Quantitation of human herpesvirus 6 DNA in infant with exanthem subitum by microplate PCR‐hybridization assay

Abstract

Quantitative analysis of human herpesvirus 6 (HHV-6) genome is important for monitoring active virus infection. The purpose of our study is to evaluate the reliability of a hybridization-based microtiter plate assay (polymerase chain reaction enzyme-linked immunosorbent assay (PCR ELISA)) for quantifying the virus genome. Semiquantitative analysis of the virus genome was carried out in 31 (18 male and 13 female) infants with primary HHV-6 infection. If the HHV-6 virus could be isolated from the peripheral blood mononuclear cells (PBMC), the infants were defined as being infected with HHV-6. The PCR ELISA method was used to determine the virus load. A titration of the virus was also carried out in the samples obtained during the acute phase of exanthem subitum. Specificity of the method was demonstrated by a lack of amplification of human herpesvirus 7 and cytomegalovirus DNA. The upper and lower detection limits of the method were 58 and 5800 copies of the virus genome, respectively. The quantity of HHV-6 DNA in the PBMC during the acute phase (879 +/- 975 copies/10(4) PBMC) was significantly higher than during the convalescent phase (54 +/- 76 copies/10(4) PBMC). Furthermore, the virus load in acute phase plasma (53 +/- 75 copies/microL) was also significantly higher than in the convalescent phase samples (2 +/- 9 copies/microL). Virus load in both PBMC and plasma gradually increased after the onset of exanthem subitum until about day 3 to 4 of the illness, but then decreased quickly. However, there was no significant association between virus load and the numbers of infected cells. Virus load in both PBMC and plasma gradually increased after the onset of exanthem subitum until about day 3 and day 4 of the illness, respectively, then it decreased quickly. These results indicate that our PCR ELISA system is reliable for monitoring active HHV-6 infection in vivo.