Production of Oryctes nudivirus in DSIR-HA-1179 insect cell cultures in roller bottle systems

Abstract

The coconut rhinoceros beetle, an economically important pest of coconut palms, is effectively managed by application of its natural pathogen, the Oryctes nudivirus (OrNV). The traditional method of producing OrNV in infected larvae is labor intensive, and suffers from inconsistencies in virus titer and purity. In vitro production of OrNV in the susceptible and permissive anchorage-dependent DSIR-HA-1179 insect cell line can overcome these disadvantages. While production of OrNV in cell monolayers grown in T-flasks or Cell Factories is possible, the approach would not be cost-effective at industrial level. Roller Bottle Systems (RBS), with larger growth surface areas and capable of holding larger culture volumes in a more dynamic environment, would be a better option for scaling up purposes. DSIR-HA-1179 cells grown in 490 cm2 RBS filled with 60 ml of TC-100 culture medium added with 10% fetal bovine serum yielded a maximum of 1.5 x 106 cells/ml, which was comparable to yields reached in T-flasks under the same culture conditions (1.7 x 106 cells/ml). Infection in RBS of cells in their early exponential growth phase (5 x 105 cells/ml) at multiplicity of infection of 1 TCID50/cell, produced OrNV volumetric and cell-specific yields of 2.1 x 108 TCID50/ml and 250 TCID50/cell respectively, which was an improvement over those produced under identical culture and infection conditions in T-flasks (volumetric and cell-specific yields of 6.8 x 107 TCID50/ml and 102 TCID50/cell, respectively). Thus, the roller bottle system has proven to be a robust alternative for the industrial production of OrNV.