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Open AccessArticle

Tracing Baculovirus AcMNPV Infection Using a Real-Time Method Based on ANCHORTM DNA Labeling Technology

LGEI, IMT Mines Alès, Institut Mines-Télécom et Université de Montpellier Sud de France, 6 Avenue de Clavières, 30100 Alès, France
INRA, SPE, 400 route des Chappes BP 167, 06903 Sophia-Antipolis CEDEX, France
CNRS UPS3044 Baculovirus et Thérapie, LabEx-53, 30380 Saint Christol lèz Alès, France
Department of Biological & Medical Sciences, Oxford Brookes University, Gipsy Lane, Oxford OX3 0BP, UK
Oxford Expression Technologies Ltd. BioInnovation Hub, Oxford OX3 0BP, UK
NeoVirTech SAS, 1 Place Pierre Potier, 31000 Toulouse, France
Institute for Advanced Life Science Technology; ITAV USR3505, 1 Place Pierre Potier, 31000 Toulouse, France
Authors to whom correspondence should be addressed.
Present address: Bioline agroscience France, 1306 Route de Biot, 06560 Valbonne, France.
Viruses 2020, 12(1), 50;
Received: 4 December 2019 / Revised: 23 December 2019 / Accepted: 25 December 2019 / Published: 2 January 2020
(This article belongs to the Special Issue Insect Viruses and Pest Management)
Many steps in the baculovirus life cycle, from initial ingestion to the subsequent infection of all larval cells, remain largely unknown; primarily because it has hitherto not been possible to follow individual genomes and their lineages. Use of ANCHORTM technology allows a high intensity fluorescent labelling of DNA. When applied to a virus genome, it is possible to follow individual particles, and the overall course of infection. This technology has been adapted to enable labelling of the baculovirus Autographa californica Multiple NucleoPolyhedroVirus genome, as a first step to its application to other baculoviruses. AcMNPV was modified by inserting the two components of ANCHORTM: a specific DNA-binding protein fused to a fluorescent reporter, and the corresponding DNA recognition sequence. The resulting modified virus was stable, infectious, and replicated correctly in Spodoptera frugiperda 9 (Sf9) cells and in vivo. Both budded viruses and occlusion bodies were clearly distinguishable, and infecting cells or larvae allowed the infection process to be monitored in living cells or tissues. The level of fluorescence in the culture medium of infected cells in vitro showed a good correlation with the number of infectious budded viruses. A cassette that can be used in other baculoviruses has been designed. Altogether our results introduce for the first time the generation of autofluorescent baculovirus and their application to follow infection dynamics directly in living cells or tissues. View Full-Text
Keywords: real-time imaging; fluorescence labelling; baculovirus infection real-time imaging; fluorescence labelling; baculovirus infection
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MDPI and ACS Style

Hinsberger, A.; Graillot, B.; Blachère Lopez, C.; Juliant, S.; Cerutti, M.; King, L.A.; Possee, R.D.; Gallardo, F.; Lopez Ferber, M. Tracing Baculovirus AcMNPV Infection Using a Real-Time Method Based on ANCHORTM DNA Labeling Technology. Viruses 2020, 12, 50.

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