A multiplex one-step real-time RT-PCR assay for influenza surveillance
| Autor: | Regina Konrad, H Campe, Ingrid Huber, D Sebah, C Hartberger, Ulrich Busch, M Bayer, Andreas Sing |
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| Rok vydání: | 2011 |
| Předmět: |
Genes
Viral Epidemiology Pcr assay Biology medicine.disease_cause Sensitivity and Specificity law.invention Influenza A Virus H1N1 Subtype law Virology Germany Multiplex polymerase chain reaction Influenza Human Influenza A virus medicine Humans Multiplex Polymerase chain reaction DNA Primers Reverse Transcriptase Polymerase Chain Reaction Public Health Environmental and Occupational Health Pandemic influenza Gene Amplification virus diseases Influenza pandemic Influenza B virus Real-time polymerase chain reaction Population Surveillance RNA Viral Reagent Kits Diagnostic |
| Zdroj: | Euro surveillance : bulletin Europeen sur les maladies transmissibles = European communicable disease bulletin. 16(7) |
| ISSN: | 1560-7917 |
| Popis: | For surveillance purposes real-time PCR assays for influenza viruses had to be adapted to the pandemic influenza A(H1N1)2009 strain. We combined published primers and probes for influenza A, influenza B and an internal amplification control with a detection system for influenza A(H1N1)2009 to set up a rapid, reliable, simple and cost-effective high-throughput multiplex one-step real-time RT-PCR. The workflow also includes automated sample preparation for high-throughput screening. The lower limit of detection of the multiplex assay was 3.5x102 RNA copies per PCR reaction. The diagnostic sensitivity of the multiplex assay was 87.7%, but increased to 99.4% for influenza-positive samples yielding Ct values of less than 34 cycles in the respective diagnostic assay. High specificity was confirmed by sequencing and correct detection of 15 reference samples from two quality assurance studies. The multiplex PCR was introduced for surveillance of samples from a network of general practitioners and paediatricians in Bavaria, Germany during the influenza pandemic of 2009. Comparison with surveillance data from reported cases proved the reliability of the multiplex assay for influenza surveillance programmes. |
| Databáze: | OpenAIRE |
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