Pattern of Cytopathic Effects Produced by Recent Bangladeshi Field Isolates of PPR Virus in Vero Cells and Preparation of Purified Viral Antigen.

BACKGROUND

This study was conducted to explore the pattern of appearance of cytopathic effect (CPE) produced by a Bangladeshi strain of Peste des petits ruminant's virus (PPRV) in Vero cells during serial passaging, to estimate the virus titer and concentration of PPR viral RNA at different passage levels, and to find out a suitable passage level for viral antigen purification.

METHODS

Five isolates of PPR virus from local outbreaks were isolated, confirmed with conventional RT-PCR, and then used as viral inoculant in Vero cells and lamb kidney cell culture (LKC). Morphology of CPE's and pattern of development of CPE were studied and recorded at different passage levels and days postinfection (dpi). Tissue culture fluid (TCF) was used to prepare purified viral antigen by ultracentrifugation and for quantitation by spectrophotometry. Viral titers were determined following an endpoint dilution assay.

RESULTS

The study revealed that the CPE was not perceptible in all isolates until 11 dpi for the first six passages. The virus was not detected in TCF of both infected and control flasks by RT-PCR up to the 6th passage. The first CPE appeared on 5 dpi at the 7th passage level for all isolates. The CPE was characterized by initial cell rounding, followed by the formation of small syncytia, progression to total cell infection, and eventual complete detachment of the cell monolayer. Maximum titer (3.5 Log10TCID50) and RNA concentration (260 ng/µL) were found at the 30th passage level by one isolate (Isolate-F).

CONCLUSION

It is concluded that initiation of CPE production in Vero cells by the Bangladeshi field strain of PPR virus occurs at the 7th passage level. Virus titer and concentration of PPR viral RNA increased with the advancement of passaging, and these are highly related to each other. TCF at higher passage levels, such as the 60th passage, is most suitable for purification of PPR viral antigen.