Detection of Human Cytomegalovirus (HCMV) — DNA from Granulocytes and Peripheral Blood Mononuclear Cells (PBMNC) of Immunosuppressed Patients by Nested PCR: Lack of Correlation to Isolation of Infectious Virus

  • K. Hamprecht
  • H.-J. Gerth
Chapter

Abstract

A rapid and very sensitive nested PCR method with primer oligonucleotides from the immediate early gene region of HCMV laboratory strain AD 169 was used to detect HCMVDNA from PBMC, granulocytes, urine, CSF, bronchoalveolar lavage and autopsy samples from bone marrow or kidney transplant recipients with clinically suspected primary or recurrent CMV-infection. Results of nested DNA-PCR were compared to isolation of infectious virus by conventional cell culture and culture-immunoperoxidase staining. Prior to amplification DNA was isolated from all specimens. Native urine was inhibitory to the action of Taq polymerase. Therefore urine samples were first concentrated with PEG 6000/ NaCl. In total 60 different specimens from 20 patients were analyzed. In 43 samples HCMV-DNA could be detected. Detection rates of HCMV-DNA from PBMC (14/21) and granulocytes (11/17) were nearly identical (67% and 65% respectively). Infectious virus however could be isolated in only one case. The initial detection rate of virus isolation from leukocytes prior to ganciclovir treatment was 19% (4/21). As shown for three patients ganciclovir treatment had no influence on HCMV-DNA detection from granulocytes, PBMC and urine (within the indicated time) but detection of initially infectious virus failed after ganciclovir administration. These results clearly demonstrate that detection of HCMV-DNA from granulocytes and PBMC (containing up to 20% monocytes) from immunosuppressed patients by DNA-PCR does not correlate to viremia, which has been established as predictive indicator for HCMV disease in renal and bone marrow transplantation. In this context the persistence of HCMVDNA in monocytes and granulocytes of asymptomatic seropositive subjects has to be discussed.

Keywords

Hepatitis Chloroform Glycol Sedimentation Electrophoresis 

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Copyright information

© Springer Science+Business Media New York 1994

Authors and Affiliations

  • K. Hamprecht
    • 1
  • H.-J. Gerth
    • 1
  1. 1.Department of Medical Virology and Epidemiology of Viral Diseases, Hygiene-InstituteUniversity of TübingenGermany

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