Therapeutically exploring persister metabolism in bacteria

Abstract

8Bacterial persisters are rare phenotypic variants that are temporarily tolerant to high 9 concentrations of antibiotics. We have previously discovered that persisters are mostly 10 derived from stationary-phase cells with high redox activities that are maintained by 11 endogenous protein and RNA degradation. This intracellular degradation resulted in self- 12 inflicted damage that transiently repressed the cellular functions targeted by antibiotics. 13 Leveraging this knowledge, we developed an assay integrating a degradable fluorescent 14 protein system and a small library, containing FDA-approved drugs and antibiotics, to detect 15 chemicals that target persister metabolism. We identified several metabolic inhibitors, 16 including anti-psychotic drugs, that can reduce Escherichia coli persistence. These chemical 17 inhibitors also reduce Pseudomonas aeruginosa persistence, potentially verifying the 18 existence of similar mechanisms in a medically relevant organism. 19 20 21 22 42high redox activities that are maintained by endogenous protein and RNA degradation (12). 43 We speculated that this intracellular degradation (i.e., self-digestion or autophagy) not only 44 provides energy to bacterial cells in a non-nutritive environment, but also produces self-45 inflicted damage that renders the cells less fit for rapid resumption of growth. Inhibiting 46 stationary-phase respiratory activities chemically (treatment with potassium cyanide or 47 nitric oxide to suppress cellular respiration), environmentally (culturing under anaerobic 48 3 conditions), or even genetically (genes encoding redox enzymes such as ubiF, sucB, mdh, 49 aceE, sdhC, and acnB) reduced persister levels by preventing digestion of endogenous 50 proteins and RNA, yielding cells that were more capable of translation and replication and 51 thus susceptible to cell death when exposed to antibiotics (12, 13). This reduction in 52 persister levels was not found to be associated with the inhibition of RNA and protein 53 synthesis or elimination of reactive oxygen species (ROS) (12). These results also suggest 54 persisters harbor ETC activities associated with bacterial cytochromes, oxidoreductases and 55 PMF, which was supported by previous studies, where "aminoglycoside (AG) potentiation 56 assays" were used (14-16). Our current study further provides strong support for the notion 57 that persister metabolism is a rich source of novel antipersister strategies. Using a high-58 throughput screening approach and a small chemical library (Biolog Phenotype Arrays 59 containing FDA-approved drugs and antibiotics) in the current study, we identified a subset 60 of drugs that can reduce persistence in Gram-negative bacteria by targeting their 61 metabolism. 62 RESULTS 63Chlorpromazine pretreatment can reduce E. coli persistence 64 As effective sterilization methods for treating chronic and recurrent infections remain 65 scarce, identifying novel targets, together with medicinally relevant inhibitors, is becoming 66 an urgent priority to im...

Identifying Metabolic Inhibitors to Reduce Bacterial Persistence

“…We would like to thank Dr. Aslan Massahi for providing assistance in persister assays. This manuscript has been released as a Pre-Print at bioRxiv (Mohiuddin et al, 2019).…”

Single-cell Raman spectroscopy identifies Escherichia coli persisters and reveals their enhanced metabolic activities

“…In order to effectively tackle microbial persisters, it is of great importance to understand how they form and figure out how they resuscitate (Song and Wood, 2020a). Traditional approaches investigate the resuscitation of persister cells by detecting the changes in their turbidity in broth (Curry et al, 2021), recording colony formation units on an agar plate (Mohiuddin et al, 2020a;Song and Wood, 2020b;Yamasaki et al, 2020), observing the resuscitation using time-lapse microscopy (Sulaiman and Lam, 2020), and performing flow cytometry (Mohiuddin et al, 2020b). In the present study, we have explored the changes in cellular components within E. coli persisters after their resuscitation using SCRS.…”