Strategies to Detect Novel Influenza Virus Infections in Humans with the CDC rRT-PCR Diagnostic Panel Pan American Health Organization Meeting Strengthening Influenza Inter-Sectoral Surveillance and Response Washington, D.C. October 2-3. 2017 Stephen Lindstrom, Ph.D. Diagnostics Development Team Virus Surveillance and Diagnosis Branch Influenza Division Centers for Disease Control and Prevention The findings and conclusions in this presentation are those of the author(s) and do not necessarily represent the views of the Centers for Disease Control and Prevention National Center for Immunization and Respiratory Diseases Influenza Division

Overview Strategies to detect novel influenza virus infections in humans Situation update for variant influenza swine origin H3v & H1v in US

Influenza Viruses in Humans - 2017 Avian H5 Avian H6 Avian H7 Avian H9 Potential Pandemic strains Avian H10 H3v Type A H1v H1 H3 H2 H1 2009 1998/9 Seasonal Strains 1918 1940 1957 1968 1977 1997 2003 2017 B/Yam Type B B/Vic

Avian influenza HPAI H5 and H7N9 H5N1/H5Nx, H7N9 Poultry

H5N8 HPAI outbreaks officially reported in Asia, Europe and Africa since 1 January 2017 http://www.fao.org/ag/againfo/programmes/en/empres/h5n8/situation_update.html

Cumulative number of confirmed human cases for avian influenza A(H5N1) reported to WHO, 2003-2017

H7N9 human cases and positive findings in birds or the environment http://www.fao.org/ag/againfo/programmes/en/empres/h7n9/situation_update.html

Incidence of officially reported human cases of H7N9 5th Wave Cases >650 Official HPAI Cases 22 as of Aug 23, 2017 (FAO) http://www.fao.org/ag/againfo/programmes/en/empres/h7n9/situation_update.html

CDC Influenza Virus Real-time RT-PCR Reagent Kits Influenza A/B Typing Panel (InfA, Inf B, RP) Influenza A Subtyping Panel (InfA, H1, H3, pdmInfA, pdmH1) Influenza B genotyping Panel (InfB, Yam, Vic, RP) Influenza A H5 (Asian lineage) Subtyping Panel (VER3) (InfA, H5a, H5b, RP) Influenza A EuH7 Assay (EuH7)

CDC Influenza rRT-PCR Reagent distribution: 2017 117 countries received CDC rRT-PCR reagents

H3N2 (Asian avian origin) Other Animal Viruses with Potential Pandemic Other Potential Pandemic Viruses Swine Avian H3N2v H1N1v/H1N2v H9N2 N. Am. H7 (HPAI & LPAI) Canine Feline (avian) H3N2 (Asian avian origin) H3N8 (equine origin) N. Am. H7N2 (LPAI)

Human cases of “variant” Influenza infection in the United States H3N2v, H1N1v, H1N2v Pigs

H3N2v Results Interpretation Result Interpretation CDC rRT-PCR Assay: H3N2v Results Interpretation InfA H3 pdm InfA pdmH1 RP Result Interpretation + - ± Influenza A detected; Presumptive positive for Influenza A(H3N2) variant virus

Infections with Variant Influenza Viruses by State since December 2005 - 2016 H3N2v H1N1v H1N2v Total Detected Total cases 372 20 9 401 Total states 15 5 18

Detected U.S. Infections with Variant Influenza Viruses by State 2017…so far H3N2v H1N1v H1N2v Total Detected Total cases 54 3 57 Total states 6

Fair Season 2017…so far!! A(H3N2)v: 54 human cases Ohio – 16 cases / 3counties North Dakota – 2 cases / 1 count Maryland – 35 cases / 3 counties Michigan – 1 case / 1 county A(H1N2)v: Ohio – 1 case / 1 county

Swine flu in the news… “it's been difficult, especially for children whose animals will be slaughtered”

County Fairs and Illness Onset Cases by illness onset July 8 July 15 July 22 July 29 Aug 5 Clark OH (Jul 21- 28) Clinton, OH (July 8-15) Ward ND (July 21-29) Warren, OH (July 17-22) Union, OH (Jul 23-29) County Fares

