IMPROVED ANIMAL ROTAVIRUS ISOLATION IN MA104 CELLS USING DIFFERENT TRYPSIN CONCENTRATIONS

Abstract

ABSTRACT Rotavirus is one of the most important etiological agents in infant gastroenteritis and diarrhea in neonatal animals. The purpose of this study was the optimization of isolation of rotaviruses in MA104 (fetal kidney Rhesus monkey) cells by the comparison of culture, taking into account the number of samples, different treatments with trypsin, number of serial passages, and positivity by polyacrylamide gel electrophoresis (PAGE). Twenty diarrheic fecal samples of calves and piglets from São Paulo, State, Brazil, all positive for rotavirus in ELISA and PAGE, were submitted to four isolation protocols: in treatment 1, the inoculum was left on the monolayer and the maintenance medium was trypsin-free; treatment 2 was the same as treatment 1 except that the inoculum was discarded; in treatment 3 the inoculum was left on the monolayer and the maintenance medium had 5 µg/mL of trypsin; treatment 4 was the same as the monolayer treatment 3, except that the maintenance medium had 10 µg/mL of trypsin. Fifteen samples were successfully isolated, representing an efficacy of 75% (15/20). Treatment 3 was the most suitable because the lower number of serial passages and lower cytotoxity to the cells, with statistical significance (p < 0.001) using Freedman's and Wilcoxon's tests.