Tracking the emergence and global spread of foot-and-mouth disease (FMD)

A Symposium to Mark the 50th Anniversary of the Designation of the Pirbright Laboratory of the Institute for Animal Health as FAO World Reference Laboratory for FMD

9:00-9:10 Wellcome Prof Martin Shirley, IAH Director9:10-9:30 WRLFMD overview Dr David Paton, Head of WRL9:30-10:00 Global diversity of FMDV Mr Nick Knowles, IAH

10:00-10:30 Coffee

10:30-11:00 FMD in Africa Dr Wilna Vosloo, Ondestepoort11:00-11:30 FMD viral evolution Dr Esteban Domingo, Madrid11:30-12:00 Tracing FMDV in UK outbreaks Dr Eleanor Cottam, IAH12:00-12:30 FMD virus structure Prof Dave Stuart, Oxford

12:30-13:30 Lunch

13:30-14:00 Antigenic cartography Dr Derek Smith, Cambridge14:00-14:30 Cross-protection between FMD strains Dr Bernd Haas, Riems14:30-15:00 Virus detection methods Dr Donald King, IAH

15:00-15:30 Tea

15:30-16:00 FMD virus transmission dynamics Prof Mark Woolhouse, Edinburgh16:00-16:20 European collaboration on FMD Dr Kris de Clercq, Brussels16:20-16:40 Future prospects for FMD control Dr Keith Sumption, Rome

17:00 Close

Tracing FMDV in UK outbreaks

The rationale behind using complete genome sequencing of Foot-and-Mouth Disease Virus as an epidemiological modelling tool

9 May 2008

E.Cottam@uea.ac.uk

• Rapid replication rate

• Large population sizes

• High error rate of viral polymerase in the order of 10-3 to 10-5 misincorporations per nucleotide copied

FMDV evolution

Fast adaptation and evolution

UK 2001 Outbreak

• 2030 infected premises culled

• 8131 culled as dangerous contacts

• > 6.5 million animals culled

• economic cost exceeding £6 billion

• Pan Asia O serotype• Identified 20th Feb 2001 in

Cheale’s abattoir Essex• Source farm identified as Heddon-

on-the-Wall.• Airborne spread to nearby sheep

farm.• Movement of sheep through

markets one week before disease identified

• Subsequent outbreaks throughout the UK

UK 2001 Outbreak

• The transmission of FMDV is poorly characterised, and information on the direction of farm to farm spread would greatly enhance our understanding and hence control.

• Broad scale genetic tracing of FMDV using VP1 sequences (633nt) between countries

• Attempt in Denmark at intra-epidemic tracing of FMDV using VP1 sequences only*

• Forensic genetic tracing for HIV, Hep C, SARS, Rhinovirus, Norovirus…….

*Christensen, L.S., P. Normann, S. Thykier-Nielsen, J. H. Sorensen, K. de Stricker, and S. Rosenorn. 2005. Analysis of the epidemiological dynamics during the 1982-1983 epidemic of foot-and-mouth disease in Denmark based on molecular high-resolution strain identification. J. Gen. Virol. 86: 2577-2584

Previous work

• Development of method for sequencing complete genome

• Investigation of FMDV evolution during the UK 2001 Outbreak

• Use of complete genome sequence data for forensic genetics of FMDV transmission

