AWWA WQTC62553

Human noroviruses (NoVs) are the leading cause of non-bacterial gastroenteritis worldwide with numerous outbreaks being attributed to consumption of NoV-contaminated water. Based on epidemiological outbreak investigations and limited human volunteer challenge studies, it has been reported that NoVs are resistant to environmental degradation and chemical inactivation. The absence of a cell culture infectivity model for human NoVs has required the use of molecular methods to monitor changes in NoV nucleic acid persistence in conjunction with the use of viral surrogates amenable to cell culture. To evaluate the stability of human NoVs in source water used by water treatment plants, the prototype genogroup I NoV; Norwalk virus (NV) and the viral surrogates feline calicivirus (FCV), poliovirus (PV) and MS2 bacteriophage were seeded into surface (n=3) and ground (n=2) waters used as source water for drinking water treatment plants. Infectivity of the viral surrogates and reduction of viral nucleic acid for all the viruses was monitored using cell culture-based viral infectivity and quantitative reverse transcription-PCR (qRT-PCR), respectively. qRT-PCR measures the increase of PCR product during each amplification cycle by continuously monitoring the intensity of a target-specific fluorescent reporter oligoprobe. The oligoprobe produces a signal proportional to the amount of generated PCR product. Template concentration is determined based on the threshold positive cycle value. The threshold cycle (i.e. Ct) is the cycle at which the fluorescent reporter signal is first observed over background. PCR product detection occurs in real-time during the exponential phase of the reaction rather than following completion of the total reaction (30-40 cycles) at the plateau stage. Therefore, qRT-PCR is less affected by amplification limiting components during amplification and better represents initial template concentration. Moreover, the amplicon detection and confirmation are conducted at the same time by incorporating a fluorescent reporter probe saving time and effort during amplicon confirmation.