DETECTION OF GANCICLOVIR RESISTANCE MUTATIONS AND QUANTITATION OF CYTOMEGALOVIRUS (CMV) DNA IN LEUKOCYTES OF PATIENTS WITH FATAL DISSEMINATED CMV DISEASE

Cytomegalovirus (CMV) UL97 mutations associated with ganciclovir resis tance at codons 460, 594, and 595 were detected by polymerase chain re action (PCR) followed by restriction enzyme analysis in CMV blood isol ates and directly in polymorphonuclear leukocyte (PMNL) DNA extracts o f 4 subjects who died of progressive disseminated CMV disease due to g anciclovir-resistant CMV strains. The CMV DNA load was also serially d etermined in leukocyte fractions of these patients using a quantitativ e-competitive PCR assay, There was excellent concordance between speci fic UL97 mutations in blood culture isolates and those detected in PMN L fractions for all patients, Emergence of such UL97 mutations during ganciclovir therapy was associated with an increasing CMV DNA burden i n leukocytes of the 2 patients with AIDS but not in the 2 subjects wit h chronic lymphocytic leukemia, Rapid molecular strategies, including detection of common CMV UL97 mutations and CMV DNA quantitation, can b e used directly in leukocytes of immunocompromised subjects with CMV d isease to monitor antiviral therapy.