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Journal of Clinical Microbiology, April 1998, p. 1090-1095, Vol. 36, No. 4
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Nested PCR Assay for Detection of Granulocytic Ehrlichiae

Robert F. Massung,1,* Kim Slater,1 Jessica H. Owens,1 William L. Nicholson,1 Thomas N. Mather,2 Victoria B. Solberg,3 and James G. Olson1

National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 303331; Center for Vector-Borne Disease, University of Rhode Island, Kingston, Rhode Island 028812; and Department of Entomology, Walter Reed Army Institute of Research, Washington, D.C. 20307-51003

Received 6 August 1997/Returned for modification 10 November 1997/Accepted 12 January 1998

A sensitive and specific nested PCR assay was developed for the detection of granulocytic ehrlichiae. The assay amplifies the 16S rRNA gene and was used to examine acute-phase EDTA-blood and serum samples obtained from seven humans with clinical presentations compatible with human granulocytic ehrlichiosis. Five of the seven suspected cases were positive by the PCR assay using DNA extracted from whole blood as the template, compared with a serologic assay that identified only one positive sample. The PCR assay using DNA extracted from the corresponding serum samples as the template identified three positive samples. The sensitivity of the assay on human samples was examined, and the limit of detection was shown to be fewer than 2 copies of the 16S rRNA gene. The application of the assay to nonhuman samples demonstrated products amplified from template DNA extracted from Ixodes scapularis ticks collected in Rhode Island and from EDTA-blood specimens obtained from white-tailed deer in Maryland. All PCR products were sequenced and identified as specific to granulocytic ehrlichiae. A putative variant granulocytic ehrlichia 16S rRNA gene sequence was detected among products amplified from both the ticks and the deer blood specimens.

* Corresponding author. Mailing address: Centers for Disease Control and Prevention, 1600 Clifton Rd., MS G-13, Atlanta, GA 30333. Phone: (404) 639-1075. Fax: (404) 639-4436. E-mail: rfm2{at}cdc.gov.

Journal of Clinical Microbiology, April 1998, p. 1090-1095, Vol. 36, No. 4
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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Copyright © 1998 by the American Society for Microbiology. All rights reserved.