Bergoin, Max

Pham, Hanh T.

Tijssen, Peter

Yu, Qian

English

OA OpenAgrar repository

    89  extracted and amplified through rolling circle amplification to enrich for small circular ssDNA templates. The concatenated circular genomes were then digested with restriction enzymes and the resulting products (~1– 4 kb) were cloned and sequenced. Thirteen distinct ssDNA viral genomes were recovered from five crab species and three shrimp species. Putative encoded proteins share less than 60% identity with known viral proteins from members of the Circoviridae. The detected genomes exhibit four different genomic architectures revealing an incredible diversity of ssDNA viruses in shrimp and crabs. Ongoing work aims to propagate these viruses in insect cell lines and develop a system to assess viral infectivity and modes of transmission.    Contributed paper. Wednesday, 15:00 188   Remarkable diversity of endogenous viruses in the genome of an isopod crustacean Julien Thézé, Sébastien Leclercq, Bouziane Moumen, Richard Cordaux, Clément Gilbert Université de Poitiers, Laboratoire Ecologie et Biologie des Interactions - UMR CNRS 7267, Equipe Ecologie Evolution Symbiose, 86073 Poitiers Cedex 9, France Address for Correspondence: theze.julien@gmail.com  Recent studies in paleovirology have uncovered myriads of endogenous viral elements (EVEs) integrated in the genome of their eukaryotic hosts. These fragments result from endogenization, i.e., integration of the viral genome into the host germline genome followed by vertical inheritance. So far, most studies have used a virus-centred approach, whereby endogenous copies of a particular group of viruses were searched in all available sequenced genomes. Here we follow a host-centred approach whereby the genome of a given species (the crustacean isopod Armadillidium vulgare) is comprehensively screened for the presence of EVEs using all viral sequences available as queries. This search and downstream evolutionary analyses revealed that 56 EVEs corresponding to 11 different viral lineages belonging to 5 viral families (Bunyaviridae, Circoviridae, Parvoviridae, Nimaviridae, Totiviridae) and one viral order (Mononegavirales) became endogenized in A. vulgare. We show that viral endogenization occurred recurrently during the evolution of isopods, that A. vulgare viral lineages were involved in multiple host-switches that took place between widely divergent taxa. Furthermore, 32 A. vulgare EVEs have uninterrupted open reading frames, suggesting they result from recent endogenization of viruses likely to be currently infecting isopod populations. Overall, our work shows that isopods have been and are still infected by a large variety of viruses. It also extends the host range of several families of viruses and brings new insights into their evolution. More generally, our results underline the power of paleovirology in characterizing the viral diversity currently infecting eukaryotic taxa.    Contributed paper. Wednesday, 15:15 189   Iteraviruses (Densovirinae) from monarch and black swallowtail butterflies and slug caterpillar moths and characterization of their expression strategies Qian Yu, Max Bergoin, and Peter Tijssen INRS-Institut Armand-Frappier, 531 boul. des Prairies, Laval, QC, Canada H7V1B7 Address for correspondence: peter.tijssen@iaf.inrs.ca  Iteraviruses belong to a separate genus of the Densovirinae subfamily of the Parvoviridae family and includes three densoviruses, i.e. Casphalia extranea Densovirus (CeDNV), Dendrolimus punctatus Densovirus (DpDNV) and Bombyx mori  Densovirus (BmDNV). In this study, we used a Sequence-Independent Single-Primer Amplification (SISPA) method to detect the pathogens of larvae from three additional insect species (Papilio polyxenes, Sibine fusca and Danaus plexippus), killed by some unknown pathogen. Sequencing of the clones that were obtained and BLAST analysis revealed the existence of three previously unknown densoviruses (provisionally named PpDNV, SfDNV and DppIDV). The genome of the new densoviruses were cloned into pCR2.1-topo or pBluescript(SK-) vectors. These virus sequences (including ITRs) have high identities with CeDNV and BmDNV. The identical genome organizations indicated that these three new densoviruses should be classified in the Iteravirus genus. Together with the infectious clones of CeDNV and BmDNV, we investigated the expression strategies of five different iteraviruses (PpDNV, SfDNV, CeDNV, BmDNV, DppIDV). Total RNA was obtained both from LD cell line transfected by infectious clones of the iteraviruses and virus infected larvae (Papilio polyxenes). RACE methods were used to identify the 5’ and 3’ transcription ends. The nonstructural (NS) and structural (VP) genes were located on the same strand of the genome. The NS cassette consists of two genes with NS1 and overlapping NS2. The NS2 transcripts all start at 7 nts downstream of the NS1 start codon. Transcription starts for NS1 genes are close to the AUG of NSS1. NS and VP transcripts do not overlap. The four VPs were similarly generated by leaky scanning translation of unspliced mRNA. The VP transcripts just start 2nts downstream of the poly (A) motif for NS transcripts. Interestingly, poly (A) signals for VP transcripts all overlap with the stop codons of the VP genes.     Contributed paper. Wednesday, 15:30 190   Remarkable genetic diversity of single-stranded DNA viruses in cultured shrimps and crickets Hanh T. Pham, Qian Yu, Max Bergoin, Peter Tijssen INRS-Institut Armand-Frappier, Université du Québec, Laval, QC, Canada Address for Correspondence: peter.tijssen@iaf.inrs.ca  Single-stranded DNA viruses are among the smallest viruses and include members of the Parvoviridae and Circoviridae families (linear and circular ssDNA viruses). In the past decades, PstDNV and AdDNV have been well-known viruses that have caused a severe impact on cultured shrimps and crickets. Here, we report the discovery and genome characterization of numerous novel denso- and denso-like viruses and, for the first time, new circoviruses from these hosts. During the last years, we received many cricket samples from North America that were negative for AdDNV. However, denso-like particles have been observed by EM. Complete sequence of different viral genome have been isolated and cloned including one circular ssDNA viruses of 2.5 kb, an ambisense densovirus of 4.9 kb and a segmented brevidenso-like virus (3.3 kb). Meantime, large numbers of new ssDNA viruses were also isolated from cultured shrimp from Vietnam. Characterization of these viruses revealed 3 different, unrelated circoviruses of 1.7, 1.7 (the latter is not using the standard genetic code and may have been ingested) and 1.3 kb. We also discovered a new shrimp parvovirus of about 4.1 kb that is phylogenetically poorly related to any known parvovirus. Near-atomic structures of some cricket and shrimp parvoviruses were obtained by X-ray crystallography as well as their transcription strategy. These results demonstrate a great diversity of ssDNA-viruses infecting these economically important animals. Future work will be focused on molecular features of theses viruses for a further insight into the evolution and classification of ssDNA viruses.     

Remarkable genetic diversity of single-stranded DNA viruses in cultured shrimps and crickets

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Remarkable genetic diversity of single-stranded DNA viruses in cultured shrimps and crickets

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