Armored RNA as virus surrogate in a real-time reverse transcriptase PCR assay proficiency panel

S. K. Hietala, Beate Crossley

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

In recent years testing responsibilities for high-consequence pathogens have been expanded from national reference laboratories into networks of local and regional laboratories in order to support enhanced disease surveillance and to test for surge capacity. This movement of testing of select agents and high-consequence pathogens beyond reference laboratories introduces a critical need for standardized, noninfectious surrogates of disease agents for use as training and proficiency test samples. In this study, reverse transcription-PCR assay RNA targets were developed and packaged as armored RNA for use as a noninfectious, quantifiable synthetic substitute for four high-consequence animal pathogens: classical swine fever virus; foot-and-mouth disease virus; vesicular stomatitis virus, New Jersey serogroup; and vesicular stomatitis virus, Indiana serogroup. Armored RNA spiked into oral swab fluid specimens mimicked virus-positive clinical material through all stages of the reverse transcription-PCR testing process, including RNA recovery by four different commercial extraction procedures, reverse transcription, PCR amplification, and real-time detection at target concentrations consistent with the dynamic ranges of the existing real-time PCR assays. The armored RNA concentrations spiked into the oral swab fluid specimens were stable under storage conditions selected to approximate the extremes of time and temperature expected for shipping and handling of proficiency panel samples, including 24 h at 37°C and 2 weeks at temperatures ranging from ambient room temperature to -70°C. The analytic test performance, including the reproducibility over the dynamic range of the assays, indicates that armored RNA can provide a noninfectious, quantifiable, and stable virus surrogate for specific assay training and proficiency test purposes.

Original languageEnglish (US)
Pages (from-to)67-70
Number of pages4
JournalJournal of Clinical Microbiology
Volume44
Issue number1
DOIs
StatePublished - Jan 2006

Fingerprint

RNA Viruses
Reverse Transcriptase Polymerase Chain Reaction
Real-Time Polymerase Chain Reaction
RNA
Reverse Transcription
Temperature
Vesicular stomatitis New Jersey virus
Vesicular stomatitis Indiana virus
Surge Capacity
Classical swine fever virus
Viruses
Foot-and-Mouth Disease Virus
Polymerase Chain Reaction

ASJC Scopus subject areas

  • Microbiology (medical)
  • Microbiology

Cite this

Armored RNA as virus surrogate in a real-time reverse transcriptase PCR assay proficiency panel. / Hietala, S. K.; Crossley, Beate.

In: Journal of Clinical Microbiology, Vol. 44, No. 1, 01.2006, p. 67-70.

Research output: Contribution to journalArticle

@article{4bc4b4603de24c10959d136d9b73bfa0,
title = "Armored RNA as virus surrogate in a real-time reverse transcriptase PCR assay proficiency panel",
abstract = "In recent years testing responsibilities for high-consequence pathogens have been expanded from national reference laboratories into networks of local and regional laboratories in order to support enhanced disease surveillance and to test for surge capacity. This movement of testing of select agents and high-consequence pathogens beyond reference laboratories introduces a critical need for standardized, noninfectious surrogates of disease agents for use as training and proficiency test samples. In this study, reverse transcription-PCR assay RNA targets were developed and packaged as armored RNA for use as a noninfectious, quantifiable synthetic substitute for four high-consequence animal pathogens: classical swine fever virus; foot-and-mouth disease virus; vesicular stomatitis virus, New Jersey serogroup; and vesicular stomatitis virus, Indiana serogroup. Armored RNA spiked into oral swab fluid specimens mimicked virus-positive clinical material through all stages of the reverse transcription-PCR testing process, including RNA recovery by four different commercial extraction procedures, reverse transcription, PCR amplification, and real-time detection at target concentrations consistent with the dynamic ranges of the existing real-time PCR assays. The armored RNA concentrations spiked into the oral swab fluid specimens were stable under storage conditions selected to approximate the extremes of time and temperature expected for shipping and handling of proficiency panel samples, including 24 h at 37°C and 2 weeks at temperatures ranging from ambient room temperature to -70°C. The analytic test performance, including the reproducibility over the dynamic range of the assays, indicates that armored RNA can provide a noninfectious, quantifiable, and stable virus surrogate for specific assay training and proficiency test purposes.",
author = "Hietala, {S. K.} and Beate Crossley",
year = "2006",
month = "1",
doi = "10.1128/JCM.44.1.67-70.2006",
language = "English (US)",
volume = "44",
pages = "67--70",
journal = "Journal of Clinical Microbiology",
issn = "0095-1137",
publisher = "American Society for Microbiology",
number = "1",

