Pyrosequencing — a robust method for detecting antiviral resistance in seasonal and 2009 A (H1N1) influenza A viruses circulating with Department of Defense populations


Resistance to antiviral agents in influenza A viruses has steadily increased globally. The Centers for Disease Control (CDC) has reported incidences of 100% adamantane resistance in some influenza A subtypes and >98% of previous seasonal A/H1N1 is resistant to Oseltamivir. Single nucleotide polymorphisms (SNPs) with matrix (M2) and neuraminidase (NA) genes correlate well with in vitro and in vivo evidence of antiviral resistance. Pyrosequencing can be optimized to produce short, targeted genetic sequences that contain non-synonymous nucleotide changes that correlate with resistance to antiviral chemotherapeutics. The US Air Force School of Aerospace Medicine has employed Pyrosequencing to detect molecular markers of antiviral resistance within M2 and NA genes of influenza A viruses collected from Department of Defense (DoD) populations. A quick, highly sensitive, and cost-effective alternative to Sanger sequencing, Pyrosequencing allows rapid detection of common SNPs associated with adamantane and neuraminidase inhibitor (NAI) resistance respectively. Monitoring antiviral susceptibility within DoD populations complements influenza surveillance efforts and allows timely information exchange with the Armed Forces Health Surveillance Center, DoD sentinel sites, public health agencies, and collaborative partners.

This presentation provides:

An overview of the PyroMark Pyrosequencing based detection platform
An introduction to the DoD sentinel-based influenza surveillance program
A description of efforts to incorporate assays to detect M2-blocker and NAI susceptibilities in seasonal A/H3N2 viruses into the DoD program

Frank Fischinger MS, MBA

Frank Fischinger is currently a senior member of QIAGEN's North American PyroMark Field Applications Group. Frank is one of QIAGEN's Pyrosequencing experts providing field support to researchers and clinicians with their PyroMark systems and Pyrosequencing applications. Frank has five years experience in methylation assay design, assay optimization, and assay troubleshooting using Pyrosequencing technology. In addition, Frank has managed the Clinical Neuroimmunology lab at the University of Chicago and has provided field technical support for Abbot Molecular Diagnostics Fluorescent In-Situ Hybridization and Microarray platforms. Frank was awarded an MS/MBA dual degree in Molecular Pharmacology and Marketing from Loyola University Chicago.
Jason L. Garner

Jason L. Garner is a Senior Molecular Biologist with the Henry M. Jackson Foundation for the Advancement of Military Medicine and US Air Force School of Aerospace Medicine (USAFSAM) at Brooks City-Base in Texas. Mr. Garner's career in research, molecular biology, and clinical diagnostics spans almost 10 years, supplemented by over 10 years in operations management. Currently, Mr. Garner's primary activities include high-throughput detection, isolation, and molecular characterization of global seasonal/pandemic influenza A & B viruses within Department of Defense communities. Mr. Garner leads analyses efforts with USAFSAM that contribute to FDA/VRBPAC annual flu vaccine selection considerations and has developed, validated, and implemented novel molecular-based clinical diagnostic assays for Department of Defense flu surveillance efforts. Mr. Garner is a Tier II Honors graduate from the University of Texas at San Antonio with a BS in Biology and MS in Cell/Molecular Biology as well as an NIH graduate fellow. Mr. Garner has authored and co-authored over 20 journal articles, posters, and oral presentations.