Identification of rotaviruses by dot-blot hybridization using an alkaline phosphatase-conjugated synthetic oligonucleotide probe

We have evaluated a recently-developed dot-blot hybridization assay for the detection of human rotaviruses using an alkaline-phosphatase conjugated oligonucleotide probe. The lower detection limit of this assay was 1 ng (approximately 5 x 10(7) copies) of the double-stranded (ds) RNA, when a purified preparation from serotype 1 human rotavirus was used but appeared to be much higher when applied on clinical specimens. This assay could detect dsRNA from rotavirus strains belonging to serotypes 1 through 6, 8 and 9. A total of 235 stool specimens were used to evaluate the oligonucleotide probe assay in comparison with polyacrylamide gel electrophoresis and latex agglutination assay (Rotalex). When polyacrylamide gel electrophoresis was used as a gold standard, sensitivity, specificity and positive and negative predictive values of the probe assay were 84, 100, 100 and 89%, respectively. These values were slightly better than those of Rotalex assay which is commonly used in clinical laboratories in Japan. Although the probe assay requires more hands-on time than the immunoassays, the high specificity of this probe assay recommends it as a confirmatory test in the clinical laboratory setting.