Isolation of Rotaviruses from Calves with Acute Enteritis and Their Cultivation in Vitro

Abstract

Dvorak R., J. Stepanek, I. P§ikal, J. Franz: Isolation of Rotaviruses from Calves with Acute Enteritis and Their Cultivation in vitro. Acta vet. Brno, 62, 1993: 71-77. Five bovine rotavh-us strains were isolated in monolayers of idA-I04 cell culture from faeces of 3-to ll-day-old calves suffering from gastroenteritis. Cytophatic effect, accompanied by the release of cells from the glass surface of cultivation flasks was ()bs~ed,from the 5th to the 6th passage. After the stabilization of the cytopathic effect, the highest virus concentration was observed in cell cultures that had been, frozen 18-36 hrs after inoculation. Electron microscopy revealed particles with typical rotavirus morphology. Antigenic relationship of the isolates with the reference strain Lincolil was' confirmed by ELIS:A. The isolates were classified as members of the serological group A by ELISA and immunofluorescence assay. The identity of the rotavirus isolates was also confirmed by electrophoresis in agarose gel. The assay confirmed that their genome consists of segmented viral RNA, which produced in the electric field migration patterns, electrophorogrammes, typical of rotaviruses. In 4 isolates the migration speed of the segments in the electric field was identical with that ()f the reference strain. A different electropherotype was identified in the remaining isolate. Bovine rotavirus, diarrhoea, cell culture, isolation Rotaviruses are at present ~nsidered as important Causal agents of diarrheic infection~ i~ many domestic and wild ruminants, swine, monkey, horse, cat, dog, mouse, rabbit, poultry, fish, and also man (Wyatt et al. 1974; Flewett and Woode 1978; Fulton et al. 1981; Hos,hino et al. 1981; Theil et al. 1985; Thouless et al. 1986; Korych 1987; Marshall 1990). 'They are found especially in newborn farm animals (neonatal diarrhoea) and children. Rotaviruses ,have been detected in various climatic zones, mostly in connection with an outbreak of gastro'intestinal infections with various intensity of clinical manifestation, ranging from asymptomatic infection to severe, even lethal cases (Dea et al. 1985; Korych 1987; Ruuska and Vesikari 1990). Causal relation of rotaviruses to gastroenteritis in newborn calves was first described by Me b u Ii -et al. (1969), who induced gastroenteritis in calves by peroral administration of bacteria-frc;e filtrates of faeces from scouring animals. Reovirus-like agentS were detected in the gut contents ,of infected calves (Fernelius et al. 1972). After a series of unsuccessful attempts, the virus 'was isolated and adapted to the growth in cultures of bovine embryonic kidney cells (Fernelius et al. 1972). The isolate was identified as bovine rotavirus, different from reovirus (Flewett :and Woode 1978). The new information concerning the aetiology of scours was confirmed by 'Several authors in the U.S.A. (Ekern et al. 1981), United Kingdom (Woode et al. 1974; McNulty et al. 1977) and other countries including Czechoslovakia (Scherrer et al. 1976; Marsolais -et al. 1978; Wyn-Jones et al. 1978; Kaves 1979; Smid et al. 1980; Castrucci, et al. 1983; ,Barboiet al. 1987). Most studies, however, focused on the, methods of rotavirus demonstration .in faeces of calves suffering from gastroenteritis (Rodger and Holmes 1979; Smid et al. 1980; Ekern et al. 1981). Early attempts to isolate rotaviruses in cell or organ cultures derived from various animal species were unsuccessful. Studies with established cell lines of various origins were carried out with the aim of selecting suitable ones for the growth of rotavirus strains