Inactivation conditions for human norovirus measured by an in situ capture-qRT-PCR method

Human norovirus (HuNoV) is a leading cause of foodborne gastroenteritis. Unfortunately, the inactivation parameters for HuNoV in clinical, food and environmental samples have not been established. Due to the inability to cultivate HuNoV in vitro, quantitative real-time RT-PCR (qRT-PCR) is widely-used for detecting HuNoVs. However, qRT-PCR does not indicate viral infectivity. Our method employs histo-blood group antigens (HBGAs) as viral receptors/co-receptors and container-affixed capture agents to concentrate HuNoVs. The captured viruses are denatured and its genome is amplified in the same module by in situ capture qRT-PCR (ISC-qRT-PCR). Greater than three log10 reduction in the receptor-captured viral genomic signal (RCVGS) was observed when HuNoV was treated by heat at 72 °C for 4 min, by chlorine at a final concentration of 16 ppm in less than 1 min, and by UV irradiation at 1J/cm². Treatment of low-titer HuNoV (<10³ copies/sample) with 70% ethanol for 20 s reduced the RCVGS of HuNoV by two log10. However, ethanol had a limited effect on high-titer samples of HuNoV (>10³ copies/sample). The results demonstrate that ISC-qRT-PCR method could be used as an alternative method to measure encapsidated viral RNA and indirectly indicate the inactivation status of HuNoV caused by physical treatment such as heat, and chemical treatment such as chlorine, that damage the ability of the virus to bind to its receptor.