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Author (up) Monteiro, S.; Santos, R. doi  openurl
  Title Enzymatic and viability RT-qPCR assays for evaluation of enterovirus, hepatitis A virus and norovirus inactivation: Implications for public health risk assessment Type Journal Article
  Year 2018 Publication Journal of applied microbiology Abbreviated Journal J Appl Microbiol  
  Volume 124 Issue 4 Pages 965-976  
  Corporate Author Thesis  
  Address Laboratorio Analises, Instituto Superior Tecnico, Lisbon, Portugal  
  Keywords Caliciviridae Infections/virology; Enterovirus/*chemistry/genetics/growth & development/isolation & purification; Enterovirus Infections/virology; Hepatitis A/virology; Hepatitis A virus/*chemistry/genetics/growth & development/physiology; Hot Temperature; Humans; Norovirus/*chemistry/genetics/growth & development/physiology; Real-Time Polymerase Chain Reaction/*methods; Reverse Transcriptase Polymerase Chain Reaction; Ribonucleases/chemistry; Virus Inactivation; enzymatic PCR; heat inactivation; infectivity; public health; viability PCR; viruses  
  Abstract AIM: To assess the potential of a viability dye and an enzymatic reverse transcription quantitative PCR (RT-qPCR) pretreatment to discriminate between infectious and noninfectious enteric viruses. METHODS AND RESULTS: Enterovirus (EntV), norovirus (NoV) GII.4 and hepatitis A virus (HAV) were inactivated at 95 degrees C for 10 min, and four methods were used to compare the efficiency of inactivation: (i) cell culture plaque assay for HAV and EntV, (ii) RT-qPCR alone, (iii) RT-qPCR assay preceded by RNase treatment, and (iv) pretreatment with a viability dye (reagent D (RD)) followed by RT-qPCR. In addition, heat-inactivated NoV was treated with RD coupled with surfactants to increase the efficiency of the viability dye. No treatment was able to completely discriminate infectious from noninfectious viruses. RD-RT-qPCR reduced more efficiently the detection of noninfectious viruses with little to no removal observed with RNase. RD-RT-qPCR method was the closest to cell culture assay. The combination of surfactants and RD did not show relevant improvements on the removal of inactivated viruses signal compared with viability RT-qPCR, with the exception of Triton X-100. CONCLUSION: The use of surfactant/RD-RT-qPCR, although not being able to completely remove the signal from noninfectious viral particles, yielded a better estimation of viral infectivity. SIGNIFICANCE AND IMPACT OF THE STUDY: Surfactant/RD-RT-qPCR may be an advantageous tool for a better detection of infectious viruses with potential significant impact in the risk assessment of the presence of enteric viruses.  
  Publisher Place of Publication Editor  
  Language English Summary Language Original Title  
  Series Editor Series Title Abbreviated Series Title  
  Series Volume Series Issue Edition  
  ISSN 1364-5072 ISBN Medium  
  Area Expedition Conference  
  Notes PMID:28833965 Approved no  
  Call Number NCSU @ edshirle @ Serial 3608  
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