FMD-NSP Epizone reportEU FMD-NSP workshop 2 Contents Page Abstract 3 EU FMD-NSP workshop organising...

EU FMD-NSP workshop 1

EPIZONE, EUFMD (FAO) &

EU COORDINATION ACTION FMD-CSF

Workshop on the design and interpretation of post Foot-and-Mouth Disease (FMD)-vaccination

serosurveillance by NSP tests Part I (dense cattle-pig countries)

January 23-25, 2007

Report

VAR-CODA-CERVA Leuvensesteenweg 17

B-3080 Tervuren Belgium

DG RESEARCH

DG SANCO

EUFMD


Contents Page Abstract 3 EU FMD-NSP workshop organising committee 6 EU FMD-NSP workshop supervisory team 6 Participants 6 Observers 6 Sponsors 6 Objectives 6 Report 7

Opening session 8 Session I: Review on FMD vaccination and NSP testing 8 Session II and III: Results from working groups 25 Session IV: Discussion of results and models for serosurveillance – Summary, Conclusions, Recommendations and Observations

41

Acknowledgments 49 Annex A: FMD-NSP workshop program 50 Annex B: presentation of scenario 1 by group 1 51 Annex C: presentation of scenario 1 by group 2 59 Annex D: presentation of scenario 2 by group 3 65 Annex E: presentation of scenario 2 by group 4 68 Annex F: presentation of scenario 3 by group 5 76 Annex G: presentation of scenario 3 by group 6 83 Annex H: Post NSP-workshop thoughts – Dr. David Paton 92 Annex I: Literature 94


Abstract Given the current FMD-free status without vaccination in Europe and the possibility of a future outbreak with vaccinate-to-live used as an emergency measure, followed-up by a post-vaccination serosurveillance, to return to the status ‘free from infection without vaccination’, this workshop, for countries with area’s of dense animal populations, concentrated on:

1. The design and implementation of a survey to substantiate free from infection with a certain degree of confidence after vaccination has been performed.

2. The guidance to the interpretation on the follow-up on seropositive animals/herds/flocks;

3. The guidance to the use of laboratory test results in decision-making; 4. The identification of the resources (laboratory, veterinarians) required.

Three different exercise scenarios were written, including

(a) one scenario with a limited number of outbreaks where only bovine were vaccinated,

(b) a second scenario with a limited number of outbreaks where all animals (bovine, pigs, sheep and goats) were vaccinated, and

(c) a third scenario with a large number of outbreaks where only bovine were vaccinated.

Twelve invited countries, including Spain, Ireland, the United Kingdom, Germany, the Netherlands, Belgium, France, Italy, Slovenia, Denmark, Portugal and Lithuania (of which Portugal did not participate), and observers of OIE, EUFMD and EC were divided in 6 groups, each group having to make the best possible survey design (which was not necessarily the design proposed by OIE Guidelines or EU Directives), to follow-up the seropositive herds/animals/flocks and to do an identification of the necessary resources for their own scenario. Summary of workshop conclusions: Despite the use of different working groups/countries, the approaches taken showed a clear degree of similarity. Summary of conclusions:

1. The vaccination-to-live policy with subsequently substantiating freedom from infection by a survey system including NSP testing is a realistic and achievable option in FMD control. However, stamping out will always remain part of the control policy;

2. Because NSP assays are not sensitive enough (especially for carriers), conclusions on the infection status of the vaccinated herds can only be based on a combination of clinical and serological surveys and epidemiological investigations such as cluster analysis;

3. Proving freedom from infection for vaccinated animals is impossible, in contrast to substantiating freedom from virus circulation or freedom from infection in non-vaccinated animals;


4. The current EU Directive (2003/85/EC) for the control of FMD mentions two surveys, i.e.: (a) A survey for detecting the presence of FMD virus (Article 56) in the

vaccination zone, which should be a combination of clinical, epidemiological and serological investigations with high overall system sensitivity, which includes: • a survey of the non-vaccinated animals; • a serosurveillance of all herds with vaccinated animals (NSP tests). Within

herds sampling of all vaccinated ruminants and their non-vaccinated off-spring; For large numbers of other species, sampling should be based on a 5% prevalence with 95% confidence.

(b) A survey, to regain freedom from infection after emergency vaccination(Article 61), which must have a high specificity. This survey might include a second serosurveillance, but the first serosurvey could serve the purpose of this one (cf. OIE Guidelines in App. 3.8.7. of the TAHC);

5. The follow-up of herds with seroreactors by serological investigation has to be based on NSP assays (Paton et al., 2006) with well-defined performance characteristics.

6. If specificity of the serological test system were known, only seroreactor rates above the Herd Cut Point could be considered, but this is not compatible with EU directive 2003/85/EC.

7. A clinical surveillance combined with paired serology can detect holdings where virus circulation is ongoing. Since there is no possibility of detecting each and every carrier within sub-clinically infected herds, all ruminants should be tested. Evidence of virus circulation would lead to herd slaughter, but evidence of carriers would lead to slaughter of these reactor animals only;

8. Testing all animals in the vaccinated population as prescribed in the EU Directive 2003/85/EC, is considered as not achievable in areas with a dense pig population or within big pig herds, if such pigs have been vaccinated.

9. Vaccination of small herds remains a controversial item. Two possible options were discussed: (a) a non-vaccination policy for small herds with integration in the survey system

as sentinels or (b) a vaccination policy for small herds because of their contribution in achieving

the necessary level for population protection and because of political reasons. 10. This kind of workshop should be done for other veterinary diseases, like CSF and

AI also (for all EU members, EUFMD countries and EU neighbours).

Summary of recommendations: 1. Conclusions on the infection status of the herds after FMD outbreaks in a vaccinated

population should only be based on a survey system, including at least clinical, serological and epidemiological investigations;

2. The performance characteristics of the survey system should be determined; 3. The term ‘demonstrate absence’ should be replaced by ‘substantiate absence’; 4. Contingency plans should include a clear flow chart for the follow-up of seropositive

herds (minimum requirements of App. 3.8.7. of TAHC)


5. All large ruminants should be tested to substantiate freedom from infection in a vaccinated population after FMD outbreaks. While evidence of virus circulation must lead to the declaration of an outbreak, consensus should be sought on the slaughter of reactor animals only, in case it is evident that these animals are carriers;

6. A change in the definition of an outbreak in OIE guidelines and EU Directives is needed where carriers are concerned;

7. The relative confidence attainable with “herd-based” and “individual” certification needs to be explored for different herd sizes and prevalence;

8. Consideration should be given to an amendment of the Directive in order to allow a within-herd sampling scheme based on a 5% prevalence and 95% confidence for vaccinated pigs;

9. The vaccination of small herds should be further discussed; 10. To refine the application of NSP tests, more work could be done in predicting the

expected prevalence of infection within and amongst vaccinated herds; 11. Functional FMD expert groups should be created in every country;


EU FMD-NSP Workshop Organizing Committee See Annex A: FMD-NSP workshop program

EU FMD-NSP Workshop Supervisory Team See Annex A: FMD-NSP workshop program

Participants Twelve countries, including Spain, Ireland, the United Kingdom, Germany, the Netherlands, Belgium, France, Italy, Slovenia, Denmark, Portugal and Lithuania were invited. Except for Portugal, every invited country had a team that participated to the workshop. Participant list: Annex A: FMD-NSP workshop program

Observers See Annex A: FMD-NSP workshop program

Sponsors

The workshop was kindly sponsored by the EU Coordination action FMD-CSF project, the EU EPIZONE project Work Package 4.3 (DIVA), FAO EUFMD, the Veterinary and Agrochemical Research Centre and the Federal Agency for the Safety of the Food Chain in Belgium.

Objectives

Given the current FMD-free status without vaccination in Europe and the possibility of a future outbreak with vaccinate-to-live used as an emergency measure, being followed-up by post-vaccination serosurveillance, to return to the favoured status of ‘free from infection without vaccination’:

1. Design and implementation of a survey to substantiate free from infection (i.e. free from infection with a certain degree of confidence, according to the EU directives and the OIE general and specific serosurveillance guidelines). So a country has an outbreak (or different outbreaks), vaccination of the surrounding herds is performed, the outbreaks are stopped, what will this country do to regain the ‘free from FMD without vaccination’ status?

2. Guidance to the interpretation on follow-up on seropositive animals/herds/flocks, for example by using 3 different exercise scenario’s, where we will have 3 different conclusions on the last day.

3. Guidance to the use of laboratory test results in decision-making 4. Identification of the resources required


Report Opening session: Dr. Preben Willeberg, CVO of Denmark, Secretary General of the OIE Scientific Commission for animal disease, member of the FAO EUFMD executive committee I would like to welcome you all to this workshop on the application of NSP-tests in post-vaccination sero-surveillance. Thank you all for coming along for what undoubtedly will be an interesting two-days program, and a special thanks to the organizers, donors, supporting international organizations and to the host laboratory for inviting us all and for making this event possible. I am also grateful for being asked to open this workshop and to be part of the Supervisory Team, and I would like to make just a few observations to underline why I think that this event is a very important and in fact a much needed initiative. Firstly, we all know the background for the present situation following the 2001 outbreaks of FMD in Europe: The public and political pressure to avoid in future situations the culling and destruction of massive numbers of apparently healthy animals, and the move in European and international regulations to make provisions for the use of emergency vaccination to live by application of purified vaccines and DIVA tests. Although the principles of this technology have been generally accepted for some time, there has until now been a shortage of results from applied research and development to document exactly how this would and should work in practice. There would have been serious consequences associated with this lag-time of 4 – 5 years between agreed principles and documented evidence if we had experienced a new emergency situation in Europe – fortunately it did not happen – and now we are much closer to having comfortable insight into details of e.g. sensitivity and specificity of the available NSP-tests. This is thanks to a small group of dedicated, hard working and very skilled individuals, many of whom are present here today. The course material sent to us in the last week documents convincingly what a lot of work has been done: in collecting and analyzing serum samples, comparing and reporting laboratory results from a battery of tests and publishing scientific papers with a wealth of pertinent observations. Secondly, I have had the privilege to follow these activities from the sideline and to encourage and prioritize the work, e.g. in the EUFMD Executive Committee and Research Group meetings, as well as in meetings in the OIE and in the EU. One particular challenge for me has been to encourage cooperation between FMD laboratory experts and analytical epidemiologists to ensure that design and analysis of laboratory test results for surveillance to document freedom from FMD infection following emergency vaccination would be exploited to its full potential – and I think that the


course material which we have received for this workshop also documents that this has been successfully achieved. Finally, this workshop seems to complete the built-up of experience with the applicability of the NSP test principles by bringing in the risk managers, i.e. the CVOs, the Veterinary Services experts and field epidemiologists. As one representative of this segment I appreciate the initiative – again mainly thanks to the same group of people. This is quite a unique event in that it brings together all parties involved in the various aspects of FMD control around a common goal or challenge. Because this workshop involves different disciplines and expertise, some of us may go away tomorrow – still confused or even frustrated – but at least confused or frustrated at a higher level – and I am also sure that we will all have learned something or even a lot, which will prove to be valuable for implementing national contingency plans and control strategies applying emergency vaccination. Thanks again to Kris De Clercq and his group for organizing this, and I wish you all an interesting and challenging workshop. Session I: Review on FMD vaccination and NSP testing The Science behind Non-Structural Protein (NSP) testing, Dr. Kris De Clercq, Belgium

Differentiation of

infected – vaccinated animals

Kris De Clercq

A virus infects a cell, the RNA is read by the cell system and produces structural proteins (SPs) to make the structure of the virus; other parts of the RNA are decoded in proteins to help to make the structure of the virus or help to duplicate the RNA: these are the non-structural proteins (NSPs)


The virus cycle goes on and more and more virus is made but also more and more NSPs. When the cell is destroyed, virus is liberated as well as the NSPs. Part comes in the blood, so the animal will make antibodies against the SPs and the NSPs A vaccine producer mimics in fact what happens in nature by adding virus to a cell culture. The virus cycle goes exactly in the same way…


but then he filters the vaccine to have a pure virus culture. The virus is inactivated so that a virus cycle (and the formation of NSPs) becomes impossible. So, when the animal is vaccinated it will make only antibodies against the SPs. Summary of the reaction of the animal after infection or vaccination or no infection/vaccination


ELISA

StructuralProteins

Non-StructuralProteins

+ +

+ -

- -

ELISA

StructuralProteins


+ +

+ -

Translation to the laboratory using two different ELISAs: one classical ELISA detecting antibodies against SPs and one NSP-ELISA, showing how you can differentiate between infected and/or vaccinated animals. If an animal gets infected after the vaccination, normally the infected animals will react but however some animals will not react, causing a false negative. If a vaccine is not well purified you will have NSPs in the vaccine.


ELISA

StructuralProteins


+ +

+ -

So, after one injection you will normally get no reaction against NSPs but after several vaccinations with this kind of vaccine you will get false positives.

Emergency vaccination: the FMD vaccinate-to-live policy - objectives and conditions for re-applying for freedom from FMD, Dr. Alf Füssel, EU DG SANCO, Head of Trade and Zootechnics Sector 1.

