HCMV strain comparison by restriction analysis of genomic virus DNA and of a PCR-amplified DNA fragment

Abstract

background: Human cytomegalovirus (HCMV) is ubiquitous and a major cause of disease in the immunocompromised which include the unborn fetus. HCMV strains are closely related and may be difficult to differentiate. Objectives: To analyze new HCMV isolates and highly passaged strains by conventional restriction fragment length polymorphism (RFLP) analysis of HCMV DNA and restriction analysis (RA) of a PCR-amplified a-sequence fragment to identify epidemiological relatedness. Study design: DNA from 14 low-passaged clinical isolates and the high-passaged strains S348 and AD169 were analyzed by RFLP analysis of BamHI, PstI or HpaII digestion as well as by PCR a-sequence amplification. The laboratory results were compared according to epidemiological findings. Results: Of the 14 clinical isolates 13 could be distinguished from each other by RFLP analysis. Isolates from an infant and its mother had identical restriction fragment patterns. Eight of 13 clinical isolates were differentiated from each other by comparing a-sequence PCR fragments according to length and the presence of cleavage sites for 6 different restriction enzymes. The mother-infant isolates were identical by the a-sequence PCR criteria. All 14 low-passaged isolates differed from the high-passaged S384 and AD169 strains by both distinguishing approaches. Conclusions: Both RFLP and PCR-amplified a-sequence fragment analysis clearly differentiated between low- and high-passaged HCMV isolates and are useful epidemiological tools for determining viral strain relatedness.