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In vitro replication of Macrobrachium rosenbergii nodavirus and extra small virus in C6/36 mosquito cell line
R Sudhakaran1, V Parameswaran, A S Sahul Hameed
1Aquaculture Biotechnology Division, Department of Zoology, C. Abdul Hakeem College, Melvisharam 632509, Vellore Dist, Tamil Nadu, India.
Abstract:
White tail disease (WTD) is a serious problem in hatcheries and nursery ponds of Macrobrachium rosenbergii in India and many parts of the world. The pathogenic agents have been identified as M. rosenbergii nodavirus (MrNV) associated with extra small virus (XSV), which is 27nm and 15nm in diameter, respectively. Replication of MrNV and XSV was investigated in apparently healthy C6/36 subclone of Aedes albopictus cell line. The results revealed that C6/36 cells were susceptible to these viruses. The replication of these viruses in C6/36 cells was confirmed by RT-PCR, acridine orange staining, infectivity study and electron microscopy. Cytoplasmic ribonucleic acid (RNA) stained by acridine orange increased by 48h, and by 72h larger proportion of cells which indicated alterations in quantity and localization of RNA in the infected cells. Post-larvae, challenged by immersion method using inoculum prepared from infected cells, exhibited lethargy, anorexia and opaqueness of abdominal muscle and 100% mortality was observed at 6 days post-infection. Experimentally infected C6/36 cells and post-larvae showed positive by RT-PCR, whereas control cells and healthy post-larvae showed negative. This is the first study to report the multiplication of MrNV and XSV in C6/36 cell line.
Insights
White tail disease (WTD) in shrimp is caused by Macrobrachium rosenbergii nodavirus (MrNV) and extra small virus (XSV). These viruses can replicate in Aedes albopictus cells, offering a new avenue for studying WTD.
Area of Science:
- Aquaculture
- Virology
- Cell Biology
Background:
- White tail disease (WTD) significantly impacts Macrobrachium rosenbergii aquaculture globally.
- The causative agents are identified as Macrobrachium rosenbergii nodavirus (MrNV) and extra small virus (XSV).
Purpose of the Study:
- To investigate the replication of MrNV and XSV in the C6/36 Aedes albopictus cell line.
- To confirm viral replication using molecular and microscopic techniques.
Main Methods:
- Infection of C6/36 cells with MrNV and XSV.
- Confirmation of replication via RT-PCR, acridine orange staining, infectivity assays, and electron microscopy.
- Challenge studies on M. rosenbergii post-larvae.
Main Results:
- C6/36 cells supported the replication of MrNV and XSV.
- Viral replication was evidenced by increased cytoplasmic RNA, altered RNA localization, and positive RT-PCR results.
- Infected post-larvae exhibited WTD symptoms and 100% mortality within 6 days.
Conclusions:
- The C6/36 Aedes albopictus cell line is susceptible to MrNV and XSV.
- This study demonstrates the successful multiplication of MrNV and XSV in an insect cell line for the first time.
- This provides a valuable model for studying WTD pathogenesis and developing control strategies.
