Pathogenesis of Junonia coenia densovirus in Spodoptera frugiperda: A route of infection that leads to hypoxia

AbstractTo evaluate densovirus potential against lepidopteran pests and their capacity to invade new hosts, we have characterised in vivo the infection and pathogenesis of the Junonia coenia densovirus (JcDNV) in the noctuid pest Spodoptera frugiperda. Here we show that infection starts with the ingestion of viral particles that cross the midgut epithelium without replicating. By quantitative PCR we established the kinetic and the route of infection, from virus ingestion to replication in visceral tracheae and hemocytes. JcDNV has a high particle-to-infection ratio mostly due to the barrier function of the midgut. Pathology and cytopathology suggested that infection of tracheal cells impairs oxygen delivery to demanding tissues leading to cytopathic effects in all the tissues. Finally, larval death results from several physiological shocks, including molting arrest and anoxia

Establishment and analysis of a reference transcriptome for Spodoptera frugiperda

International audienceBackground Spodoptera frugiperda (Noctuidae) is a major agricultural pest throughout the American continent. The highly polyphagous larvae are frequently devastating crops of importance such as corn, sorghum, cotton and grass. In addition, the Sf9 cell line, widely used in biochemistry for in vitro protein production, is derived from S. frugiperda tissues. Many research groups are using S. frugiperda as a model organism to investigate questions such as plant adaptation, pest behavior or resistance to pesticides.ResultsIn this study, we constructed a reference transcriptome assembly (Sf_TR2012b) of RNA sequences obtained from more than 35 S. frugiperda developmental time-points and tissue samples. We assessed the quality of this reference transcriptome by annotating a ubiquitous gene family - ribosomal proteins - as well as gene families that have a more constrained spatio-temporal expression and are involved in development, immunity and olfaction. We also provide a time-course of expression that we used to characterize the transcriptional regulation of the gene families studied.ConclusionWe conclude that the Sf_TR2012b transcriptome is a valid reference transcriptome. While its reliability decreases for the detection and annotation of genes under strong transcriptional constraint we still recover a fair percentage of tissue-specific transcripts. That allowed us to explore the spatial and temporal expression of genes and to observe that some olfactory receptors are expressed in antennae and palps but also in other non related tissues such as fat bodies. Similarly, we observed an interesting interplay of gene families involved in immunity between fat bodies and antennae

Contribution à l'étude du potentiel d'utilisation des Densovirus en lutte microbiologique

La lutte microbiologique connaît un regain d'intérêt suite aux problématiques soulevée spar les pesticides chimiques en agriculture. L'objectif de ces travaux de thèse est d'étudier le potentiel d'une ressource virale alternative, les Densovirus pathogènes de Lépidoptères,appartenant à la sous famille des Densovirinae, strictement inféodée aux Arthropodes. Nos travaux ont été axés sur trois points (i) décrire le spectre d'hôtes de deux Densovirus candidats et identifier des déterminants de spécificité pour comprendre l'évolution du spectre d'hôtes ; (ii) caractériser des mécanismes de transmission horizontale pour analyser la dynamique de l'infection et (iii) valider des outils de détection des Densovirinae afin d'étudier la diversité génétique virale dans la nature. Le premier axe nous a permis d'identifier (i) un Densovirus candidat qui ne semble être virulent que pour des Lépidoptères ravageurs de culture, et (ii) des déterminants de spécificité à la surface de la capside impliqués le franchissement de la barrière intestinale. Concernant la transmission, nos résultats montrent qu'un Densovirus se propage rapidement dans une population hôte en induisant un comportement cannibale probablement développé par les individus non infectés. Nous avons également caractérisé deux mécanismes de transmission horizontale,par morsures et par un vecteur endoparasitoïde. Enfin, pour étudier la prévalence virale,nous avons mis au point un protocole basé sur le principe d'une PCR nichée que nous avons testé sur des insectes collectés dans la nature. Ceci nous a permis de dresser un inventaire de la diversité d'insectes, sans toutefois révéler à ce jour la présence de Densovirinae. Ces résultats constituent les premières connaissances pour l'élaboration d'un modèle épidémiologique qui nous permettra d'évaluer l'impact sur l'environnement d'utiliser un Densovirus en tant qu'agent de lutte microbiologique.Microbial control has a renewed interest due to issues raised by chemical pesticides inagriculture. The aim of this work is to propose a new viral resource, Densoviruses pathogenicfor Lepidoptera belonging to Densovirinae sub family restricted to arthropods. We focus onthree axes, (i) we described the host range of two Densovirus candidates, and identified hostspecificity determinants to understand host range evolution; (ii) we characterized themechanisms driving the horizontal transmission of Densovirus to model the dynamic of theinfection; and (iii) we validated tools to detect Densovirinae to study the viral geneticdiversity in natura. The first point lead us to identify (i) a Densovirus potentially pathogeniconly for lepidopteran pests, and (ii) few determinants of specificity localized at the surface ofthe capsid that are essential for the midgut recognition, although this barrier of specificity isnot the only one. Concerning horizontal transmission, our results show that a Densovirus canspread rapidly within a host population, inducing a cannibalistic behavior probablydeveloped by non infected individuals. We also characterize two mechanisms involved intransmission, by biting and by an endoparasitoïd vector. To study the densoviral prevalence,we develop a nested PCR and test it on insect sampling. This allowed us to describe theinsect diversity. No Densovirinae have been detected so far. These results are the first steptoward building an epidemiology model that may allow to evaluate the impact of using aDensovirus as a biological control tool.MONTPELLIER-BU Sciences (341722106) / SudocSudocFranceF

