Use of Sequence-Independent, Single-Primer-Amplification (SISPA) for rapid detection, identification, and characterization of avian RNA viruses

Abstract Current technologies with next generation sequencing have revolutionized metagenomics analysis of clinical samples. To achieve the non-selective amplification and recovery of low abundance genetic sequences, a simplified Sequence-Independent, Single-Primer Amplification (SISPA) technique in...

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Bibliographic Details
Published in:Virology (New York, N.Y.) N.Y.), 2017-09, Vol.509, p.159-166
Main Authors: Chrzastek, Klaudia, Lee, Dong-hun, Smith, Diane, Sharma, Poonam, Suarez, David L, Pantin-Jackwood, Mary, Kapczynski, Darrell R
Format: Article
Language:English
Subjects:
Animals
Avian influenza virus
DNA Primers - genetics
Infectious bronchitis virus
Infectious bronchitis virus - genetics
Infectious bronchitis virus - isolation & purification
Infectious Disease
Influenza A virus - genetics
Influenza A virus - isolation & purification
Metagenomics - methods
Newcastle disease virus
Newcastle disease virus - genetics
Newcastle disease virus - isolation & purification
Next generation sequencing
Nucleic Acid Amplification Techniques - methods
Poultry
Poultry Diseases - virology
Random amplification
RNA Virus Infections - veterinary
RNA Virus Infections - virology
RNA viruses
Sequence Analysis, DNA - methods
Sequence-independent single-primer amplification
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Summary:Abstract Current technologies with next generation sequencing have revolutionized metagenomics analysis of clinical samples. To achieve the non-selective amplification and recovery of low abundance genetic sequences, a simplified Sequence-Independent, Single-Primer Amplification (SISPA) technique in combination with MiSeq platform was applied to target negative- and positive-sense single-stranded RNA viral sequences. This method allowed successful sequence assembly of full or near full length avian influenza virus (AIV), infectious bronchitis virus (IBV), and Newcastle disease virus (NDV) viral genome. Moreover, SISPA analysis applied to unknown clinical cases of mixed viral infections produced genome assemblies comprising 98% NDV and 99% of IBV genomes. Complete or near complete virus genome sequence was obtained with titers at or above 104.5 EID50 /ml (50% embryo infectious dose), and virus identification could be detected with titers at or above 103 EID50 /ml. Taken together, these studies demonstrate a simple template enrichment protocol for rapid detection and accurate characterization of avian RNA viruses.
ISSN:0042-6822
1096-0341
DOI:10.1016/j.virol.2017.06.019