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. 1989 Apr;83(4):1109–1115. doi: 10.1172/JCI113990

Primer-mediated enzymatic amplification of cytomegalovirus (CMV) DNA. Application to the early diagnosis of CMV infection in marrow transplant recipients.

S A Cassol 1, M C Poon 1, R Pal 1, M J Naylor 1, J Culver-James 1, T J Bowen 1, J A Russell 1, S A Krawetz 1, R T Pon 1, D I Hoar 1
PMCID: PMC303796  PMID: 2539389

Abstract

A nucleic acid amplification procedure, the polymerase chain reaction (PCR), has been used to establish a diagnostic assay for the identification of cytomegalovirus (CMV) immediate-early sequences in clinical specimens. Preliminary testing against virus-infected cell cultures indicated that the PCR assay was highly CMV-specific, recognizing both wild-type and laboratory strains of CMV. There was no cross-reactivity with human DNA or with DNA from other herpes viruses. The sensitivity of the assay, using cloned CMV AD169 Eco RI fragment-J as template, was 1 viral genome per 40,000 cells. In a prospective study of CMV infection in bone marrow transplant recipients, the PCR assay correctly identified four patients with confirmed CMV infection. In three of these patients who were followed longitudinally, correlation of DNA reactivity with CMV culture and CMV antibody status over time indicated that DNA was the most sensitive marker for the diagnosis of CMV infection.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Akrigg A., Wilkinson G. W., Oram J. D. The structure of the major immediate early gene of human cytomegalovirus strain AD169. Virus Res. 1985 Mar;2(2):107–121. doi: 10.1016/0168-1702(85)90242-4. [DOI] [PubMed] [Google Scholar]
  2. Ambinder R. F., Charache P., Staal S., Wright P., Forman M., Hayward S. D., Hayward G. S. The vector homology problem in diagnostic nucleic acid hybridization of clinical specimens. J Clin Microbiol. 1986 Jul;24(1):16–20. doi: 10.1128/jcm.24.1.16-20.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Augustin S., Popow-Kraupp T., Heinz F. X., Kunz C. Problems in detection of cytomegalovirus in urine samples by dot blot hybridization. J Clin Microbiol. 1987 Oct;25(10):1973–1977. doi: 10.1128/jcm.25.10.1973-1977.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chou S., Merigan T. C. Rapid detection and quantitation of human cytomegalovirus in urine through DNA hybridization. N Engl J Med. 1983 Apr 21;308(16):921–925. doi: 10.1056/NEJM198304213081603. [DOI] [PubMed] [Google Scholar]
  5. Duggan D. B., Ehrlich G. D., Davey F. P., Kwok S., Sninsky J., Goldberg J., Baltrucki L., Poiesz B. J. HTLV-I-induced lymphoma mimicking Hodgkin's disease. Diagnosis by polymerase chain reaction amplification of specific HTLV-I sequences in tumor DNA. Blood. 1988 Apr;71(4):1027–1032. [PubMed] [Google Scholar]
  6. Emanuel D., Peppard J., Stover D., Gold J., Armstrong D., Hammerling U. Rapid immunodiagnosis of cytomegalovirus pneumonia by bronchoalveolar lavage using human and murine monoclonal antibodies. Ann Intern Med. 1986 Apr;104(4):476–481. doi: 10.7326/0003-4819-104-4-476. [DOI] [PubMed] [Google Scholar]
  7. Embury S. H., Scharf S. J., Saiki R. K., Gholson M. A., Golbus M., Arnheim N., Erlich H. A. Rapid prenatal diagnosis of sickle cell anemia by a new method of DNA analysis. N Engl J Med. 1987 Mar 12;316(11):656–661. doi: 10.1056/NEJM198703123161103. [DOI] [PubMed] [Google Scholar]
  8. Fleckenstein B., Müller I., Collins J. Cloning of the complete human cytomegalovirus genome in cosmids. Gene. 1982 Apr;18(1):39–46. doi: 10.1016/0378-1119(82)90054-3. [DOI] [PubMed] [Google Scholar]
  9. Hoar D. I., Bowen T., Matheson D., Poon M. C. Hepatitis B virus DNA is enriched in polymorphonuclear leukocytes. Blood. 1985 Dec;66(6):1251–1253. [PubMed] [Google Scholar]
  10. Hoar D. I., Haslam D. B., Starozik D. M. Improved direct molecular diagnosis and rapid fetal sexing. Prenat Diagn. 1984 Jul-Aug;4(4):241–247. doi: 10.1002/pd.1970040402. [DOI] [PubMed] [Google Scholar]
  11. Jackson J. B., Orr H. T., McCullough J. J., Jordan M. C. Failure to detect human cytomegalovirus DNA in IgM-seropositive blood donors by spot hybridization. J Infect Dis. 1987 Dec;156(6):1013–1016. doi: 10.1093/infdis/156.6.1013. [DOI] [PubMed] [Google Scholar]
  12. Kogan S. C., Doherty M., Gitschier J. An improved method for prenatal diagnosis of genetic diseases by analysis of amplified DNA sequences. Application to hemophilia A. N Engl J Med. 1987 Oct 15;317(16):985–990. doi: 10.1056/NEJM198710153171603. [DOI] [PubMed] [Google Scholar]
