Characterization of Escherichia coli Persisters from Biofilm Culture: Multiple Dormancy Levels and Multigenerational Memory in Formation

Persister cells (PCs), a subpopulation occurring within normal cells, exhibit a transient tolerance to antibiotics because of their dormant state. PCs are categorized into two types: type I PCs, which emerge during the stationary phase, and type II PCs, which emerge during the logarithmic phase. Using the conventional colony-forming method, we previously demonstrated that type I PCs of <i>Escherichia coli</i> form more frequently in air-solid biofilm culture than in liquid culture. In the current study, we modified a cell filamentation method as a more efficient and rapid alternative for quantifying PCs. This modified method yielded results consistent with those of the conventional method with 10³-10⁴ times higher sensitivity and less detection time, within several hours, and further revealed the existence of multiple levels of type I PCs, including a substantial number of deeply dormant cells. This study also discovered a potential epigenetic memory mechanism, spanning several generations (four or six cell divisions), which influences type II PC formation based on prior biofilm experience in <i>E. coli</i>.