We report here the sequences for all three segments of a novel RNA virus (LepmorLBV1) from the insect trypanosomatid parasite Leptomonas moramango. This virus belongs to a newly discovered group of bunyavirus-like elements termed Leishbunyaviruses (LBV), the first discovered from protists related to arboviruses infecting humans

Akopyants, Natalia S

Beverley, Stephen M

Dobson, Deborah E

Lukes, Julius

Lye, Lon-Fye

PubMed

ABSTRACT
          
            We report here the sequences for all three segments of a novel RNA virus (LepmorLBV1) from the insect trypanosomatid parasite
            Leptomonas moramango
            . This virus belongs to a newly discovered group of bunyavirus-like elements termed Leishbunyaviruses (LBV), the first discovered from protists related to arboviruses infecting humans.
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Natalia S. Akopyants

Lon-Fye Lye

Deborah E. Dobson

Julius Lukeš

Stephen M. Beverley

Crossref

Genome Announcements

A Novel Bunyavirus-Like Virus of Trypanosomatid Protist Parasites

Digital Commons@Becker

Washington University School of MedicineDigital Commons@BeckerOpen Access Publications2016A novel bunyavirus-like virus of trypanosomatidprotist parasitesNatalia S. AkopyantsWashington University School of Medicine in St. LouisLon-Fye LyeWashington University School of Medicine in St. LouisDeborah E. DobsonWashington University School of Medicine in St. LouisJulius LukesUniversity of South BohemiaStephen M. BeverleyWashington University School of Medicine in St. LouisFollow this and additional works at: http://digitalcommons.wustl.edu/open_access_pubsThis Open Access Publication is brought to you for free and open access by Digital Commons@Becker. It has been accepted for inclusion in OpenAccess Publications by an authorized administrator of Digital Commons@Becker. For more information, please contact engeszer@wustl.edu.Recommended CitationAkopyants, Natalia S.; Lye, Lon-Fye; Dobson, Deborah E.; Lukes, Julius; and Beverley, Stephen M., ,"A novel bunyavirus-like virus oftrypanosomatid protist parasites." Genome Announcements.4,4. e00715-16. (2016).http://digitalcommons.wustl.edu/open_access_pubs/5283A Novel Bunyavirus-Like Virus of Trypanosomatid Protist ParasitesNatalia S. Akopyants,a Lon-Fye Lye,a Deborah E. Dobson,a Julius Lukeš,b,c Stephen M. BeverleyaDepartment of Molecular Microbiology, Washington University School of Medicine, St. Louis, Missouri, USAa; Institute of Parasitology, Biology Centre, and Faculty ofSciences, University of South Bohemia, Cˆeské Budeˇjovice, Czech Republicb; Canadian Institute for Advanced Research, Toronto, Ontario, CanadacWe report here the sequences for all three segments of a novel RNA virus (LepmorLBV1) from the insect trypanosomatid para-site Leptomonas moramango. This virus belongs to a newly discovered group of bunyavirus-like elements termed Leishbunyavi-ruses (LBV), the first discovered from protists related to arboviruses infecting humans.Received 30 May 2016 Accepted 17 June 2016 Published 4 August 2016Citation Akopyants NS, Lye L-F, Dobson DE, Lukeš J, Beverley SM. 2016. A novel bunyavirus-like virus of trypanosomatid protist parasites. Genome Announc 4(4):e00715-16.doi:10.1128/genomeA.00715-16.Copyright © 2016 Akopyants et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license.Address correspondence to Stephen M. Beverley, beverley@wusm.wustl.edu.Bunyaviridae comprise 350 species, often arthropod-borneand including serious human, animal, and plant pathogens(1). In