collection), with the inactivation of the typical AmpC hyperproduction-related gene mpl being the most frequent underlying mechanism. The likely silent dissemination of this kind of strains, for which an important fitness cost-related contention barrier does not seem to exist, is envisaged as a neglected threat for most ß–lactams effectiveness including recently introduced combinations. Analyzing whether this phenomenon is applicable to other transferable ß–lactamases/species as well as determining the levels of conferred resistance, poses an essential topic to be approached.