11

FMDFMD

NSP Tests inNSP Tests in

PostPost--VaccinationVaccination SurveillanceSurveillance

Dr. Alf-Eckbert FüsselEuropean Commission - DG Health and Consumer Protection

Directorate D - Animal health and welfareUnit D1 - Animal health and Standing Committees

This presentation does not necessarily represent the views of the Commission 3.

3

3 3 PhasesPhases of of PostPost--vaccinationvaccination

MeasuresMeasures

1. from the beginning of emergency vaccination until at least 30 days have elapsed following the completionof such vaccination (Article 54)

2. from emergency vaccination until the survey and theclassification of holdings are completed (Article 55)

1. Clinical and serological survey in the zone (Article 56)

2. Classification of herds in the zone (Article 57)

3. after the completion of the survey and theclassification of holdings until FMD free status isrecovered (Article 58)

2.

2

EmergencyEmergency vaccinationvaccination

►► CouncilCouncil DirectiveDirective 2003/85/EC 2003/85/EC providesprovides forforemergencyemergency vaccinationvaccination in in casecase of of moremore thanthan oneoneoutbreakoutbreak of FMDof FMD

►► TypesTypes of of vaccinationvaccination�� ProtectiveProtective ((vaccinatevaccinate--toto--livelive))

�� Suppressive (Suppressive (vaccinatevaccinate--toto--killkill))

►► VaccinationVaccination isis carriedcarried out in out in accordanceaccordance withwith�� ArticleArticle 52(1) 52(1) -- vaccinationvaccination zonezone

�� ArticleArticle 52(2) 52(2) -- surveillancesurveillance areaarea surroundingsurrounding thethevaccinationvaccination zonezone

4.

4

Sheep and cattle notvaccinated

Sheep not vaccinated

Herd vaccinated


5.

5

Clinical and serological survey

►Aim : identification of herds within the vaccination zone with contact to the FMDV without showing overt clinical signs

►Content: clinical inspection of all animals in all herds

and

laboratory testing►Sampling:

� herds not vaccinated with species not showing clinical signs 5 % prevalence, 95 % confidence

and

� all vaccinated animals and their non- vaccinated offspring in all vaccinated herds

7.

7

RecoveryRecovery of FMD of FMD freefree statusstatus

followingfollowing emergencyemergency vaccinationvaccination

► all the measures provided for in Articles 36, 44, 54, 55, 56 and 57 have been completed,

► at least one of the following conditions applies:� OIE rules as in force, or� 3 months after slaughter of vaccinated animals + serological

surveillance in accordance with OIE guidelines, or� 6 months after the last outbreak and last vaccination + serological

survey for NSP demonstrated absence of infection in vaccinated animals

► a Decision has been adopted to re-establish the free status

9.

9

Objectives Objectives

of postof post--vaccination measuresvaccination measures►►Risks from products from vaccinated Risks from products from vaccinated animals can be mitigatedanimals can be mitigated

butbut

►►PostPost--vaccination surveillance is carried out vaccination surveillance is carried out to ensure that after the recovery of the to ensure that after the recovery of the status vaccinated ruminants can be moved status vaccinated ruminants can be moved safely out of the previous vaccination zone safely out of the previous vaccination zone and continue their productive life within the and continue their productive life within the vaccinating Member Statevaccinating Member State

6.

6

Classification of holdingsClassification of holdings

► holdings with at least 1 animal positive in the NSP

test

� Presence of FMDV = outbreak

� Previous contact with FMDV, no actual virus circulation

►all animals are killed and the carcasses processed, or

►animals are classified:

� the animals positive to at least one of the approved tests are killed and their carcasses processed, and

� the remaining animals are slaughtered under conditions authorised by the competent authorities;

► fresh meat from ruminants – de-boning and maturation,

►meat from porcine animals - heat treated products

8.

8

ModifiedModified recoveryrecovery of of freefree statusstatus

Following emergency vaccination, and by way of derogation, it may be decided byComitology, to withdraw the restrictionsapplied in the vaccination zone after theclinical and serological survey and theclassification of herds have been completedand confirmed the absence of foot-and-mouthdisease virus infection

Risk assessment taking into account different test strategies

10.

10

ConclusionConclusion

►►When the free status is recovered there When the free status is recovered there

should only be herds which are should only be herds which are ►►either either serosero--negative negative

OrOr

►►SeroSero--positive exclusively from the administration of positive exclusively from the administration of

an inactivated vaccine an inactivated vaccine

►►No danger of carriers or undetected disease No danger of carriers or undetected disease


Presentation of adopted FMD vaccination plans - scenarios prepared for the workshop, Dr. Phillippe Houdart, Belgium 1.

Federal Agency for the Safety of the Food Chain

INTRODUCTION TO THE EXERCISE

Federal Agency for the Safetyof the Food Chain

3.


Approach

• epizootic in dense region (Belgium)

• 3 clusters of outbreaks:

– cluster 1 = index case + neighbouring outbreaks

– cluster 2 = via infected pig transport from index case

– cluster 3 = via infected sheep from index case

• different control strategy but always vaccination

• exercise starts > 28 days after last outbreak

5.


Cluster 1

• SZ surrounding index case + 5 neighbouring outbreaks

• size = 656 km2

• density: – cattle = medium density (85 / km2)– pigs = low density (147 / km2)

– sheep/goat = low density (11 / km2)

2.


Aim of the exercise

• design and interpret a post-vaccination FMD surveillance scheme including the use of NSP tests

• objective: substantiate a claim to return to the status free of infection

• vaccination to live policy � vaccinated animals are not killed

• starting point: EU and OIE guidelines• same type of workshop/exercise in regions with high

density of susceptible animals and in regions with low density of susceptible animals

4.


6.


Cluster 2

• SZ surrounding 2 pig outbreaks

• size = 407 km2

• density: – cattle = medium to high density (166 / km2)

– pigs = medium to high density (668 / km2)– sheep/goat = low density (8 / km2)


7.


Cluster 3

• vaccination area surrounding some 20 outbreaks following movement of infected sheep

• size = 770 km2

• density: – cattle = medium to high density (188 / km2)– pigs = high density (1.187 / km2)– sheep/goat = low density (15 / km2)

• includes buffer area of 5 km (670 km2) around vaccination area with slightly lesser density

9.


scenario 1

• deals only with clusters 1 and 2

• culling: – cluster 1: all animals in 500 m– cluster 2: cattle in 500 m, pig and sheep/goat in

1.500 m

• vaccination of cattle only:– cluster 1: in 3.000 m radius– cluster 2: in 1.500 m radius

• groups:– ES + IR– IT + OIE + FAO

11.


scenario 3

• deals only with cluster 3

• culling of outbreaks only

• vaccination of cattle only in vaccination area:

• groups:– UK + SL

– DK + LI

8.


Methodology

• 3 different scenarios

• each approached by 2 groups of 2 countries

• tasks: – design surveillance scheme for vaccinated area

(both vaccinated and non vaccinated animals)– design surveillance scheme for rest of SZ and

buffer area– interpret set of results of surveillance scheme– design follow up for positive results

10.


scenario 2

• deals only with clusters 1 and 2

• culling: – cluster 1: all animals in 500 m– cluster 2: all animals in 500 m

• vaccination of all animals:– cluster 1: in 3.000 m radius– cluster 2: in 1.500 m radius

• groups:– NL + DE– BE + FR

12.


documentation

• publications about FMD testing and in particular NSP testing

• tables with composition of the different clusters


Background information about the epizootic In the second half of November an outbreak of FMD strain A Iran 96 is observed in a mixed holding, rearing pigs and sheep. On the holding, the construction of a new pig stable is in its final stage. The most likely source of infection are leftovers of infected food brought back from his homeland by a foreign employee of the construction firm. The employee has dropped these leftovers about 2 weeks prior to the detection of the outbreak in the grassland next to the stable, where some 30 sheep were kept. Due to the epizootic of BT serotype 8 that is at its height in the region, the discrete symptoms shown by the infected sheep in the initial stages are mistakenly taken for a BT infection. They are therefore ignored by the owner, all the more because an open house is planned to show of the new stable to the farming community in the area and the owner does not want any fussing about by the authorities at the time. Three major routes are responsible for the subsequent spreading of the disease:

- A week prior to the detection of the outbreak, the last of the pigs present in two other pig stables on the holding are transported to the slaughterhouse. After loading the pigs, the driver visits the new stable and passes through the pasture with the infected sheep. The next day the driver collects pigs of 2 other pig farms some 40 kilometres to the north, wearing the same boots and infecting both farms in this way.

- Around the same time, most of the sheep are sold to a sheep trader to clear the pasture next to the new stable in view of the open house. The animals remaining on the premise, including some showing more marked symptoms, are moved by the owner and his brother (also a farmer) to a neighbouring pasture a kilometre away. The animals that are sold are moved by the trader to his premises about 100 km west. In the following days, he sells most of the animals on a local market and directly from his stable to several sheep holders.

- Finally, on the occasion of the open house that is organised just prior to the detection of the outbreak to show off the new stable, several local farmers enter the pasture previously infected by the sheep and take some virus home on their footwear.

The day following the open house and in view of the deteriorating condition of the sheep, the owner finally consults his veterinarian. The case is still reported as a BT suspicion and samples are sent in to the laboratory. The diagnosis of FMD is finally made by the reference lab. In the weeks following the detection of the case, several outbreaks are diagnosed grouped into 3 clusters (see map). Half December, after about 20 days, all 3 clusters of outbreaks are finally managed by different combinations of culling and preventive vaccination of cattle and/or small ruminants and/or pigs.


Characteristics of cluster 1 Cluster 1 is comprised of the 10 km surveillance zones surrounding the index case and 5 other outbreaks in the same area (amongst which the farm of the owner’s brother and a neighbouring farm):

- general composition: medium density area for cattle, low density area for small ruminants and pigs;

- size: 656 km2;

- composition: see tables. The index case is detected late, the other outbreaks in the cluster pop up in less than a week. Characteristics of cluster 2 Cluster 2 is comprised of the 10 km surveillance zones surrounding the 2 pig outbreaks in the north:

- general composition: medium to high density area for pigs and cattle, low density area for small ruminants;

- size: 407 km2;

- composition: see tables. Both outbreaks are detected in an early stage and subsequently no more outbreaks are recorded in this area. Characteristics of cluster 3 Cluster 3 is comprised of a large zone that surrounds about 20 outbreaks and their 10 km surveillance zones in a large area around the traders stable. It includes an additional 5 km zone that is added around the this large zone as a buffer zone with the non infected areas.

- general composition: high density area for pigs, medium to high density area for cattle and low density area for small ruminants;

- size: 1.440 km2; - composition: see tables.

Several of the outbreaks have no clear link with the trader’s movements or are recorded after vaccination in this cluster has started.

Scenario’s The time setting for the exercise is about 1 month after the last outbreak has occurred. Surveillance since has been based on clinical surveillance. Apart from the holdings that are culled, no serological surveillance has been performed yet.


The aim of the exercise is to device and interpret a post-vaccination FMD surveillance scheme including the use of NSP tests. The surveillance scheme must substantiate a claim to return to the status free of infection (i.e. freedom of infection with a certain degree of confidence, according to the EU guidelines an the OIE general and specific serosurveillance guidelines) taking into account the characteristics of the tests chosen. The participants are divided into 3 working groups that will device the surveillance scheme for one of the scenarios.

- Since the exercise focuses on the use of NSP test, the scheme for dealing with both the vaccinated and non vaccinated animals in the vaccination areas is the principal assignment.

- Where it is relevant and sufficient time is left, the scheme for the rest of the surveillance zone can also be elaborated.

- Finally, based on the schemes presented, the groups will be presented with some results of the serosurveillance scheme chosen. They will have to interpret these results and present a suitable follow up of the seropositive animals/herds/flocks. A sample is considered positive when it will be twice positive in a double, successive testing with the same NSP-kit.

In each of the scenarios all outbreaks and direct contact herds are culled. The classic 3 km protection zone and 10 km surveillance zone Scenario 1 Scenario 1 focuses on clusters 1 and 2 taking into account the following control strategy:

- culling: - cluster 1: culling of all susceptible animals (cattle, small ruminants and pigs) in a

500 m radius; - cluster 2: culling of all pigs and small ruminants in a 1.500 m radius and of all cattle

in a 500 m radius;

- vaccination: - cluster 1: all cattle in a 3.000 m radius around the outbreaks (from 500 m to 3.000 m

around the outbreaks); - cluster 2: all cattle in a 1.500 m radius around the outbreaks (from 500 m to 1.500 m

around the outbreaks);

- size of the vaccination area: 162 km2. Scenario 2 Scenario 2 focuses on clusters 1 and 2 taking into account the following control strategy:

- culling in clusters 1 and 2: culling of all susceptible animals (cattle, small ruminants and pigs) in a 500 m radius;

- vaccination: - cluster 1: all susceptible species (cattle, small ruminants and pigs) in a 3.000 m

radius around the outbreaks (from 500 m to 3.000 m around the outbreaks); - cluster 2: all susceptible species (cattle, small ruminants and pigs) in a 1.500 m


radius around the outbreaks (from 500 m to 1.500 m around the outbreaks);

- size of the vaccination area: 162 km2. Scenario 3 Scenario 3 focuses only on cluster 3 (vaccination area and buffer area surrounding the vaccination area), taking into account the following control strategy.

