Giuseppe
Emeritus
J Clin Microbiol. 2008 Dec 3. [Epub ahead of print]
Design and validation of a microarray for detection, haemagglutinin subtyping and pathotyping of avian influenza viruses.
Gall A, Hoffmann B, Harder T, Grund C, Hoeper D, Beer M. - Institute of Diagnostic Virology, Friedrich-Loeffler-Institut, S?dufer 10, 17493 Greifswald - Insel Riems, Germany.
Continuing threats of devastating outbreaks in poultry and of human infections by highly pathogenic avian influenza virus (HPAIV) H5N1 emphasize the need for further development of rapid and reliable methods of virus detection and characterization.
Here we report the design and comprehensive validation of a low-density microarray for AIV as a diagnostic tool.
The array consists of one probe for the conserved matrix gene and 97 probes targeting the HA0 cleavage site region.
Following fragment amplification by a generic PCR approach, the array enables AIV detection, HA subtyping and pathotyping within a single assay.
For validation, a panel of 92 influenza A viruses was used, which included 43 reference strains representing all 16 HA-subtypes.
All reference strains were correctly typed with respect to their HA-subtypes and pathotypes, including HPAIV H5N1/Asia, which had caused outbreaks in Germany in 2006 and 2007. In addition, differentiation of Eurasian and North American lineage strains of the H5- and H7-subtypes was possible. Sensitivity of the microarray for the M gene is comparable to real-time RT-PCR. It is, however, 10- to 100-fold lower than real-time RT-PCR with respect to HA subtyping/pathotyping.
Specificity of the array was excellent as no pathogens relevant for differential diagnosis yielded a positive reaction. Validation with field samples included 19 cloacal swabs from wild and domestic birds. Influenza A virus was verified in all samples whereas HA-subtypes could be determined for 14 samples.
The results demonstrate that the described microarray complements current methods and can accelerate diagnosis and characterization of AIV.
PMID: 19052173 [PubMed - as supplied by publisher
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Design and validation of a microarray for detection, haemagglutinin subtyping and pathotyping of avian influenza viruses.
Gall A, Hoffmann B, Harder T, Grund C, Hoeper D, Beer M. - Institute of Diagnostic Virology, Friedrich-Loeffler-Institut, S?dufer 10, 17493 Greifswald - Insel Riems, Germany.
Continuing threats of devastating outbreaks in poultry and of human infections by highly pathogenic avian influenza virus (HPAIV) H5N1 emphasize the need for further development of rapid and reliable methods of virus detection and characterization.
Here we report the design and comprehensive validation of a low-density microarray for AIV as a diagnostic tool.
The array consists of one probe for the conserved matrix gene and 97 probes targeting the HA0 cleavage site region.
Following fragment amplification by a generic PCR approach, the array enables AIV detection, HA subtyping and pathotyping within a single assay.
For validation, a panel of 92 influenza A viruses was used, which included 43 reference strains representing all 16 HA-subtypes.
All reference strains were correctly typed with respect to their HA-subtypes and pathotypes, including HPAIV H5N1/Asia, which had caused outbreaks in Germany in 2006 and 2007. In addition, differentiation of Eurasian and North American lineage strains of the H5- and H7-subtypes was possible. Sensitivity of the microarray for the M gene is comparable to real-time RT-PCR. It is, however, 10- to 100-fold lower than real-time RT-PCR with respect to HA subtyping/pathotyping.
Specificity of the array was excellent as no pathogens relevant for differential diagnosis yielded a positive reaction. Validation with field samples included 19 cloacal swabs from wild and domestic birds. Influenza A virus was verified in all samples whereas HA-subtypes could be determined for 14 samples.
The results demonstrate that the described microarray complements current methods and can accelerate diagnosis and characterization of AIV.
PMID: 19052173 [PubMed - as supplied by publisher
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