Regan J M, Oldenburg P S, Park H D, Harrington G W, Noguera D R
Department of Civil and Environmental Engineering, The Pennsylvania State University, University Park, PA 18802, USA.
Water Sci Technol. 2003;47(5):123-8.
A protocol for simultaneously interrogating bacterial viability and identity using in situ, culture-independent methods is described. Viability is assayed using ethidium monoazide (EMA) staining of cells with compromised membranes, and identity is determined using fluorescent in situ hybridization (FISH). Experiments with planktonic cultures were used to demonstrate the compatibility of EMA staining and FISH after covalently bonding EMA to nucleic acids by photoreaction. Applications to biofilm samples showed that diffusion limitations in the biofilm matrix were not problematic and that effective discrimination of viable target cells within a mixed microbial community was possible.
本文描述了一种使用原位、非培养方法同时检测细菌活力和鉴定细菌身份的方案。使用叠氮溴化乙锭（EMA）对细胞膜受损的细胞进行染色来检测活力，使用荧光原位杂交（FISH）来确定身份。通过光反应使EMA与核酸共价结合后，对浮游培养物进行的实验证明了EMA染色和FISH的兼容性。对生物膜样品的应用表明，生物膜基质中的扩散限制不成问题，并且能够有效区分混合微生物群落中的活靶细胞。
