Walters Sarah P, Yamahara Kevan M, Boehm Alexandria B
Stanford University, Department of Civil and Environmental Engineering, 473 Via Ortega, M-06, Stanford, CA 94305, USA.
Water Res. 2009 Nov;43(19):4929-39. doi: 10.1016/j.watres.2009.05.047. Epub 2009 Jun 13.
In the last decade, the use of culture-independent methods for detecting indicator organisms and pathogens in recreational waters has increased and has led to heightened interest in their use for routine water quality monitoring. However, a thorough understanding of the persistence of genetic markers in environmental waters is lacking. In the present study, we evaluate the persistence of enterococci, enterovirus, and human-specific Bacteroidales in seawater microcosms. Two microcosms consisted of seawater seeded with human sewage. Two additional seawater microcosms were seeded with naked Enterococcus faecium DNA and poliovirus RNA. One of each replicate microcosm was exposed to natural sunlight; the other was kept in complete darkness. In the sewage microcosms, concentrations of enterococci and enterovirus were measured using standard culture-dependent methods as well as QPCR and RT-QPCR respectively. Concentrations of human-specific Bacteroidales were determined with QPCR. In the naked-genome microcosms, enterococci and enterovirus markers were enumerated using QPCR and RT-QPCR, respectively. In the sewage microcosm exposed to sunlight, concentrations of culturable enterococci fell below the detection limit within 5 days, but the QPCR signal persisted until the end of the experiment (day 28). Culturable enterococci did not persist as long as infectious enteroviruses. The ability to culture enteroviruses and enterococci was lost before detection of the genetic markers was lost, but the human-specific Bacteroidales QPCR signal persisted for a similar duration as infectious enteroviruses in the sewage microcosm exposed to sunlight. In the naked-genome microcosms, DNA and RNA from enterococci and enterovirus, respectively, persisted for over 10d and did not vary between the light and dark treatments. These results indicate differential persistence of genetic markers and culturable organisms of public health relevance in an environmental matrix and have important management implications.
在过去十年中，用于检测娱乐用水中指示生物和病原体的非培养方法的使用有所增加，并引发了人们对将其用于常规水质监测的浓厚兴趣。然而，目前仍缺乏对环境水体中遗传标记物持久性的全面了解。在本研究中，我们评估了海水微宇宙中肠球菌、肠道病毒和人类特异性拟杆菌的持久性。两个微宇宙由接种了人类污水的海水组成。另外两个海水微宇宙分别接种了裸露的粪肠球菌DNA和脊髓灰质炎病毒RNA。每个重复微宇宙中的一个暴露在自然阳光下；另一个则置于完全黑暗中。在污水微宇宙中，分别使用标准培养方法以及QPCR和RT-QPCR测量肠球菌和肠道病毒的浓度。使用QPCR测定人类特异性拟杆菌的浓度。在裸露基因组微宇宙中，分别使用QPCR和RT-QPCR对肠球菌和肠道病毒标记物进行计数。在暴露于阳光的污水微宇宙中，可培养肠球菌的浓度在5天内降至检测限以下，但QPCR信号一直持续到实验结束（第28天）。可培养肠球菌的持续时间不如传染性肠道病毒长。在遗传标记物检测消失之前，培养肠道病毒和肠球菌的能力就已经丧失，但在暴露于阳光的污水微宇宙中，人类特异性拟杆菌QPCR信号的持续时间与传染性肠道病毒相似。在裸露基因组微宇宙中，来自肠球菌和肠道病毒的DNA和RNA分别持续了超过10天，并且在光照和黑暗处理之间没有差异。这些结果表明，在环境基质中，与公共卫生相关的遗传标记物和可培养生物的持久性存在差异，具有重要的管理意义。