Establishing source of infection: Finding the “smoking pig” Collaboration between animal/human health important to confirm virologic link and ensure effectiveness of diagnostic tests

Evolutionary Relationships Among Influenza A(H3N2)v HA, 2017 N145K A/swine/Illinois/A01409675/2014 H3N2 A/swine/Illinois/A01475266/2014 H3N2 A/swine/Illinois/A01409717/2014 H3N2 A/swine/Illinois/A01409611/2014 H3N2 A/swine/Illinois/A00843193/2014 H3N2 A/Indiana/19/2013 H3N2v A/Illinois/03/2013 H3N2v N145R A/Indiana/09/2013 H3N2v A/Ohio/05/2013 H3N2v A/Indiana/10/2013 H3N2v A/swine/South Dakota/A01480341/2014 H3N2 A/swine/Nebraska/A01491325/2014 H3N2 H3N2v vaccine candidate in red CDC HI reference viruses in blue Recent US H3N2v virus Viruses collected in 2015/2016 (*) Amino acid differences are relative to Human-like lineage - A/Ohio/28/2016 Cluster IV lineage - A/Minnesota/11/2010 A/swine/Minnesota/A01280462/2013 H3N2 A/Iowa/04/2013 Oct H3N2v A/Ohio/86/2012 H3N2v A/swine/Iowa/A01410633/2014 H3N2 P289S A/Ohio/77/2012 H3N2v A/Hawaii/03/2012 H3N2v A/swine/Indiana/A00968373/2012 H3N2 A/Ohio/13/2012 H3N2v A/Indiana/21/2012 H3N2v A/swine/Illinois/A01502053/2014 H3N2 A/swine/Iowa/A01410748/2014 H3N2 A/West Virginia/06/2011 H3N2v A/swine/North Carolina/A01410136/2014 H3N2 A/swine/North Carolina/A01475277/2014 H3N2 F79L A/swine/Illinois/A01241469/2012 H3N2 A/swine/NY/A01104005/2011 H3N2 A/Utah/10/2012 H3N2 A/Indiana/10/2011 H3N2v A/Indiana/10/2011 X-213 H3N2v G49S F79S V88I A/swine/Illinois/A01411097/2014 H3N2 A/Michigan/39/2015 H3N2v A/Texas/105/2017_H3N2v H3-IV(A) E83K V112I Q261L T48I K207R P289S 16TOSU5235 HA A11T A/Michigan/94/2016 H3N2v A/Michigan/84/2016 H3N2v A/swine/North Carolina/A01410877/2014 H3N2 A/swine/Iowa/A01410847/2014 H3N2 A/New Jersey/53/2015 H3N2v A/swine/Iowa/A01410995/2014 H3N2 A/swine/Iowa/A01501525/2014 H3N2 A/Wisconsin/24/2014 H3N2v A/swine/Iowa/A01410752/2014 H3N2 A/swine/Iowa/A01410600/2014 H3N2 A/Pennsylvania/40/2010 H3N2v A/swine/Iowa/A01501048/2014 H3N2 A/swine/Iowa/A01410971/2014 H3N2 A/Minnesota/11/2010 H3N2v A/Wisconsin/12/2010 H3N2v A/Minnesota/11/2010 X-203 H3N2v D133G A/swine/Michigan/A01259000/2012 H3N2 A/swine/Wyoming/A01444562/2013 H3N2 Y155H K189R A/swine/Minnesota/00709/2005 H3N2 A/Indiana/08/2011 H3N2v D275G E276N N117T H273P A/swine/Minnesota/01146/2006 H3N2 T107S N158D K299R A/swine/Minnesota/00484/2005 H3N2 A/swine/Minnesota/1300/2007 H3N2 A/swine/Iowa/01700/2007 H3N2 A/swine/Minnesota/5947/2007 H3N2 T107S G142K K299R M10T A106V S124N K173N V196I N278K A304V A/swine/Pennsylvania/A01076777/2010 H3N2 A/Pennsylvania/14/2010 H3N2v K189R N158D A/swine/Minnesota/02782/2009 H3N2 A/Kansas/13/2009 H3N2v A/swine/Illinois/02907/2009 H3N2 A/swine/Minnesota/18/2012 H3N2 E83K K299R N53S N117T S135A N156S R208W A/swine/Minnesota/A01327922/2012 H3N2 S46T T48K R50E A138S N193S V196I D275G E276N A/swine/Arkansas/A01554688/2015 H3N2 A/swine/Iowa/A01822205/2015 H3N2 A/swine/Iowa/A02077555/2015 