Coding-Complete Genome Sequence of a Partitivirus Isolated from Pine Processionary Moth Eggs

International audienceTwo coding-complete nucleotide sequences of a partitivirus (family Partitiviridae) were discovered in transcriptomic data sets obtained from eggs of the Lepidoptera Thaumetopoea pityocampa. Each segment encodes a single open reading frame, and these two segments are predicted to encode an RNA-dependent RNA polymerase and a coat protein, respectively

Coding-Complete Genome Sequences of an Iteradensovirus and an Alphapermutotetra-Like Virus Identified from the Pine Processionary Moth ( Thaumetopoea pityocampa ) in Portugal

International audienceThe coding-complete genome sequences of an iteradensovirus (family Parvoviridae) and an alphapermutotetra-like virus (family Permutotetraviridae) were discovered from transcriptomic data sets obtained from Thaumetopoea pityocampa larvae collected in Portugal. Each of the coding-complete genome sequences of these viruses contains three main open reading frames (ORFs)

Genome-wide identification of lncRNAs associated with viral infection in Spodoptera frugiperda

International audienceThe role of lncRNAs in immune defence has been demonstrated in many multicellular and unicellular organisms. However, investigation of the identification and characterization of long non-coding RNAs (lncRNAs) involved in the insect immune response is still limited. In this study, we used RNA sequencing (RNA-seq) to investigate the expression profiles of lncRNAs and mRNAs in the fall armyworm Spodoptera frugiperda in response to virus infection. To assess the tissue- and virus-specificity of lncRNAs, we analysed and compared their expression profiles in haemocytes and fat body of larvae infected with two entomopathogenic viruses with different lifestyles, i.e. the polydnavirus HdIV ( Hyposoter didymator IchnoVirus) and the densovirus JcDV ( Junonia coenia densovirus). We identified 1883 candidate lncRNAs, of which 529 showed differential expression following viral infection. Expression profiles differed considerably between samples, indicating that many differentially expressed (DE) lncRNAs showed virus- and tissue-specific expression patterns. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment and target prediction analyses indicated that DE-LncRNAs were mainly enriched in metabolic process, DNA replication and repair, immune response, metabolism of insect hormone and cell adhesion. In addition, we identified three DE-lncRNAs potentially acting as microRNA host genes, suggesting that they participate in gene regulation by producing miRNAs in response to virus infection. This study provides a catalogue of lncRNAs expressed in two important immune tissues and potential insight into their roles in the antiviral defence in S. frugiperda . The results may help future in-depth functional studies to better understand the biological function of lncRNAs in interaction between viruses and the fall armyworm

Four Amino Acids of an Insect Densovirus Capsid Determine Midgut Tropism and Virulence

International audienceDensoviruses are insect parvoviruses that are orally infectious for Lepidoptera. To assess the mechanisms underlying their specificity and their virulence, we investigated the role of eight candidate residues in the densovirus capsid. We showed that the substitutions of four amino acids were associated with decreased virulence due to a decreased ability to cross the host midgut epithelium, without an effect on viral replication in other tissues

Can Virus-like Particles Be Used as Synergistic Agent in Pest Management?

Among novel strategies proposed in pest management, synergistic agents are used to improve insecticide efficacy through an elevation of intracellular calcium concentration that activates the calcium-dependent intracellular pathway. This leads to a changed target site conformation and to increased sensitivity to insecticides while reducing their concentrations. Because virus-like particles (VLPs) increase the intracellular calcium concentration, they can be used as a synergistic agent to synergize the effect of insecticides. VLPs are self-assembled viral protein complexes, and by contrast to entomopathogen viruses, they are devoid of genetic material, which makes them non-infectious and safer than viruses. Although VLPs are well-known to be used in human health, we propose in this study the development of a promising strategy based on the use of VLPs as synergistic agents in pest management. This will lead to increased insecticides efficacy while reducing their concentrations