Abstract
Drug resistance traits are rapidly disseminated across bacteria by horizontal gene transfer, especially through plasmids. Plasmid curing agents that are active both in vitro and in vivo will resensitize Multi Drug Resistant (MDR) bacteria to antimicrobial agents. Pectin capped platinum nanoparticles (PtNPs) at sub MIC (20 µM) concentration was effective, in causing loss of Extended Spectrum Beta Lactamase (ESBL) harboring plasmid as evidenced by, absence of plasmid in agarose gel and by a concomitant (16–64 fold) drop in MIC for cell wall inhibitors ceftriaxone and meropenem, in carbapenem resistant Escherichia coli (CREC). Interestingly, the plasmid cured strain exhibited small colony morphology and displayed slower growth both in vitro and in vivo. Complementation of cured strain with plasmid from the wild type strain restored resistance towards meropenem and ceftriaxone. Relative to wild type, plasmid cured strain displayed 50% reduction in biofilm formation. Plasmid curing also occurred in vivo in infected zebrafish with curing efficiency of 17% for nanoparticle + meropenem treatment. PtNPs + meropenem reduced bioburden of CREC in infected zebrafish by 2.4 log CFU. Mechanistic studies revealed that nanoparticle interacted with cell surface and perturbed inner membrane integrity. PtNPs did not induce ROS, yet it caused plasmid DNA cleavage, as evidenced by gyrase inhibition assay. Our study for the first time reveals that PtNPs as plasmid curing agent can resensitize MDR bacteria to selective antimicrobial agents in vivo.
Highlights
Multi Drug Resistant (MDR) pathogens pose severe threat to public health, which has prompted WHO to declare 12 infectious agents as priority pathogens, urging researchers across the globe to devise urgent measures in the form of new antimicrobial agents or resistance modulatory agents to curtail these pathogens
Our results showed that pectin capped platinum nanoparticle can function at sub Minimum Inhibitory Concentration (MIC) as plasmid curing agent in MDR clinical isolate of E. coli both in vitro and in vivo in a zebrafish infection model
Attempts to explore the mechanism of curing using various studies like TEM imaging, membrane permeability, Reactive Oxygen Species (ROS), membrane potential and membrane integrity indicate that sub MIC levels of platinum nanoparticles (PtNPs) interacted with cell surface and compromised the inner membrane integrity without affecting cell viability
Summary
Multi Drug Resistant (MDR) pathogens pose severe threat to public health, which has prompted WHO to declare 12 infectious agents as priority pathogens, urging researchers across the globe to devise urgent measures in the form of new antimicrobial agents or resistance modulatory agents to curtail these pathogens. PtNPs rescues the fish from infection and promotes adaptive immune response against the pathogen, so much so that zebrafish is able to survive repetitive infection[17] Inspired from these studies, the plasmid curing effect of PtNPs was investigated for the first time. The objective of the present study is to evaluate potential of sub MIC levels of PtNPs to cure plasmid in vitro and in vivo to curtail drug resistance and explore the mechanism of curing mediated by PtNPs. Our results showed that pectin capped platinum nanoparticle can function at sub Minimum Inhibitory Concentration (MIC) as plasmid curing agent in MDR clinical isolate of E. coli both in vitro and in vivo in a zebrafish infection model. Gyrase inhibition assay revealed that PtNP treatment, both in presence and absence of gyrase, induced DNA cleavage even at sub MIC concentrations which might account for plasmid curing ability of PtNPs
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