Use of a portable real-time reverse transcriptasepolymerase chain reaction assay for rapid detection of foot-and-mouth disease virus

Johnny D. Callahan Tetracore Inc, 11 Firstfield Rd, Gaithersburg, MD 20878.
Department of Pathology, School of Medicine, University of Maryland, Baltimore, MD, 21201.

Search for other papers by Johnny D. Callahan in
Current site
Google Scholar
PubMed
Close
 MS
,
Fred Brown Plum Island Animal Disease Center, Agricultural Research Service, US Department of Agriculture, PO Box 848, Greenport, NY 11944.

Search for other papers by Fred Brown in
Current site
Google Scholar
PubMed
Close
 PhD
,
Fernando A. Osorio Department of Veterinary and Biomedical Sciences, College of Agricultural Sciences and Natural Resources, University of Nebraska, Lincoln, NE 68583.

Search for other papers by Fernando A. Osorio in
Current site
Google Scholar
PubMed
Close
 MV, PhD, DACVM
,
Jung H. Sur Plum Island Animal Disease Center, Agricultural Research Service, US Department of Agriculture, PO Box 848, Greenport, NY 11944.

Search for other papers by Jung H. Sur in
Current site
Google Scholar
PubMed
Close
 DVM, PhD
,
Ed Kramer Plum Island Animal Disease Center, Agricultural Research Service, US Department of Agriculture, PO Box 848, Greenport, NY 11944.

Search for other papers by Ed Kramer in
Current site
Google Scholar
PubMed
Close
,
Gary W. Long Tetracore Inc, 11 Firstfield Rd, Gaithersburg, MD 20878.

Search for other papers by Gary W. Long in
Current site
Google Scholar
PubMed
Close
 PhD
,
Juan Lubroth Animal and Plant Health Inspection Service, US Department of Agriculture, PO Box 848, Greenport, NY 11944.

Search for other papers by Juan Lubroth in
Current site
Google Scholar
PubMed
Close
 DVM, PhD
,
Stefanie J. Ellis Tetracore Inc, 11 Firstfield Rd, Gaithersburg, MD 20878.

Search for other papers by Stefanie J. Ellis in
Current site
Google Scholar
PubMed
Close
 BS
,
Katina S. Shoulars Tetracore Inc, 11 Firstfield Rd, Gaithersburg, MD 20878.

Search for other papers by Katina S. Shoulars in
Current site
Google Scholar
PubMed
Close
 MS
,
Kristin L. Gaffney Tetracore Inc, 11 Firstfield Rd, Gaithersburg, MD 20878.

Search for other papers by Kristin L. Gaffney in
Current site
Google Scholar
PubMed
Close
 MS
,
Daniel L. Rock Plum Island Animal Disease Center, Agricultural Research Service, US Department of Agriculture, PO Box 848, Greenport, NY 11944.

Search for other papers by Daniel L. Rock in
Current site
Google Scholar
PubMed
Close
 PhD
, and
William M. Nelson Tetracore Inc, 11 Firstfield Rd, Gaithersburg, MD 20878.

Search for other papers by William M. Nelson in
Current site
Google Scholar
PubMed
Close
 MD

Abstract

Objective—To evaluate a portable real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assay designed to detect all 7 viral serotypes of footand- mouth disease virus (FMDV).

Design—Laboratory and animal studies.

Study Population—Viruses grown in tissue culture and animals experimentally infected with FMDV.

Procedure—1 steer, pig, and sheep were infected with serotype O FMDV. Twenty-four hours later, animals were placed in separate rooms that contained 4 FMDV-free, healthy animals of the same species. Oral and nasal swab specimens, oropharyngeal specimens obtained with a probang, and blood samples were obtained at frequent intervals, and animals were observed for fever and clinical signs of foot-and-mouth disease (FMD). Samples from animals and tissue cultures were assayed for infectious virus and viral RNA.

Results—The assay detected viral RNA representing all 7 FMDV serotypes grown in tissue culture but did not amplify a panel of selected viruses that included those that cause vesicular diseases similar to FMD; thus, the assay had a specificity of 100%, depending on the panel selected. The assay also met or exceeded sensitivity of viral culture on samples from experimentally infected animals. In many instances, the assay detected viral RNA in the mouth and nose 24 to 96 hours before the onset of clinical disease.

Conclusions and Clinical Relevance—The assay reagents are produced in a vitrified form, which permits storage and transportation at ambient temperatures. The test can be performed in 2 hours or less on a portable instrument, thus providing a rapid, portable, sensitive, and specific method for detection of FMDV. (J Am Vet Med Assoc 2002;220:1636–1642)

Abstract

Objective—To evaluate a portable real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assay designed to detect all 7 viral serotypes of footand- mouth disease virus (FMDV).

Design—Laboratory and animal studies.

Study Population—Viruses grown in tissue culture and animals experimentally infected with FMDV.

Procedure—1 steer, pig, and sheep were infected with serotype O FMDV. Twenty-four hours later, animals were placed in separate rooms that contained 4 FMDV-free, healthy animals of the same species. Oral and nasal swab specimens, oropharyngeal specimens obtained with a probang, and blood samples were obtained at frequent intervals, and animals were observed for fever and clinical signs of foot-and-mouth disease (FMD). Samples from animals and tissue cultures were assayed for infectious virus and viral RNA.

Results—The assay detected viral RNA representing all 7 FMDV serotypes grown in tissue culture but did not amplify a panel of selected viruses that included those that cause vesicular diseases similar to FMD; thus, the assay had a specificity of 100%, depending on the panel selected. The assay also met or exceeded sensitivity of viral culture on samples from experimentally infected animals. In many instances, the assay detected viral RNA in the mouth and nose 24 to 96 hours before the onset of clinical disease.

Conclusions and Clinical Relevance—The assay reagents are produced in a vitrified form, which permits storage and transportation at ambient temperatures. The test can be performed in 2 hours or less on a portable instrument, thus providing a rapid, portable, sensitive, and specific method for detection of FMDV. (J Am Vet Med Assoc 2002;220:1636–1642)

Advertisement