Cargando…
Identification and complete genome sequencing of paramyxoviruses in mallard ducks (Anas platyrhynchos) using random access amplification and next generation sequencing technologies
BACKGROUND: During a wildlife screening program for avian influenza A viruses (AIV) and avian paramyxoviruses (APMV) in Belgium, we isolated two hemagglutinating agents from pools of cloacal swabs of wild mallards (Anas platyrhynchos) caught in a single sampling site at two different times. AIV and...
Autores principales: | , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2011
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3219605/ https://www.ncbi.nlm.nih.gov/pubmed/21978491 http://dx.doi.org/10.1186/1743-422X-8-463 |
_version_ | 1782216858891452416 |
---|---|
author | Rosseel, Toon Lambrecht, Bénédicte Vandenbussche, Frank van den Berg, Thierry Van Borm, Steven |
author_facet | Rosseel, Toon Lambrecht, Bénédicte Vandenbussche, Frank van den Berg, Thierry Van Borm, Steven |
author_sort | Rosseel, Toon |
collection | PubMed |
description | BACKGROUND: During a wildlife screening program for avian influenza A viruses (AIV) and avian paramyxoviruses (APMV) in Belgium, we isolated two hemagglutinating agents from pools of cloacal swabs of wild mallards (Anas platyrhynchos) caught in a single sampling site at two different times. AIV and APMV1 were excluded using hemagglutination inhibition (HI) testing and specific real-time RT-PCR tests. METHODS: To refine the virological identification of APMV2-10 realized by HI subtyping tests and in lack of validated molecular tests for APMV2-10, random access amplification was used in combination with next generation sequencing for the sequence independent identification of the viruses and the determination of their genomes. RESULTS: Three different APMVs were identified. From one pooled sample, the complete genome sequence (15054 nucleotides) of an APMV4 was assembled from the random sequences. From the second pooled sample, the nearly complete genome sequence of an APMV6 (genome size of 16236 nucleotides) was determined, as well as a partial sequence for an APMV4. This APMV4 was closely related but not identical to the APMV4 isolated from the first sample. Although a cross-reactivity with other APMV subtypes did not allow formal identification, the HI subtyping revealed APMV4 and APMV6 in the respective pooled samples but failed to identify the co-infecting APMV4 in the APMV6 infected pool. CONCLUSIONS: These data further contribute to the knowledge about the genetic diversity within the serotypes APMV4 and 6, and confirm the limited sensitivity of the HI subtyping test. Moreover, this study demonstrates the value of a random access nucleic acid amplification method in combination with massive parallel sequencing. Using only a moderate and economical sequencing effort, the characterization and full genome sequencing of APMVs can be obtained, including the identification of viruses in mixed infections. |
format | Online Article Text |
id | pubmed-3219605 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2011 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-32196052011-11-18 Identification and complete genome sequencing of paramyxoviruses in mallard ducks (Anas platyrhynchos) using random access amplification and next generation sequencing technologies Rosseel, Toon Lambrecht, Bénédicte Vandenbussche, Frank van den Berg, Thierry Van Borm, Steven Virol J Research BACKGROUND: During a wildlife screening program for avian influenza A viruses (AIV) and avian paramyxoviruses (APMV) in Belgium, we isolated two hemagglutinating agents from pools of cloacal swabs of wild mallards (Anas platyrhynchos) caught in a single sampling site at two different times. AIV and APMV1 were excluded using hemagglutination inhibition (HI) testing and specific real-time RT-PCR tests. METHODS: To refine the virological identification of APMV2-10 realized by HI subtyping tests and in lack of validated molecular tests for APMV2-10, random access amplification was used in combination with next generation sequencing for the sequence independent identification of the viruses and the determination of their genomes. RESULTS: Three different APMVs were identified. From one pooled sample, the complete genome sequence (15054 nucleotides) of an APMV4 was assembled from the random sequences. From the second pooled sample, the nearly complete genome sequence of an APMV6 (genome size of 16236 nucleotides) was determined, as well as a partial sequence for an APMV4. This APMV4 was closely related but not identical to the APMV4 isolated from the first sample. Although a cross-reactivity with other APMV subtypes did not allow formal identification, the HI subtyping revealed APMV4 and APMV6 in the respective pooled samples but failed to identify the co-infecting APMV4 in the APMV6 infected pool. CONCLUSIONS: These data further contribute to the knowledge about the genetic diversity within the serotypes APMV4 and 6, and confirm the limited sensitivity of the HI subtyping test. Moreover, this study demonstrates the value of a random access nucleic acid amplification method in combination with massive parallel sequencing. Using only a moderate and economical sequencing effort, the characterization and full genome sequencing of APMVs can be obtained, including the identification of viruses in mixed infections. BioMed Central 2011-10-06 /pmc/articles/PMC3219605/ /pubmed/21978491 http://dx.doi.org/10.1186/1743-422X-8-463 Text en Copyright ©2011 Rosseel et al; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Rosseel, Toon Lambrecht, Bénédicte Vandenbussche, Frank van den Berg, Thierry Van Borm, Steven Identification and complete genome sequencing of paramyxoviruses in mallard ducks (Anas platyrhynchos) using random access amplification and next generation sequencing technologies |
title | Identification and complete genome sequencing of paramyxoviruses in mallard ducks (Anas platyrhynchos) using random access amplification and next generation sequencing technologies |
title_full | Identification and complete genome sequencing of paramyxoviruses in mallard ducks (Anas platyrhynchos) using random access amplification and next generation sequencing technologies |
title_fullStr | Identification and complete genome sequencing of paramyxoviruses in mallard ducks (Anas platyrhynchos) using random access amplification and next generation sequencing technologies |
title_full_unstemmed | Identification and complete genome sequencing of paramyxoviruses in mallard ducks (Anas platyrhynchos) using random access amplification and next generation sequencing technologies |
title_short | Identification and complete genome sequencing of paramyxoviruses in mallard ducks (Anas platyrhynchos) using random access amplification and next generation sequencing technologies |
title_sort | identification and complete genome sequencing of paramyxoviruses in mallard ducks (anas platyrhynchos) using random access amplification and next generation sequencing technologies |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3219605/ https://www.ncbi.nlm.nih.gov/pubmed/21978491 http://dx.doi.org/10.1186/1743-422X-8-463 |
work_keys_str_mv | AT rosseeltoon identificationandcompletegenomesequencingofparamyxovirusesinmallardducksanasplatyrhynchosusingrandomaccessamplificationandnextgenerationsequencingtechnologies AT lambrechtbenedicte identificationandcompletegenomesequencingofparamyxovirusesinmallardducksanasplatyrhynchosusingrandomaccessamplificationandnextgenerationsequencingtechnologies AT vandenbusschefrank identificationandcompletegenomesequencingofparamyxovirusesinmallardducksanasplatyrhynchosusingrandomaccessamplificationandnextgenerationsequencingtechnologies AT vandenbergthierry identificationandcompletegenomesequencingofparamyxovirusesinmallardducksanasplatyrhynchosusingrandomaccessamplificationandnextgenerationsequencingtechnologies AT vanbormsteven identificationandcompletegenomesequencingofparamyxovirusesinmallardducksanasplatyrhynchosusingrandomaccessamplificationandnextgenerationsequencingtechnologies |