This document summarizes research on the growing diversity of H3N2 influenza A virus in US swine and its impact. It finds that the H3 subtype is evolving into multiple genetic clusters with evidence of reassortment. Antibody studies show inconsistent cross-reactivity between clusters, which may impact vaccine effectiveness. Nearly 350 human cases of influenza from exposure to infected fair pigs have been reported since 2005, primarily in children. Continued genetic monitoring, timely vaccine updates, and partnerships across animal and human health are needed to minimize the impact.

1. Growing diversity of H3N2
influenza A virus in swine:
Impact on control in swine and at
the human animal interface.
Amy L. Vincent, DVM PhD
Research Veterinary Medical Officer
National Animal Disease Center
USDA-ARS
USDA-NADC Team
USDA-NVSL
USDA-NAHLN Laboratories
USDA-NSU
US Swine Producers and Veterinarians

2. USDA IAV Surveillance in Swine
• Implemented in 2009
– Samples began being submitted in November
• 39 participating NAHLN labs
– Approved diagnostic assays
•
•
•
•
Matrix FluA screening PCR
HA and NA subtyping PCR
Virus Isolation – Isolates sent to NVSL repository
Sequencing – priority on HA, NA and M genes
– Sequences deposited to GenBank public database
• Passive Surveillance
– Primary source of samples are from diagnostic samples
from clinically affected pigs, voluntary and anonymous
– Small number of samples sent directly to NVSL associated
with zoonotic events as identified by Public Health: H3N2v
2

3. Systematic approaches to monitor IAV in swine
Genetic Analysis
Sequencing
Antigenic
Analysis
Swine
Studies
• Regional/national efforts – USDA, CEIRS, ESNIP, etc.
• International networks – CEIRS, ESNIP, OFFLU

4. U.S. swine IAV surveillance results
• What does the USDA data tell us ?
– Trends in HA and NA dynamics
• Subtypes: H1N1, H1N2, and H3N2
• Phyloclusters: defined using genetic distance of the HA
– H1s with alpha, beta, gamma, pdm09, and delta (human
seasonal origin)
– H3s with historical Clusters I, II and III, along with a rapidly
expanding contemporary Cluster IV
– Evidence of reassortment
• Matrix Lineage: Classical swine origin or pandemic matrix gene
as an indicator of reassortment and virus evolution.
• Viruses for further research (NADC and others)
• Viruses for vaccine development

5. H3 & H1 Phyloclusters in 2009-2013
200
180
Number of Swine Isolates
160
140
H3N2
120
no data
H1N1 pdm
100
H1N2 delta2
H1N2 delta1
80
H1N1 gamma
H1N1 beta
60
H1N1 alpha
40
20
0
09Q410Q110Q210Q310Q411Q1 11Q2 11Q3 11Q412Q112Q212Q312Q413Q113Q213Q3
Year by quarter

6. Phylogenetic relationships of H3 swine IAV
Phylogenetic clade
Cluster I
Cluster II
Cluster III
Cluster IV
6 clusters evolving
forward into 2013
HA associated with H3N2v
0.02
Between clade amino acid identity ~94%: within clade amino acid identity ~98%
Tavis Anderson, IRV accepted

7. 200
180
160
140
120
100
80
60
40
20
0
09Q4
10Q1
10Q2
10Q3
10Q4
11Q1
11Q2
11Q3
11Q4
12Q1
12Q2
12Q3
12Q4
13Q1
13Q2
13Q3
Number of Swine isolates
Matrix lineage 2009-2012
No data
Year by quarter
North American
Pandemic
• pdm09 Matrix gene currently ~75%
0.02
Tavis Anderson, IRV accepted

8. Reassortment generates new viruses

9. Reassortment patterns of H3N2 virus in U.S. swine
Genotypes associated with H3N2v
(Kitikoon et al., 2013, J Gen Virol)

10. Are the H3 genetic clusters impacting cross-reactivity?
H3 antibodies had inconsistent cross-reactivity among sub-clusters and with vaccines
D
F
TX/2011
IA/2011
IA/2011
IL/2011
IN/2011
IN/2012
NY/2011
IN/2011
PA/2010
IV
I&IV
I
Vaccine 3
C
Vaccine 2
A
IL/2009
MN/2009
(IV)
IA/2007
(I)
IV
MN/2006
TX/1998
Virus (cluster)
TX/1998
MN/2006
IA/2007
MN/2009
IL/2009
PA/2010
IN/2011
NY/2011
IN/2012
IL/2012
MI/2012
MN/2012
MN/2012
MN/2012
IA/2012
IN/2011
IL/2011
IA/2011
IA/2011
IA/2012
IA/2011
TX/2011
IL/2012
IA/2012
NE/2012
I
Vaccine 1
Sera
Homologous
< 4-fold reduction
= 4-8-fold reduction
> 8-fold reduction
(A)
(B)
(C)
(D)
(F)
Pravina Kitikoon, IRV accepted

11. Cause or Consequence?
• What is driving the observed increase in diversity?
– Reassortment?
– Pig movement?
– Ineffective vaccines?
– Unknown factors?
• With all this antigenic diversity, how well are control
measures working on a national level?
• Can we develop a system for more efficient updates of
commercial vaccines?
• What are the implications for human health?

