• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of jcmPermissionsJournals.ASM.orgJournalJCM ArticleJournal InfoAuthorsReviewers
J Clin Microbiol. Jan 1995; 33(1): 41–44.
PMCID: PMC227876

Comparison of three methods for extraction of viral nucleic acids from blood cultures.

Abstract

Reliable nucleic acid extraction techniques for blood specimens are required for the sensitive detection of viral DNA. Standardized procedures for processing blood specimens for the molecular detection of herpesviruses (cytomegalovirus [CMV], herpes simplex virus, varicella-zoster virus, and Epstein-Barr virus [EBV]) have not been established. Three methods were used to extract DNA from blood specimens from healthy donors and asymptomatic immunocompromised patients: (i) IsoQuick treatment of whole blood, (ii) extraction of the peripheral blood leukocytes by lysis (lysis buffer and proteinase K), and (iii) extraction of peripheral blood leukocytes with phenol-chloroform (sodium docecyl sulfate solution and proteinase K). All blood specimens from 25 healthy blood donors were negative for CMV, herpes simplex virus and varicella-zoster virus nucleic acid sequences, regardless of the extraction method, while three samples (12%) extracted by the lysis technique were positive for EBV DNA. Of 25 blood samples from asymptomatic immunocompromised patients, CMV and EBV each were detected in nine specimens by lysis extraction, four each by IsoQuick and four (CMV) and six (EBV) by the phenol-chloroform method. Our results indicate that the lysis method is optimal for the detection of CMV and EBV DNA sequences by PCR from the leukocytic fraction of blood specimens. DNA of these viruses is frequently present in blood specimens from asymptomatic immunocompromised patients and occasionally from healthy donors.

Full Text

The Full Text of this article is available as a PDF (165K).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Bitsch A, Kirchner H, Dupke R, Bein G. Failure to detect human cytomegalovirus DNA in peripheral blood leukocytes of healthy blood donors by the polymerase chain reaction. Transfusion. 1992 Sep;32(7):612–617. [PubMed]
  • Brice SL, Stockert SS, Jester JD, Huff JC, Bunker JD, Weston WL. Detection of herpes simplex virus DNA in the peripheral blood during acute recurrent herpes labialis. J Am Acad Dermatol. 1992 Apr;26(4):594–598. [PubMed]
  • Brytting M, Xu W, Wahren B, Sundqvist VA. Cytomegalovirus DNA detection in sera from patients with active cytomegalovirus infections. J Clin Microbiol. 1992 Aug;30(8):1937–1941. [PMC free article] [PubMed]
  • Chirgwin JM, Przybyla AE, MacDonald RJ, Rutter WJ. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. [PubMed]
  • Delgado R, Lumbreras C, Alba C, Pedraza MA, Otero JR, Gómez R, Moreno E, Noriega AR, Payá CV. Low predictive value of polymerase chain reaction for diagnosis of cytomegalovirus disease in liver transplant recipients. J Clin Microbiol. 1992 Jul;30(7):1876–1878. [PMC free article] [PubMed]
  • Drouet E, Boibieux A, Michelson S, Ecochard R, Biron F, Peyramond D, Colimon R, Denoyel G. Polymerase chain reaction detection of cytomegalovirus DNA in peripheral blood leukocytes as a predictor of cytomegalovirus disease in HIV-infected patients. AIDS. 1993 May;7(5):665–668. [PubMed]
  • Espy MJ, Smith TF, Persing DH. Dependence of polymerase chain reaction product inactivation protocols on amplicon length and sequence composition. J Clin Microbiol. 1993 Sep;31(9):2361–2365. [PMC free article] [PubMed]
  • Gozlan J, Salord JM, Chouaïd C, Duvivier C, Picard O, Meyohas MC, Petit JC. Human cytomegalovirus (HCMV) late-mRNA detection in peripheral blood of AIDS patients: diagnostic value for HCMV disease compared with those of viral culture and HCMV DNA detection. J Clin Microbiol. 1993 Jul;31(7):1943–1945. [PMC free article] [PubMed]
  • Gerna G, Zipeto D, Parea M, Percivalle E, Zavattoni M, Gaballo A, Milanesi G. Early virus isolation, early structural antigen detection and DNA amplification by the polymerase chain reaction in polymorphonuclear leukocytes from AIDS patients with human cytomegalovirus viraemia. Mol Cell Probes. 1991 Oct;5(5):365–374. [PubMed]
  • Gleaves CA, Smith TF, Shuster EA, Pearson GR. Comparison of standard tube and shell vial cell culture techniques for the detection of cytomegalovirus in clinical specimens. J Clin Microbiol. 1985 Feb;21(2):217–221. [PMC free article] [PubMed]
  • Jordan ML, Hrebinko RL, Jr, Dummer JS, Hickey DP, Shapiro R, Vivas CA, Simmons RL, Starzl TE, Hakala TR. Therapeutic use of ganciclovir for invasive cytomegalovirus infection in cadaveric renal allograft recipients. J Urol. 1992 Nov;148(5):1388–1392. [PMC free article] [PubMed]
  • Kwok S, Higuchi R. Avoiding false positives with PCR. Nature. 1989 May 18;339(6221):237–238. [PubMed]
  • Meyers JD, Ljungman P, Fisher LD. Cytomegalovirus excretion as a predictor of cytomegalovirus disease after marrow transplantation: importance of cytomegalovirus viremia. J Infect Dis. 1990 Aug;162(2):373–380. [PubMed]
  • Myers MG. Viremia caused by varicella-zoster virus: association with malignant progressive varicella. J Infect Dis. 1979 Aug;140(2):229–233. [PubMed]
  • Saiki RK, Gelfand DH, Stoffel S, Scharf SJ, Higuchi R, Horn GT, Mullis KB, Erlich HA. Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science. 1988 Jan 29;239(4839):487–491. [PubMed]
  • Shibata D, Martin WJ, Appleman MD, Causey DM, Leedom JM, Arnheim N. Detection of cytomegalovirus DNA in peripheral blood of patients infected with human immunodeficiency virus. J Infect Dis. 1988 Dec;158(6):1185–1192. [PubMed]
  • Telenti A, Marshall WF, Smith TF. Detection of Epstein-Barr virus by polymerase chain reaction. J Clin Microbiol. 1990 Oct;28(10):2187–2190. [PMC free article] [PubMed]
  • Telenti A, Smith TF, Ludwig J, Keating MR, Krom RA, Wiesner RH. Epstein-Barr virus and persistent graft dysfunction after liver transplantation. Hepatology. 1991 Aug;14(2):282–286. [PubMed]
  • Vilmer E, Mazeron MC, Rabian C, Azogui O, Devergie A, Perol Y, Gluckman E. Clinical significance of cytomegalovirus viremia in bone marrow transplantation. Transplantation. 1985 Jul;40(1):30–35. [PubMed]
  • Wagner HJ, Bein G, Bitsch A, Kirchner H. Detection and quantification of latently infected B lymphocytes in Epstein-Barr virus-seropositive, healthy individuals by polymerase chain reaction. J Clin Microbiol. 1992 Nov;30(11):2826–2829. [PMC free article] [PubMed]

Articles from Journal of Clinical Microbiology are provided here courtesy of American Society for Microbiology (ASM)

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...