CMV PCR monitoring in leucocytes of transplant patients

Abstract

Background: The presumed latency of cytomegalovirus (CMV) in leucocytes and the sensitivity of the polymerase chain reaction (PCR) raise a question of its clinical value. Objectives: To develop and standardize a CMV PCR as a diagnostic tool for CMV infection in solid organ and bone marrow transplant patients by comparing it to a likewise standardized isolation, rapid isolation and to clinical symptoms. Study design: The material comprised 822 EDTA peripheral blood samples from 96 solid organ and 119 bone marrow transplant patients. One sample from each of 21 healthy bone marrow donors and 25 blood donors were used as controls. Two million leucocytes were lysed and one-tenth of a volume was used in a nested PCR employing immediate early gene primers. Results: The limit of detection was ≈ 10 gene copies of a CMV DNA clone and 1 TCID 50 of extracted DNA from a cell suspension. The specificity was ≥0.99 when tested in CMV seronegative individuals. The positive and negative predictive values were 0.62 and 1.00, respectively. When PCR was compared to virus isolation/rapid culture in individual patients, PCR was positive more frequently in solid organ transplant patients than was CMV isolation/rapid culture, but the difference was not significant in bone marrow transplant patients. In isolation-positive patients, PCR became positive in samples taken 1–2 weeks earlier. In 54 solid organ transplant patients with PCR-positive samples, CMV-associated symptoms were present in 29 31 patients with CMV isolated from blood but in only 5 23 patients without viraemia. In 17 bone marrow transplant patients treated with ganciclovir, PCR became negative during or immediately after treatment in 14 20 (70%) episodes. This was true of 5 12 (42%) solid organ transplant patients. Conclusion: Screening of transplant patients with CMV PCR can be standardized at a clinically relevant level so that antiviral therapy can be instituted early.