Demonstration of the protective effects of fluorescent proteins in baculoviruses exposed to ultraviolet light inactivation

Autographa californica multiple nucleopolyhedrovirus (AcMNPV) recombinants, namely AcRFP produced by fusion of the red fluorescent protein (RFP) gene with the polyhedrin gene, and a recombinant (pAcUW21-23GFP) carrying the green fluorescent protein (GFP) in its viral envelope, were evaluated for their resistance to inactivation by ultraviolet light. AcRFP recombinants produced incomplete polyhedra with low infectivity for Trichoplusia ni larvae, whereas AcuW21-23GFP produced normal polyhedra with high infectivity. Electron microscopy of AcRFP CL14 showed the incorporation of very few viral particles into polyhedrin matrix protein material. The LC(50) for AcuW21-23GFP was 0.10 occlusion bodies/mm(2), whereas the LC(50) values for several AcRFP recombinants ranged from 20 to 329 occlusion bodies/mm(2). When both the RFP and GFP recombinants were exposed to ultraviolet light (UV-B 280–320 nm), the results support the conclusion that these fluorescent proteins afford some protection against its damaging effects. Abbreviation: / AcMNPV: Autographa californica multiple nucleopolyhedrovirus BV: budded virus CPE: cytopathogenic effect ECV: extracellular virus OB: occlusion body ODV: occlusion derived virus RFP: red fluorescent protein GFP: green fluorescent protein TCID(50): tissue culture infective dose at the 50 % level UV-B: ultraviolet light of 280–320 n

Electron photomicrograph of infected TN-CL1 cells with AcRFP14 showing the presence of numerous viral particles and incompletely formed polyhedra

<p><b>Copyright information:</b></p><p>Taken from "Demonstration of the protective effects of fluorescent proteins in baculoviruses exposed to ultraviolet light inactivation"</p><p>Journal of Insect Science 2004;4():-.</p><p>Published online 5 Oct 2004</p><p>PMCID:PMC1081559.</p><p>Copyright © 2004. Open access; copyright is maintained by the authors.</p

(A) larvae infected with AcRFP14 at 4 d following inoculation and (B) un-inoculated control larva

<p><b>Copyright information:</b></p><p>Taken from "Demonstration of the protective effects of fluorescent proteins in baculoviruses exposed to ultraviolet light inactivation"</p><p>Journal of Insect Science 2004;4():-.</p><p>Published online 5 Oct 2004</p><p>PMCID:PMC1081559.</p><p>Copyright © 2004. Open access; copyright is maintained by the authors.</p> Magnification 7X

Schematic diagram illustrating the construction of the AcMNPV recombinant (AcRFP) from the fusion of the RFP gene with the polyhedrin gene and co-transfection of amplicon with wt AcMNPV-HPP

<p><b>Copyright information:</b></p><p>Taken from "Demonstration of the protective effects of fluorescent proteins in baculoviruses exposed to ultraviolet light inactivation"</p><p>Journal of Insect Science 2004;4():-.</p><p>Published online 5 Oct 2004</p><p>PMCID:PMC1081559.</p><p>Copyright © 2004. Open access; copyright is maintained by the authors.</p

An electron photomicrograph at higher magnification of showing few viral particles incorporated into the polyhedrin matrix protein and several virus particles without a membrane on the periphery of the protein matrix material

<p><b>Copyright information:</b></p><p>Taken from "Demonstration of the protective effects of fluorescent proteins in baculoviruses exposed to ultraviolet light inactivation"</p><p>Journal of Insect Science 2004;4():-.</p><p>Published online 5 Oct 2004</p><p>PMCID:PMC1081559.</p><p>Copyright © 2004. Open access; copyright is maintained by the authors.</p

Electron micrograph of AcRFPCL14 in infected TN-CL1 cells showing the presence of irregular shaped polyhedrin matrix protein material (grey structures) with some occluded viral particles

<p><b>Copyright information:</b></p><p>Taken from "Demonstration of the protective effects of fluorescent proteins in baculoviruses exposed to ultraviolet light inactivation"</p><p>Journal of Insect Science 2004;4():-.</p><p>Published online 5 Oct 2004</p><p>PMCID:PMC1081559.</p><p>Copyright © 2004. Open access; copyright is maintained by the authors.</p> The circular grey structure in upper right is the virogenic stroma