Genetic recombination during coinfection of two mutants of human respiratory syncytial virus

Spann, Kristen, Collins, Peter L. and Teng, Michael N. (2003) Genetic recombination during coinfection of two mutants of human respiratory syncytial virus. Journal of Virology, 77 20: 11201-11211. doi:10.1128/JVI.77.20.11201-11211.2003

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Author Spann, Kristen
Collins, Peter L.
Teng, Michael N.
Title Genetic recombination during coinfection of two mutants of human respiratory syncytial virus
Journal name Journal of Virology   Check publisher's open access policy
ISSN 0022-538X
1098-5514
Publication date 2003-10-01
Year available 2003
Sub-type Article (original research)
DOI 10.1128/JVI.77.20.11201-11211.2003
Open Access Status File (Publisher version)
Volume 77
Issue 20
Start page 11201
End page 11211
Total pages 11
Place of publication Washington, DC, United States
Publisher American Society for Microbiology
Language eng
Abstract Recombination between coinfecting viruses had not been documented previously for a nonsegmented negative-strand RNA virus (mononegavirus). We investigated the potential of intermolecular recombination by respiratory syncytial virus (RSV) by coinfecting HEp-2 cells with two recombinant RSV (rRSV) mutants lacking either the G gene (DeltaG/HEK) or the NS1 and NS2 genes (DeltaNS1/2). These viruses replicate inefficiently and form pinpoint plaques in HEp-2 cells. Therefore, potential recombined viruses with a growth and/or plaque formation advantage should easily be identified and differentiated from the two parental viruses. Further identification of potential recombinants was aided by the inclusion of point mutation markers in the F and L genes of DeltaG/HEK and the design of reverse transcription-PCR (RT-PCR) primers capable of detecting these markers. Independent coinfections and control single infections by these two rRSV mutants were performed. In one of six coinfections, an RSV variant was identified that produced plaques slightly larger than those of wild-type RSV in HEp-2 cells. RT-PCR and sequencing provided evidence that this variant was a recombined RSV (rec-RSV). The rec-RSV appeared to have been generated by a polymerase jump from the DeltaG/HEK genome to that of DeltaNS1/2 and back again in the vicinity of the SH-G-F genes. This apparently involved nonhomologous and homologous recombination events, respectively. The recombined genome was identical to that of the DeltaG/HEK mutant except that all but the first 12 nucleotides of the SH gene were deleted and replaced by an insert consisting of the last 91 nucleotides of the G gene and its downstream intergenic region. This insert could have come only from the coinfecting DeltaNS1/2 virus. This resulted in the formation of a short chimeric SH:G gene. Northern and Western blot analysis confirmed that the rec-RSV did not express the normal SH and G mRNAs and proteins but did express the aberrant SH:G mRNA. This provides an experimental demonstration of intermolecular recombination yielding a viable, helper-independent mononegavirus. However, the isolation of only a single rec-RSV under these optimized conditions supports the idea that RSV recombination is rare indeed.
Keyword Virology
Replication In-vitro
Rna Recombination
Dengue Virus
Glycoprotein
Reassortment
Proteins
Rsv
Transcription
Evolution
Deletion
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status Unknown

Document type: Journal Article
Sub-type: Article (original research)
Collections: Excellence in Research Australia (ERA) - Collection
Clinical Medical Virology Centre Publications
 
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Citation counts: TR Web of Science Citation Count  Cited 60 times in Thomson Reuters Web of Science Article | Citations
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Created: Fri, 25 Jan 2008, 16:01:39 EST