  13. Kwok S., Mack D. H., Mullis K. B., Poiesz B., Ehrlich G., Blair D., Friedman-Kien A., Sninsky J. J. Identification of human immunodeficiency virus sequences by using in vitro enzymatic amplification and oligomer cleavage detection. J Virol. 1987 May;61(5):1690–1694. doi: 10.1128/jvi.61.5.1690-1694.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Loche M., Mach B. Identification of HIV-infected seronegative individuals by a direct diagnostic test based on hybridisation to amplified viral DNA. Lancet. 1988 Aug 20;2(8608):418–421. doi: 10.1016/s0140-6736(88)90412-6. [DOI] [PubMed] [Google Scholar]
  15. Meyers J. D., Flournoy N., Thomas E. D. Nonbacterial pneumonia after allogeneic marrow transplantation: a review of ten years' experience. Rev Infect Dis. 1982 Nov-Dec;4(6):1119–1132. doi: 10.1093/clinids/4.6.1119. [DOI] [PubMed] [Google Scholar]
  16. Ou C. Y., Kwok S., Mitchell S. W., Mack D. H., Sninsky J. J., Krebs J. W., Feorino P., Warfield D., Schochetman G. DNA amplification for direct detection of HIV-1 in DNA of peripheral blood mononuclear cells. Science. 1988 Jan 15;239(4837):295–297. doi: 10.1126/science.3336784. [DOI] [PubMed] [Google Scholar]
  17. Peden K., Mounts P., Hayward G. S. Homology between mammalian cell DNA sequences and human herpesvirus genomes detected by a hybridization procedure with high-complexity probe. Cell. 1982 Nov;31(1):71–80. doi: 10.1016/0092-8674(82)90406-8. [DOI] [PubMed] [Google Scholar]
  18. Pritchett R. F. DNA nucleotide sequence heterogeneity between the Towne and AD169 strains of cytomegalovirus. J Virol. 1980 Oct;36(1):152–161. doi: 10.1128/jvi.36.1.152-161.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Rasmussen L., Kelsall D., Nelson R., Carney W., Hirsch M., Winston D., Preiksaitis J., Merigan T. C. Virus-specific IgG and IgM antibodies in normal and immunocompromised subjects infected with cytomegalovirus. J Infect Dis. 1982 Feb;145(2):191–199. doi: 10.1093/infdis/145.2.191. [DOI] [PubMed] [Google Scholar]
  20. Rüger R., Bornkamm G. W., Fleckenstein B. Human cytomegalovirus DNA sequences with homologies to the cellular genome. J Gen Virol. 1984 Aug;65(Pt 8):1351–1364. doi: 10.1099/0022-1317-65-8-1351. [DOI] [PubMed] [Google Scholar]
  21. Saiki R. K., Scharf S., Faloona F., Mullis K. B., Horn G. T., Erlich H. A., Arnheim N. Enzymatic amplification of beta-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science. 1985 Dec 20;230(4732):1350–1354. doi: 10.1126/science.2999980. [DOI] [PubMed] [Google Scholar]
  22. Saltzman R. L., Quirk M. R., Jordan M. C. Disseminated cytomegalovirus infection. Molecular analysis of virus and leukocyte interactions in viremia. J Clin Invest. 1988 Jan;81(1):75–81. doi: 10.1172/JCI113313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Schrier R. D., Nelson J. A., Oldstone M. B. Detection of human cytomegalovirus in peripheral blood lymphocytes in a natural infection. Science. 1985 Nov 29;230(4729):1048–1051. doi: 10.1126/science.2997930. [DOI] [PubMed] [Google Scholar]
  24. Schuster V., Matz B., Wiegand H., Traub B., Kampa D., Neumann-Haefelin D. Detection of human cytomegalovirus in urine by DNA-DNA and RNA-DNA hybridization. J Infect Dis. 1986 Aug;154(2):309–314. doi: 10.1093/infdis/154.2.309. [DOI] [PubMed] [Google Scholar]
  25. Shibata D. K., Arnheim N., Martin W. J. Detection of human papilloma virus in paraffin-embedded tissue using the polymerase chain reaction. J Exp Med. 1988 Jan 1;167(1):225–230. doi: 10.1084/jem.167.1.225. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  27. Spector S. A., Rua J. A., Spector D. H., McMillan R. Detection of human cytomegalovirus in clinical specimens by DNA-DNA hybridization. J Infect Dis. 1984 Jul;150(1):121–126. doi: 10.1093/infdis/150.1.121. [DOI] [PubMed] [Google Scholar]
  28. Spector S. A., Spector D. H. The use of DNA probes in studies of human cytomegalovirus. Clin Chem. 1985 Sep;31(9):1514–1520. [PubMed] [Google Scholar]
  29. Virtanen M., Syvänen A. C., Oram J., Söderlund H., Ranki M. Cytomegalovirus in urine: detection of viral DNA by sandwich hybridization. J Clin Microbiol. 1984 Dec;20(6):1083–1088. doi: 10.1128/jcm.20.6.1083-1088.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]

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