H3N2 A/swine/Illinois/A01554952/2015 H3N2 H3-IV(B) I203T Q164L E163A G142E V223I V144E T107S A/Ohio/02/2014 H3N2v A/Minnesota/38/2015 H3N2v A/swine/Iowa/A01300195/2012 H3N2 A/Ohio/4319/2014 H3N2v T107S V144D H273P I217V S228G I260M G263R H273P E276N K299R S9N T107S A131S A138T G142K T192I N193S A/swine/Illinois/A01201606/2011 H3N2 A/swine/Minnesota/A01432433/2013 H3N2 A/swine/Minnesota/A01501738/2014 H3N2 N117T Y155H D275G S279F A/swine/Michigan/A01203498/2012 H3N2 A/swine/Iowa/A01203751/2012 H3N2 Y105H V144D N145K N156K K173N D188N K189E R201K D275G A/swine/Iowa/A01202889/2011 H3N2 A/swine/Iowa/A01203196/2012 H3N2 I34V D78E K82T A106S T107S N117T N126D R140K Y155H E163A Q164L K173N K189R T192A I203T I229V S262N K299R A/swine/Iowa/A01202613/2011 H3N2 A/swine/Iowa/A01203197/2012 H3N2 R50G Y137R A138S N158G Y159H K173N K189E N193S H273S A/turkey/Ontario/FAV-9/2011 H3N2 A/swine/Iowa/A01049750/2011 H3N2 A/swine/Nebraska/A01241171/2012 H3N2 T107S N117T N156H E163A Q164L E276N K299R V144G A/swine/Indiana/A01202866/2011 H3N2 A/swine/Michigan/A01259034/2012 H3N2 A/swine/Illinois/A01411363/2014 H3N2 N53S E83K T121N Y137N G142E V144N N145K Y159N N193Y S199P I203T V204I I217V A/Ontario/RV3236/2016 H3N2 K207R H273P N278D A/swine/Indiana/A01260983/2015 A / H3N2 A/swine/Indiana/A01812045/2015 A / H3N2 A/swine/Indiana/A01260967/2015 A / H3N2 A/swine/North Carolina/A01945244/2015 A / H3N2 A/swine/Iowa/A02025409/2015 A / H3N2 A/swine/Iowa/A01945156/2015 A / H3N2 A/swine/North Carolina/A01797533/2015 A / H3N2 A/swine/Iowa/A01730108/2016 A / H3N2 A/swine/North Carolina/A01945800/2015 A / H3N2 A/Michigan/82/2016 * H3N2v A/Ohio/32/2016 * H3N2v A/Ohio/27/2016 * H3N2v A/Ohio/35/2016 * H3N2v 16SW4788 N158D A/Michigan/87/2016 * H3N2v A/Michigan/83/2016 * H3N2v A01104095 lung 16TOSU4788 16SW4783 A01104095 A/Michigan/90/2016 * H3N2v A/Michigan/96/2016 * H3N2v 16TOSU5097 A/Michigan/89/2016 * H3N2v 16TOSU5081 A/Michigan/93/2016 * H3N2v A/Ohio/28/2016 * H3N2v 16TOSU4783 A/Ohio/29/2016 * H3N2v A/Michigan/91/2016 * H3N2 A/swine/Iowa/A01727249/2015 * H3N2 A/swine/Iowa/A01796498/2015 * H3N2 A/swine/Iowa/A01797426/2015 * H3N2 A/swine/Minnesota/A01941510/2015 * H3N2 A/swine/Iowa/A01795441/2015 * A A/swine/Minnesota/A01677935/2016 * H3N2 A/swine/Minnesota/A01941094/2015 * H3N2 A/swine/Minnesota/A01677360/2016 * H3N2 I242T A/swine/Minnesota/A01676585/2016 * H3N2 A/swine/North_Carolina/A01728953/2016 * H3N2 A/swine/Iowa/A01733867/2016 * H3N2 A/swine/Iowa/A01775848/2016 * H3N2 A/swine/Iowa/A01729570/2016 * H3N2 S114T A/swine/Illinois/A01795739/2015 * H3N2 A/swine/Iowa/A01775597/2016 * H3N2 Seasonal human-like H3 A/swine/Iowa/A01847868/2015 * H3N2 A/Michigan/95/2016 * H3N2v_partial A/swine/Iowa/A01728010/2015 * H3N2 A/swine/Iowa/A01895860/2016 * H3N2 A/swine/Missouri/A01847626/2015 * H3N2 A/Ohio/33/2016 * H3N2v_partial A/swine/Missouri/A01795668/2015 * H3N2 A/swine/Ohio/A01798226/2015 * H3N2 16TOSU5240_HA A/swine/Ohio/A01795737/2015 * H3N2 A/swine/Indiana/A01796207/2015 * H3N2 A/swine/Missouri/A01770005/2015 * H3N2 A/swine/Arkansas/A01797066/2015 * H3N2 A/swine/Indiana/A01730580/2016 * H3N2 A/swine/Ohio/A01728296/2015 * H3N2 S219Y A/swine/Minnesota/A01847237/2015 *H3N2 A/swine/Missouri/A01822838/2015 * H3N2 A/swine/Missouri/A01798099/2015 * H3N2 A/swine/Missouri/A01554556/2015 * H3N2 A/swine/Iowa/A01944136/2015 * H3N2 N216D A/swine/Illinois/A01796457/2015 * H3N2 A/swine/Missouri/A01943095/2016 * H3N1 A/swine/Missouri/A01840163/2014_H3N2 A/swine/Missouri/A01840324/2015 * H3N2 A/swine/Missouri/A01840170/2014_H3N2 A/swine/Missouri/A02076058/2015 * H3N2 A/swine/Missouri/A01554320/2015 * A A/Ohio/14/2017 4 A/swine/Ohio/17TOSU1389/2017 HA A/Ohio/13/2017 4 A01354299 swab HA A/swine/Ohio/17TOSU1395/2017 HA H56Y A138S K145N S209N A/Ohio/15/2017 4 A/Ohio/17/2017 4 A/Ohio/21/2017 4 A/Ohio/19/2017 4 A/Ohio/20/2017 4 A/Ohio/22/2017 4 A/swine/Iowa/A01944889/2015 * A A/Ohio/23/2017 4 A/swine/Missouri/A01476458/2013_H3N1 A/swine/Iowa/A02077646/2015 * H3N2 L3I F46S N82K D133N S144N I230V A/swine/Missouri/A01410819/2014_H3N1 A/swine/Missouri/A01410818/2013_H3N1 A/swine/Iowa/A01476914/2014_H3N2 F46S N82K D133N S144N I230V N290S S262G A/swine/Minnesota/A01940176/2015 * H3N2 A/swine/Minnesota/A01941831/2015 * H3N2 A/swine/Iowa/A01728956/2016 * H3N2 A/swine/Iowa/A01566613/2014_H3N2 A186G I203T I230V R238K I323V A135T K140I S144N K145N E165N N9S M10T F46S N82K D133N A/swine/Iowa/A01941549/2015 * H3N2 T48I Q57R A/swine/Minnesota/A01943420/2016 * H3N2 A/swine/Minnesota/A01941693/2015 * H3N2 A/South_Dakota/02/2010_H3N2 A/swine/Missouri/A01476459/2012_H3N2 A/swine/Iowa/A01476471/2014_H3N1 K83T A/swine/Minnesota/A01942171/2016 * H3N2 0.01 A/Florida/24/2011_H3N2 A/Peru/PER373/2010_H3N2 D225N A/Chile/72/2011_H3N2 A199S I203T V223I R238K A280E N312S I323V H94Y D133N A135T S144N E165N S193F N9S M10T S45N F46S T48I N53D N82K S193F A/Rhode_Island/04/2011_H3N2 A/California/22/2011_H3N2 A/District_of_Columbia/03/2011_H3N2 D225N A/Mississippi/05/2011_H3N2 A/Tennessee/F2011/2011_H3N2 A/Argentina/676/2011_H3N2 A/Boston/DOA75/2011_H3N2 A/Santiago/18454/2011_H3N2 A/Mississippi/03/2011_H3N2 A/Minnesota/09/2011_H3N2 A/Tennessee/F2018/2011_H3N2 Seasonal human H3 D188Y A199S R201I I203T A212T R238K A280E I323V A135K A138S R142G S144D F159Y E165K Q173K L3F N9S I25M E50G E62G N82K L111I D133N A/Victoria/361/2011_H3N2 A/Texas/50/2012_H3N2 A/Switzerland/9715293/2013_H3N2 A/Wisconsin/67/2005_H3N2 A/Brisbane/10/2007_H3N2 A/Perth/16/2009_H3N2 Q33R N278K A/swine/Binh_Duong/03_08/2010_H3N2 A/swine/Hong_Kong/NS2439/2011_H3N2 A/Vietnam/85/2015 * H3N2v S137A K145N H56Y V182A S209T A/swine/Hong_Kong/2503/2011_H3N2 A/Fujian/411/2002_H3N2 H94Y K189N A/swine/Guangxi/NS2783/2010_H3N2 A/Vietnam/302/2011_H3N2 A/California/32/1999_H3N2 A/swine/Iowa/H03LDH5/2003_H3N2 A/Moscow/10/99_H3N2 A/Wuhan/359/1995_H3N2 A/swine/Texas/4199-2/1998_H3N2 A/Sydney/5/1997_H3N2

