Applicability of integrated cell culture reverse transcriptase quantitative PCR (ICC-RTqPCR) for the simultaneous detection of the four human enteric enterovirus species in disinfection studies

• Published in: Journal of virological methods Vol. 258; pp. 35 - 40
• Main Authors: Ryu, Hodon, Schrantz, Karen A., Brinkman, Nichole E., Boczek, Laura A.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.08.2018
• Subjects: Disinfection - methods; Enterovirus - growth & development; Enterovirus - isolation & purification; Enteroviruses; Environmental Microbiology; Humans; Multiplex integrated cell culture quantitative PCR; Multiplex Polymerase Chain Reaction - methods; Real-Time Polymerase Chain Reaction - methods; Reverse Transcriptase Polymerase Chain Reaction - methods; Ultraviolet Rays; UV disinfection; Virus Cultivation; Enteroviruses; UV disinfection; Multiplex integrated cell culture quantitative PCR
• ISSN: 0166-0934; 1879-0984
• DOI: 10.1016/j.jviromet.2018.05.008

•The ICC-RTqPCR method detects four infectious enteroviruses simultaneously.•This multiplex approach is successfully applied for use in disinfection studies.•A high-throughput format leads to more statistically sound data sets. A newly developed integrated cell culture reverse transcriptase quantitative PCR (ICC-RTqPCR) method and its applicability in UV disinfection studies is described. This method utilizes a cell culture system coupled with four RTqPCR assays to detect four species of human enterovirus (e.g., Enterovirus A, Enterovirus B, Enterovirus C and Enterovirus D). Evaluation of the RTqPCR assays was conducted with coxsackievirus A10, echovirus 30, poliovirus 1 and enterovirus 70 and resulted in 100% specificity for the tested assays. A comparison of ICC-RTqPCR between the individual enteroviruses and a mixture of all four viruses resulted in an approximate 1:1 correlation, demonstrating a lack of competition during incubation in cell culture and RTqPCR. The simultaneous detection of multiple human enterovirus species within mixed cultures is relevant to many applications, including virus disinfection studies. This high-throughput, multiplex approach costs less in money and time. By helping with data collection, this approach will lead to more statistically sound data sets that directly compare the inactivation rates of enteroviruses tested.