Project Introduction

• Direct from epithelium

• RT-PCR– 5 overlapping fragments– 84 sequencing reactions

• Use of QIAGEN robot

• Complete sequence within 3 days

Complete genome sequencing

Cottam et al. 2006 J Virol 80(22), 11274-82

C 7038-2001 FAS

C2 7039-2001 FAS

O 5681-2001

M 9443-2001

D 9161A FAS

D 9161B

B 4141-2001

E 7299-2001 FAS

L 4998-2001

K 4014-2001

F 7675-2001 FAS

G 9327-2001 FAS

I 9788-2001 FAS

J 9964-2001 FAS

Darlington Cluster

9011-2001 FAC

14476-2001 FASWest Yorkshire

Cumbria14391-2001 FAS

14603-2001 FAS

15101-2001 FAS

14524-2001 FAS

Northumberland

Powys14339-2001 FAS

Staffordshire621-2001 FAS

Northampton220-2001 FAS

Northumberland4569-2001 FAS

Devon173-2001 FAS

Ireland438-2001 FAS

Devon11676-2001 FAS

A 3952-2001 FAS

N 5470-2001 Darlington 2Cheales Abbattoir11-2201 FAS

Ponteland150-2001 FAS

126-2001 FAS

127-2001 FAS

128-2001 FASHeddon on the Wall

100

100

100

92

99

91

99

99

99

98

92

66

89

72

86

64

83

87

87

61 Genetic relationship of34 virus sequences

Maximum likelihood phylogenetic tree10,000 Bootstrap

• 52 nucleotide substitutions between source and final case– 8 were amino acid changes

• 83 variant nucleotides between Wales and Northumberland

Observed Variation

Cottam et al 2006 J Virol 80(22), 11274-82

Molecular clock

Nucleotide changes accrue linearly with time and are inherited

2.26 x 10-5 / site / day

0

10

20

30

40

50

60

0 50 100 150 200 250Day of Outbreak

Nuc

leot

ide

subs

titut

ions

from

inde

x ca

se x

Cottam et al 2006 J Virol 80(22), 11274-82

Early animal movement transmission events between 5 farms.

7088

7109

8091

Farm 223/02/01

7088

7109

4382

5277

8107

3428

4808

Farm 426/02/01

7088

7109

4382

5277

8107

3428

4808

4553

4982

5303

Farm 501/03/01

Farm 324/02/01

7088

7109

4382

5277

8107

2420

25/02/01 28/02/0127/02/01

7088

Farm 122/02/01

time

Known chain of events

4 5

2

3

1

1

Cottam et al 2006 J Virol 80(22), 11274-82

0 9 184.5 Kilometers

Pos + sequenced

Pos not sequenced

Negative

No samples submitted for diagnosis

A

N

O

F

GI

P

ECB

L

DM

J

K

Durham cluster of Infected Premises

Cottam et al. 2008 Proc Biol Sci. 22;275(1637):887-95.

45

2

3

C1

C2

D

F

G

I

JE

B

A1b1a

k

L

M

N

O

P

Difficult to determine transmission tree –839808 different trees consistent with genetic data

Statistical parsimony analysis of sequence data from IPs by TCS

Cottam et al. 2008 Proc Biol Sci. 22;275(1637):887-95.

0 10 20 30 40 50 60 70 80 90 100 110

Day of outbreak

A

B

C

D

M

E

N

G I J

F

K

L

1

2

3

54 O

P

• Restricted by date of cull

• Individual farm infection profiles?

• Statistical weighting for transmission tree – how confident are we?

Cottam et al. 2008 Proc Biol Sci. 22;275(1637):887-95.

Start of outbreak

*Most likely infection date

(mode)

2 dIi(t) probability that i th farm was infected at time t (discrete beta-distribution)

Day of cull

*date of confirmation minus oldest lesion minus 5d incubation

∑ ∑=

−

=⎟⎟⎠

⎞⎜⎜⎝

⎛⎟⎟⎠

⎞⎜⎜⎝

⎛⋅=

iC

j

jt

kii kLjItF

0 1)()()(

Probability that the ith farm is a source of infection at time t :

5d

L(k) probability of incubation for K days prior to becoming infectious, gamma-distribution, 95% probability between 2 and 12 days

2d 12d

Cottam et al. 2008 Proc Biol Sci. 22;275(1637):887-95.

Start of outbreak

*Most likely infection date

(mode)

2 dIi(t) probability that ith farm infected at time t (discrete beta-distribution)

Fi(t) probability that the ith farm is a source of infection at time t

Day of cull

AB

C

D

E

F

KLNO

GMIJ

AB

C

D

E

F

KLNO

GMIJ

TIME

Likelihood of infection

Likelihood of being infectious

Infection profiles of farms

Cottam et al. 2008 Proc Biol Sci. 22;275(1637):887-95.

∑∑∑≠= ==

⋅⋅=n

ikk

k

C

tij

C

tiij tFtItFtI

kj

1 00)()()()(l

Likelihood that farm i infected farm j :

From these infection profiles we can calculate a likelihood for each hypothesised transmission event

It is now possible to determine the transmission tree with the highest likelihood from all the trees consistent with the genetic data

Cottam et al. 2008 Proc Biol Sci. 22;275(1637):887-95.

Most likely tree

Cottam et al. 2008 Proc Biol Sci. 22;275(1637):887-95.

Num

ber o

f con

secu

tive

farm

infe

ctio

ns

0

1

2

3

4

5

1 2 3 4 5 6 7 8 9 10

Number of variant nucleotides between consecutive farm infections

Mean 4.2 (SD 2.1) nucleotide changes per transmssion event

Thus if the number of changes seen is outside this distribution, a missing intermediate farm would be suspected

Distribution of the number of changes that occur upon transmission of virus between farms (n=16)

Cottam et al. 2008 Proc Biol Sci. 22;275(1637):887-95.

0 10 20 30 40 50 60 70 80 90 100 110

Day of outbreak

A

B

C

D

M

E

N

G I J

F

K

L1

2

3

54 O

P (IP1515)

IP14

Rate of nucleotide change seems to vary…….

2.26 x 10-5 / site / day

0

10

20

30

40

50

60

0 50 100 150 200 250

Day of Outbreak

Nuc

leot

ide

subs

titut

ions

from

inde

x ca

se x

• Previously linked mutation rate with time

• Was the rate of genetic evolution of FMDV in the epidemic due in part to the rate of transmission and spread of the virus?

Bottleneck theory

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Genetic evolution of FMDV

2007 UK FMDV Outbreaks

Cottam et al. 2008 PLoS Pathogens 4(4): e1000050

2007 UK FMDV Outbreaks

IAH2

IAH1

AY593815

IP2b

IP2c

IP1b(1)

IP1b(2)MAH

Synonymous substitution

Non-synonymous substitution

Non-synonymous substitution important in cell culture adaptation

August

Cottam et al. 2008 PLoS Pathogens 4(4): e1000050

IAH2

IAH1

AY593815

IP2b

IP2c

IP3bIP1b(1)

IP1b(2)MAH

SeptemberAugust

Cottam et al. 2008 PLoS Pathogens 4(4): e1000050

9 substitutions

Date

IP2b

IP2c

IP3b

IP1b(2)

IP1b(1)

29-J

ul-0

7

19-A

ug-0

7

12-A

ug-0

7

05-A

ug-0

7

26-A

ug-0

7

02-S

ep-0

7

09-S

ep-0

7

16-S

ep-0

7

23-S

ep-0

7

22-J

ul-0

7

15-J

ul-0

7

Lesionspresent

Infection window

IAH2

IAH1

AY593815

IP2bIP2c

IP3c

IP4b

IP3b

IP5

IP6b

IP1b(1)

IP1b(2)

IP7

MAHIP8

29-J

ul-0

7

19-A

ug-0

7

12-A

ug-0

7

05-A

ug-0

7

26-A

ug-0

7

02-S

ep-0

7

09-S

ep-0

7

16-S

ep-0

7

23-S

ep-0

7

Date

IP2b

IP2c

IP3c

IP4b

IP3b

IP5

IP1b(2)IP1b(1)

IP6b

IP7

22-J

ul-0

7

15-J

ul-0

7

30-S

ep-0

7

IP8

Cottam et al. 2008 PLoS Pathogens 4(4): e1000050

Summary• Complete genome sequencing data can assist in

our understanding of the spread of FMDV

• Combining genetic and epidemiological data gives greater resolution.

• More research is needed to help refine the use of these data in real time.

Don KingDavid PatonNick KnowlesNigel FerrisGeoff HutchingsAndrew ShawJemma WadsworthJohn Gloster

Thank you

Dan HaydonGael Thébaud

John Wilesmith

Sam Mansley