- culling of the outbreaks, direct contact holdings and some neighbouring holdings;

- vaccination: all cattle in the whole of the vaccination area;

- size of the vaccination area: 770 km2. Annexes The following tables reproduce:

- the surface of the different zones,

- the composition of the various clusters with a stratification by radius,

- the composition of the various clusters with a stratification by size.

Surface area:

area surface (km2)

cattle farm

density (per km2)

cattle density

(per km2)

sheep/goat farm

density (per km2)

sheep/goat density

(per km2)

pig farm density

(per km2)

pig density

(per km2)

cluster 1 - vaccinatie

149 - - - - - -

cluster 1 - 10.000 m 656 1,59 85 0,94 11 0,23 147 cluster 2 - vaccination

13 - - - - - -

cluster 2 - 10.000 m 407 1,75 166 0,89 8 0,63 668 cluster 3 - vaccination

770 2,84 188 1,77 15 1,40 1.187

cluster 3 - buffer 670 2,05 127 1,32 10 1,01 806


Cluster composition: Cluster 1:

zone number of

cattle holdings

total number of cattle

number of sheep/goat holdings

total number of sheep/goat

number of pig holdings

total number of

pigs

500 m 9 556 1 13 3 714

3.000 m 151 6.856 91 1.299 14 14.283

10.000 m 884 48.455 523 6.196 137 81.448

1.044 55.867 615 7.508 154 96.445 Cluster 2:

zone number of

cattle holdings





total number of

pigs

500 m 6 392 1 366 2 1.411

1.500 m 33 4.317 23 140 14 17.198

3.000 m 76 9.809 49 486 32 36.247

10.000 m 596 53.102 291 2.367 208 216.977

711 67.620 364 3.359 256 271.833 Cluster 3:

zone number of

cattle holdings





total number of

pigs

vaccination 2.188 144.872 1.365 11.780 1.075 914.155

buffer 1.372 85.007 885 6.771 677 540.289

3.560 229.879 2.250 18.551 1.752 1.454.444


Cluster size: Cluster 1 – 3000 m

size number of cattle holdings


average number of cattle



average number of sheep/goat

number of pig

holdings

total number of pigs

average number of pigs

1 to 4 54 125 2 42 116 3 0 0 -

5 to 10 37 266 7 27 213 8 0 0 -

11 to 25 20 327 16 14 206 15 0 0 -

26 to 50 10 373 37 3 119 40 0 0 -

51 to 100 7 438 63 3 247 82 1 90 90 101 to 250

17 2.521 148 1 118 118 1 124 124

251 to 500

4 1.340 335 1 280 280 2 876 438

501 to 1000

2 1.466 733 0 0 - 3 2.432 811

1001 to 0 0 - 0 0 - 7 10.761 1.537

151 6.856 91 1.299 14 14.283 Cluster 1 – 10000 m







number of pig

holdings



1 to 4 299 642 2 211 593 3 13 27 2

5 to 10 160 1.111 7 195 1.351 7 4 26 7

11 to 25 122 1.975 16 87 1.388 16 3 45 15

26 to 50 69 2.493 36 14 502 36 6 214 36

51 to 100 117 8.595 73 7 406 58 6 458 76 101 to 250

81 12.182 150 6 890 148 25 4.178 167

251 to 500

14 5.363 383 3 1.066 355 23 8.375 364

501 to 1000 20 13.855 693 0 0 - 28 20.479 731

1001 to 2 2.239 1.120 0 0 - 29 47.646 1.643

884 48.455 523 6.196 137 81.448


Cluster 2 – 1500 m







number of pig

holdings



1 to 4 2 6 3 9 21 2 0 0 -

5 to 10 3 26 9 9 55 6 1 9 9

11 to 25 2 44 22 5 64 13 0 0 -

26 to 50 5 208 42 0 0 - 0 0 -

51 to 100 7 489 70 0 0 - 0 0 - 101 to 250

11 1.467 133 0 0 - 0 0 -

251 to 500

0 0 - 0 0 - 2 684 342

501 to 1000

3 2.077 692 0 0 - 5 3.741 748

1001 to 0 0 - 0 0 - 6 12.764 2.127

33 4.317 23 140 14 17.198








number of pig

holdings



1 to 4 6 16 3 19 48 3 0 0 -

5 to 10 3 22 7 19 129 7 0 0 -

11 to 25 13 237 18 8 123 15 3 55 18

26 to 50 6 214 36 1 38 38 0 0 -

51 to 100 21 1.590 76 2 148 74 0 0 - 101 to 250

21 3.101 148 0 0 - 1 220 220

251 to 500

1 400 400 0 0 - 5 1.713 343

501 to 1000

4 2.700 675 0 0 - 8 5.856 732

1001 to 1 1.529 1.529 0 0 - 15 28.403 1.894

76 9.809 49 486 32 36.247









number of pig

holdings



1 to 4 71 182 3 138 359 3 1 1 1

5 to 10 64 439 7 111 745 7 2 18 9

11 to 25 61 1.059 17 32 505 16 0 0 -

26 to 50 68 2.506 37 7 223 32 5 199 40

51 to 100 133 9.982 75 1 75 75 4 353 88 101 to 250

172 25.303 147 1 116 116 27 5.078 188

251 to 500

15 5.347 356 1 344 344 26 10.186 392

501 to 1000

11 7.076 643 0 0 - 54 38.528 713

1001 to 1 1.208 1.208 0 0 - 89 162.614 1.827

596 53.102 291 2.367 208 216.977


Cluster 3 – vaccination zone







number of pig

holdings



1 to 4 331 746 2 748 1.949 3 19 44 2

5 to 10 215 1.594 7 395 2.675 7 9 60 7

11 to 25 280 4.663 17 158 2.433 15 14 225 16

26 to 50 278 10.599 38 38 1.281 34 14 497 36

51 to 100 564 42.031 75 17 1.251 74 49 3.839 78 101 to 250

471 68.997 146 8 991 124 134 23.970 179

251 to 500

45 14.054 312 0 0 - 198 73.245 370

501 to 1000

4 2.188 547 0 0 - 289 214.091 741

1001 to 0 0 - 1 1.200 1.200 349 598.184 1.714

2.188 144.872 1.365 11.780 1.075 914.155 Cluster 3 – buffer zone







number of pig

holdings



1 to 4 225 519 2 456 1.252 3 33 76 2

5 to 10 147 1.054 7 279 1.870 7 9 63 7

11 to 25 202 3.517 17 123 1.961 16 13 223 17

26 to 50 207 7.861 38 15 534 36 17 623 37

51 to 100 294 22.116 75 9 640 71 22 1.714 78 101 to 250

261 38.891 149 3 514 171 91 16.519 182

251 to 500

36 11.049 307 0 0 - 144 54.444 378

501 to 1000 0 0 - 0 0 - 157 115.983 739

1001 to 0 0 - 0 0 - 191 350.644 1.836

1.372 85.007 885 6.771 677 540.289


Session II and III: Results from working groups Scenario 1: 8 outbreaks – 2 clusters

Only bovine vaccinated All bovine/sheep/goats/pigs culled 0 - 500 m

Group 1 = Spain & Ireland (see annex B) Group 2 = Italy + FAO + OIE (see annex C)

Vaccination zone (VZ) Cluster 1:

• Vaccination of bovine in 500 m – 3 km zone: • bovine: 151 herds – 6,856 animals

• No vaccination in 500 m – 3 km zone for: • Pigs: 14 herds – 14,283 animals • Sheep/Goats: 91 herds – 1,299 animals

Design: G1 – option 1(1) G1 – option 2 G1 – option 3(2) G2 Bovine Sample all herds/all

animals Sample only herds ≥5 animals/(95:2) animals;150

Sample all herds ≥5 animals/all animals;

Sample all herds/all animals

Sheep/ goats

Sample all herds/(95:5) animals;60

Sample only herds ≥5 animals/(95:5) animals;60

Sample all herds ≥5 animals/(95:5) animals;60

Sample all herds/(95:5) animals

Pigs Sample all herds/(95:5) animals;60

Sample only herds ≥5 animals/(95:5) animals;60

Sample all herds ≥5 animals/(95:5) animals;60

Clin. surv. (CS) all herds/all animals

(1)according to EU directives (2003/85/EC) (2)Preferred option of Group 1 Figures: G1 – option 1 G1 – option 2 G1 – option 3 G2 Bovine 151 herds/6,856

samples 97 herds/5,675 samples

97 herds/N1 samples(3)

151 herds/6,856 samples

Sheep/ goats

91 herds/954 samples 49 herds/838 samples 49 herds/838 samples 91 herds/876 samples

Pigs 14 herds/840 samples 14 herds/840 samples 14 herds/840 samples CS 14 herds/14,283 animals

Cluster 2:

• Vaccination of all bovine in 500 m – 1.5 km zone: • Bovine: 33 herds – 4,317 animals

• Sheep/goats and pigs culled in 500 m – 1.5 km zone: • Pigs: 14 herds – 17,198 animals • Sheep/Goats: 23 herds – 140 animals


Design: G1 – option 1 G1 – option 2 G1 – option 3 G2 Bovine Sample all herds/all

animals Sample only herds ≥5 animals/(95:2) animals;150

Sample only herds ≥5 animals/all animals;

Sample all herds/all animals

Sheep/ goats

Culled Culled Culled Culled

Pigs Culled Culled Culled Culled

Figures: G1 – option 1 G1 – option 2 G1 – option 3 G2 Bovine 33 herds/4,317

samples 31 herds/2,684 samples

31 herds/ N2 samples(3)

33 herds/4,317 samples

Sheep/ goats

/ / / /

Pigs / / / / (3) N1 + N2 = 11042 Resources: G1 – option 1 G1 – option 2 G1 – option 3 G2 Vet teams

12 (5 herds x 5 days) 8 8

Lab Bovine NSP-IB Sheep SP Ceditest

Protection zone (PZ) Cluster 1:

• Vaccination zone = protection zone

Cluster 2: • No vaccination of animals in 1.5 km – 3 km zone:

• Bovine: 76 herds – 9,809 animals • Sheep/goats: 49 herds – 486 animals • Pigs: 32 herds – 36,247 animals

Design: G1 (only 1 option) G2 Bovine Clin. surv. all herds/all animals Clin. surv. all herds/all animals

Sheep/ goats

Sample all herds/(95:5) animals Sample all herds/(95:5) animals

Pigs Clin. surv. all herds/all animals Clin. surv. all herds/all animals


Figures: G1 G2 Bovine CS 76 herds/9,809 animals CS 76 herds/9,809 animals

Sheep/ goats

49 herds/458 samples 49 herds/415 samples

Pigs CS 32 herds/36,247 animals CS 32 herds/36,247 animals

Resources: G1 G2 Vet teams 2

Lab SP Ceditest

Surveillance zone (SZ) Cluster 1:

• No vaccination of animals in 3 km – 10 km zone: • Bovine: 884 herds – 48,455 animals • Sheep/goats: 523 herds – 6,196 animals • Pigs: 137 herds – 81,448 animals

Design: G1 G2 Bovine Clin. surv. all herds/all animals Clin. surv. all herds/all animals Sheep/ goats

Sample (95:2) of herds/(95:5) animals Sample (95:2) of herds/(95:5) animals

Pigs Clin. surv. all herds/all animals Clin. surv. all herds/all animals Figures: G1 G2 Bovine CS 884 herds/48,455 animals CS 884 herds/48,455 animals

Sheep/ goats

149 herds/1,388 samples 131 herds/1,207 samples


Cluster 2:




Sample (95:2) of herds/(95:5) animals Sample (95:2) of herds/(95:5) animals


Sheep/ goats

149 herds/1,176 animals 119 herds/912 animals


Resources: G1 G2 Vet teams 12

Lab SP Ceditest

Initial laboratory results (given by the supervisory team, based on the design made by the group) Cluster 1:

G1 G2 • 1 bovine farm with 1 NSP positive

(herd size 611, sample size 150); VZ

• 2 sheep/goat farms with 1 NSP positive each (herd size 280, sample size 60 and herd size/sample size 19); VZ

• 4 pig farms with 1 NSP positive each (sample size 60); VZ

• 1 bovine farm with 6 NSP+ (herd size 611, sample size 611); VZ

• 1 sheep farm with 1 NSP+ (herd size 19, sample size 19); VZ

• 1 sheep farm with 2 NSP+ (herd size 280, sample size 100); VZ

• 1 sheep/bovine farm with 1 NSP+ sheep (herd size 25, sample size 25); SZ

Cluster 2:

G1 G2 • / • 1 bovine/sheep herd with 7 NSP+

bovine (herd size 861, sample size 861); VZ

• 1 bovine herd with 1 NSP+ (herd size 111, sample size 111), VZ

• 1 sheep herd with 1 NSP+ (herd size 116, sample size 64); SZ



Interpretation and follow-up

G1 G2 • Analysis of test results:

o Any evidence of clustering of NSP+

o Magnitude of NSP+ • Laboratory follow-up:

o NSP+ bovine: retest (IB or other) → retest negative

o NSP+ non-vaccinates: retest (SPCE/VNT) → 2 NSP+ pigs retest positive → single reactors from a sample of large herds

Field follow-up

G1 G2 • Revisit farms with NSP+:

o Clinical surveillance of all animals

o EPI investigation of all animals o Resample NSP+ animals (if

identifiable) o Sample all herd mates

(2003/85/EC) o Sampling of contact herds: not

yet

• Revisit farms with seropositives: o vaccinated bovine:

• serological retesting of head

• Probang test of positive head

• Serological testing of un-vaccinated animals in farm

o un-vaccinated animals: • Retesting of farm (95:5) • Clin. surv. of all animals

in farm Group conclusions

G1 G2 • Option 3 could provide enough

confidence and it needs les vet teams than required by the EU Directive (2003/85/EC)

• Option 2 and 3 need similar resources with a higher confidence in option 3


Scenario 2: 8 outbreaks – 2 clusters All animals vaccinated All bovine/sheep/goats/pigs culled 0 - 500 m

Group 3 = The Netherlands + Germany (see annex D) Group 4 = Belgium + France (see annex E)


• Vaccination of all animals 500 m – 3 km: • Bovine: 151 herds – 6,856 animals • Pigs: 14 herds – 14,283 animals • Sheep/goats: 91 herds – 1,299 animals

Design: G3 G4 Bovine Sample all herds/all animals Sample all herds/(95:5) animals

Sheep/ goats

Sample all herds/all animals Sample all herds/all animals

Pigs Sample all herds/(95:5) animals (x3?)(2) Sample all herds/(95:5) animals Figures: G3 G4 Bovine 151 herds/6,856 samples 151 herds/4,879 samples

Sheep/ goats

91 herds/1,299 samples 91 herds/1,299 samples

Pigs 14 herds/N1 samples(1) (x3?)(2) 14 herds/1,415 samples (1) N1 + N2 = 1,629 (2) in a worst case scenario the number of pig samples should be tripled (when 3 epidemiological units/pig farm) Cluster 2:

• Vaccination of all animals 500 m – 1.5 km • Bovine: 33 herds – 4,317 animals • Pigs: 14 herds – 17,198 animals • Sheep/goats: 23 herds – 140 animals

Design: G3 G4 Bovine Sample all herds/all animals Sample all herds/(95:5) animals

Sheep/ goats

Sample all herds/all animals Sample all herds/all animals

Pigs Sample all herds/(95:5) animals (x3? ?)(2) Sample all herds/(95:5) animals


Figures: G3 G4 Bovine 33 herds/4,317 samples 33 herds/2,675 samples

Sheep/ goats

23 herds/140 samples 23 herds/140 samples

Pigs 14 herds/ N2 samples(1) (x3?)(2) 14 herds/1,363 samples (1) N1 + N2 = 1629 (2) in a worst case scenario the number of pig samples should be tripled (when 3 epidemiological units/pig farm) Resources: G3 G4 Vet teams

Lab Cedi NSP (99.2% Sp)(1) Cedi NSP for bovine Sp 99.5%(2)/Sn 63.6% For pigs Sp 100%/Sn 55.5% 200,000 test/week available

(1) Sp of 99.2% is based on Brocchi et al. (2006) after retesting of initial positive bovine results regardless of their vaccination status (2) Sp of 99.5% is based on Brocchi et al. (2006) on initial test results for vaccinated bovine Protection zone (PZ) Cluster 1:

• Vaccination zone = protection zone

Cluster 2: • No vaccination of animals in 1.5 km – 3 km zone:

• Bovine: 76 herds – 9,809 animals • Sheep/goats: 49 herds – 486 animals • Pigs: 32 herds – 36,247 animals

Design: G3 G4 Bovine Clin. surv. all herds/all animals Clin. surv. all herds/all animals

Sheep/ goats

Sample all herds/(95:5) animals;60 Sample all herds/all animals

Pigs Clin. surv. all herds/all animals Clin. surv. all herds/all animals


Sheep/ goats

49 herds/N3 Samples(3) 49 herds/486 samples

Pigs CS 32 herds/36,247 animals CS 32 herds/36,247 animals (3) N3 + N4 + N5 = 7264


Resources: G3 G4 Vet teams

Lab SP test SPC ELISA (Sn 100%/Sp 99.5%)




Sample all herds/(95:5) animals;60 Sample all herds/all animals (SPCE)


Sheep/ goats

523 herds/N4 Samples(3) 523 herds/6,196 samples

Pigs CS 137 herds/81,448 animals CS 137 herds/81,448 animals (3) N3 + N4 + N5 = 7264 Cluster 2:



Sample all herds/(95:5) animals;60 Sample all herds/all animals (SPCE)


Sheep/ goats

291 herds/N5 Samples(3) 291 herds/2,367 animals

Pigs CS 208 herds/216,977 animals CS 208 herds/216,977 animals (3) N3 + N4 + N5 = 7264



Lab SP test SPC ELISA (Sn 100%/Sp 99.5%)

Initial laboratory results (given by the supervisory team, based on the design made by the group) Cluster 1:

G3 G4 • 2 cattle farms with 2 NSP+ (herd

size 855/284, sample size 60); VZ • 1 sheep/goat farm with 1 NSP+

(herd size 19, sample size 19); VZ • 1 sheep/cattle farm with 1 NSP+

sheep (herd size 25/sample size 25); SZ

• 1 sheep/cattle farm with 1 NSP+ sheep(herd size 25, sample size 25); SZ

• 2 cattle farms with 2 NSP+ each(herd size 855/284, sample size 164/154); VZ

Cluster 2:

G3 G4 • 2 sheep farms with 2 NSP+ each

(herd size 116/344, sample size 79/99); SZ

• 1 cattle/pig farm with 1 NSP cattle positive and 3 NSP pig positives (herd size 51/951, sample size 60/51); VZ



• 1 pig/cattle herd with 3+1 NSP+ (herd size 951/51, sample size 227/51); VZ



o Much lower no. of positive farms than expected: 114 NSP+ expected in VZ (13 NSP+ pigs); worst case scenario: 140 NSP+ (39 NSP+ pigs)

• Laboratory follow-up: o NSP+ vaccinates: retest

(Cedi or other) o SP+ non-vaccinates: retest

(SPCE)

• Analysis of test results: o SZ: 12 SPCE+ = lot less

than expected (45) o VZ: 5 NSP+ cattle = lot less

than expected (46) o VZ: 3 NSP+ pigs = more

than expected (0), but Sp for pigs is not really 100%, so this is possible


(Cedi + Svanova) o SP+ non-vaccinates: retest

(Cedi NSP)


Initial testing vaccination area all ruminants and pigs 5% 95%

pos neg

Rebleed positive animals at least 4 days later Same animal positive again

neg

Remove positive animals

Rebleed in ruminants whole herd at least 4 days later, for pigs penmates and adjacent pens (looking for seroconversion)

pos neg

? Outbreak

Exclude evidence of virus circulation, for ruminants in addition exclude presence of carriers

Rebleed positive animals at least 4 days later

Same animal positive again

neg

In all cases clustering of positive samples indicates virus circulation then the CVO can decide to call it an outbreak in an earlier stage

Field follow-up

G3 G4 • Revisit non-vaccinated farms with SP+:

o Clinical surveillance of all animals

o Resample (95:5) o Check for epidemiological link to

known outbreak o Revisit farms ASAP o Further: see appendix 2

• Revisit vaccinated farms with NSP+ (figure 1)

• SZ: retest positive animals (CEDI NSP)

→ if + : • Kill positives • Epidem. survey in positive herds • 2 weeks of CS in positive herds • Serosurvey in herd after 2 weeks

(NSP Cedi) • VZ: retest positives (Cedi/Cedi/

Svanova) o if - : kill initial positives o if + : kill herd

Figure 1 :


Group conclusions (Recommendations)

G3 G4 • EU labs should develop an independent

confirmatory test • The CEDI-Test has 99.2 % specifity in

the repeated once scenario. In consequence 353 animals could be false positive (approximately one positive per herd!) if all vaccinated animals are sampled.

• It is possible from a capacity point of view to test all vaccinated herds and animals, but alternative scenarios should be developed, because testing all animals will not be possible in areas with a higher animal density and a regime is difficult to change in a new situation.

• Necessity to rediscuss Directive 2003/85/EC (Art. 56): testing all vaccinated animals (pigs)?

• What to do with NSP positives? Exclude virus circulation by cluster analysis (risk assessment of NSP positive herds, or virus detection) and culling of positive herds? Alternative measures to culling herds (only culling of positive animals)?

• Different serological tests for different zones (SP tests in surveillance zone and NSP as confirmatory test; NSP tests in vaccination zone)

• Continuation of NSP test validation for pigs, sheep and goats in experimental and field settings

• Improvement of existing NSP tests (Sn and Sp) and development of confirmatory assays?

• Vaccination strategy is a good but complex option (follow-up is harder, need for further harmonisation of interpretation of NSP positives))


Scenario 3: 25 outbreaks – 3 clusters Only bovine vaccinated All bovine/sheep/goats/pigs culled 0 - 500 m Only cluster 3 considered

Group 5 = Slovenia + UK (see annex F) Group 6 = Denmark + Lithuania (see annex G)


• Vaccination of only bovine 500 m – 3 km: • Bovine: 2,188 herds – 144,872 animals • pigs: 1,075 herds – 914,155 animals • sheep/goats: 1,365 herds –11,780 animals

Design: G5 G6 Bovine Clinical surveillance +

Sample all herds ≤50/all animals Sample all herds >50/(95:5) animals Or Sample all herds ≤50/all animals Sample (95:2) herds >50/(95:5) animals Or Sample all herds ≤50/all animals Sample (95:1) herds >50/(95:5) animals Or Subdivide region into three geographical areas containing approximately equal numbers of cattle herds and test within each area so as to detect a 2% prevalence of infection with 95% confidence*.

Sample all herds /all animals

Sheep/ goats

Clinical surveillance + Sample all herds/(95:5) animals

Sample all herds/(95:5) animals , thus: Sample all herds ≤50/all animals Sample all herds >50 and <101/51-77 animals Sample all herds >100/77-125 animals

Pigs Clinical surveillance all herds/all animals Clinical surveillance all herds/all animals Remark: herds < 51 animals = spatial clusters analysis *preferred option, but impossible to calculate no. of samples because there are not enough outputs Figures: G5 G6 Bovine 2,188 herds/89,713 samples

Or 1,245 herds/26,823 samples Or 1,362 herds/34,770 samples Or 3x121 herds/N samples*

2,188 herds/144,872 samples

Sheep/ goats

1,365 herds/9,389 samples 1365 herds/appr. 10,000 samples

Pigs CS 1,075 herds – 914,155 animals CS 1,075 herds – 914,155 animals

* cannot be calculated: not enough outputs


Resources: G5 G6 Vet teams 2 teams for bleeding, 3 weeks, 5 days

2 teams for CS (pigs), 3.5 weeks, 5 days

Lab Sheep: SPCE ELISA; NSP or VNT for confirmation Cattle: NSP ELISA (Cedi/Cedi/Svanova, assuming Se 66.7% and Sp 99.99%)

Sheep: SP ELISA (Se 99-100%; Sp 100%) Cattle: NSP Cedi/ NSP Cedi (Se 81.8%; Sp 99.2%) 12 technicians + 2 staff members (3 weeks) 155,000 NSP Cedi-Cedi 310 Svanova


• No vaccination in 3 km – 10 km zone: • Bovine: 1,372 herds – 85,007 animals • Sheep/goats: 885 herds – 6,771 animals • Pigs: 677 herds – 540289 animals

Design: G5 G6 Bovine Clin. surv. all herds/all animals Clin. surv. all herds/all animals + early

warning (milkdrop) Sheep/ goats

CS + Sample flocks with sheep/goats only (95:2) + 7 largest flocks/(95:5) animals

Clinical surveillance

Pigs Clin. surv. all herds/all animals Clin. surv. all herds/all animals + early warning (increased mortality)


Sheep/ goats

140 herds/1,213 samples CS 885 herds/6,771 animals



Lab Sheep: SPCE ELISA, NSP or VNT for confirmation (100% Se and Sp)


Initial laboratory results (given by the supervisory team, based on the design made by the group)

G5 G6 • 3 cattle herds with 1 NSP+ each

(herd size 171/227/477, sample size 60); VZ

• 4 sheep/goat herd with 1 NSP positive each (herd sizes 1200/114/36/25, sample sizes 60/60/36/25); VZ

• 1 sheep/cattle herd with 3 NSP+ sheep(herd size 1200, sample size 125); SZ

• 2 cattle herds with 2 NSP+ each(herd size 855/284, sample size 164/154); VZ

• 1 sheep herd with 1 NSP+ (herd size 36, sample size 36)

• 1 sheep herd with 2 NSP+ (herd size 114, sample size 77)

• 1 sheep/cattle herd with 1 NSP+ sheep herd size 25, sample size 25)

• 303 cattle farms with 1 NSP+ each (herd sizes 71-1000, sample sizes ??)



o Any evidence of clustering of positives → no, perhaps 1 sheep/cattle farm in the middle

o Small cattle herds are negative: uncertain of attainment of desired confidence

o Sheep: 4 SP+, 5 expected o Cattle: 3 NSP+, 9 expected


(Cedi/Svanova) o SP+ non-vaccinates: retest

(NSP or VNT)

• Analysis of test results: o Cedi-Cedi: in all cases the

number of positives is below the herd cut point


(Svanova) → 3 farms with 1 NSP+ each → if epidemiological link with true outbreak: article 57, if not: FP

o NSP+ non-vaccinates: retest (Svanova) → no positives


Field follow-up

G5 G6 • Revisit and check all farms with

seropositives • Sheep:

– Cull (and resample / test singleton) positives (n=4)

– If positive, retest same cohort as before to determine if positives at 95/5

– And test rest of flock to estimate the prevalence of seropositives assuming true prevalence is 10% +/- 5% (all from 2 small flocks, 39 of 54 (114), 123 of 1140 (1200))

– No probangs – Classify flocks based on results

• Cattle: – Cull (and resample / test

singleton) positives (n=3) – If positive, resample cohort and

rest of herd to estimate prevalence at 95% confidence assuming true prevalence is 1% +/- 0.5%

– No probangs – Classify herds based on results

• No slaughter if herds classified as uninfected


Group conclusions (Remarks)

G5 G6 • VZ: The need to test all sheep flocks is

difficult to assess without a map showing location of all IPs and associated zones.

