Dataset Information

Detection of monkeypox virus with real-time PCR assays.

ABSTRACT:

Background

Human monkeypox, a zoonotic disease, was first reported outside of Africa during the 2003 US outbreak.

Objectives

We present two real-time PCR assays critical for laboratory diagnosis of monkeypox during the 2003 US outbreak.

Study design

A TaqMan-based assay (E9L-NVAR) targets the orthopoxvirus DNA polymerase gene and detects Eurasian orthopoxviruses other than Variola. A hybridization assay, utilizing a MGB Eclipsetrade mark (Epoch Biosciences) probe, targets an envelope protein gene (B6R) and specifically detects monkeypox virus (MPXV). Assays were validated using coded orthopoxvirus DNA samples and used to evaluate lesion samples from five confirmed US monkeypox cases.

Results

E9L-NVAR did not detect variola (48 strains), North American orthopoxviruses (2), or DNA derived from non-poxviral rash illnesses. The assay reproducibly identified various concentrations of 13 Eurasian orthopoxvirus strains and was sensitive to 12.5 vaccinia genomes. The B6R assay recognized 15 different MPXV strains, while other orthopoxvirus (9) and bacteria (15) strains did not cross-react. Of the 13 human samples tested from confirmed cases, both assays identified 100% as containing MPXV DNA.

Conclusions

E9L-NVAR and B6R assays demonstrate 100% specificity for non-variola Eurasian orthopoxvirus and MPXV, respectively. Using two discrete viral gene targets, these assays together provide a reliable and sensitive method for quickly confirming monkeypox infections.

SUBMITTER: Li Y

PROVIDER: S-EPMC9628957 | biostudies-literature | 2006 Jul

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Detection of monkeypox virus with real-time PCR assays.

Li Yu Y Olson Victoria A VA Laue Thomas T Laker Miriam T MT Damon Inger K IK

Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology 20060530 3

<h4>Background</h4>Human monkeypox, a zoonotic disease, was first reported outside of Africa during the 2003 US outbreak.<h4>Objectives</h4>We present two real-time PCR assays critical for laboratory diagnosis of monkeypox during the 2003 US outbreak.<h4>Study design</h4>A TaqMan-based assay (E9L-NVAR) targets the orthopoxvirus DNA polymerase gene and detects Eurasian orthopoxviruses other than Variola. A hybridization assay, utilizing a MGB Eclipsetrade mark (Epoch Biosciences) probe, targets a ...[more]

PMID: 16731033

Similar Datasets

Project description:During the summer of 2003, an outbreak of human monkeypox occurred in the Midwest region of the United States. In all, 52 rodents suspected of being infected with monkeypox virus were collected from an exotic pet dealer and from private homes. The rodents were euthanized and submitted for testing to the United States Army Medical Research Institute of Infectious Diseases by the Galesburg Animal Disease Laboratory, Illinois Department of Agriculture. The rodent tissue samples were appropriately processed and then tested by using an integrated approach involving real-time polymerase chain reaction (PCR) assays, an antigen-detection immunoassay, and virus culture. We designed and extensively tested two specific real-time PCR assays for rapidly detecting monkeypox virus DNA using the Vaccinia virus F3L and N3R genes as targets. The assays were validated against panels of orthopox viral and miscellaneous bacterial DNAs. A pan-orthopox electrochemiluminescence (ECL) assay was used to further confirm the presence of Orthopoxvirus infection of the rodents. Seven of 12 (58%) animals (seven of 52 (15%) of all animals) tested positive in both monkeypox-specific PCR assays and two additional pan-orthopox PCR assays (in at least one tissue). The ECL results showed varying degrees of agreement with PCR. One hamster and three gerbils were positive by both PCR and ECL for all tissues tested. In addition, we attempted to verify the presence of monkeypox virus by culture on multiple cell lines, by immunohistology, and by electron microscopy, with negative results. Sequencing the PCR products from the samples indicated 100% identity with monkeypox virus strain Zaire-96-I-16 (a human isolate from the Congo). These real-time PCR and ECL assays represent a significant addition to the battery of tests for the detection of various orthopoxviruses. In light of the recent monkeypox virus transmissions, early detection of the virus is crucial for both natural outbreaks and potential acts of bioterrorism.

Dataset Information

Detection of monkeypox virus with real-time PCR assays.

Background

Objectives

Study design

Results

Conclusions

Publications

Detection of monkeypox virus with real-time PCR assays.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets