Selection of and Updating

Avian Influenza Vaccine Seed

Strains to Maintain Vaccine

Efficacy in the Face of Field

Virus Drift

David E. Swayne

OIE Collaborating Centre for Research On

Emerging Avian Diseases, FAO Reference Centre

for Avian Influenza, and Exotic & Emerging

Avian Viral Diseases Research Unit

SEPRL, ARS, USDA, Athens, Georgia, USA

Vaccine/Vaccination Terminology

• Efficacy: vaccine passes a lab challenge test

• Effectiveness: vaccine provides protection under field

conditions (implies proper application & coverage)

• Potency: adequate amount of antigen (HA) to

provide a strong immune response in field

– Direct (Ag [HA] content, e.g. 0.3-7.8 μg or 512 HA

units/dose) or indirect assessment (HI titers in immunized

poultry, e.g. ≥ 1:32)

– Determine mean protective dose (PD50); e.g. minimal 50

PD50 per dose (50 PD50)

• Surveillance: looking for H5 virus in field

• Monitoring: looking at vaccinated flocks for

serotiters indicating protection and coverage

Historically - Broad cross protection within a

hemagglutinin subtype; i.e. H5N2, H5N9, etc.

protects against H5N1 HPAI viruses (inactivated

AI and live recombinant vaccines)

Experimental protection studies support the claim

Field success in using existing H5 vaccines in H5N1

HPAI control in Asia: e.g. Mexican, USA and

European H5 vaccine strains in Asia

Introduction

• Historical H5 Vaccines –

Similar antigenicity: wild birds

viruses and initial AI outbreak

viruses in poultry (Swayne et al., Av Dis

45:355-365, 2001)

• As the virus lineage becomes

enzootic in gallinaceous

poultry, the HA drifts away

from root

(A/ck/WIJ-PWT/06)

(A/ck/SMI-HAMD/06)

HA Subtype Cross Protection

Antigenic Cartography

(Updated from Swayne et al., Vaccine 18:1088-1095. 2000)

* challenge viruses

Homosubtypic HA Protection by AI Vaccines

• Fowl pox with H5

AIV gene insert

• Different challenge

viruses (87.3-100% aa

sequence similarity)

TK/Ireland/1378/83

Dk/Ireland/113/83

CK/Scotland/59

Tern/South Africa/61

Peking Duck/Malaysia/645/97

TK/England/50-92/91

CK/Hong Kong/220/97

Hong Kong/156/97

Whooper Swan/Mongolia/244/2005

CK/Korea/ES/03

CK/Vietnam/2004

CK/Italy/1485/97

CK/PA/13609/93

Emu/Tx/39442/93

Tk/Ontario/7732/66

CK/PA/1370/83

TK/WI/68

Dk/Michigan/80

Ruddyturnstone/NJ/2242/00

Ck/Queretaro/7653-20/95 5 changes

N. Amer.

Eurasian * *

*

* *

*

* *

*

*

*

Chickens vaccinated SQ 1d with rFP-H5-Ire/83 and IN challenged at 3

wks with 105-6 EID50 of HPAIV

Decreasing AIV Susceptibility:

Biotechnology to Solve Genetic Drift

Ck/Scotland/59 H5N1 92 0/10

Tern/S. Afr/61 H5N3 93.1 0/10

Tk/Ontario/66 H5N9 89.1 0/10

Ck/PA/83 H5N2 87.3 0/10

Tk/Ireland/83 H5N8 100 0/10

Tk/England/91 H5N1 94.2 0/10

Ck/Queretaro/95 H5N2 89.3 0/10

HK/156/97 H5N1 90.2 0/10

Ck/S. Korea/03 H5N1 89.9 0/10

ck/Vietnam/04 H5N1 88.4 est 0/22

WS/Mongolia/05 H5N1 89.7 1/8

Control - - 80-100%

SubtypeChallenge Virus

HA

Similarity

to Tk/Ire/83

Mortality

(MDT)

