Enterovirus isolation from children with acute respiratory infections and presumptive identification by a modified microplate method

Abstract Objective: To evaluate a modified microplate method, utilizing HEF, HEp-2, Vero, MDCK and newly introduced RD-18S and GMK cell lines, for virus isolation. Methods: From June to October 2001, 723 throat swab specimens taken from children with acute respiratory infections (ARIs) were inoculated onto these cells. To analyze cell sensitivity, we also inoculated 20 serotypes of stocked enteroviruses. Results: During the period, we isolated 40 Coxsackie A2 (CoxA2), 13 CoxA4, 16 CoxA16, 1 CoxB2, 11 CoxB3, 2 CoxB5, 54 echo16, 2 entero7l and 1 polio2. By observing a cell sensitivity pattern with HEF, HEp-2, Vero, RD-18S, and GMK, we could finally differentiate five enterovirus groups: CoxA except for CoxA16, CoxA16/entero7l, CoxB, echovirus, and poliovirus. Conclusions: With this system, the RD-18S cell line enabled us to isolate CoxA virus, except for CoxA16, for the first time. Differentiation of five enterovirus groups by cell sensitivity simplified the specific identification by neutralization test as a presumptive identification. A modified microplate method may be an appropriate cell combination for virus isolation, especially for enteroviruses, and is expected to be used routinely for virologic diagnosis and to clarify the epidemiology of ARI in children.