Antibiotic tolerance, persistence, and resistance of the evolved minimal cell, Mycoplasma mycoides JCVI-Syn3B

Abstract

Summary Antibiotic resistance is a growing problem, but bacteria can evade antibiotic treatment via tolerance and persistence. Antibiotic persisters are a small subpopulation of bacteria that tolerate antibiotics due to a physiologically dormant state. Hence, persistence is considered a major contributor to the evolution of antibiotic-resistant and relapsing infections. Here, we used the synthetically developed minimal cell Mycoplasma mycoides JCVI-Syn3B to examine essential mechanisms of antibiotic survival. The minimal cell contains only 473 genes, and most genes are essential. Its reduced complexity helps to reveal hidden phenomenon and fundamental biological principles can be explored because of less redundancy and feedback between systems compared to natural cells. We found that Syn3B evolves antibiotic resistance to different types of antibiotics expeditiously. The minimal cell also tolerates and persists against multiple antibiotics. It contains a few already identified persister-related genes, although lacking many systems previously linked to persistence (e.g. toxin-antitoxin systems, ribosome hibernation genes).

Escherichia coli cells are primed for survival before lethal antibiotic stress

“…We did a similar FT with clones grown from a single cell and treated them with a lethal Amp concentration (0.1 mg/mL). Percent survival was determined for 1 h and 3 h of treatment using persister assays ( 27 , 28 , 38 ). Short-term tolerance at 1 h does not indicate the level of persistence at 3 h with lethal Amp; no correlation at 1 h vs 3 h population ( r 2 = 0.02).…”

Escherichia coli cells are primed for survival before lethal antibiotic stress

“…We did a similar FT with clones grown from a single cell and treated them with a lethal Amp concentration (0.1 mg/mL). Percent survival was determined for 1 h and 3 h of treatment using persister assays ( 27 , 28 , 38 ). Short-term tolerance at 1 h does not indicate the level of persistence at 3 h with lethal Amp; no correlation at 1 h vs 3 h population ( r 2 = 0.02).…”

Escherichia coli cells are primed for survival before lethal antibiotic stress

“…We did a similar FT with clones grown from a single cell and treated them with a lethal Amp concentration (0.1 mg/mL). % Survival was determined for 1 h and 3 h of treatment using persister assays 26,27,34 , and short-term tolerance at 1 h does not indicate the level of persistence at 3 h with lethal Amp; no correlation at 1 h vs 3 h population (r 2 = 0.02). If the primed cells are advantageous for long-term survival, we expect the high persister populations observed at 3 h treatment to stay high with more prolonged antibiotic exposure.…”

Escherichia coli cells are primed for survival before lethal antibiotic stress

“…We did a similar FT with clones grown from a single cell and treated them with a lethal Amp concentration (0.1 mg/mL). % Survival was determined for 1 h and 3 h of treatment using persister assays 26, 27, 34 , and short-term tolerance at 1 h does not indicate the level of persistence at 3 h with lethal Amp; no correlation at 1 h vs 3 h population (r 2 = 0.02). If the primed cells are advantageous for long-term survival, we expect the high persister populations observed at 3 h treatment to stay high with more prolonged antibiotic exposure.…”

Escherichia coli cells are primed for survival before lethal antibiotic stress

“…We did a similar FT with clones grown from a single cell and treated them with a lethal Amp concentration (0.1 mg/mL). Percent survival was determined for 1 h and 3 h of treatment using persister assays ( 27 , 28 , 38 ). Short-term tolerance at 1 h does not indicate the level of persistence at 3 h with lethal Amp; no correlation at 1 h vs 3 h population ( r 2 = 0.02).…”

Escherichia coli cells are primed for survival before lethal antibiotic stress

“…We did a similar FT with clones grown from a single cell and treated them with a lethal Amp concentration (0.1 mg/mL). % Survival was determined for 1 h and 3 h of treatment using persister assays 26,27,34 , and short-term tolerance at 1 h does not indicate the level of persistence at 3 h with lethal Amp; no correlation at 1 h vs 3 h population (r 2 = 0.02). If the primed cells are advantageous for long-term survival, we expect the high persister populations observed at 3 h treatment to stay high with more prolonged antibiotic exposure.…”

Escherichia coli cells are primed for survival before lethal antibiotic stress

“…We did a similar FT with clones grown from a single cell and treated them with a lethal Amp concentration (0.1 mg/mL). % Survival was determined for 1 h and 3 h of treatment using persister assays 26, 27, 34 , and short-term tolerance at 1 h does not indicate the level of persistence at 3 h with lethal Amp; no correlation at 1 h vs 3 h population (r 2 = 0.02). If the primed cells are advantageous for long-term survival, we expect the high persister populations observed at 3 h treatment to stay high with more prolonged antibiotic exposure.…”

Escherichia coli cells are primed for survival before lethal antibiotic stress

“…We did a similar FT with clones grown from a single cell and treated them with a lethal Amp concentration (0.1 mg/mL). Percent survival was determined for 1 h and 3 h of treatment using persister assays ( 27 , 28 , 38 ). Short-term tolerance at 1 h does not indicate the level of persistence at 3 h with lethal Amp; no correlation at 1 h vs 3 h population ( r 2 = 0.02).…”

Escherichia coli cells are primed for survival before lethal antibiotic stress

“…We did a similar FT with clones grown from a single cell and treated them with a lethal Amp concentration (0.1 mg/mL). % Survival was determined for 1 h and 3 h of treatment using persister assays 26,27,34 , and short-term tolerance at 1 h does not indicate the level of persistence at 3 h with lethal Amp; no correlation at 1 h vs 3 h population (r 2 = 0.02). If the primed cells are advantageous for long-term survival, we expect the high persister populations observed at 3 h treatment to stay high with more prolonged antibiotic exposure.…”

Escherichia coli cells are primed for survival before lethal antibiotic stress

“…We did a similar FT with clones grown from a single cell and treated them with a lethal Amp concentration (0.1 mg/mL). % Survival was determined for 1 h and 3 h of treatment using persister assays 26, 27, 34 , and short-term tolerance at 1 h does not indicate the level of persistence at 3 h with lethal Amp; no correlation at 1 h vs 3 h population (r 2 = 0.02). If the primed cells are advantageous for long-term survival, we expect the high persister populations observed at 3 h treatment to stay high with more prolonged antibiotic exposure.…”