Evaluation of two current generation enzyme immunoassays and an improved isolation-based assay for the rapid detection and isolation of rotavirus from stool

Background: Rapid and accurate rotavirus testing is important in decisions involving patient care and management. Quality assurance testing needs to b e periodically performed, especially among widely used assays having a dire ct impact on patient care. Objectives: To evaluate the current generation K allestad Pathfinder Direct antigen Detection system (PTH), and the widely u sed Rotaclone((R)) Rotavirus EIA Diagnostic Kit (RTC), in comparison with a n improved cell culture amplification-antigen detection (CCA-Ag) isolation- based assay. Study design: Two hundred stool specimens (specimen stored at greater than or equal to - 75 degreesC), which had been previously tested b y PTH, were tested by RTC and CCA-Ag. Discordant specimens were retested by PTH, blocking assay, polyacrylamide gel electrophoresis (PAGE), and/or ele ctron microscopy (EM). Results: Among 200 stool specimens, 197 were in acco rd by PTH, RTC and CCA-Ag. The sensitivity, specificity, positive and negat ive predictive values for RTC, PTH and CCA-Ag were, 100, 99, 99, 100, 100, 99, 99, 100; and 98, 100. 100, 98%, respectively. Among five initially disc ordant specimens, two required a period of 10 days to affect isolation. A n on-cultivatable (CCA-Ag negative) but true positive specimen, was identifie d as rotavirus group A serotype G2 by RT-PCR. Four true positive but discor dant specimens were blocking assay negative using one or both EIA kits. Con clusions: PTH and RTC are excellent rotavirus detection system. However, PT H is more expensive (ca. $3.50 vs. $2.00 per test), mandates a slightly lon ger turn-around time (ca. 1 vs. 1.5 h), and necessitates slightly more hand s on manipulative/preparative steps. Blocking assay was not a reliable conf irmatory test for the resolution of specimen discordancy. A combination of CCA-Ag, PAGE, EM, and/or perhaps RT-PCR, is recommended as an appropriate t est panel for the resolution of discordant results during assay evaluation. The newly modified and simplified 48-h rotavirus isolation-based assay may serve as a base line methodology in laboratory evalaution studies, as a la boratory support methodology during drug/vaccine efficacy trials, or for th e testing of sources (e.g., biopsy/autopsy tissues) not approved for assay by commercial rotavirus kits. (C) 2000 Elsevier Science B.V. All rights res erved.