Inhibition of multi-drug resistant Klebsiella pneumoniae: Nanoparticles induced photoinactivation in presence of efflux pump inhibitor

Abstract

In the current scenario, frontline antibiotics are losing effectiveness against multidrug-resistant (MDR) bacteria because of the single mode of action. The accumulation of mutations and spread of antibiotic resistance markers among the bacteria results into the severe threat to community health. Now, there is an urgent requirement for the development of an alternate and as well as multiple-targeted action of drugs to stop the spread of resistance in bacteria. Here, we showed an alternative nanoparticle based photodynamic therapy (PDT) targeting the bacterial efflux pumps and its cell wall. The dextran capped gold nanoparticles (GNPDEX) were localized to the bacterial surface by nanoparticle attached Concanavalin-A (ConA), where GNPDEX attached methylene blue (MB) photosensitizer as an MB@GNPDEX-ConA formulation induced the killing of MDR Klebsiella pneumoniae clinical isolates in no time. The intervention of efflux pump inhibitor (EPI) further improved the MB@GNPDEX-ConA treatment modality and displayed the maximum bactericidal cytoplasmic phototoxicity. The CCCP EPI (carbonyl cyanide m-chlorophenylhydrazone) with the PDT increased the bacterial killing by>3 log10 as compared with or without EPI intervention. Further, the fractionated (two light treatment after long dark phase) PDT treatment modality decreased the bacterial biofilm growth up to ~90%. The microscopic as well as ROS fluorescent probes showed the singlet oxygen mediated cytotoxicity. The mode of interactions and genomic DNA photo-toxicity confirmed that EPI enhanced the killing mediated by singlet oxygen generation. The multi-targeted (Cell wall, DNA and efflux pump) modality of MB@GNPDEX-ConA in presence of EPI is an effective and alternative therapeutic approach against most potent Klebsiella MDR infections.