Detection of cytomegalovirus in urine by nonisotopic DNA hybridization.

Abstract

Cytomegalovirus (CMV) is an opportunistic pathogen that is responsible for significant morbidity and mortality in newborns and immunocompromised patients. Approximately l^o of all newborns in the United States are congenitally infected, and identification of these infants would define a population at risk for developmental and hearing abnormalities [1]. Hepatitis, pneumonitis, and chorioretinitis due to CMV are also devastating illnesses in recipients of bone marrow, renal, and cardiac transplants and in patients with AIDS [2-5]. A rapid, inexpensive test for active CMV infection would identify patients in need of prompt therapeutic intervention. The reference procedure for identification of CMV is tissue culture. Because of its expense, however, tissue culture is not a practical method for routine screening of newborns. Also, because up to six weeks may be necessary to identify CMV in tissue culture, it may not be rapid enough to allow prompt therapeutic intervention in the immunocompromised patient. Isotopic DNA hybridization assays for rapid detection of CMV in urine and buffy coat specimens have adequate sensitivity and specificity to identify infected newborns and immunocompromised patients [6, 7]. The use of radioisotopes, however, has several disadvantages, such as the need to use the short-lived radioisotope 32P and the time and expense required for autoradiography. These disadvantages have made clinical application impractical for the majority of laboratories. Biotin-labeled DNA probes, linked through avidin to an enzyme such as horseradish peroxidase or alkaline phosphatase, can be used to detect DNA bound to nitrocellulose. Biotin-labeled DNA and RNA probes have the advantage of a long shelf life and are reusable in hybridization assays. We have developed a nonisotopic DNA hybridization assay capable of detecting as little as 10 pg of isolated CMV DNA. This report shows an application of this assay for the rapid detection of CMV in clinical urine spec