JCM Accepts, published online ahead of print on 29 July 2009

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J. Clin. Microbiol. doi:10.1128/JCM.00999-09
Copyright (c) 2009, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Rapid semi-automated Subtyping of influenza during the 2009 swine-origin influenza A H1N1 virus epidemic in Milwaukee, Wisconsin.

Michael E. Bose, Eric T. Beck, Nate Ledeboer, Sue C. Kehl, Lisa A. Jurgens, Teresa Patitucci, Lorraine Witt, Elizabeth LaGue, Patrick Darga, Jie He, Jiang Fan, Swati Kumar, and Kelly J. Henrickson^*

Midwest Respiratory Virus Program (MRVP), Department of Pediatrics and Pathology, Medical College of Wisconsin, Children's Research Institute, Children's Hospital of Wisconsin, Dynacare Laboratories, Milwaukee, Wisconsin, USA

^* To whom correspondence should be addressed. Email: Khenrick{at}mcw.edu.

In spring of 2009 a novel influenza A (H1N1) virus [swine-origin influenza virus (S-OIV)] emerged and began causing a large outbreak in Milwaukee, WI. Our laboratory developed a semi-automated NucliSENS® easyMAG^TM (bioMérieux, Durham, NC) and Raider (Handylab Inc., Ann Arbor, MI)] real-time multiplex RT-PCR (Seasonal) assay that typed influenza A, influenza B, RSV and subtyped influenza A into currently circulating H1 and H3 subtypes and a similar assay that identified H1 of S-OIV. These assays demonstrated analytical limits of detection of <50 TCID₅₀/ml and 3-30 input copies respectively. The analytical specificity demonstrated no cross-reactivity against 39, and 27 different common organisms tested and outstanding reproducibility. Clinical testing demonstrated 95% sensitivity against influenza A and influenza B and 95% and 97% specificity compared to tissue culture. Compared to other molecular tests positive agreement for the Seasonal and H1 S-OIV assays were 99% and 100% and negative agreement was 98% and 100%.

This study has demonstrated the use of a semi-automated system for sensitive, specific, and rapid detection of influenza A, influenza B, and RSV and subtyping of influenza A into human H1 and H3 and S-OIV strains. This assay/system performed well in clinical testing of regular seasonal influenza subtypes and has been outstanding during the current Milwaukee S-OIV outbreak. This recent outbreak of a novel influenza A (H1N1) virus also demonstrates the importance of quickly distributing bioinformatics on new agents and in having rapid influenza subtyping assays widely available for clinical and public health decisions.

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