• Liked to have seen all IPs in PZ, SZ and VZ drawn out on a map to properly consider whether or not all sheep flocks in the vaccination zone needed to be sampled.

• Needed spatial distribution of all cattle herds subdivided into two herd sizes (greater or smaller than 50 cattle)

• Needed qualitative and quantitative results for all positive test results

• Needed all serology results by herd or flock size

• Needed all positive herds or flocks – distance to nearest infected premises and date that IP was confirmed

• Needed all positive herds or flocks – distance to one another (scale map?)

• VZ: Sampling cattle herds/(95:2) animals: desired confidence not achieved

• VZ: Sampling cattle herds/(95:5) animals: desired confidence not achieved below 90 cattle on the farm

• VZ: therefore sample all cattle herds/all animals


Session IV: Discussion of results and models for serosurveillance: Summary, Conclusions, Recommendations and Observations Summary Vaccination zone: Total no. of samples/herds: G1 – option 1 G1 – option 3 G2 Bovine (vacc.) Sheep/goats (non-vacc.) Pigs (non-vacc.)

11,173/184 3,976/438

840/14

11,042/128 3,887/438

840/14

11,173/184 3,410/438

CS G3 G4 Bovine (vacc.) Sheep/goats (vacc.) Pigs (vacc.)

11,173/184 1,439/114 1,629/28

7,534/184 1,439/114 2,778/28

G5 (option 3) G6 Bovine (vacc.) Sheep/goats (non-vacc.) Pigs (non-vacc.)

34,770/1,362 9,389/1365

CS

144,872/2188 10,000/1,365

CS • Vaccinated bovine:

o group 1 considered option 3 to be the best option (cf. sample only herds ≥ 5 animals/all animals → approximately same sample size as according to EU Directive 2003/85/EC, but less vet teams).

o Groups 2, 3 and 6 propose testing all vaccinated bovine (based on the risk of becoming a carrier).

o Groups 4 and 5 propose (95:5) sampling within herds for vaccinated bovine (according to EU Directive 2003/85/EC, all herds/all animals have to be sampled), but then there is the small herds problem: confidence cannot be achieved when sampling (95:5) within herd. Group 5 tries to solve this by doing spatial cluster analysis.

• Non-vaccinated pigs: o Only clinical surveillance (CS) of non-vaccinated pigs proposed by groups 2,

5 and 6. o Group 1 proposes sampling of all herds ≥ 5 animals and 95:5 sampling within

these herds. • Vaccinated pigs:

o Smaller sample size recommended (95:5 sampling) instead of all animals (as according to EU directive 2003/85/EC) proposed by groups 3 and 4.

• Non-vaccinated and vaccinated sheep and goats: agreement on sampling (following EU Directive 2003/85/EC).

• Some groups calculated the Herd Cut Point to see if SP positive results should be considered as significant.


Protection zone + surveillance zone: Total no. of samples/herds: G1 – option 1 G2 Bovine (non-vacc.) Sheep/goats (non-vacc.) Pigs (non-vacc.)

CS 3,022/347

CS

CS 2,534/299

CS G3 G4 Bovine (non-vacc.) Sheep/goats (non-vacc.) Pigs (non-vacc.)

CS 7,264/863

CS

CS 9,049/863

CS G5 G6 Bovine (non-vacc.) Sheep/goats (non-vacc.) Pigs (non-vacc.)

CS 1,213/140

CS

CS CS CS

• Non-vaccinated bovine and pigs:

o All groups agree on clinical surveillance only • Non-vaccinated sheep and goats:

o Group 6 proposes clinical surveillance only o Groups 1, 2, 3, 4 and 5 follow EU Directive 2003/85/EC

• The in advance calculation of necessary lab resources and vet teams to execute the survey was very useful.

• Calculation of expected positive herds and expected positives per herd was done by groups 3, 4, 5 and 6

• Sometimes different sample sizes obtained with the same procedure: due to different Test Sp and Se used to calculate the sample sizes.


Conclusions Despite the use of different working groups/countries, the approaches taken showed a clear degree of similarity. The results were extensively discussed and following conclusions were made: 1. The vaccination-to-live policy with subsequently substantiating freedom from

infection by a survey system including NSP testing is a realistic and achievable option in FMD control. However, it must be clear that in order to regain a free status without vaccination within a reasonable time, i.e. in accordance with Article 2.2.10.7 paragraph (1)(c) of the Terrestrial Animal Code, a stamping out policy, as defined by OIE, remains part of the control policy such as in the beginning of an epidemic.

2. NSP assays are not sensitive enough to detect all infected animals (especially carriers)

in a vaccinated population and therefore conclusions on the status of the herds can only be on a herd basis and in combination of results from clinical and serological surveys and epidemiological investigations such as cluster analysis.

3. Demonstrating absence of infection, in particular in a vaccinated population, is

impossible and therefore the term ‘demonstrate absence’ should be replaced by ‘substantiate absence’.

4. The current EU Directive (2003/85/EC) for the control of FMD mentions two

surveys: (a) A first survey, for detecting the presence of FMD virus in the vaccination zone

(Article 56), should be a combination of clinical, epidemiological and serological investigations with high overall system sensitivity (Martin et al., 20071). Article 56 takes into account that the vaccination status within a vaccination zone may not be homogeneous, i.e. that within the declared vaccination zone there might be vaccinated and non-vaccinated animals. Consequently this survey includes: • a survey of non-vaccinated animals for detecting FMD virus infection in the

same manner as in a protection zone without vaccination; • a serosurveillance of all herds with vaccinated animals for detecting FMD

virus infection by use of NSP tests. Within the herds all vaccinated ruminants and their non-vaccinated off-spring must be sampled and other species kept in large numbers making individual testing impractical shall be sampled based on a 5% prevalence with 95% confidence.

(b) A second survey, to regain the freedom from infection status after emergency vaccination (Article 61), must have a high specificity. This survey might include a second serosurveillance. However, the serosurvey described in Article 56 could serve the purpose of the serosurvey required in Article 61 of the Directive, referring to surveillance guidelines to be laid down and in fact being the OIE Guidelines in Appendix 3.8.7 of the Terrestrial Animal Health Code2.

1 Martin PA, Cameron AR, Greiner M. Demonstrating freedom from disease using multiple complex data sources 1: a new methodology based on scenario trees. PrevVet Med 2007 Jan 12; [Epub ahead of print] 2 In 2003 there were no Guidelines of OIE and it was discussed at OIE not at all to lay down Guidelines. A decision is in preparation to lay down that the OIE Guidelines should be the Guidelines mentioned in


5. The follow-up of herds with seroreactors by serological investigation has to be based

on a combination of NSP assays (Paton et al., 2006) with well-defined performance characteristics and those assays should preferably be conditional independent. The final conclusion must take into account the specificity of the overall ‘test system’.

6. If the specificity of the ‘serological test system’ used were known, then one approach

would be to anticipate the rate of false positive test reactions and only consider seroreactor rates above this expected number or proportion as significant (Herd Cut Point - Paton et al., 20063). However, this is not considered as compatible with the OIE Code and the EU Directive 2003/85/EC, which require all herds with seroreactors to be followed up and classified as either containing or free from infection. Therefore re-sampling and testing herds with seroreactors should be applied. The latter approach would enable active virus circulation to be confirmed or ruled out by use of paired serology to look for changes in antibody status or titer rises and/or by evaluating results using Likelihood Ratios (Dekker et al., submitted4). Contingency plans should include a clear flow chart for the follow-up of seropositive herds.

7. A clinical surveillance combined with paired serology can detect holdings where

virus circulation is ongoing. However, there is no possibility of detecting each and every carrier within sub-clinically infected herds if, as seems likely, they are present in few herds and at a low level. Therefore all ruminants should be tested. Evidence of virus circulation would lead to the declaration of an outbreak and the stamping out of the herd, but evidence of carriers (in particular if at a low number within a herd) should, different to the current requirements, lead to slaughter of these reactor animals only, but not of the whole herd.

The advantages of this approach would be:

(a) The test specificity can be lowered since the consequence of false positive results is now individual animal rather than whole herd slaughter.

(b) This in turn leads to an increased test system sensitivity (c) The “small herd” problem is decreased

(cf. Annex H). 8. Testing all animals in the vaccinated population as prescribed in the EU Directive

2003/85/EC, is certainly the way forward in case of vaccinated bovine, it is however considered as not achievable in areas with a dense pig population or within big pig herds, if such pigs have been vaccinated. In addition is the risk of developing a carrier state in pigs considered to be minimal, if not negligible. It is proposed to replace Article 56(3) (b) of Directive 2003/85/EC by the following: ‘… testing for antibodies against NSP of the FMD virus shall be carried out on

Article 61. The current OIE Guidelines also do not specify the period that must elapse between the last vaccination and the beginning of the survey. 3 Application of non-structural protein antibody tests in substantiating freedom from foot-and-mouth disease virus infection after emergency vaccination of cattle, Paton DJ et al., 2006 4 Comparison of ELISAs for antibodies against foot-and-mouth disease virus non-structural proteins in cattle sera base don the continuous results, Dekker A et al., submitted, see Annex 9 of FMD-NSP program.


samples taken from all vaccinated large ruminants and their non-vaccinated offspring and from all vaccinated pig herds with a within herd sampling based on a 5% prevalence with 95% confidence …’.

9. Vaccination of small herds remains a controversial item. Two possible options were

discussed: (a) A non-vaccination policy for small herds and integration in the survey system

as sentinels. This option is based on the fact that the 95% confidence cannot be achieved in small herds (considering a realistic within herd prevalence) and that small herds slow down the emergency vaccination campaign. Spatial cluster analysis should be applied to evaluate the serological survey results of the sentinels.

(b) Vaccinate small herds and test all animals in all of these small herds. This option is considered because vaccination of small herds contributes to an increase in vaccination coverage and thus to achieving the necessary level of protection for the population, considering the fact that vaccinating all animals with a homologous vaccine will only protect about 90% of the animals. It might also be politically difficult to deny vaccination to owners of small herds and thus exposing the animals to infection and possible stamping out.

10. This kind of workshop should also be done for other veterinary diseases such as

Classical Swine Fever or Avian Influenza (for all EU members, EUFMD countries and EU neighbours).


Recommendations 1. Conclusions on the infection status of the herds after FMD outbreaks in a vaccinated

population should only be based on a survey system including at least clinical, serological and epidemiological investigations;

2. The performance characteristics of the survey system should be determined. When the desired confidence level cannot be reached, spatial clusters analysis should be considered;

3. The term ‘demonstrate absence of infection’ should be replaced by ‘substantiate absence of infection’;

4. Contingency plans should include a clear flow chart for the follow-up of seropositive herds, that must take into account at least the requirements of Appendix 3.8.7. of the Terrestrial Animal Health Code. Serological investigation should be based on a combination of NSP assays with well-defined performance characteristics;

5. All large ruminants should be tested to substantiate freedom from infection in a vaccinated population after FMD outbreaks. While evidence of virus circulation must lead to the declaration of an outbreak, consensus should be sought on the slaughter of reactor animals only, in case there should be evidence that these animals are carriers, instead of the whole herd removal as currently required in Article 57(3). Consideration should be given to finding out if there would be a consensus on the latter;

6. A change in the definition of an outbreak in OIE Guidelines and EU Directives is needed where carriers are concerned;

7. The relative confidence attainable with “herd-based” and “individual” certification needs to be explored for different herd sizes and prevalence;

8. Consideration should be given to an amendment of the Directive in order to allow a within-herd sampling scheme based on a 5% prevalence and a 95% confidence for vaccinated pigs;

9. The vaccination of small herds should be further discussed;

10. To refine the application of NSP tests, more work could be done in predicting the expected prevalence of infection within and amongst vaccinated herds;

11. Functional FMD expert groups should be created in every country (cf. article 78 of Council Directive 2003/85/EC for the EU Member States5), including in non-EU countries.