• Excellent protection

from mortality using

diverse H5 AIV

• Reduction in virus

replication and

shedding from

respiratory and

digestive tracts

(Swayne et al., Vaccine 18:1088-1095. 2000)

• Higher HA identify between vax and challenge AIV

• Respiratory shedding – significant reduction

• Alimentary shedding – no association in reduction

(Swayne et al., Vaccine 18:1088-1095. 2000)

Oropharangeal Swabs - Peak Shedding

1 1.5 2 2.5 3 3.5 4 4.5

Cloacal Swabs - Peak Shedding

1 1.5 2 2.5 3 3.5 4 4.5

rs = 0.783, P = 0.009 rs = -0.100, P = 0.780

Reduction in Titer (Log 10)

% H

ema

gglu

tin

in I

den

tity

Homosubtypic HA Protection by AI Vaccines

rFP-H5 vaccines, pre-2005 challenge HPAI viruses

• H5 viruses are diverse – N.

Amer. and Eurasian lineages

• H5N1 HPAI is not a single

virus, but a family of viruses

with 10 lineages and many

sublineages

• Virus is drifting as other

influenza A viruses have

done –from natural

infections and “vaccine

pressure”

Changing Genetic

Diversity

WHO WHO

Antigenic Diversity of H5N1 Avian

Influenza Virus: AgC-H5 (Ferret Serum)

Source: Ron Fouchier

Antigenic Diversity of H5N1 Avian Influenza

Virus: AgC-H5 (Chicken Serum)

Source: Erica Spackman

• Historical H5 Vaccines –

Similar antigenicity

• Drifting of HA away from

root

• Good protection:

Ck/HK/220/97, Ck/Legok/03,

VN/1203/04, Ck/WJ/HAMD/06

• Intermediate protection:

Ck/Papua/06

• Poor protection: PWT/06 (A/ck/WIJ-PWT/06)

(A/ck/SMI-HAMD/06)

G

I

P

Impact of Drift on Protection

Antigenic Cartography

Other Examples of Antigenic Drift

• Egypt (2006-): Field HPAIV resistance to immunity from

Mexico/94 and Re-1 vaccines (Cattoli et al, J.Virol. 85:8718-8724, 2011

and Grund et al., Vaccine 29:5567-5573, 2011)

• China (2004-) (Chen et al., OIE Rev Sci Tech 28:267-274, 2009):

– Tk/England/1973 [H5N2]: (2004-6)

– Re-1 (rg A/gs/Guangdong/1/1996 [H5N1] (0): 2004-8

– Re-4 (rgA/ck/Shanxi/2006 [H5N1](7): 2006-7

– Re-5 (rgA/dk/Anhui/1/2006 [H5N1](2.3.4): 2008-12

– Re-6 (rgA/dk/Guangdong/S1322/2010 [H5N1] (2.3.2): 2012-

• Vietnam: 2011 2.3.2.1B resistant to immunity from Re-1 &

Re-5 (now use Re-6) (Cha et al. Vaccine 31:4953-4960, 2013)

• Hong Kong (2008): clade 2.3.4 on 1 farm (Leung et al., Vaccine

31:3536-3542, 2013)

– Mexico (LPAI): North American H5N2 (Lee et al.,

J.Virol. 78:8372-8381, 2004; Eggert et al., Vaccine 28:4609-4615,

2010 )

Antigenic drift must be continually addressed &

vaccine seed strains updated; future will require

reverse genetic LPAI seed strains based on

hemagglutinin of recent field HPAIV

How do you update?

Ongoing and adequate surveillance and monitoring in

vaccinated and non-vaccinated populations

• Passive surveillance for mortality events in all vaccinated

flocks and subset of non-vaccinated flocks – VI and

sequencing of isolates

• Active surveillance in subset of vaccinated flocks with normal

mortality profile – VI on deads as routine with sequencing of

isolates

• Ongoing genetic analysis of all H5 viruses (phylogeny)

• Subset of genetically diverse isolates are HI tested &

analyzed for antigenic divergence (cartography)

• Challenge testing of subset of antigenic divergent viruses

• Proposed seed strain change based on frequency of

antigenic variant viruses in field

What Does This Mean?