12. Human Infections with variant IAV - USA
Case Count: Detected U.S. Infections with
Variant Influenza Viruses by State since December 2005
Reporting State
Hawaii
Illinois
Indiana
Iowa
Kansas
Maine
Maryland
Michigan
Minnesota
Missouri
Ohio
Pennsylvania
South Dakota
Texas
Utah
West Virginia
Wisconsin
Total
H3N2v
1
5
154
5
1
2
12
8
8
108
16
Total
H1N1v
H1N2v
2011
1
2
3
3
H3N2v
2012
1
4
138
1
2013*
1
14
2
2
2
2
12
6
5
1
4
3
107
11
2
1
1
1
1
5
21
2
2
347
14
5
366
1
3
20
12
309
18
339
*2013 human cases recently report associated with county fair swine exposure
http://www.cdc.gov/flu/swineflu/variant-cases-us.htm

13. H3N2v, 2011-13
• >339 cases in humans primarily associated with children at
swine barns at agricultural fairs in summers of 2011, 2012 and
again in 2013
– All genotype 1 H3N2
– Nearly all HA cluster type A
– Nearly all NA 2002 lineage
– Low serologic cross-reactivity in children and teenage
population
• Many questions regarding recommendations to fair officials,
exhibitors, and exhibited stock
– How to screen stock before entering exhibit
– Recommendations on vaccines for kids and pigs
– Animal movement practices at fairs/venues
– Multi-disciplinary team drafted guidance documents early
2013

14. Epi-Curve for Confirmed H3N2v Cases
State of Isolation
July - Sept 28, 2012 (N=306)
MN (n=4)
40
MD (n=12)
35
PA (n=11)
30
WI (n=20)
25
MI (n=5)
WV (n=3)
20
IL (n=4)
15
OH (n=106)
10
IN (n=138)
HI (n=1)
5
0
7/9
7/16
7/23
7/30
8/6
8/13
8/20
8/27
9/3
Illness Onset Date
(Includes illness onset dates imputed from specimen collection dates and report dates)
http://www.cdc.gov/vaccines/acip/meetings/downloads/slides-oct-2012/02-influenza-Finelli.pdf
9/10

15. Proportion of Persons with Hemagglutination Inhibition
Antibody Titer >40 to Influenza A (H3N2)v Virus
by Year of Birth
NORWAY
CANADA
USA
90
80
Proportion
70
60
50
40
30
20
10
0
2011 2001 1991 1981 1971 1961 1951 1941 1931 1921 1911
Slide courtesy of Dr. Nancy Cox, CDC
Skowronski DM. et al. JID. 2012 August 12; CDC, Unpublished data, 2012; Waalen et al., Euro Surveill 17:pii=20170, 2012

16. Future Challenges and Gaps for Swine Exhibits
• Swine IAVs in a period of frequent reassortment and HA evolution.
– Continued monitoring and timely dissemination of information required.
• Currently available vaccines for swine do not prevent nasal shedding, some
performed better than others.
• There is no uniform system to select swine IAV for vaccine updates.
– Mass vaccinating has been shown to dramatically decrease viral shedding.
– Mixed immune populations comingled together will continue to be
difficult to manage.
• A sensitive and specific pen-side test is not available and clinical signs are not
sufficient for identifying shedding pigs at points of concentration.
• Pig movement and mixing is difficult to track with fair and show pigs.
• There is variability among exhibition venues and markets for animal
movement and show requirements.
• Guidelines & recommendations for swine IAV implemented in a number of
states for the 2013 Fair season.
Partnerships between animal health, human health, and the swine industry
are critical to minimize the impact of these events.

17. Acknowledgments
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NADC
Michelle Harland
Gwen Nordholm
Daniela Rajao
Alessandra Dias
Matt Sandbulte
Brian Campbell
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Kelly Lager
Crystal Loving
Mark Kehrli
Jason Huegel
Jason Crabtree
Tyler Standley
Darrell Bayles
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Pravina Kitikoon
Tavis Anderson
Phil Gauger
Jamie Henningson
Alessio Lorusso
Doug Braucher
Janice Zanella
Eraldo Zanella
Sabrina Swenson
John Korslund
Marie Gramer
Nicola Lewis
Daniel Perez
Martha Nelson
Richard Webby
James Roth
Ruben Donis
Hana Golding
Surender Khurana
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U.S. Producers and
Swine Vets
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Funding:
USDA-ARS
USDA-APHIS
National Pork Board
CDC
Many thanks to
USDA NCAH Visual Services