Sanguinarine synergistically potentiates aminoglycoside-mediated bacterial killing.

Aminoglycosides are one of the oldest classes of antimicrobials that are being used in current clinical practice, especially on multi-drug resistant Gram-negative pathogenic bacteria. However, the serious side effects at high dosage such as ototoxicity, neuropathy and nephrotoxicity limit their applications in clinical practice. Approaches that potentiate aminoglycoside killing could lower down their effective concentrations to a non-toxic dosage for clinical treatment. In this research, we screened a compound library and identified sanguinarine that acts synergistically with various aminoglycosides. By checkerboard and dynamical killing assay, we found that sanguinarine effectively potentiated aminoglycoside killing on diverse bacterial pathogens, including Escherichia coli, Acinetobacter baumannii, Klebsiella pneumonia and Pseudomonas aeruginosa. The mechanistic studies showed an elevated intracellular ROS and DNA oxidative level in the bacterial cells treated by a combination of sanguinarine with aminoglycosides. Furthermore, an enhanced level of sanguinarine was observed in bacteria in the presence of aminoglycosides, suggesting that aminoglycosides promote the uptake of sanguinarine. Importantly, sanguinarine was shown to promote the elimination of persister cells and established biofilm cells both in vivo and in vitro. Our study provides a novel insight for approaches to lower down the clinical dosages of aminoglycosides.

Non-walled spherical Acinetobacter baumannii is an important type of persister upon ß-lactam antibiotic treatment.

Bacterial persistence is one of the major causes of antibiotic treatment failure and the step stone for antibiotic resistance. However, the mechanism by which persisters arise has not been well understood. Maintaining a dormant state to prevent antibiotics from taking effect is believed to be the fundamental mechanistic basis, and persisters normally maintain an intact cellular structure. Here we examined the morphologies of persisters in Acinetobacter baumannii survived from the treatment by three major classes of antibiotics (i.e. ß-lactam, aminoglycoside, and fluoroquinolone) with microcopy and found that a fraction of enlarged spherical bacteria constitutes a major sub-population of bacterial survivors from ß-lactam antibiotic treatment, whereas survivors from the treatment of aminoglycoside and fluoroquinolone were less changed morphologically. Further studies showed that these spherical bacteria had completely lost their cell wall structures but could survive without any osmoprotective reagent. The spherical bacteria were not the viable-but-non-culturable cells and they could revive upon the removal of ß-lactam antibiotics. Importantly, these non-walled spherical bacteria also persisted during antibiotic therapy in vivo using Galleria mellonella as the infection model. Additionally, the combinational treatment on A. baumannii by ß-lactam and membrane-targeting antibiotic significantly enhanced the killing efficacy. Our results indicate that in addition to the dormant, structure intact persisters, the non-wall spherical bacterium is another important type of persister in A. baumannii. The finding suggests that targeting the bacterial cell membrane during ß-lactam chemotherapy could enhance therapeutic efficacy on A. baumannii infection, which might also help to reduce the resistance development of A. baumannii.