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Published on: August 23, 2018
Stability of a Spodoptera frugiperda nucleopolyhedrovirus deletion recombinant during serial passage in insects
Oihane Simón1, Trevor Williams, Robert D Possee
1Instituto de Agrobiotecnología, CSIC-Gobierno de Navarra, 31192 Mutilva Baja, Navarra, Spain.
Abstract:
The stabilities of the Spodoptera frugiperda multiple nucleopolyhedrovirus (SfMNPV) complete genome bacmid (Sfbac) and a deletion recombinant (Sf29null) in which the Sf29 gene was replaced by a kanamycin resistance cassette were determined during sequential rounds of per os infection in insect larvae. The Sf29 gene is a viral factor that determines the number of virions in occlusion bodies (OBs). The Sf29null bacmid virus was able to recover the Sf29 gene during passage. After the third passage (P3) of Sf29null bacmid OBs, the population was observed to reach an equilibrium involving a mixture of those with a kanamycin resistance cassette and those with the Sf29 gene. The biological activity of Sf29null bacmid OBs at P3 was similar to that of Sfbac OBs. The recovered gene in the Sf29null virus was 98 to 100% homologous to the Sf29 genes of different SfMNPV genotypes. Reverse transcription-PCR analysis of uninoculated S. frugiperda larvae confirmed the expression of the SfMNPV ie-0 and Sf29 genes, indicating that the insect colony harbors a covert SfMNPV infection. Additionally, the nonessential bacterial artificial chromosome vector was spontaneously deleted from both viral genomes upon passage in insects.
Insights
The Spodoptera frugiperda multiple nucleopolyhedrovirus (SfMNPV) Sf29 gene can be recovered by a deletion recombinant virus during insect infection. This SfMNPV virus eventually reaches an equilibrium, maintaining biological activity and deleting the bacterial artificial chromosome vector.
Area of Science:
- Virology
- Insect Pathology
- Molecular Biology
Background:
- Spodoptera frugiperda multiple nucleopolyhedrovirus (SfMNPV) is an important insect pathogen.
- The Sf29 gene influences the number of virions within occlusion bodies (OBs).
- Understanding viral genome stability during infection is crucial for baculovirus applications.
Purpose of the Study:
- To determine the stability of SfMNPV bacmid genomes during sequential per os infections in insect larvae.
- To investigate the ability of a deletion recombinant virus (Sf29null) to recover the deleted Sf29 gene.
- To analyze the impact of gene recovery and vector deletion on viral biological activity.
Main Methods:
- Sequential per os infection of insect larvae with SfMNPV bacmids (Sfbac and Sf29null).
- Passage experiments to monitor viral genome changes and gene recovery.
- Reverse transcription-PCR (RT-PCR) to confirm gene expression in larvae.
- Homology analysis of recovered Sf29 genes.
Main Results:
- The Sf29null bacmid virus spontaneously recovered the Sf29 gene during passage in insect larvae.
- Viral populations reached an equilibrium by the third passage (P3), containing both Sf29-repaired and kanamycin-resistant viruses.
- Biological activity of Sf29null OBs at P3 was comparable to the wild-type Sfbac.
- The bacterial artificial chromosome vector was spontaneously deleted from both viral genomes.
- A covert SfMNPV infection was detected in the uninoculated insect colony.
Conclusions:
- The SfMNPV Sf29 gene is not essential for viral replication and can be recovered during serial passage.
- The SfMNPV genome exhibits plasticity, allowing for gene recovery and vector deletion.
- The findings have implications for the development and stability of SfMNPV-based expression systems and biocontrol agents.
