Controlling FMD and PPR

by vaccination

Satya Parida

The Pirbright Institute

Oxford, University

BBSRC institute

~380 science staff

+ research students

+ visiting scientists

The Pirbright Institute

. Virus biology, gene functions, evolution

Virus-host interactions in

infection models in natural hosts

Immune responses to

virus infections & vaccines

Our Research

Role of arthropod vectors in virus transmission

Diagnostics, disease surveillance, mathematical

modelling

. • Reference laboratories

• New & improved diagnostic tests

• Disease preparedness and emergencies

• Strong international links

• Training

Surveillance, early detection and characterisation is

crucial for emerging and re-emerging viruses

Highly contagious disease Causative agent:+ve strand RNA virus Spread- Direct contact and airborne route Persistent infection: in ruminants Due to wide antigenic variation of virus, disease is difficult to control Shorter duration of immunity (~3-4M)

Foot-and-mouth disease

Endemic pools

• Maintain specific FMD virus strains • Distribution of FMDV serotypes in the endemic pools is not

equal • Control via (tailored) vaccination and supporting diagnostics

• In addition to circulation of local strains, long-distance “trans-

pool” movements of FMDV are frequently observed

Conjectured global status

Challenges for FMD control in endemic settings through vaccination

Killed inactivated oil adjuvanted vaccines are in use. Biannual vaccinations (3 PD50 vaccines) are practised to control the disease in endemic settings One of the major challenges- short lived immunity (~3 to 4 months max) which allows at least 2 month window for bringing back the disease before 2nd vaccination. Main aim: To increase the duration of immunity, at least to cover the window between biannual vaccination

VNT and IFN-γ responses in serotype A and SAT2 vaccinated cattle on 21dpv

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100

150

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full dose 1/4 dose 1/16 dose control

IFN

-γ (n

g/

ml)

VN

T t

itre

A Malaysia 97 (VNT) SAT2 Eritrea (VNT)

A Malaysia 97 (IFN-γ) SAT2 Eritrea (IFN-γ)

Oh et al 2012

.

How does the new generation

adjuvants help?

• Activate the innate immune system via ligand binding to PRR

• Enhance the anti-

viral environment • Enhance the

adaptive response (CMI and humoral) • Improve memory

responses

Final screening of potent adjuvants using Type A antigen (sub-optimal dose) in cattle

Non-Vaccinates control

A22 Iraq+ ISA-206 control

A22 Iraq+ ISA-206 + poly (I:C) adjuvant

A22 Iraq+ ISA-206 + AbISCO adjuvant

A22 Iraq+ ISA-206 + R848 adjuvant

A22 Iraq+ ISA-206 + MPLA adjuvant

A22 Iraq+ ISA-206 + R848+ MPLA adjuvant

Serotype A Vaccine Trial Design

0

A22 Iraq

Vaccination

(sub-optimal dose)

7 14 21 28

A22 Iraq

Challenge Cull

30 14 0 7 21 28

Sampling

Daily Rectal Temp

Swabs

Clotted blood

Heparinized blood

FACS analysis- IFN-γ from CD4+ and

CD8+ cells

Poly (I:C) and AbISCO had significantly

increased neutralizing antibodies on 28dpv

compared to the ISA-206 control group

• Achieved an increased potency as indicated by

lack of clinical symptoms in Poly I:C and AbISCO groups (although not able to prevent sub-clinical infection)

• Significantly elevated neutralizing antibodies in Abisco and Poly I:C group cattle

• Some indication of IFN-g upregulation by CD4+ and

CD8+ cells both in Poly I:C and AbISCO groups

Summary-1

ISA 206 + Poly I:C 11 cattle

ISA 206 ( conventional) 11 cattle

Type O antigen full dose- 10 microgram

Vaccinated cattle were monitored for 225 days (7.5 months)

Serum, PBMC and antigen specfic stimulated whole blood plasma were analysed at Pirbright

Longer duration of immunity study

Virus neutralisation titres

• Titres significantly higher in cattle when vaccinated with poly

I:C (blue) compared with conventional adjuvant (red)

Whole blood Interferon-γ

• Significantly higher levels in cattle when vaccinated with poly

I:C (blue) compared with conventional adjuvant (red)

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Gr-1 Conv. vaccine Gr-2 Poly:IC vaccine

Mean-CD4-IFNg+

Mean-CD8-IFNg+

IFN-γ production -flow cytometry on 120 dpv

% I

FN

-γ+

cell

s

% of CD4 or CD8 cells from the total live population showing

antigen specific IFNγ expression

• Significantly higher VN titre in cattle vaccinated with poly I:C compared with conventional adjuvant

• By 4th month many of the conventional vaccinated animals lost the protective level of antibody titre

• IFNγ levels significantly higher in cattle when vaccinated with poly I:C when compared with conventional adjuvant

• Indication of IFN-gamma upregulation

by CD4+ and CD8+ cells were observed in Poly I:C group in FACS analysis.

• Duration of immunity can be enhanced up to 6 months post-vaccination that covers the window of susceptibility before 2nd vaccination

• Boosting after first vaccination?

Summary-2

Acknowledgements

The Pirbright campus is being redeveloped

Katie Lloyds-Jones Mana Mahapatra Krupali Parekh Katy Moffat Becky Herbert Aravindh Babu Simon Gubbins David Paton Geraldine Taylor

Funding:

BBSRC

DFID/UKAIDS

Oxford, University

Indian Immunological

Dr K Anandkumar Dr V.A Srinivasan Dr S B NagendraKumar Dr M Madhanmohan Dr R Lingala

Satya Parida

Vaccine Differentiation Group, The Pirbright Institute

Oxford, University

PPR Vaccines that Differentiate Infection from Vaccinated

Animals (DIVA)

Highly contagious and infectious viral disease of small ruminants – Domestic: Goat, Sheep – Wild life: Gazelle and Capra sp.

Taxonomy – Order Mononegavirales

– Family Paramyxoviridae

– Genus Morbillivirus (PPRV, RPV, MeV, CDV)

Huge burden on the developing world where it prevents the development of sustainable agriculture

Peste des petits ruminants (PPR)

Susceptible hosts 2: Wildlife

Documented disease in captive wild ungulates: Dorcas gazelle (Gazelle dorcas), Thomson's gazelles (Gazella thomsoni) Nubian ibex (Capra ibex nubiana) Laristan sheep (Ovis gmelini laristanica) Gemsbok (Oryx gazella)

Susceptible hosts (Continued)

Wild Goats Capra aegagrus aegagrus

Saiga antelope- Euroasia to Mongolia

• The outbreak of PPRV in Kenya in 2006 caused severe socioeconomic consequences for food security

• 2006 to 2008 more than 5 million animals across 16 Kenyan districts were affected (>50% mortality rates)

• Annual loss attributed to PPR in Kenya > 1 billion Kenyan shillings (US $15 million, £10.5 million). •

Impact of PPRV in livestock, Kenya- 2006

• PPR Eradication- 2030 • Global and regional strategies and road maps be developed and monitored (notably regional approaches for surveillance and vaccination). • Understanding of disease epidemiology/ecology in the context of the socio-economic and farming systems dimensions for targeted intervention be improved;

• Preventive measures such as vaccination be supported: very efficient live attenuated vaccine, providing protection for the life of all small ruminants- but can not work as DIVA . • Combine with other campaigns to improve small ruminant flock health and disease prevention and thus maximise available resources.

FAO/OIE Strategy

Live attenuated vaccines- provides sterile immunity

Differentiate between naturally Infected and Vaccinated Animals (DIVA) vaccine: not possible Marker vaccine associated with a diagnostic test (mainly serological) allows serological differentiation of vaccinated animals from the naturally recovered animals.

Existing PPR vaccines – Nigeria 75/1

and Sungri 96

Design for PPRV Sungri 96 full length clone

N, P and L protein

virus genome as RNP

N P

L

Fowl pox /T7

Cytoplasmic T7 transcription (+ve) 5' 3'

N, P and L mRNA

Encapsidation

virus N, P and L dependent replication

5' (-ve) 3'

cDNA

T7

VDS cells

PPRV Sungri96

PPRV Sungri 96

Parida et al Patent application filed

Manipulation of full-length DNA by Reverse genetics to generate virus

Recovery of Sungri 96 PPR DIVA vaccine

viruses from DNA

RT-PCR

Sequencing

Confirmation of identity CPE following transfection

Growth characteristic: same as parent

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Hours post infection

Lo

g10 (

TC

ID50/m

l)

Vaccine-challenge experiment

PPR-DIVA PPR Control

Challenge (i/n): 28 dpv

Clinical signs: 14 dpc Temperature: daily Samples: daily/alternate day

Samples: Blood, nasal, occular and saliva swabs

Following vaccination (s/c):

No rise in body temp. No leucopenia

Temp. following challenge

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Re

cta

l te

mp

era

ture

(°C

)

Days post challenge

PPR-DIVA PPR Control

WBC count following challenge

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Days post challenge

PPR-DIVA PPR Control

WB

C (

in 1

03)/

mm

3 o

f b

loo

d

4

16

8

12

Genome detection in nasal

secretions following challenge

Similar results for occular and salivary secretions

Ct-

valu

es

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0 dpc 2 dpc 4 dpc 5 dpc 6 dpc 7 dpc 8 dpc 9 dpc 10 dpc 12 dpc

Control

PPR-DIVA

PPR

Neutralising antibody on 28 dpv

VN

tit

re (

log

10)

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PPR-DIVA PPR

Antibody against N-protein (IDVET)

on 28 dpv P

I valu

es

PI values < 50% are positive

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25

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75

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0 dpv 7 dpv 14 dpv 21 dpv 28 dpv

PPR-DIVA

PPR

OD

valu

es

DIVA test DIVA vaccinated

animals

Conventional /

infected animals

PPRV antigen - +

Proprietary test

antigen

+ -

Antibody detection by DIVA tests

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PPRV antigen Proprietary test antigen

PPR-DIVA

PPR

Control

• DIVA vaccines and DIVA tests are in place

• Growth and Production of DIVA vaccines are same as existing vaccines

• Tested in group of fives goats in pilot studies for safety, stability and potency and much comparable to existing vaccines

• IP has been protected-patent application has been filed

• Received interest from vaccine industries

• Pirbright decided to provide as many as industries with non-exclusive agreement

Conclusions

Vaccine differentiation group (Pirbright)

• Muni Selvaraj • Mana Mahapatra

TANUVAS

• Dhinakar Raj • KG Thirumurugan

IVRI

• R P Singh

• Muthu Chelvan Royal Veterinary College Animal Isolation Units (Pirbright)

• Richard A Kock All the staffs

Acknowledgement

Acknowledgements

The Pirbright campus is being redeveloped

Oxford, University

Big thanks to BIOVET,

Bangalore for sponsoring my

visit to IVA meeting

Pirbright Institute