Genome Comparisons of H3N2v 2005-2017 Classical Swine – North American lineage Avian – North American lineage Seasonal H3N2 (Cluster IV; 1997/1998 lineage) Seasonal H3N2 (human-like; 2010/2011 lineage Eurasian swine lineage (from H1N1 pdm09 virus) Gene derived from H1N1 pdm09 virus Majority of 2011-2012 (n=300) and 2013 from Indiana (n=13) A/Minnesota/38/2015 A/Michigan/39/2015 A/New Jersey/53/2015 2016 (n=2) Michigan IV-A 2013 (n=5) Ohio, Illinois, Indiana, Michigan 2013 (n=1) Iowa 2005-2010 PB2 PB2 PB2 PB2 PB1 PB1 PB1 PB1 PA PA PA PA HA HA HA HA NP NP NP NP NA NA NA NA MP MP MP MP NS NS NS NS 2014 (n=3) Ohio(IV-B), Wisconsin(IV-A) 2017 (n=1) Texas (IV-A) 2016 (n=16) Ohio, Michigan, human-like 2016 (n=1) Ontario, Canada IV-B 2017 (n=7) Ohio human-like PB2 PB2 PB2 PB2 PB1 PB1 PB1 PB1 PA PA PA PA HA HA HA HA NP NP NP NP NA NA NA NA MP MP MP MP NS NS NS NS

Evolutionary Relationships Among Influenza A (H1v) HA Genes Multiple genotypes of H1 in US swine: “Classical swine” H1 Pre-2009 human H1 (H1)pdm2009 Eurasian avian H1 WHO WER No12, 24 Mar 2017

Diagnostics Development Team Lakshmi Malapati Ji Liu LaShondra Berman Shannon Emery Bo Shu Bill Davis Christine Warnes Brian Lynch Janna Murray Kai-Hui Wu

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