Global marketing of a single H5 vaccine seed strain does not

always meet the local (national/regional) needs for efficacy;

i.e. science and not marketing should prevail

Solutions:

• Best: Updating vaccine seed strains to match antigenic drift in

field viruses (reverse genetic seeds to match field virus)

• Adjunct: Increase antigen mass in inactivated vaccine – HA

protein quantity (HA units or 50 PD50) (Source: CLEVB Egypt, challenge

tests for vaccine batch release)

• Adjunct: Multiple vaccinations to enhance immunity in field – use

of heterologous H5 vaccine seeds (bivalent or multiple

heterogenous monovalent)

o Prime-Boost usage

o More use of live recombinants in hatchery

• Adjunct: Adjuvants to enhance immune response (current

[proprietary] & future [oils, surfactants & cytokines])

Increasing Antigen Content

Current National Minimum Standard: 1:40 (1:32)

hemagglutination inhibition (HI) titer (SPF WL lab

conditions): Mortality protection

• HI serology titers of GMT ≥ 8 (Swayne et al., Av Pathol 28:245-255, 1999) or

≥10 (Kumar et al., Av Dis 51:481-483, 2007) were associated with survival

when had antigenically similar seed and challenge viruses

• HI serology titers of GMT ≥ 40 (Kumar et al., Av Dis 51:481-483, 2007) were

associated with prevention of challenge virus shedding from the

oropharynx in the majority of vaccinated chickens and GMT ≥ 128

(Swayne, et al., Av Path 35:141-146, 2006) were associated with preventing

challenge virus shedding from the oropharynx in all vaccinated

chickens

• Newer Data: HI serology titers (challenge virus antigen) of GMT ≥

32 associated with survival and GMT ≥ 64 associated with

preventing respiratory replication

Increasing Antigen Content

Proposed National Minimum Standard: GMT ≥ 120

(128) HI titer as optimal (using vaccine seed as HI

test antigen)

If HI test using challenge or antigenic drift variants

produce mean titers of GMT ≤ 32, vaccine seed

change should be further examined

– Challenge studies to assess protection

– Examine other field viruses in HI tests to determine how

common the antigenic variant viruses are in the field

Other Related Issues

Pre-develop logistics for vaccination and exercise

vaccination plan for emergency or ‘routine’ usage

Vaccine availability: commercially or vaccine bank within

the country

• Completed vaccine licensing/registration process, or have

emergency authority

• Stockpiles available in country through manufacturing capacity

or importation to meet need

License issues:

• Specific seed strain for each license submission

• Replacing seed strain requires additional data and varies with

each country: safety to full efficacy and potency

• Cassette concept – seed strain replacement with safety

and seropotency data

Conclusions

• Genetic and antigenic variant HPAI/LPAI field viruses,

resistant to licensed H5 AI vaccines, have been

identified in Mexico, Indonesia, Egypt, Vietnam, Hong

Kong and China

• Updating vaccine seed strains is needed to assure the

most efficacious and effective vaccines are used in the

field

• More rg inactivated seed strains and recombinant

vaccines will be needed for antigenic matching

• Need more rapid means of licensing updates in vaccine

seed strains through national regulatory processes

Conclusions

• Increasing antigen content in vaccines, using multi-

vaccinations with antigenically diverse seeds, or single

vaccination with bivalent vaccines (with antigenically

diverse seeds) can compensate for some antigenic

diversity of field viruses, but will not replace the

periodic need to update seed strains

• Need to update the minimum potency of all H5 and H7

AI vaccines

Merci Beaucoup!

Thanks: OIE: K. Hamilton, G.

Pavade. B. Vallat,

FAO: G. Dauphine, J.

McGrane, J Lubroth, M.

Kim

SEPRL: D. Suarez, E.

Spackman, Mary Jackwood,

D. Kapczynski

Others: T. Prajitno, D.

Smith, R. Fouchier