(639g) Z-Ring Formation As a Resuscitation Biomarker in Escherichia coli Persisters

Bacterial persisters, a subpopulation of cells that transiently survive antimicrobial treatment, are an ever-present and growing concern in the medical field. Alongside the more commonly known problem of bacterial resistance, persisters play a significant role in recurring infectious diseases [1]. A persister cell can exit the antibiotic-tolerance state after treatment (a process called resuscitation), repopulate the area and cause a recurring infection [2]. While research has been focused on preventing the mechanisms by which persistence develops, another viable option is inhibiting the pathways by which persisters resuscitate. Unfortunately, studying persisters cells is challenging due to their transient state and rarity, as well as the abundance of ‘viable but nonculturable’ cells, which hinders the isolation of persister cells. Using a persister-specific biomarker is one way to address this issue. A previous study in our lab revealed a plausible biomarker in the formation of Z-rings (mediated by the FtsZ protein) during the growth-inhibited state of Escherichia coli persisters [3]. However, as our study only examined the ofloxacin antibiotic, further investigation is needed to determine if the Z-ring can serve as a universal biomarker for E. coli persisters.

In this study we looked to confirm the formation of Z-rings for various persister cells obtained from a diverse range of antibiotics. E. coli cells ̶ harboring Z-ring reporter, in which the ftsZ gene is fused to a green fluorescent protein gene (gfp) ̶ treated with antibiotics Gentamicin, Streptomycin, Ampicillin, and Fosfomycin and then monitored on slides coated with agar medium using fluorescent microscopy. Images in both phase contrast and green fluorescence were taken to monitor the resuscitation of the cells (see Figure 1). Phase contrast was used for a visual of the individual cells, while green fluorescence was used to characterize Z-rings. Data shows a clear correlation between the number of organized Z-rings in a cell and its ability to resuscitate (see Figure 2). This approach successfully demonstrated that Z-rings are a reliable biomarker for identifying cells capable of resuscitation after treatment. With the Z-ring biomarker, we can directly characterize the persister cell physiology by isolating them from highly heterogeneous antibiotic-treated cultures.

[1] K. Lewis, “Persister cells,” Annual Review of Microbiology, vol. 64, no. 1, pp. 357–372, Jun. 2010.

[2] K. Lewis, “Persister cells, dormancy and infectious disease,” Nature Reviews Microbiology, vol. 5, no. 1, pp. 48–56, 2006.

[3] S. G. Mohiuddin, A. Massahi, and M. A. Orman, “lon deletion impairs Persister cell resuscitation in escherichia coli,” mBio, vol. 13, no. 1, 2022.