Giuseppe
Emeritus
Virus discovery by deep sequencing and assembly of virus-derived small silencing RNAs (Proc Natl Acad Sci USA, abstract, edited)
Virus discovery by deep sequencing and assembly of virus-derived small silencing RNAs
1. Qingfa Wu a, 2. Yingjun Luo a, 3. Rui Lu a, 4. Nelson Lau b, 5. Eric C. Lai c, 6. Wan-Xiang Li a and 7. Shou-Wei Ding a,1
Author Affiliations
1. aDepartment of Plant Pathology and Microbiology, Institute for Integrative Genome Biology, University of California, Riverside, CA 92521
2. bDepartment of Biology, Brandeis University, Waltham, MA 02454
3. cDepartment of Developmental Biology, Sloan?Kettering Institute, New York, NY 10065
1. Edited by Peter Palese, Mount Sinai School of Medicine, New York, NY, and approved December 3, 2009 (received for review October 1, 2009)
Abstract
In response to infection, invertebrates process replicating viral RNA genomes into siRNAs of discrete sizes to guide virus clearance by RNA interference. Here, we show that viral siRNAs sequenced from fruit fly, mosquito, and nematode cells were all overlapping in sequence, suggesting a possibility of using siRNAs for viral genome assembly and virus discovery. To test this idea, we examined contigs assembled from published small RNA libraries and discovered five previously undescribed viruses from cultured Drosophila cells and adult mosquitoes, including three with a positive-strand RNA genome and two with a dsRNA genome. Notably, four of the identified viruses exhibited only low sequence similarities to known viruses, such that none could be assigned into an existing virus genus. We also report detection of virus-derived PIWI-interacting RNAs (piRNAs) in Drosophila melanogaster that have not been previously described in any other host species and demonstrate viral genome assembly from viral piRNAs in the absence of viral siRNAs. Thus, this study provides a powerful culture-independent approach for virus discovery in invertebrates by assembling viral genomes directly from host immune response products without prior virus enrichment or amplification. We propose that invertebrate viruses discovered by this approach may include previously undescribed human and vertebrate viral pathogens that are transmitted by arthropod vectors.
* arboviruses
* piRNAs
* siRNAs
* viral immunity
* Massively parallel sequencing
Footnotes
* 1To whom correspondence should be addressed at: Department of Plant Pathology and Microbiology, Institute for Integrative Genome Biology, University of California, Riverside, 900 University Avenue, Riverside, CA 92521. E-mail: shou-wei.ding@ucr.edu.
* Author contributions: Q.W. and S.-W.D. designed research; Q.W., Y.L., R.L., and W.-X.L. performed research; N.L. and E.C.L. contributed new reagents/analytic tools; Q.W., Y.L., and S.-W.D. analyzed data; and Q.W. and S.-W.D. wrote the paper.
* This article is a PNAS Direct Submission.
* Data deposition: The sequences reported in this paper have been deposited in the GenBank database [accession nos. GQ342961 (DTV), GQ342962 (DBV-A), GQ342963 (DBV-B), GQ342964 (DTrV), GQ342965 (ANV-RNA1), GQ342966 (ANV-RNA2), and GU144510 (MNV)].
* The authors are declare no conflict of interest.
* This article contains supporting information online at www.pnas.org/cgi/content/full/0911353107/DCSupplemental.
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<cite cite="http://www.pnas.org/content/early/2009/12/15/0911353107.abstract">Virus discovery by deep sequencing and assembly of virus-derived small silencing RNAs ? PNAS</cite>1. Qingfa Wu a, 2. Yingjun Luo a, 3. Rui Lu a, 4. Nelson Lau b, 5. Eric C. Lai c, 6. Wan-Xiang Li a and 7. Shou-Wei Ding a,1
Author Affiliations
1. aDepartment of Plant Pathology and Microbiology, Institute for Integrative Genome Biology, University of California, Riverside, CA 92521
2. bDepartment of Biology, Brandeis University, Waltham, MA 02454
3. cDepartment of Developmental Biology, Sloan?Kettering Institute, New York, NY 10065
1. Edited by Peter Palese, Mount Sinai School of Medicine, New York, NY, and approved December 3, 2009 (received for review October 1, 2009)
Abstract
In response to infection, invertebrates process replicating viral RNA genomes into siRNAs of discrete sizes to guide virus clearance by RNA interference. Here, we show that viral siRNAs sequenced from fruit fly, mosquito, and nematode cells were all overlapping in sequence, suggesting a possibility of using siRNAs for viral genome assembly and virus discovery. To test this idea, we examined contigs assembled from published small RNA libraries and discovered five previously undescribed viruses from cultured Drosophila cells and adult mosquitoes, including three with a positive-strand RNA genome and two with a dsRNA genome. Notably, four of the identified viruses exhibited only low sequence similarities to known viruses, such that none could be assigned into an existing virus genus. We also report detection of virus-derived PIWI-interacting RNAs (piRNAs) in Drosophila melanogaster that have not been previously described in any other host species and demonstrate viral genome assembly from viral piRNAs in the absence of viral siRNAs. Thus, this study provides a powerful culture-independent approach for virus discovery in invertebrates by assembling viral genomes directly from host immune response products without prior virus enrichment or amplification. We propose that invertebrate viruses discovered by this approach may include previously undescribed human and vertebrate viral pathogens that are transmitted by arthropod vectors.
* arboviruses
* piRNAs
* siRNAs
* viral immunity
* Massively parallel sequencing
Footnotes
* 1To whom correspondence should be addressed at: Department of Plant Pathology and Microbiology, Institute for Integrative Genome Biology, University of California, Riverside, 900 University Avenue, Riverside, CA 92521. E-mail: shou-wei.ding@ucr.edu.
* Author contributions: Q.W. and S.-W.D. designed research; Q.W., Y.L., R.L., and W.-X.L. performed research; N.L. and E.C.L. contributed new reagents/analytic tools; Q.W., Y.L., and S.-W.D. analyzed data; and Q.W. and S.-W.D. wrote the paper.
* This article is a PNAS Direct Submission.
* Data deposition: The sequences reported in this paper have been deposited in the GenBank database [accession nos. GQ342961 (DTV), GQ342962 (DBV-A), GQ342963 (DBV-B), GQ342964 (DTrV), GQ342965 (ANV-RNA1), GQ342966 (ANV-RNA2), and GU144510 (MNV)].
* The authors are declare no conflict of interest.
* This article contains supporting information online at www.pnas.org/cgi/content/full/0911353107/DCSupplemental.
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