searching for novel viruses from trypanosomatid protists(Kinetoplastida, supergroup Excavata), we found a new viral ele-ment within the insect parasite Leptomonas moramango (2).RNA was prepared from an uncloned strain of parasites (2)using TRIzol reagent (Thermo Fisher), treatment with DNase I(Thermo Fisher), and purified with RNA Clean & Concen-trator-25 (Zymo Research). Replicative viral double-strandedRNAs (dsRNAs) were visualized following treatment with S1 nu-clease (Thermo Fisher), separation by agarose gel electrophoresis,and staining by ethidium bromide (3). Three prominent bandswith sizes of about 6, 1.3, and 0.7 kb were observed (L, M, and S,respectively). Total RNA was depleted of rRNA with the Ribo-Zero kit (Illumina), fragmented to 200 to 600 nucleotides (nt),and used as a template for the generation of multiplexed TruSeqcDNA libraries. Thesewere sequenced (2 101 cycles, paired-endreads) on the HiSeq 2500 (Illumina). A total of 45,411,994 readswere obtained, of which 722,520 (1.6%) were assigned to viralsegments (CLC Genomics Workbench).For each segment, we identified two related similarly sized con-tigs (L, 5,982 and 6,029 nt; M, 1,205 and 1,154 nt; S, 662 and820 nt). Within each pair, one was more abundant, as judged byread depth coverage (L, 1,634 or 471 nt; M, 12,453 or 225 nt; S,53,243 or 622 nt); we arbitrarily grouped the more abundant seg-ments as “a” and refer to the viruses as LepmorLBV1a and 1b. TheGC compositions of LepmorLBV1s were similar to each otherand to that of the meta-transcriptomic assembly (38 to 41%). Wewere able to detect 9-nt “panhandle” repeats (4, 5) on 6/12 terminiwithin the metatranscriptomic contigs.Comparisons between the LepmorLBV1a/b segment openreading frames (ORFs) showed significant nucleotide and aminoacid identity (52 to 54% and 37 to 50%, respectively). L segmentsdisplayed a single ORF encoding 1,979 or 1,982 amino acids (aa),bearing motifs found typically in bunyavirus RNA-dependentRNA polymerases (RdRp) (6, 7). M segments encoded a singleORF of 322 or 321 aa; no database hits were found, but the pro-tein(s) were predicted to have a hydrophobic N-terminal signalsequence. S segments predicted proteins of 165 and 168 aa, whichshowed structural similarity to the nucleocapsid of other bunya-viruses (4, 5, 8).Phylogenetic analysis of the RdRp domain showed thatLepmorLBV1s grouped together with bunyaviruses, relative toother viral outgroups, but as a clearly distinct clade well separatedfrom Phlebovirus or other genera. Additional affinities to Bunya-viridae include a negative single-stranded RNA genome, terminalrepeated panhandle sequences important for replication and tran-scription, and three segments typically encoding the viral RdRp,envelope glycoproteins, and the nucleocapsid (4, 5, 7). TheL. moramango virus thus resembles a group of related viruses dis-covered recently in the closely related human parasite Leishmania,which we have termed Leishbunyavirus (LBV).L. moramango may bear cytoplasmic virus-like particles (2)that could correspond to LepmorLBV1. Future studies will estab-lish the evolutionary distribution of LBVs and their potential rolein host pathogenicity (9).Accession number(s). The sequences of all segments ofLepmorLBV1a and 1b were deposited in GenBank under the ac-cession numbers KX280012 to KX280017.ACKNOWLEDGMENTSWe thank members of our laboratories, Vyacheslav Yurchenko, and Ni-colas Fasel for discussions.This work was supported by NIH AID R56 AI099364 and AI29646 toS.M.B. Next-generation sequencing was performed