Abstract:

Under the stimulation of traditional disinfection techniques, bacteria could become viable but nonculturable (VBNC) for survival. When external stress is eliminated, VBNC bacteria that are not removed by disinfection technology could resuscitate during water distribution and storage, and resuscitated bacteria are likely to enter the human body and cause serious diseases. However, the formation and resuscitation mechanisms of VBNC bacteria induced by traditional disinfection techniques are still unknown. Therefore, this review conducted a systematic analysis based on 95 relevant articles, coupled with our research about disinfection control and bacterial dormancy of antibiotic resistant bacteria. First, the formation of VBNC bacteria under different water disinfection techniques was introduced and the potential formation mechanisms of VBNC bacteria were elucidated, which mainly involved stringent response, energy metabolism, general stress response system and toxin-antitoxin system. In addition, the resuscitation risks of VBNC pathogen and 13 methods for VBNC bacteria to resuscitate were summarized. Moreover, three different resuscitation mechanisms were summarized in this review, including natural resuscitation, resuscitation promoting factor（Rpf）resuscitation, and self-inducer resuscitation. After repairing cell damage and restoring redox balance and metabolic activity, VBNC bacteria could resuscitate spontaneously. VBNC bacteria could reshape cell walls and restore their cultivability with the help of Rpf. Due to the automatic inducer 2, intercellular communication in microbial populations and the expression of katG were increased, which reduced toxicity of hydrogen peroxide and triggered the resuscitation of VBNC bacteria. Finally, prospects for further studies were discussed, and the frontier technologies that could be used to study the resuscitation mechanisms of VBNC bacteria were introduced, such as microfluidics technology, stable isotope tracing method for metabolic activity detection, single-cell heavy water-labeled Raman spectroscopy method and Förster resonance energy transfer technology. This review provides important guidance for answering questions such as “how much dose of disinfection technology can inactivate VBNC bacteria and inhibit their resuscitation?” and reverted to those of wild-type bacteria?”. This review also provides a theoretical basis for evaluating the microbial safety in a water treatment process and developing more effective disinfection strategies.

Key words: antibiotic-resistant bacteria, viable but non-culturable state, resuscitation mechanisms, resuscitation promoting factor (Rpf), quorum sensing

摘要：

在传统水体消毒技术刺激下，细菌会进入活的不可培养（viable but nonculturable，VBNC）状态来提高自身存活率。在撤去外部压力时，未被消毒技术完全去除的VBNC细菌可在水储存和分配期间发生一定程度的复苏，而这些复苏的细菌很可能进入人体并导致严重疾病。然而，目前仍不清楚水体消毒过程中活的不可培养细菌的形成与复苏机制。该文通过检索了95篇相关文献并结合课题组在抗生素耐药菌方面的消毒控制和细菌休眠方面的研究进行了系统分析。首先，介绍了在不同水消毒技术下VBNC细菌的形成并阐述了其潜在的形成机制，其主要包括了严谨反应、能量代谢、一般应激反应系统和毒素-抗毒素系统。其次，介绍了VBNC致病菌复苏的风险和总结了13种复苏方法。该文还综述了自然复苏、复苏促进因子（Rpf）与自诱导剂复苏等3种复苏机制的不同。在修复了细胞损伤并恢复氧化还原平衡和代谢活性之后，VBNC细菌才能发生自然复苏。Rpf能够帮助VBNC细菌重塑细胞壁，这有助于VBNC细菌恢复可培养能力。自诱导剂-2能够促进微生物种群中的细胞间通讯并增加katG的表达来降低过氧化氢毒性，从而促进VBNC细菌的复苏。最后，对今后的研究方向进行了展望，介绍了微流控技术、稳定同位素示踪代谢活性分析方法、单细胞重水标记拉曼光谱方法和荧光能量共振转移技术等可以用于研究VBNC细菌复苏机制的前沿技术。该综述能为回答“多大剂量的消毒技术能够灭活VBNC细菌并避免其复苏”和“复苏的VBNC细菌生理特性是否都恢复到正常水平”等问题提供参考，为水处理过程中微生物安全性评估和制定更有效的消毒策略提供理论依据。

关键词: 抗生素耐药菌, 活的不可培养状态, 复苏机制, 复苏促进因子, 群体感应