}

TY - JOUR

T1 - Armored RNA as virus surrogate in a real-time reverse transcriptase PCR assay proficiency panel

AU - Hietala, S. K.

AU - Crossley, Beate

PY - 2006/1

Y1 - 2006/1

N2 - In recent years testing responsibilities for high-consequence pathogens have been expanded from national reference laboratories into networks of local and regional laboratories in order to support enhanced disease surveillance and to test for surge capacity. This movement of testing of select agents and high-consequence pathogens beyond reference laboratories introduces a critical need for standardized, noninfectious surrogates of disease agents for use as training and proficiency test samples. In this study, reverse transcription-PCR assay RNA targets were developed and packaged as armored RNA for use as a noninfectious, quantifiable synthetic substitute for four high-consequence animal pathogens: classical swine fever virus; foot-and-mouth disease virus; vesicular stomatitis virus, New Jersey serogroup; and vesicular stomatitis virus, Indiana serogroup. Armored RNA spiked into oral swab fluid specimens mimicked virus-positive clinical material through all stages of the reverse transcription-PCR testing process, including RNA recovery by four different commercial extraction procedures, reverse transcription, PCR amplification, and real-time detection at target concentrations consistent with the dynamic ranges of the existing real-time PCR assays. The armored RNA concentrations spiked into the oral swab fluid specimens were stable under storage conditions selected to approximate the extremes of time and temperature expected for shipping and handling of proficiency panel samples, including 24 h at 37°C and 2 weeks at temperatures ranging from ambient room temperature to -70°C. The analytic test performance, including the reproducibility over the dynamic range of the assays, indicates that armored RNA can provide a noninfectious, quantifiable, and stable virus surrogate for specific assay training and proficiency test purposes.

AB - In recent years testing responsibilities for high-consequence pathogens have been expanded from national reference laboratories into networks of local and regional laboratories in order to support enhanced disease surveillance and to test for surge capacity. This movement of testing of select agents and high-consequence pathogens beyond reference laboratories introduces a critical need for standardized, noninfectious surrogates of disease agents for use as training and proficiency test samples. In this study, reverse transcription-PCR assay RNA targets were developed and packaged as armored RNA for use as a noninfectious, quantifiable synthetic substitute for four high-consequence animal pathogens: classical swine fever virus; foot-and-mouth disease virus; vesicular stomatitis virus, New Jersey serogroup; and vesicular stomatitis virus, Indiana serogroup. Armored RNA spiked into oral swab fluid specimens mimicked virus-positive clinical material through all stages of the reverse transcription-PCR testing process, including RNA recovery by four different commercial extraction procedures, reverse transcription, PCR amplification, and real-time detection at target concentrations consistent with the dynamic ranges of the existing real-time PCR assays. The armored RNA concentrations spiked into the oral swab fluid specimens were stable under storage conditions selected to approximate the extremes of time and temperature expected for shipping and handling of proficiency panel samples, including 24 h at 37°C and 2 weeks at temperatures ranging from ambient room temperature to -70°C. The analytic test performance, including the reproducibility over the dynamic range of the assays, indicates that armored RNA can provide a noninfectious, quantifiable, and stable virus surrogate for specific assay training and proficiency test purposes.

UR - http://www.scopus.com/inward/record.url?scp=30744446773&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=30744446773&partnerID=8YFLogxK

U2 - 10.1128/JCM.44.1.67-70.2006

DO - 10.1128/JCM.44.1.67-70.2006

M3 - Article

VL - 44

SP - 67

EP - 70

JO - Journal of Clinical Microbiology

JF - Journal of Clinical Microbiology

SN - 0095-1137

IS - 1

ER -