Real-time PCR-based infectivity assay for the titration of turkey hemorrhagic enteritis virus, an adenovirus, in live vaccines.

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  • Additional Information
    • Source:
      Publisher: Elsevier/North-Holland Biomedical Press Country of Publication: Netherlands NLM ID: 8005839 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1879-0984 (Electronic) Linking ISSN: 01660934 NLM ISO Abbreviation: J Virol Methods Subsets: MEDLINE
    • Publication Information:
      Original Publication: [Amsterdam] : Elsevier/North-Holland Biomedical Press, 1980-
    • Subject Terms:
      Viral Vaccines*; Adenoviridae/*immunology ; Adenoviridae/*isolation & purification ; Antigens, Viral/*analysis ; Real-Time Polymerase Chain Reaction/*methods; Adenoviridae/genetics ; Animals ; Antigens, Viral/genetics ; Cell Culture Techniques ; Sensitivity and Specificity ; Turkeys/virology ; Vaccines, Attenuated
    • Abstract:
      The current in vitro titration method for turkey hemorrhagic enteritis virus (THEV) is the end-point dilution assay (EPD) in suspension cell culture (CC). This assay is subjective and results in high variability among vaccine lots. In this study, a new in vitro infectivity method combining a SYBR Green I-based qPCR assay and CC was developed for titration of live hemorrhagic enteritis (HE) CC vaccines. The qPCR was used to determine the virus genome copy number (vGCN) of the internalized virus particles following inoculation of susceptible RP19 cells with 1 vaccine label dose. The measured vGCN represents the number of infectious viral particles (IVP) per 1 dose. This method was used to compare 9 vaccine lots from 3 companies in the United States. Significant lot-to-lot variations within the same company and among the various companies were found in genomic and qPCR-based infectious titer per label dose. A positive linear relationship was found between qPCR infectious titer and genomic titer. Further, considerable variations in CCID 50 titers were found among tested vaccine lots, indicating the high variability of the current titration methods. The new method provides an alternative to classical titration assays and can help reduce variation among HE vaccine products.
      (Copyright © 2016 Elsevier B.V. All rights reserved.)
    • Contributed Indexing:
      Keywords: Cell culture; Hemorrhagic enteritis; PCR; Turkey; Vaccine titration
    • Accession Number:
      0 (Antigens, Viral)
      0 (Vaccines, Attenuated)
      0 (Viral Vaccines)
    • Publication Date:
      Date Created: 20161110 Date Completed: 20171016 Latest Revision: 20171222
    • Publication Date:
      20221213
    • Accession Number:
      10.1016/j.jviromet.2016.11.002
    • Accession Number:
      27829121