5 In article 78 it is stated that the group shall compose of epidemiologists, veterinary scientists and virologists in a balanced way


Observations made by FAO EUFMD, Dr. Tom Murray, APO FAO EUFMD I emphasise that this workshop, because of the unique mix of researchers, epidemiologists and decision makers presented an excellent opportunity for progress in the use of serosurveillance post vaccination to demonstrate disease freedom. It was also highlighted that such workshops were important for initiating and developing the necessary cross border relationships during “peace-time”. Future outbreaks of FMD are likely to cross international boundaries and such relationships would help cooperation and coordination. I also encourage members of participating countries to use the many available websites now available, including that of Epizone, The EUFMD Commission, Coordination Action FMD CSF and Improcon describing the latest developments in FMD research. The advantages of real-time alert exercises which would test the complete vaccination process from the decision to vaccinate to demonstration of freedom was mentioned. I also want to highlight the need to use all sources of evidence to assist in the substantiation of freedom from disease. If all available sources of evidence are factored in, then this would help deal with the complex problem of small numbers of test positive animals. Observations made by FAO EUFMD, Dr. Keith Sumption, FAO EUFMD Dr. Keith Sumption noticed that we have come a long way since the development of the VIA test 40 years ago to the NSP tests which are now available. He highlighted the importance of this workshop for the application FMD-NSP tests in emergency vaccination and stressed the necessity to organise part B (Balkan countries) and C (Northern European countries) of the workshop. For him the thing to bring home from this workshop is the shift in thinking of automatically slaughtering a whole herd when a carrier animal is detected to individual slaughter of the carrier, based on risk analysis.


Observations made by OIE, Dr. Gideon Bruckner 1. Found the workshop very stimulating and worthwhile (and excellently organized)! 2. Many of the difficulties experienced and questions that came forward in the exercise

also regularly are heard when evaluating country applications for freedom from disease. The workshop was therefore also very relevant for OIE!

3. Clear indication that areas that the OIE need to re-assess, to determine if current Code

requirements give sufficient guidelines to countries are: • Harmonization of follow-up tests in the event of positive results at initial

screening (both in vaccinated and unvaccinated herds); • Does the Code give sufficient guidance on the interpretation or harmonization

on the interpretation of NSP tests? • Guidance in the decision-making process in applying the findings of test

results; • Guidance on survey design e.g. if statistical significance cannot be reached

with the sampling design – what then? 4. About 120/167 OIE Member Countries are from developing and in-transition

countries. The delegates at this workshop were all demonstrating high expertise in the subject matter. There is a worry and urgent need to do similar exercises in these developing countries to at least put them on the right pathway with decision-making. Perhaps a recommendation in this regard could be made on how these developing countries could be helped to come on board with the requirements of decision-making in FMD control.


Acknowledgments

DG RESEARCH

DG SANCO

VAR Belgium Food Agency Belgium

EUFMD


Annex A see FMD-NSP workshop program


Annex B

Annex BGroup 1 – Scenario 1

Spain & Ireland

Summary of scenario 1

CLUSTER 1 6IPs ; all culled within 500m• Vaccination of cattle 500m – 3km• Surveillance area surrounding vacc. zone

CLUSTER 2 2IPs• Cattle culled 0-500m; vaccinated 500m – 1.5km• sheep/goats & pigs culled 0 – 1.5km• NO non-vaccinated animals in vacc. zone• PZ (3km) and SZ (10km) surrounding vacc. zone


Vaccination Zone –serosurveillance sampling options

OPTION 1 comply with 2003/85/EC

• ALL herds;

• ALL animals in vaccinated herds;

• Random sample of non-vaccinated herds (95:5); 60

OPTION 2 alternative compromise

• ONLY Herds ≥ 5 animals;

• Random sample of vaccinated herds (95:2); 150

• Random sample of non-vaccinated herds (95:5); 60

Option 1

Cluster 1 Herds Samples

VaccinatedCattle 151 6856

Non-vaccinatedPigs 14 840Sheep/Goats 91 954

Cluster 2



Option 2

Cluster 1 Herds Samples


Non-vaccinatedPigs 14 840Sheep/Goats 49 838

Cluster 2


Initial laboratory results

CATTLE 97/151 herds; ≤ 150 sera/herd; 5675 sera

• 1/97 herds with 1 NSP+ bovine; 1/5675 NSP+ sera

• NSP+ from a sample of 150 (herd size = 611)

SHEEP/GOATS 49/91 herds; ≤ 60 sera/herd; 838 sera

• 2/49 herds with 1 NSP+ sheep each; 2/838 NSP+ sera

• 1/60 (herd size = 280) and 1/ 19

PIGS All 14 herds; 60 sera/herd; 840 sera

• 4/14 herds with 1 NSP+ pig; 4/840 NSP+ sera


Interpretation & follow-up

Analysis of test results:any evidence of clustering of NSP+s; magnitude of NSP+s

Laboratory follow-upNSP+ cattle:apply NSPE2 (± IB if available)Results: ALL NSP+ are retest NEGATIVENSP+ non-vaccinates:apply SPCE/VNT (± NSPE2)Results: 2 NSP+ pigs are retest POSITIVE; both single

reactors from a sample of large herdsField follow-up

Field follow-up

Revisit herds with NSP+(s):clinical inspction of ALL animals and EPI investigation

Resample NSP+ animals(if identifiable!!)

Collect serum ONLY & for God’s sake leave your probang at home!

Sample ALL* herdmates(2003/85/EC*)

Sample contact herds?? NOT YET!


Surveillance “Area” around VZ

2003/85/EC only prescribes serosurveillance in sheep and goats; ONLY clinical surveillance is required for cattle and pigs

CLUSTER 1CATTLE 884 48,455

PIGS 137 81,448

SHEEP/GOATS 523 6,196

95:2 HERDS; 95:5 149 1,388

Cluster 1 herds Percentage Number herds sampled Number animals sampled1 to 4 211 0.403441683 60 1805 to 10 195 0.372848948 56 389

11 to 25 87 0.166347992 25 39726 to 50 14 0.026768642 4 14451 to 100 7 0.013384321 2 116

101 to 250 6 0.011472275 2 103251 to 500 3 0.005736138 1 60

total 523 1 149 1388


Cluster 2 herds Percentage Number herds sampled Number animals sampled1 to 4 138 0.474226804 71 2135 to 10 111 0.381443299 57 39911 to 25 32 0.109965636 16 25626 to 50 7 0.024054983 4 12851 to 100 1 0.003436426 1 60101 to 250 1 0.003436426 1 60251 to 500 1 0.003436426 1 60total 291 1 149 1176

Surveillance “Area” around VZCLUSTER 2

PZ (3 km)CATTLE 76 9,809

PIGS 32 36,247

SHEEP/GOATS 49 486

ALL HERDS; 95:5 49 458

SZ (10 km)CATTLE 596 53,102

PIGS 208 216,977

SHEEP/GOATS 291 2,367

95:2 HERDS; 95:5 149 1,176


RESOURCESVACCINATION ZONE

8812 VET TEAMS(5 herds x 5 days)

127201013712967SAMPLES

191191289HERDS VISITED

OPTION 3>4 animalsAll animals

OPTION 2>4 animals

95-2

OPTION 1EU

DIRECTIVE

Cluster 1 andCluster 2

RESOURCES

• Protection Zone: Annex III 2.3 (95/5 s&g)– Cluster 1: VZ=PZ– Cluster 2: 49 herds s&g 486 animals (2 vet teams)

• Surveillance Zone: Annex III 2.4 (95/2 herds, 95/5 s&g)– Cluster 1: 149 herds 1388 animals– Cluster 2: 149 herds 1176 animals– 12 vet teams


• Packaging and transport of samples (special transport?)

• Laboratory testing (kits, other material, personnel, etc.)

• Follow-up resources

OTHER RESOURCES

CONCLUSIONS

• Option 2 and Option 3 need similar resources with higher confidence in Option 3

• Option 3 could provide enough confidence and it needs less Vet Teams than required by the EU Directive


Annex C

Annex CGroup 2

Scenario 1

Scenario 1

• Scenario 1 focuses on clusters 1 and 2 taking into account the following control strategy:

• Culling:• cluster 1: culling of all susceptible animals (cattle, small ruminants

and pigs) in a 500 m radius;• cluster 2: culling of all pigs and small ruminants in a 1.500 m radius

and of all cattle in a 500 m radius;• Vaccination:• cluster 1: all cattle in a 3.000 m radius around the outbreaks (from

500 m to 3.000 m around the outbreaks);• cluster 2: all cattle in a 1.500 m radius around the outbreaks (from

500 m to 1.500 m around the outbreaks);• size of the vaccination area: 162 km2.


CLUSTER 1

500 m ZONETOTAL STAMPING OUT

500-3000 m VACCINATION ZONEBOVINE ALL HERDS VACCINATED (151)ALL HERDS AND ALL BOVINE TESTED FOR NSPPIGS HERD(14) CLINICAL SURVEILLANCE ALL HERD ALL ANIMALSHEEP AND GOATS (91) ALL FLOCKS TESTED NUMBER OF ANIMAL TESTED (PREV 5% 95% CONF) Ceditest

3000-10.000 m SURVEILLANCE ZONEBOVINE HERDS AND PIG HERDS - CLINICAL SURVEILLANCE IN ALL HERDS – ALL

ANIMALSSHEEP FLOCKS SEROSURVEILLANCENUMBER OF HERDS (PREV 2% 95% CONF) NUMBER OF ANIMALS (PREV 5% -95% CONF) Ceditest

CLUSTER 2

500 M ZONETOTAL STAMPING OUT

1500 M ZONEBOVINE ALL HERDS VACCINATED ALL HERDS AND ALL BOVINE TESTED FOR NSPPIGS AND SHEEP HERDS STAMPED OUT

3000 M ZONEBOVINE HERDS AND PIG HERDS - CLINICAL SURVEILLANCE IN ALL HERDS - ALL ANIMALSSHEEP AND GOATS - ALL FLOCKS TESTED NUMBER OF ANIMAL TESTED (PREV 5% 95% CONF) Ceditest

3000-10.000 SURVEILLANCE ZONEBOVINE HERDS AND PIG HERDS - CLINICAL SURVEILLANCE IN ALL HERDS - ALL ANIMALSSHEEP FLOCKS - SEROSURVEILLANCE NUMBER OF HERDS (PREV 2% 95% CONF) NUMBER OF ANIMALS (PREV 5% -95% CONF) Ceditest


14.283148761.299916.856151

1.53710.76170-00-001001 to

8112.43230-007331.4662501 to 1000

43887625428028013351.3404251 to 500

12412414711811811482.52117101 to 250

9090112982247363438751 to 100

-0093401193373731026 to 50

-002061520614163272011 to 25

-002318213277266375 to 10

-001163116422125541 to 4



number of pig

holdings

n of samles

average number

of sheep/g

oat

total number

of sheep/g

oat

number of

sheep/goat

holdings



number of cattle holding

s

size

cluster 1: 3.000 m – N. of holdings and animals to be sampled

131

81.4481371.2071316.19652348.455884

1.64347.6462900-001.1202.23921001 to

73120.4792800-0069313.85520501 to 1000

3648.375235513551.06633835.36314251 to 500

1674.17825982148890615012.18281101 to 250

764586762584067738.59511751 to 100

36214612443650214362.4936926 to 50

1545335222161.38887161.97512211 to 25

72643434971.35119571.1111605 to 10

2271315953359321126422991 to 4

average

number of pigs


number of pig

holdings

n of samples

n of holding

s sampled

average number

of sheep/g

oat

total number

of sheep/g

oat

number of

sheep/goat

holdings




s

size



17.19814140140234.31733

2.12712.76460-00-001001 to

7483.74150-006922.0773501 to 1000

34268420-00-00251 to 500

-000-001331.46711101 to 250

-000-0070489751 to 100

-000-0042208526 to 50

-0064136452244211 to 25

99155655992635 to 10

-002122193621 to 4



number of pig

holdings

n of samples

average number

of sheep/g

oat

total number

of sheep/g

oat

number of

sheep/goat

holdings




s

size


36.24732415486499.80976

1.89428.403150-001.5291.52911001 to

7325.85680-006752.7004501 to 1000

3431.71350-004004001251 to 500

22022010-001483.10121101 to 250

-0084741482761.5902151 to 100

-00313838136214626 to 50

18553123151238182371311 to 25

-0012971291972235 to 10

-00483481931661 to 4



number of pig

holdings

n of samples

average number

of sheep/g

oat

total number

of sheep/g

oat

number of

sheep/goat

holdings




s

size

cluster 2: 3000 m – N. of holdings and animals to be sampled


117

216.9772089121192.36729153.102596

1.827162.6148900-001.2081.20811001 to

71338.5285400-006437.07611501 to 1000

39210.1862655134434413565.34715251 to 500

1885.07827471116116114725.303172101 to 250

88353441175751759.98213351 to 100

401995813322237372.5066826 to 50

-00208131650532171.0596111 to 25

91823154577451117439645 to 10

1111655533591383182711 to 4

average

number of pigs


number of pig

holdings

n of samples

n of holding

s sampled

average number

of sheep/g

oat

total number

of sheep/g

oat

number of

sheep/goat

holdings




s

size


Sampling scheme summary

3.55041311.173184

Total sheep sampledTotal sheep holdings

sampledTotal of cattle

sampledTotal cattle holdings

sampled


sheep210334410000,0010171014cluster 2

sheep16411610000,0010112913cluster 2

cattle11111111500,0010003006cluster 2

cattle786168611500,0010004789cluster 2

sheep1252524910000,0020049772cluster 1

cattle66116113000,0010115793cluster 1

sheep119193000,0070078307cluster 1

sheep21002803000,0070088787cluster 1

positive species

number of positive samples

sampledsheep/goatpigscattlezoneidcluster

Results

Strategy to deal with positives results:

