Cytotechnology

, Volume 65, Issue 6, pp 1003–1016 | Cite as

Characterization of growth and Oryctesrhinoceros nudivirus production in attached cultures of the DSIR-HA-1179 coleopteran insect cell line

  • Charlotte Pushparajan
  • Juan Daniel Claus
  • Sean David Goldie Marshall
  • Gabriel Visnovsky
JAACT Special Issue

Abstract

The DSIR-HA-1179 coleopteran cell line is a susceptible and permissive host to the Oryctesrhinoceros nudivirus (OrNV), which has been used as a biocontrol agent against the coconut rhinoceros beetle (Oryctes rhinoceros); a pest of palms in the Asia-Pacific region. However, little is known about growth and metabolism of this cell line, knowledge of which is necessary to develop an in vitro large-scale OrNV production process. The strong anchorage-dependent characteristics of the cell line, its particular fragility and its tendency to form dense clumps when manipulated, are the most likely reasons that have precluded further development of the cell line. In order to characterize DSIR-HA-1179 cells, there was first a need for a reliable technique to count the cells. A homogenous cell suspension suitable for enumeration could be produced by treatment with TrypLE Express™ with optimum mean time for cell release calculated as 30 min. The cell line was adapted to grow in four serum-supplemented culture media namely TC-100, IPL-41, Sf-900 II and Sf-900 III and cell growth, glucose consumption, lactate and ammonia production were assessed from static-batch cultures. The maximum viable cell density was reached in Sf-900 II (17.9 × 105 cells/ml), with the maximum specific growth rate observed in this culture medium as well (0.0074 h−1). Higher production of OrNV was observed in IPL-41 and TC-100 (4.1 × 107 TCID50/ml) than in cultures infected in Sf-900 III (2.0 × 107 TCID50/ml) and Sf-900 II (1.4 × 107 TCID50/ml). At the end of the growth period, glucose was completely consumed in cultures grown in TC-100, while remained in excess in the other three culture media. The cell line produced lactate and ammonia to very low levels in the TC-100 culture medium which is a promising aspect for its cultivation at large-scale.

Keywords

DSIR-HA-1179 Oryctesrhinoceros nudivirus Heteronychus arator Attachment dependent insect cell line Cell counting Metabolism 

Abbreviations

OrNV

Oryctesrhinoceros nudivirus

MOI

Multiplicity of infection

TOI

Time of infection

TCID

Tissue culture infectious dose

PDT

Population doubling time

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Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Charlotte Pushparajan
    • 1
  • Juan Daniel Claus
    • 2
  • Sean David Goldie Marshall
    • 3
  • Gabriel Visnovsky
    • 1
  1. 1.Department of Chemical and Process EngineeringUniversity of CanterburyChristchurchNew Zealand
  2. 2.Laboratorio de Virologıa, Facultad de Bioquımica y Ciencias BiologicasUniversidad Nacional del LitoralSanta FeRepublica Argentina
  3. 3.Innovative Farm Systems, AgResearch LtdLincoln Research CentreChristchurchNew Zealand

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