Umetani Miki, Fujisawa Miho, Okura Reiko, Nozoe Takashi, Suenaga Shoichi, Nakaoka Hidenori, Kussell Edo, Wakamoto Yuichi
Research Center for Complex Systems Biology, The University of Tokyo, Tokyo, Japan.
Universal Biology Institute, The University of Tokyo, Tokyo, Japan.
Elife. 2025 May 13;14:e79517. doi: 10.7554/eLife.79517.
Bacterial persistence is a phenomenon in which a small fraction of isogenic bacterial cells survives a lethal dose of antibiotics. Although the refractoriness of persistent cell populations has classically been attributed to growth-inactive cells generated before drug exposure, evidence is accumulating that actively growing cell fractions can also generate persister cells. However, single-cell characterization of persister cell history remains limited due to the extremely low frequencies of persisters. Here, we visualize the responses of over one million individual cells of wildtype to lethal doses of antibiotics, sampling cells from different growth phases and culture media into a microfluidic device. We show that when cells sampled from exponentially growing populations were treated with ampicillin or ciprofloxacin, most persisters were growing before antibiotic treatment. Growing persisters exhibited heterogeneous survival dynamics, including continuous growth and fission with L-form-like morphologies, responsive growth arrest, or post-exposure filamentation. Incubating cells under stationary phase conditions increased both the frequency and the probability of survival of non-growing cells to ampicillin. Under ciprofloxacin, however, all persisters identified were growing before the antibiotic treatment, including samples from post-stationary phase culture. These results reveal diverse persister cell dynamics that depend on antibiotic types and pre-exposure history.
细菌持留是一种现象，即一小部分同基因细菌细胞在致死剂量的抗生素作用下存活下来。尽管持久性细胞群体的耐药性传统上归因于药物暴露前产生的生长不活跃细胞，但越来越多的证据表明，活跃生长的细胞部分也能产生持留细胞。然而，由于持留细胞的频率极低，对持留细胞历史的单细胞表征仍然有限。在这里，我们将超过一百万个野生型单细胞对致死剂量抗生素的反应可视化，从不同生长阶段和培养基中采集细胞并放入微流控装置中。我们发现，当用氨苄青霉素或环丙沙星处理从指数生长期群体中采集的细胞时，大多数持留细胞在抗生素处理前正在生长。正在生长的持留细胞表现出异质的存活动态，包括具有L型形态的持续生长和分裂、反应性生长停滞或暴露后丝状化。在稳定期条件下培养细胞会增加非生长细胞对氨苄青霉素的存活频率和概率。然而，在环丙沙星作用下，所有鉴定出的持留细胞在抗生素处理前都在生长，包括来自稳定后期培养的样本。这些结果揭示了依赖于抗生素类型和暴露前历史的多种持留细胞动态。
