Department of Plant Pathology and Ecology, The Connecticut Agricultural Experiment Station, New Haven, Connecticut, USA.
Department of Plant Pathology and Environmental Microbiology, The Pennsylvania State University, University Park, Pennsylvania, USA.
mBio. 2021 Apr 13;12(2):e00161-21. doi: 10.1128/mBio.00161-21.
Antimicrobial treatment of bacteria often results in a small population of surviving tolerant cells, or persisters, that may contribute to recurrent infection. Antibiotic persisters are metabolically dormant, but the basis of their persistence in the presence of membrane-disrupting biological compounds is less well understood. We previously found that the model plant pathogen pv. 1448A () exhibits persistence to tailocin, a membrane-disrupting biocontrol compound with potential for sustainable disease control. Here, we compared physiological traits associated with persistence to tailocin and to the antibiotic streptomycin and established that both treatments leave similar frequencies of persisters. Microscopic profiling of treated populations revealed that while tailocin rapidly permeabilizes most cells, streptomycin treatment results in a heterogeneous population in the redox and membrane permeability state. Intact cells were sorted into three fractions according to metabolic activity, as indicated by a redox-sensing reporter dye. Streptomycin persisters were cultured from the fraction associated with the lowest metabolic activity, but tailocin persisters were cultured from a fraction associated with an active metabolic signal. Cells from culturable fractions were able to infect host plants, while the nonculturable fractions were not. Tailocin and streptomycin were effective in eliminating all persisters when applied sequentially, in addition to eliminating cells in other viable states. This study identifies distinct metabolic states associated with antibiotic persistence, tailocin persistence, and loss of virulence and demonstrates that tailocin is highly effective in eliminating dormant cells. Populations of genetically identical bacteria encompass heterogeneous physiological states. The small fraction of bacteria that are dormant can help the population survive exposure to antibiotics and other stresses, potentially contributing to recurring infection cycles in animal or plant hosts. Membrane-disrupting biological control treatments are effective in killing dormant bacteria, but these treatments also leave persister-like survivors. The current work demonstrates that in , persisters surviving treatment with membrane-disrupting tailocin proteins have an elevated redox state compared to that of dormant streptomycin persisters. Combination treatment was effective in killing both persister types. Culturable persisters corresponded closely with infectious cells in each treated population, whereas the high-redox and unculturable fractions were not infectious. In linking redox states to heterogeneous phenotypes of tailocin persistence, streptomycin persistence, and infection capability, this work will inform the search for mechanisms and markers for each phenotype.
细菌的抗菌治疗通常会导致一小部分存活的耐受细胞，即持久性细胞，它们可能导致反复感染。抗生素持久性细胞代谢休眠，但它们在膜破坏生物化合物存在下持续存在的基础理解较少。我们之前发现，模式植物病原体 pv. 1448A () 对乳链菌肽表现出持久性，乳链菌肽是一种具有可持续疾病控制潜力的膜破坏生物防治化合物。在这里，我们比较了与乳链菌肽和抗生素链霉素相关的持久性相关的生理特征，并确定这两种处理方法都会留下相似频率的持久性细胞。用处理过的群体进行的显微镜分析显示，虽然乳链菌肽迅速使大多数细胞穿孔，但链霉素处理会导致氧化还原和膜通透性状态的异质群体。根据代谢活性，将完整细胞按三种分数进行分类，如氧化还原感应报告染料所示。从与最低代谢活性相关的分数中培养出链霉素持久性细胞，但从与活跃代谢信号相关的分数中培养出乳链菌肽持久性细胞。可培养分数的细胞能够感染宿主植物，而非可培养分数的细胞则不能。当顺序应用时，乳链菌肽和链霉素除了消除其他存活状态的细胞外，还能有效地消除所有持久性细胞。这项研究确定了与抗生素持久性、乳链菌肽持久性和毒力丧失相关的不同代谢状态，并表明乳链菌肽在消除休眠细胞方面非常有效。遗传上相同的细菌种群包含异质的生理状态。休眠细菌的一小部分可以帮助种群在暴露于抗生素和其他压力时存活下来，这可能导致动物或植物宿主的反复感染循环。膜破坏生物防治处理在杀死休眠细菌方面非常有效，但这些处理也会留下类似于持久性细胞的幸存者。目前的工作表明，在 pv. 1448A 中，与休眠链霉素持久性细胞相比，用膜破坏乳链菌肽蛋白处理后存活的持久性细胞具有更高的氧化还原状态。联合处理对两种持久性细胞都有效。每个处理过的群体中的可培养持久性细胞与感染细胞非常接近，而高氧化还原和不可培养的细胞则不具有感染性。通过将氧化还原状态与乳链菌肽持久性、链霉素持久性和感染能力的异质表型联系起来，这项工作将为每种表型的机制和标志物的研究提供信息。
