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Journal of Clinical Microbiology, September 2009, p. 2779-2786, Vol. 47, No. 9
0095-1137/09/$08.00+0     doi:10.1128/JCM.00999-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Rapid Semiautomated Subtyping of Influenza Virus Species during the 2009 Swine Origin Influenza A H1N1 Virus Epidemic in Milwaukee, Wisconsin{triangledown}

Michael E. Bose,1,2 Eric T. Beck,1,2 Nate Ledeboer,3,6 Sue C. Kehl,1,3,4,5 Lisa A. Jurgens,1,2 Teresa Patitucci,1,2 Lorraine Witt,1,2 Elizabeth LaGue,1,2 Patrick Darga,1,2 Jie He,1,2 Jiang Fan,1,2 Swati Kumar,1,2,4,5 and Kelly J. Henrickson1,2,4,5*

Midwest Respiratory Virus Program,1 Departments of Pediatrics,2 Pathology, Medical College of Wisconsin,3 Children's Research Institute,4 Children's Hospital of Wisconsin,5 Dynacare Laboratories, Milwaukee, Wisconsin6

Received 19 May 2009/ Returned for modification 7 July 2009/ Accepted 23 July 2009

In the spring of 2009, a novel influenza A (H1N1) virus (swine origin influenza virus [S-OIV]) emerged and began causing a large outbreak of illness in Milwaukee, WI. Our group at the Midwest Respiratory Virus Program laboratory developed a semiautomated real-time multiplex reverse transcription-PCR assay (Seasonal), employing the NucliSENS easyMAG system (bioMérieux, Durham, NC) and a Raider thermocycler (HandyLab Inc., Ann Arbor, MI), that typed influenza A virus, influenza B virus, and respiratory syncytial virus (RSV) and subtyped influenza A virus into the currently circulating H1 and H3 subtypes, as well as a similar assay that identified H1 of S-OIV. The Seasonal and H1 S-OIV assays demonstrated analytical limits of detection of <50 50% tissue culture infective doses/ml and 3 to 30 input copies, respectively. Testing of the analytical specificities revealed no cross-reactivity with 41 and 26 different common organisms and demonstrated outstanding reproducibility of results. Clinical testing showed 95% sensitivity for influenza A virus and influenza B virus and 95 and 97% specificity compared to tissue culture. Comparisons of results from other molecular tests showed levels of positive agreement with the Seasonal and H1 S-OIV assay results of 99 and 100% and levels of negative agreement of 98 and 100%. This study has demonstrated the use of a semiautomated system for sensitive, specific, and rapid detection of influenza A virus, influenza B virus, and RSV and subtyping of influenza A virus into human H1 and H3 and S-OIV strains. This assay/system performed well in clinical testing of regular seasonal influenza virus subtypes and was outstanding during the 2009 Milwaukee S-OIV infection outbreak. This recent outbreak of infection with a novel influenza A (H1N1) virus also demonstrates the importance of quickly distributing information on new agents and of having rapid influenza virus subtyping assays widely available for clinical and public health decisions.

* Corresponding author. Mailing address: Pediatrics/Infectious Disease/CCC/Suite c450, Children's Corporate Center, P.O. Box 1997, Milwaukee, WI 53201-1997. Phone: (414) 337-7073. Fax: (414) 337-7093. E-mail: Khenrick{at}mcw.edu

{triangledown} Published ahead of print on 29 July 2009.

Journal of Clinical Microbiology, September 2009, p. 2779-2786, Vol. 47, No. 9
0095-1137/09/$08.00+0     doi:10.1128/JCM.00999-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • He, J., Bose, M. E., Beck, E. T., Fan, J., Tiwari, S., Metallo, J., Jurgens, L. A., Kehl, S. C., Ledeboer, N., Kumar, S., Weisburg, W., Henrickson, K. J. (2009). Rapid Multiplex Reverse Transcription-PCR Typing of Influenza A and B Virus, and Subtyping of Influenza A Virus into H1, 2, 3, 5, 7, 9, N1 (Human), N1 (Animal), N2, and N7, Including Typing of Novel Swine Origin Influenza A (H1N1) Virus, during the 2009 Outbreak in Milwaukee, Wisconsin. J. Clin. Microbiol. 47: 2772-2778 [Abstract] [Full Text]  


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