Korean Journal of Nephrology 1998;17(2):323-334.

신이식 환자에서 배양과 DNA 검출에 의한 Cytomegalocirus 감염의 진단과 IE gone 변종 검출에 관한 연구

황승덕 , 박애자 , 이희발

Abstract

Cytomegalovirus (CMV) is a ubiquitous virus and its infections occur commonly after renal transplan- tation and immunosuppressive therapy. Early and accurate laboratory diagnosis of CMV infection in renal transplant is necessary but often difficult. To find optimal diagnostic methods for CMU infection, we compared shell vial culture and polymerase chain reaction (PCR) and Southern blot of PCR products. A total of 301 specimens of urine, blood neutro- phils, tissues, or body fluids were obtained from 75 renal transplant recipients and were submitted to shell vial culture for CMV as well as DNA PCR using primers for immediate early(IE) gene of CMV, The human fibroblast cell line (MRC-5) was used to culture CMV and were examined with immuno- fluorescence staining using monoclonal antibody to the early antigen of CMV. The PCR products (274 and 379 bp) were detected by gel electrophoresis and ethidium bromide staining. When PCR products were not clearly visible on electrophoresis, PCR products were analyzed by Southern blot using IE gene probe. Sixty four(85.3%) of 75 renal transplant recipients showed CMV infection as analyzed by PCR and Southern blot as well as shell vial culture. On shell vial culture, CMV were detected in 81 specimens from 30(40%) renal transplant recipients in viremic state. On PCR and Southern blot analysis CMV were detected in 55 and 26 specimens, respectively from 59 patients. The sensitivity of culture and PCR to detect CMV infection were 42.4% and 83.3%, respectively. The results of two studies were concordant in 48%. PCR and Southern blot did not detect CMV in 10 and 5 culture proven CMV posi- tive samples, respectively. Mutant CMV were found in 3 patients which showed 5-10 bp deletion in IE gene. Moreover, DNA sequencing analysis showed 5 mutant strains among 11 strains which appeared same by PCR prodcut. These results suggest that PCR followed by Southern blot may be more sensitive, but less spe- cific than shell vial culture in the diagnosis of CMV disease. PCR followed by Southern blot may not detect mutant CMV. Combined analysis using both shell vial culture and PCR followed by Southern blot may be necessary to diagnose CMV infection in renal transplant recipients.