Rapid and High-Throughput pan-Orthopoxvirus Detection and Identification using PCR and Mass Spectrometry
ARMY MEDICAL RESEARCH INST OF INFECTIOUS DISEASES FORT DETRICK MD
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The genus Orthopoxvirus contains several species of related viruses, including the causative agent of smallpox Variola virus. Despite the eradication of smallpox, several other members of the genus are capable of causing human infection, including monkeypox, cowpox, and other zoonotic rodent-borne poxviruses. Therefore, a single assay that can accurately identify all orthopoxviruses could provide a valuable tool for rapid detection and surveillance. In this study, we developed a pan-Orthopoxvirus assay based on PCRelectrospray ionization-mass spectrometry PCRESI-MS on the Ibis T5000 system. The assay employs four PCR reactions targeting the orthopoxvirus DNA and RNA and helicase and polymerase genes. In this study, we evaluated this assay for its ability to correctly identify a panel of known orthopoxviruses and to determine the assays ability to detect and identify virus from the blood of rabbitpox virus-infected rabbits. We demonstrate that the assay can broadly detect and identify a diverse collection of orthopoxviruses and the assay is sensitive down to a few plaque-forming units per milliliter of blood and the stochastic limits of PCR.
- Medicine and Medical Research