• All the positive farms have to be re-tested– Positive Vaccinated cattle:

• Serological Re-testing of the head• Probang test of the positive head• In one farm un-vaccineted sheep are present:

serological test of the sheep

– Positive un-vaccinated sheep:• Re-testing of the farm (P: 5%, I.C. 95%)• In one farm cattle are present: clinical exame


Annex D

Annex D

Cluster 1: 151 cattle herds with 6,856 cattle91 sheep and goat holdings with 1,299 animals14 pig farms with 14,283 pigs

Cluster 2: 33 cattle herds with 4,317 cattle23 sheep and goat holdings with 140 animals14 pig farms with 17,198 pigs

In total 326 herds and 44,093 animals has to be vaccinated

Risk reduction should be same in non-vaccinated and vaccinated animals.In all scenarios the calculation of the sample size based on the minimum number samples, which has to be taken. The sampling unit is always the epidemiological unit (compartment) on the farm. Using a worst case scenario the number of samples should be tripled for pig farms.

Sampling of all cattle and sheep, but pigs are sampled to detect 5 % prevalence with 95 % confidence

no carriers in pigs14,241 animals have to be sampled on 326 herds and may produce 114 (13 pigs) false positive results (sampling fraction = 0.32)In worst case (three epidemiological units per pig farm) 17 499 animals have to be sampled resulting in approximately 140 false positive results (39 false positives on pig farms).


Surveillance ZoneThe screening of the surveillance zone does not influence the results of the elaboration of the alternatives in the vaccination areas.The clinical inspection of all cattle and pigs herds will be performed within the surveillance zone.All 863 sheep and goat holdings with 9,049 animals will be sampled with a maximum sample size of 60 (detection of 5 % with 95 % confidence). Altogether 7,264 animals have to be sampled (sampling fraction = 0.8).

vaccinated95%sheep119193000cluster 1

Insufficient numbersampled

vaccinated97%cattle2602843000cluster 1

Insufficient numbersampled

vaccinated97%

cattle2608553000cluster 1

vaccinated96%

pig+cattle

1+360+51

951511500cluster 2

Non-vaccinated96%sheep1252524910000cluster 1



percentage

negativesamples

positivespecies

number of positi

vesampl

es

sampled

sheep/goat

pigscattlezonecluster

Results

The number of positive farms is much lower than expected


Action in surveillance zone

• Non-vaccinated test initial sera in a SP test; if they are positive then

• Resample farm with 5% prevalence and 95% confidence

• Clinical inspection is rather difficult in sheep, but we should check carefully. Also check very carefully for epidemiological link to known outbreak.

• Revisit the farms as soon as possible.

Initial testing vaccination area all ruminants and pigs 5% 95%

pos neg

Rebleed positive animals at least 4 days laterSame animalpositive again

neg

Remove positive animals

Rebleed in ruminants whole herd at least 4 days later,for pigs penmates and adjacent pens (looking for seroconversion)

pos neg

? Outbreak

Exclude evidence of virus circulation, for ruminants in addition exclude presence of carriers

Rebleed positive animals at least 4 days later

Same animalpositive again

neg

In all cases clustering of positive samples indicates virus circulation then the CVO candecide to call it an outbreak in an earlier stage


Annex E

Annex E:Design & Interpretation of

post-FMD vaccination serosurveillance by NSP tests

Scenario 2Group 4

Group 4

• Belgium

Jef HooyberghsGerard Lamsens

Marc Dispas

Nesya Goris(rapporteur)

• France

Yves LeforbanBenoit Durand

François Moutou

Stéphan Zientara

(chairman)


Scenario 2

• Cluster 1

Culling: all animals in 500 m radius

Vaccination: all animals in 3000 m radius

• Cluster 2

Culling: all animals in 500 m radius

Vaccination: all animals in 1500 m radius

General information

• Cluster 3 non-existing in scenario 2• Sampling 1 month post last outbreak and

vaccination (+/- January 15th)• Availability kits: not a problem; beyond 3

week period (200 000 tests per week)• Animal population size considerable → not

according to Directive 2003/85/EC• Stratified per species


Surveillance zone

• Cluster 1 (between 3000 and 10000 m radius)– 48455 cattle– 6196 sheep/goat– 81448 pigs

• Cluster 2 (between 1500 and 10000 m radius)– 62911 cattle– 2853 sheep/goat– 253224 pigs

• Cattle & pigs: clinical surveillance• Sheep & goats:

– Serological surveillance– Most sensitive test applied: SPC ELISA– Sn = 100%, Sp = 99.5% (Paiba et al. 2004)– All animals, all herds (n = 9049)

Vaccination zone

• Serosurveillance based on NSP test using the commercially available Ceditest

• Characteristics based on Brocchi et al. 2006• Cattle

– Sn = 63.6 %– Sp = 99.5%

• Pigs– Sn = 55.5 %– Sp = 100%


Vaccination zone

• Cluster 1Sheep & goat:

all animals, all herds(n = 1299)

Cattle & pigs:all herds 5% prevalence95% confidenceFreeCalc software

• Cluster 2Sheep & goat:

all animals, all herds(n = 140)

Cattle & pigs:all herds 5% prevalence95% confidenceFreeCalc software

Cluster 1: Cattle and Pigssize # cattle holdings total # animals sample size

1 to 4 54 125 all5 to 10 37 266 all11 to 25 20 327 all26 to 50 10 373 all51 to 100 7 438 all101 to 250 17 2521 142251 to 500 4 1340 153501 to 1000 2 1466 1521001 to … 0 0 0

6856 4859

size # pigs holdings total # animals sample size1 to 4 0 0 05 to 10 0 0 011 to 25 0 0 026 to 50 0 0 051 to 100 1 90 86101 to 250 1 124 88251 to 500 2 876 100501 to 1000 3 2432 1021001 to … 7 10761 105

14283 1415


Cluster 2: Cattle and Pigssize # cattle holdings total # animals sample size

1 to 4 2 6 all5 to 10 3 26 all11 to 25 2 44 all26 to 50 5 208 all51 to 100 7 489 all101 to 250 11 1467 129251 to 500 0 0 0501 to 1000 3 2077 1611001 to … 0 0 0

4317 2675size # pigs holdings total # animals sample size

1 to 4 0 0 05 to 10 1 9 all11 to 25 0 0 026 to 50 0 0 051 to 100 0 0 0101 to 250 0 0 0251 to 500 2 684 99501 to 1000 5 3741 1041001 to … 6 12764 106

17198 1363

Laboratory results

In total

Surveillance zone: 9049 sheep/goat samples tested (SPCE)45 false positive results expected

Vaccination zone:11751 samples tested (Cedi)38 false positive cattle results expected“0” false positive pigs results expected


Laboratory results

cluster id zone cattle pigs sheep/goat samplednumber of

positive samples

positive species

cluster 2 10112913 10000 116 116 3 sheep

cluster 2 10171014 10000 344 344 8 sheep

cluster 1 20049772 10000 249 25 25 1 sheep

cluster 2 10003154 1500 51 951 227+51 3+1 pig+cattle

cluster 1 10174528 3000 855 164 2 cattle

cluster 1 10081457 3000 284 154 2 cattle

Follow-up Surveillance zone

• 3 positive herds• 12 SPCE positive animals (<< 45)


Follow-up vaccination zone

• 3 positive herds• 3 positive pigs & 5 positive cattle• Retest positives using the:

Cedi/Cedi retest/Svanova retest system– Sn = 66.7% for cattle

– Sp = 99.99% for cattle

– Paton et al. 2006

Follow-up vaccination zone


Conclusions and recommendations1. Necessity to rediscuss Directive 2003/85/EC (Art. 56):

testing all vaccinated animals (pigs)?2. What to do with NSP positives? Exclude virus circulation by

cluster analysis (risk assessment of NSP positive herds, or virus detection) and culling of positive herds? Alternative measures to culling herds (only culling of positive animals)?

3. Different serological tests for different zones (SP tests in surveillance zone and NSP as confirmatory test; NSP tests in vaccination zone)

4. Continuation of NSP test validation for pigs, sheep and goats in experimental and field settings

5. Improvement of existing NSP tests (Sn and Sp) and development of confirmatory assays?

Message for CVO

• Interest of such exercise• Still open issues• Vaccination strategy is good, but complex

option (follow-up is harder, need for furtherharmonisation of interpretation of NSP positives)

• Continued support by EU of FMD Research needed


Annex F

Annex FGroup 5: Slovenia / UK

Cluster 3

Vaccination Zone

• Pigs: Clinical surveillance only

• Sheep: Clinical plus SP serosurvey

• Cattle: Clinical plus NSP serosurvey


Vaccination Zone – Sheep SP Serosurvey

• The need to test all sheep flocks is difficult to assess without a map showing location of all IPsand associated zones.

• Decided on a serosurvey of all sheep flocks to detect 5% within herd prevalence of infection at 95% confidence.

• Using SPCE ELISA for screening and NSP ELISA or VNT for confirmation (assuming 100% Se and Sp).

Vaccination Zone – Sheep SP Serosurvey

Sampling:

93891365

571571001-

400850101-250

697174151-100

1178383126-50

2433158All11-25

2675395All5-10

1949748All1-4

Total

samples

HoldingsMean

Sample sizeFlock size


Vaccination Zone – Cattle NSP Serosurvey

• Clinical surveillance plus NSP serosurvey

• Using Cedi/Cedi/Svanova test system (assuming 66.7% Se and 99.99% Sp)

• Herds with less than or equal to 50 cattle: sample all cattle in all herds and analyse as spatial clusters

• Herds with more than 50 cattle: sample all herds and sufficient animals to detect a 5% within herd prevalence with 95% confidence

Vaccination Zone – Alternative Cattle NSP Serosurvey to reduce sampling

• Small herd sampling unchanged (could have increased Se and reduced Sp of test system)

• Herds with more than 50 cattle– Sample sufficient herds to detect a 2% prevalence of infection

with 95% confidence - 138 herds to be sampled.

– Better to ake account of non-homogeneous spread characteristics by: • 1% prevalence rate – requires 258 herds to be sampled

• Subdivide region into three geographical areas containing approximately equal numbers of cattle herds and test within eacharea so as to detect a 2% prevalence of infection with 95% confidence. This would require 3 x 121 herds to be sampled.

– As before, sample sufficient animals to detect a 5% within-herd prevalence of infection with 95% confidence.


Vaccination Zone –

Alternative Cattle NSP Serosurvey

26823124534770136289713218

8

Total

92211411716825872111108

4

Subtota

l

---1001-

841841336484501-1000

46468681136454581251-500

44376282961123485447174101-250

4236727920134332765645951-100

17602110417602110417602110

4

Subtota

l

105992781059927810599278All26-50

466328046632804663280All11-25

159421515942151594215All5-10

746331746331746331All1-4

Total samples (all small + 5% prev in large herds)

Holdings sampled (all small + 2% prev of large herds)

Total samples (all small + 5% prev in large herds)

Holdings sampled (all small + 1% prev of large herds)

Total samples (all small + 5% previn all large herds)

Holdings

Mean Sample size (for 95/5 with Se 0.67

Herd size

NB Total cattle = 144,872

Buffer Zone

• Not sure of need to sample

• If to be done:– Pigs / Cattle – clinical only

– Sheep:• Clinical surveillance plus SP serosurvey

• 691/ 885 flocks have sheep and goats only – these would be the priority.

• 134 flocks randomly selected gives 95% confidence to detect 2% prevalence by flock. Then add seven largest flocks.

• Within flocks, survey to detect 5% within herd prevalence of infection at 95% confidence.

• Using SPCE ELISA for screening and NSP ELISA or VNT for confirmation (assuming 100% Se and Sp).


Buffer Zone – Sheep SP Serosurvey

1213140690

1002*250101-250

2055*54151-100

76283126-50

3421999All11-25

31542218All5-10

17570358All1-4

Total

samples

Holdings

sampled

Total

Holdings

with only

sheep

and

goats

Mean

Sample

size

Flock size

* targeted

Outputs Needed

• Liked to have seen all IPs, PZ, SZ and vaccination zones drawn out on a map to properly consider whether or not all sheep flocks in the vaccination zone needed to be sampled.

