Early Detection of Human Cytomegalovirus DNA by PCR-ELISA

Kim, Min; Yoon, Myung; Lim, Woo Hyun; Shin, Jong Hee; Suh, Soon Pal; Ryang, Dong Wook

Abstract

BACKGROUND: Human cytomegalovirus (CMV) infections are common and occasionally severe in newborns, immunocompromised hosts, cancer patients, and recipients of organ transplant. Consequently, sensitive and rapid methods for CMV detection are of great diagnostic value since antiviral drugs have become available, which might be more effective upon early administration. We evaluated a polymerase chain reaction and enzyme-linked immunosorbent assay (PCR- ELISA) to detect human CMV infection as an aid in making a prompt diagnosis and a determination of therapeutic efficacy.
METHODS
CMV DNA was amplified by single PCR, using primers chosen from genomic regions (major immediate-early [MIE] protein coding region), and the microwell plate hybridization assay was performed for specific detection of 5'-biotinylated PCR products using CMV-specific probes labeled with digoxigenin. A total of 35 clinical specimens from 14 patients who were suspected CMV infectious state was analyzed by PCR-ELISA and its results were compared with those of serum anti-CMV IgM, shell vial culture assay and PCR.
RESULTS
The sensitivity for detection of PCR-amplified CMV DNA by the ELISA was 102 copies, which was ten-fold greater than ethidium bromide staining of agarose gels. The positive rates of 35 clinical specimens by serology, shell vial culture assay, PCR and PCR-ELISA were 37.9%, 40.0%, 60.0% and 68.6%, respectively. The OD ranges of 24 positive specimens by PCR-ELISA were from 0.042 to above 2.5. In follow-up studies of two patients with bone marrow transplantation, positive CMV results by PCR-ELISA earlier than by other methods including serologic method, shell vial culture assay and PCR.
CONCLUSIONS
These results reveal that PCR-ELISA may show higher sensitivity and positive rate than serologic method, shell vial culture assay and conventional PCR. PCR-ELISA can be useful to manage CMV infection rapidly in patients at risk.