Department of Civil and Environmental Engineering, National University of Singapore, Singapore, Singapore.
J Appl Microbiol. 2022 Mar;132(3):1788-1801. doi: 10.1111/jam.15325. Epub 2021 Oct 20.
This study aimed to investigate the physiological responses of two gram-negative bacteria (Escherichia coli and Pseudomonas aeruginosa) and two gram-positive bacteria (Enterococcus faecalis and Bacillus sphaericus) to ultraviolet (UV) and chlorine disinfection.
Bacterial inactivation by UV and chlorine disinfection were evaluated with a plate count method for culturability, FCM and PMA-qPCR for membrane integrity and DyeTox13-qPCR for enzymatic activity, respectively. Both UV and chorine disinfection caused complete loss of culturability while membrane integrity remained intact after UV disinfection. Both DyeTox13-qPCR and PMA-qPCR showed high ΔC values up to 8.9 after chlorine disinfection, indicating that both methods were able to distinguish non-treated from chlorine-treated cells. Although PMA-qPCR could not differentiate membrane integrity of cells on UV exposure, DyeTox13-qPCR showed significant differences in ΔC values of 5.05 and 10.4 for gram-negative (E. coli) and gram-positive (Enterococcus) bacteria, respectively. However, DyeTox13-qPCR for gram-negative bacteria displayed relatively small differences in ΔC values compared with gram-positive bacteria.
UV and chlorine disinfection led to changes in physiological state of gram-negative and gram-positive bacteria. Particularly, UV disinfection could induce active but non-culturable (ABNC) for gram-negative bacteria and dormant cell for gram-positive bacteria where intact cells no longer showed the enzymatic activity.
UV and chlorine are commonly used to disinfect water, food and fomites to inactivate pathogenic bacteria. However, a viable but non-culturable (VBNC) state of bacteria induced by disinfection may underestimate the health risks because of the potential resuscitation of VBNC cells. This study highlighted that bacteria could undergo different physiological (ABNC or dormant) states during UV and chlorine disinfection. In addition, viability PCR techniques could provide insight into the changes in physiological states during disinfection processes.
本研究旨在探讨两种革兰氏阴性菌（大肠杆菌和铜绿假单胞菌）和两种革兰氏阳性菌（粪肠球菌和球形芽孢杆菌）对紫外线（UV）和氯消毒的生理反应。
使用平板计数法评估细菌的UV 和氯消毒杀菌效果，采用流式细胞术（FCM）和 PMA-qPCR 评估细胞膜完整性，采用 DyeTox13-qPCR 评估酶活性。UV 和氯消毒均可完全杀灭可培养菌，但 UV 消毒后细胞膜完整性仍然完整。氯消毒后，DyeTox13-qPCR 和 PMA-qPCR 的ΔC 值均高达 8.9，表明这两种方法均能区分未经处理和经氯处理的细胞。尽管 PMA-qPCR 不能区分 UV 暴露下细胞的细胞膜完整性，但 DyeTox13-qPCR 显示革兰氏阴性（大肠杆菌）和革兰氏阳性（粪肠球菌）细菌的ΔC 值分别有显著差异，分别为 5.05 和 10.4。然而，革兰氏阴性细菌的 DyeTox13-qPCR 与革兰氏阳性细菌相比，ΔC 值差异相对较小。
UV 和氯消毒导致革兰氏阴性和革兰氏阳性细菌生理状态发生变化。特别是，UV 消毒会诱导革兰氏阴性细菌产生活跃但不可培养（ABNC）和革兰氏阳性细菌休眠细胞，此时完整的细胞不再表现出酶活性。
UV 和氯常用于水、食品和污染物的消毒，以灭活致病菌。然而，消毒诱导的细菌存活但不可培养（VBNC）状态可能会低估健康风险，因为 VBNC 细胞可能会复苏。本研究强调，细菌在 UV 和氯消毒过程中可能会经历不同的生理（ABNC 或休眠）状态。此外，存活 PCR 技术可以深入了解消毒过程中生理状态的变化。