• Spatial distribution of all cattle herds subdivided into two herd sizes (greater or smaller than 50 cattle)

• Qualitative and quantitative results for all positive test results

• All serology results by herd or flock size• All positive herds or flocks – distance to nearest infected

premises and date that IP was confirmed • All positive herds or flocks – distance to one another

(scale map?)


Results

cluster id zone cattle pigssheep/goat

sampled

number of

positive samples

positive species

cluster 3 30051155 vacc 3 1200 60 1 sheep at 6K from border of vaccination zone W and 1.5K to nearest outbreakcluster 3 31100214 vacc 114 60 1 sheep at 5K from border of vaccination zone S and 6K to nearest outbreakcluster 3 40170942 vacc 36 36 1 sheep at 2K from border of vaccination zone N and 5K to nearest outbreakcluster 3 40033902 vacc 36 25 25 1 sheep in middle of zone and 0.6K to nearest outbreakcluster 3 40014959 vacc 171 60 1 cattle at 1K from border of vaccination zone E and 4K to nearest outbreakcluster 3 40013413 vacc 227 60 1 cattle at 2K from border of vaccination zone NE and 4K to nearest outbreakcluster 3 40024547 vacc 477 60 1 cattle in middle of zone and 1K to nearest outbreak

Interpretation

• Overall results in line with expected false positive rates (or better!):

– Sheep: expected 5, got 4 singletons

– Cattle: expected 9, got 3 singletons

• No clustering evident, except perhaps one sheep and cattle farm in middle

• Small cattle herds – all negative, but uncertain attainment of desired confidence


Further sampling / testing• Revisit and check all farms with seropositives• Sheep:

– Cull (and resample / test singleton) positives (n=4)– If positive, retest same cohort as before to determine if positives

at 95/5– And test rest of flock to estimate the prevalence of seropositives

assuming true prevalence is 10% +/- 5% (all from 2 small flocks, 39 of 54 (114), 123 of 1140 (1200))

– No probangs – Classify flocks based on results

• Cattle:– Cull (and resample / test singleton) positives (n=3)– If positive, resample cohort and rest of herd to estimate

prevalence at 95% confidence assuming true prevalence is 1% +/- 0.5%

– No probangs– Classify herds based on results

• No slaughter if herds classified as uninfected


Annex G

Annex GCluster 3

• vaccination area surrounding some 20 outbreaks following movement of infected sheep

• size = 770 km2

• density: – cattle = medium to high density (188 / km2)– pigs = high density (1.187 / km2)– sheep/goat = low density (15 / km2)

• includes buffer area of 5 km (670 km2) around vaccination area with slightly lesser density

scenario 3

• deals only with cluster 3

• culling of outbreaks only

• vaccination of cattle only in vaccination area:

• groups:

– UK + SL

– DK + LI


Survey design according to directive – cluster 3- vacc area

• 1365 Sheep flocks - 11.780 sheep– Unvaccinated

– Solid Phase ELISA

– Sensitivity: 99 or 100

– Specificity: 99

– 1-50 sheep: all sheep sampled (conf. not achieved)

– 51-100 sheep: 51 - 77 samples

– 101 – 1200 sheep: 77 – 125 samples

– Appr. 10.000 samples



• 2188 cattle farms – 144.872 cattle– Vaccinated

– NSP Ceditest/Ceditest

– Sensitivity: 81.8

– Specificity: 99.2

– 144.872 samples


• 1075 pig farms – 914.155 pigs– Clinical surveillance

– 1075 herd visits


Survey design sampling cattle 95% / 2% – cluster 3- vacc area

• 2188 cattle farms – 144.872 cattle– Vaccinated– NSP Ceditest/Ceditest– Sensitivity: 81.8– Specificity: 99.2

• The desired confidence could not be achieved!

Survey design sampling cattle 95% / 5% – cluster 3- vacc area

• 2188 cattle farms – 144.872 cattle– Vaccinated

– NSP Ceditest/Ceditest

– Sensitivity: 81.8

– Specificity: 99.2

– Approximately 110.000 samples according to herd size table

– Desired level of confidence could not be reached below 90 cattle on the farm


Summary – design choice

• Pigs in vaccination zone– Clinical surveillance

• Sheep flocks in vaccination zone – 95/5 sampling - Solid Phase ELISA– Appr. 10.000 samples

• Cattle farms in vaccination zone – 95/5 sampling - NSP Ceditest/Ceditest– Appr. 110.000 samples

• Cattle farms in vaccination zone – All cattle - NSP Ceditest/Ceditest– Appr. 144.872 samples

Buffer zone intensified surveillance

• Campaign for increased awareness for clinical signs of disease

• Pigs in buffer zone (n=677 holdings)– Early warning: increase in piglet mortality by x%

• Cattle farms in buffer zone (n=1372 holdings)– Early warning: milkdrop

• Sheep flocks in buffer zone (n=885 holdings)– Nothing special (only small herds)


Sheep serology cluster 3 afterSolid Phase ELISA- Solid Phase ELISA

3125120030001

12525360004

136360003

2771140002

positivesSamples No of sheep/goats

No of cattle

Herd-id

Cattle serology cluster 3 afterCedi - Cedi (NSP ELISA)

303Total

-01001 -

14501 – 1000

154251 – 500

1211101 – 250

13471 – 100

-051-70

-00-50

No of positive per herd

No of positive holdings

Herd size cat.


Cattle serology cluster 3 afterCedi - Cedi- Svanova

3Total

-01001 -

-0501 – 1000

11251 – 500

12101 – 250

-071 – 100

-051-70

-00-50

No of positive per herd

No of positive holdings

Herd size cat.

False positives?• Sheep (7 positive samples in 4 herds)

– 3/125 pos, 2/77 pos, 1/36 pos, 1/25 pos– Retesting with Svanova test– No positives

• Vaccinated cattle – 303 positive samples in 303 herds– Retesting with Svanova test– 3 positive samples in 3 farms– If epidemiological link - article 57– No epidemiological link – false positive

• Cedi – Cedi: In all cases the number of positive samples is below the herd cut point


Ressources for screening in the vaccination zone

• Team of 2 for bleeding + clin.:– 5 farms per day– 500 samples per day– 155.000 samples in 3.553 cattle and sheep herds– 700 teamdays – 35 teams=3 weeks

• Team of 2 for clinical visits in pig farms– 1075 pig farms– 3 farms per day – 360 teamdays – 15 teams = 3.5 weeks

Ressources for intensifiedsurveillance in the buffer zone

• Number of farms– Buffer zone: 677 pig farms– Buffer zone: 1372 cattle farms– Buffer zone 885 sheep farms– 2934 farms in total

• Early warning obligation– milk drop, piglet mortality– Follow up by vet visit


Lab ressources

• 155.000 NSP test Cedi-Cedi

• 310 Sanova tests

• 3 weeks

• 12 technicians

• 2 academic staff


Annex H Post NSP Workshop thoughts The approach to post-vaccination serosurveillance set out in the EU Directive assumes that serological tests for virus infected animals will have an insufficient sensitivity to certify individual animals as infection-free, but that they could be used for herd-based certification. Accordingly, it is required that all vaccinated animals in all vaccinated herds or flocks should be sampled and tested serologically and that if infection cannot be ruled out, then the entire herds must be slaughtered and animals either disposed of into the food chain (in cases where it is considered that the probability of genuine infection is extremely low) or by destruction (in other cases). Available tests can be used to support this strategy (Paton et al., 2006) subject to certain limitations, namely that:

1. The need to slaughter entire herds where seroreactors are found makes it essential that the test system used has a very high specificity which in turn reduces the sensitivity.

2. The strategy will still lead to slaughter of some seroreactor herds of uncertain infection status.

3. The low test system sensitivity means that a low prevalence of infection cannot be reliably detected and that small herds cannot be certified as infection free, even if they have a moderate (5%) prevalence of infection.

Another problem is that the strategy espoused by the EU Directive does not distinguish between the two different aims of detecting (1) virus circulation and (2) virus carriers, a concept that has recently been introduced into the OIE Code. Demonstrating absence of virus circulation can be substantiated by paired serology and arguably, is barely necessary if there is a properly conducted campaign of vaccination and clinical surveillance. In contrast, the feasibility of demonstrating the presence of carriers is entirely dependent on their prevalence within the population and poses the main challenge for post-vaccination serosurveillance in countries wishing to substantiate freedom from infection without vaccination, after use of emergency vaccination. There is a lack of certainty about the prevalence of infection (circulating virus or carriers) that is likely to be found after use of emergency vaccination within and amongst vaccinated herds. However, limited experience of outbreaks where emergency vaccination has been used (Balkans and The Netherlands) as well as preliminary findings from modelling studies suggest that the number of infected herds or flocks will be very few and within them the prevalence of infection will also be low – perhaps one or two animals only. This is because either immunity blocks transmission or else, where immunity is insufficient such that extensive infection can occur, then clinical signs are likely to be seen allowing the herd to be diagnosed as an infected holding without serology.


In summary, clinical surveillance combined with paired serology can detect holdings where virus circulation is ongoing. However, there is no possibility of detecting each and every carrier within sub-clinically infected herds if as seems likely, they are present in few herds and at a low level. For example, even assuming a prevalence as high as 5% for carriers within herds, the approach outlined by Paton et al (2006) is only designed to detect carrier containing herds with 95% confidence, which means that 5% of herds containing carriers could expect to be missed. If the prevalence of carriers is so low amongst and between herds, that herd based testing with a relatively insensitive test cannot succeed, than an alternative testing strategy may be one based on individual animal certification. This could be combined with herd-based testing to detect circulating virus, under the assumption that evidence of virus circulation would lead to herd slaughter, but that evidence of carriers (particularly if at a low number within a herd) would lead to slaughter of these reactor animals, but not of the whole herd. This has a number of advantages:

(d) Test specificity can be lowered since the consequence of false positive results is now individual animal rather than whole herd slaughter.

(e) This in turn leads to an increased test system sensitivity (f) There is no longer a “small herd” problem nor a problem of inadequate

specificity in very large herds. For example, in cattle, a test system based on Ceditesting with repeat Ceditesting of seroreactors gives a test system sensitivity of 82% and specificity of 99.2%. If all vaccinated animals in all vaccinated herds were tested, then there is now an 82% probability of detecting all carriers, regardless of prevalence. At a specificity of 99.2%, 80 false positive seroreactors would be expected and have to be wrongly slaughtered for every 10,000 cattle tested. However, the advantage of the system is that the “unnecessary” slaughtering is shared amongst the herds so that each only suffers a little! To investigate this option further:

1. More work is needed to refine models predicting the expected prevalence of infection within and amongst vaccinated herds.

2. The relative confidence attainable with “herd-based” and “individual” certification needs to be explored for different herd sizes and prevalences.

3. The serosurveillance needed to substantiate freedom from virus circulation should be considered.

4. Consideration should be given to finding out if there would be a consensus on not slaughtering individual carrier animals within vaccinated herds where there was no evidence of virus circulation.

David Paton 6th Feb 2007


Annex I Literature a. Brocchi et al., Comparative evaluation of six ELISAs for the detection of antibodies

to the non-structural proteins of foot-and-mouth disease, Vaccine. 2006 Nov 17; 24(47-48):6966-79.

b. Paton et al., Application of non-structural protein antibody tests in substantiating freedom from foot-and-mouth disease virus infection after emergency vaccination of cattle, Vaccine. 2006 Oct 30; 24(42-43):6503-12.

c. Cagienard et al., No evidence of Bluetongue virus in Switzerland, Vet Microbiol. 2006 Aug 25; 116(1-3):13-20.

d. Appendix 3.8.1. of the Terrestrial Animal Health Code: General guidelines for animal health surveillance

e. Appendix 3.8.7. of the Terrestrial Animal Health Code: Guidelines for the surveillance of foot-and-mouth disease

f. Council Directive 2003/85/EC on Community measures for the control of foot-and-mouth disease, Official Journal of the European Union.

g. Paiba et al., Validation of a foot-and-mouth antibody screening solid-phase Competition ELISA (SPCE), J Virol Methods. 2004 Feb;115(2):145-58.

h. Greiner et al., On the surveillance for animal diseases in small herds, Preventive Veterinary Medicine 70 (2005) 223–234

i. Dekker et al., Comparison of ELISAs for antibodies against foot-and-mouth disease virus non-structural proteins in cattle sera based on the continuous results (submitted)

j. Greiner et al., Optimising sampling strategies for substantiating disease freedom in livestock populations (draft version)

k. Martin et al., Demonstrating freedom from disease using multiple complex data sources 1: a new methodology based on scenario trees, Prev Vet Med. 2007 May 16;79(2-4):71-97.

l. Martin et al., Demonstrating freedom from disease using multiple complex data sources 2: Case study – Classical Swine Fever in Denmark, Prev Vet Med. 2007 May 16; 79(2-4):98-115.

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