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Systemic and in vitro infection process of Bombyx mori nucleopolyhedrovirus
Md Masmudur Rahman1, Karumathil P Gopinathan
1Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
Abstract:
To analyse the systemic progression of infection by Bombyx mori nucleopolyhedrovirus (BmNPV) through oral ingestion by the silkworm larvae, a recombinant virus (vBmp10GFP) expressing the green fluorescent protein (GFP) under the control of the very late, viral p10 promoter (which still forms the polyhedral occlusion bodies) was constructed. Infection of B. mori derived BmN cells with the recombinant virus resulted in the expression of GFP from 12 h post infection (hpi), with maximal accumulation of the expressed protein by 60 hpi. B. mori larvae that ingested the polyhedra containing vBmp10GFP showed localized expression of GFP in the midgut epithelial cells within 24 hpi, indicating virus replication. The primary spread of the virus infection occurred through the tracheae. Viral multiplication was subsequently detected in nearly all the larval tissues including the neurons and regions of silk-glands that were in contact with the tracheae. Infection in fat bodies was widespread by 48 hpi, by which time the haemocytes also showed infection. In vitro infection of isolated organs/tissues from B. mori with the budded virions (BV) of vBmp10GFP also showed viral multiplication in the cells that were associated with the tracheae, confirming the role of tracheae in spreading the infection.
Insights
This study tracked Bombyx mori nucleopolyhedrovirus (BmNPV) infection in silkworms using a GFP-tagged virus. The virus primarily spreads through the tracheal system, infecting various tissues including neurons and silk glands.
Area of Science:
- Virology
- Insect Pathology
- Molecular Biology
Background:
- Bombyx mori nucleopolyhedrovirus (BmNPV) causes significant economic losses in sericulture.
- Understanding BmNPV systemic infection is crucial for developing control strategies.
Purpose of the Study:
- To analyze the systemic progression of BmNPV infection in silkworm larvae after oral ingestion.
- To visualize viral spread using a recombinant virus expressing Green Fluorescent Protein (GFP).
Main Methods:
- Construction of a recombinant BmNPV (vBmp10GFP) expressing GFP under the p10 promoter.
- Infection of B. mori larvae and BmN cells with vBmp10GFP.
- Monitoring GFP expression and localization using fluorescence microscopy.
Main Results:
- GFP expression initiated at 12 hpi in BmN cells, peaking at 60 hpi.
- Localized GFP expression observed in midgut epithelial cells within 24 hpi in larvae.
- Primary viral spread occurred via the tracheal system, leading to infection in various tissues including neurons and silk glands.
- Widespread infection in fat bodies and haemocytes by 48 hpi.
- In vitro studies confirmed the role of tracheae in viral dissemination.
Conclusions:
- The tracheal system is the primary route for systemic BmNPV spread in silkworm larvae.
- vBmp10GFP is an effective tool for studying BmNPV pathogenesis and dissemination.
