Love D C, Vinjé J, Khalil S M, Murphy J, Lovelace G L, Sobsey M D
Department of Environmental Sciences and Engineering, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
J Appl Microbiol. 2008 Apr;104(4):1203-12. doi: 10.1111/j.1365-2672.2007.03646.x. Epub 2007 Nov 20.
To evaluate a PCR-based detection and typing method for faecal indicator viruses (F+ RNA coliphages) in water and shellfish, and apply the method for better understanding of the ecology and microbial source tracking potential of these viruses.
Water and shellfish samples were collected over 3 years at nine estuaries in the East, West and Gulf Coasts of the USA, providing 1033 F+ RNA coliphage isolates. F+ RNA coliphage genotyping rates by reverse transcriptase-PCR-reverse line blot (RLB) hybridization ranged from 94.7% to 100% among estuaries, and were not significantly different in oysters, clams, mussels or water (P = 0.8427). Twenty samples negative by RLB were nucleotide sequenced for confirmation, and to refine RLB probes. More F+ RNA coliphages were genotyped from colder water than warmer waters, while the water salinity did not affect F+ RNA coliphage levels.
RT-PCR-RLB was a robust method for detecting and genotyping F+ RNA coliphages from diverse coastal areas, which provided new information on the ecology of F+ RNA coliphages.
This performance-validated F+ RNA coliphage method can be used for faecal indicator monitoring and microbial source tracking, to protect recreational bathers and shellfish consumers from exposure to pathogenic virus and their disease risks.
评估一种基于聚合酶链反应（PCR）的检测和分型方法，用于检测水和贝类中的粪便指示病毒（F+RNA 噬菌体），并应用该方法更好地了解这些病毒的生态学和微生物源追踪潜力。
在美国东、西和墨西哥湾沿岸的九个河口，历时 3 年采集了水和贝类样本，共获得 1033 株 F+RNA 噬菌体分离株。通过逆转录聚合酶链反应-反向线印迹（RLB）杂交进行的 F+RNA 噬菌体基因分型率在各河口之间为 94.7%至 100%，在牡蛎、蛤、贻贝或水中无显著差异（P = 0.8427）。对 20 个 RLB 检测呈阴性的样本进行核苷酸测序以进行确认，并优化 RLB 探针。从较冷水中分型得到的 F+RNA 噬菌体比温暖水中更多，而水的盐度不影响 F+RNA 噬菌体水平。
逆转录聚合酶链反应-反向线印迹（RT-PCR-RLB）是一种用于检测来自不同沿海地区的 F+RNA 噬菌体并进行基因分型的可靠方法，为 F+RNA 噬菌体的生态学提供了新信息。
这种经过性能验证的 F+RNA 噬菌体方法可用于粪便指示监测和微生物源追踪，以保护休闲沐浴者和贝类消费者免受致病病毒暴露及其疾病风险。