at the WashingtonUniversity School of Medicine, Department of Genetics Genome Tech-nology Access Center (partially supported by grants NCI Cancer CenterSupport P30 CA91842 and NCRR ICTS/CTSA UL1 TR000448).FUNDING INFORMATIONThis work, including the efforts of Natalia S. Akopyants, Lon-FyeLye, Deborah E. Dobson, and Stephen M. Beverley, was funded byHHS | NIH | National Cancer Institute (NCI) (NCI P30 CA91842).This work, including the efforts of Natalia S. Akopyants, Lon-FyeLye, Deborah E. Dobson, and Stephen M. Beverley, was funded byHHS | National Institutes of Health (NIH) (R01-AI29646). Thiswork, including the efforts of Natalia S. Akopyants, Lon-Fye Lye,Deborah E. Dobson, and Stephen M. Beverley, was funded by HHS |crossmarkGenome AnnouncementsJuly/August 2016 Volume 4 Issue 4 e00715-16 genomea.asm.org 1 on September 21, 2016 by Washington University in St. Louishttp://genomea.asm.org/Downloaded from NIH | National Center for Research Resources (NCRR) (UL1TR000448).REFERENCES1. Plyusnin A, Beaty BJ, Elliott RM, Goldbach R, Kormelink R, LundkvistKA, Schmaljohn CS, Tesh RB. 2012. Family Bunyaviridae, p 725–741. InKing AMQ, Adams MJ, Carstens EB, Lefkowitz EJ (ed), Virus taxonomy:classification and nomenclature of viruses: ninth report of the InternationalCommittee on Taxonomy of Viruses. Elsevier Academic, San Diego, CA.2. Yurchenko V, Votýpka J, Tesarˇová M, Klepetková H, Kraeva N, Jirku˚ M,Lukeš J. 2014. Ultrastructure and molecular phylogeny of four new speciesof monoxenous trypanosomatids from flies (Diptera: Brachycera) with re-definition of the genus Wallaceina. Folia Parasitol (Praha) 61:97–112.3. Beiting DP, Peixoto L, Akopyants NS, Beverley SM, Wherry EJ, Chris-tian DA, Hunter CA, Brodsky IE, Roos DS. 2014. Differential induction ofTLR3-dependent innate immune signaling by closely related parasite spe-cies. PLoS One 9:e88398. http://dx.doi.org/10.1371/journal.pone.0088398.4. Guu TS, Zheng W, Tao YJ. 2012. Bunyavirus: structure and replication.Adv Exp Med Biol 726:245–266. http://dx.doi.org/10.1007/978-1-4614-0980-9_11.5. Walter CT, Barr JN. 2011. Recent advances in the molecular and cellularbiology of bunyaviruses. J Gen Virol 92:2467–2484. http://dx.doi.org/10.1099/vir.0.035105-0.6. Reguera J, Weber F, Cusack S. 2010. Bunyaviridae RNA polymerases(L-protein) have an N-terminal, influenza-like endonuclease domain, es-sential for viral cap-dependent transcription. PLoS Pathog 6:e1001101.http://dx.doi.org/10.1371/journal.ppat.1001101.7. Gerlach P, Malet H, Cusack S, Reguera J. 2015. Structural insights intobunyavirus replication and its regulation by the vRNA promoter. Cell 161:1267–1279. http://dx.doi.org/10.1016/j.cell.2015.05.006.8. Kelley LA, Mezulis S, Yates CM, Wass MN, Sternberg MJ. 2015. ThePhyre2Web portal for protein modeling, prediction and analysis. Nat Pro-toc 10:845–858. http://dx.doi.org/10.1038/nprot.2015.053.9. Hartley MA, Drexler S, Ronet C, Beverley SM, Fasel N. 2014. Theimmunological, environmental, and phylogenetic perpetrators of meta-static leishmaniasis. Trends Parasitol 30:412– 422. http://dx.doi.org/10.1016/j.pt.2014.05.006.Akopyants et al.Genome Announcements2 genomea.asm.org July/August 2016 Volume 4 Issue 4 e00715-16 on September 21, 2016 by Washington University in St. Louishttp://genomea.asm.org/Downloaded from 

A novel bunyavirus-like virus of trypanosomatid protist parasites

http://dx.doi.org/10.1128/genomeA.00715-16

A novel bunyavirus-like virus of trypanosomatid protist parasites

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