Foot-And-mouth Disease Vaccine Potency Testing -Determination and Statistical Validation of a Model...

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Vaccine 21 (2003) 32403248

Foot-and-mouth disease vaccine potency testing: determination andstatistical validation of a model using a serological approach

Paul V. Barnett a,, Robert J. Statham a, Wilna Vosloo b, Daniel T. Haydon c

a Institute for Animal Health, Pirbright Laboratory, Ash Road, Pirbright, Woking, Surrey GU24 0NF, UKb ARC-Onderstepoort Veterinary Institute, Exotic Diseases Division, Private Bag X5, Onderstepoort 0110, So uth Africa

c Department of Zoology, University of Guelph, Guelph, Ont., Canada N1G 2W1

Received 7 October 2002; received in revised form 13 February 2003; accepted 11 March 2003

Abstract

European foot-and-mouth disease vaccine manufacturers are required to quantify the efficacy of their product in accordance with the

European Pharmacopoeia (EP). The method used most often to establish the potency of foot-and-mouth disease vaccines requires viral

challenge of vaccinated cattle. Alternative approaches, such as challenge-free serological assessments have many advantages over existing

methods and could be used if robust statistical models could be developed that related antibody titres to protection from challenge. Logistic

regression analysis of data from two independent research laboratories, representing six of the seven main serotypes of FMD, permitted the

parameterisation of these models and indicated that a significant relationship existed between antibody titre and probability of protection.

Furthermore, no significant differences were observed in the parameters of logistic models fitted to different strains within the serotypes

A, O, and SAT-3, or when strains from serotypes A, O, and Asia-1, or SAT-1 and SAT-3, were combined. However, significant differences

in the model parameters did exist between different laboratories. Using these models a bootstrap analysis suggested that for vaccines that

induced consistently high titres, as few as six to eight individual animals could be used to establish with confidence the minimum protective

doses that would protect 50% of vaccinated animals. We conclude that a serologically evaluated truncated test that eliminates the need to

virus challenge cattle is a credible alternative for quantifying vaccine potency.

2003 Elsevier Science Ltd. All rights reserved.

Keywords: Foot-and-mouth disease; Vaccine; Potency; Challenge test; Logistic regression; Antibody titre

1. Introduction

Inactivated foot-and-mouth disease virus (FMDV) is used

in vaccine preparations to control foot-and-mouth disease

(FMD), one of the most economically important diseases

affecting livestock. These vaccines are used in many parts

of the world, particularly where the disease is endemic,

including South America, Africa, the Middle East and the

Far East. In disease free countries, concentrated inactivated

FMD virus antigens are also kept in strategic reserves,

so-called antigen banks, which can be rapidly formulated

into vaccine during an emergency should an outbreak re-

quire additional control measures.

In order to assess the quality of the vaccine, all FMD

vaccine producers are obliged to instigate a series of tests

to establish the safety and efficacy of their product. These

tests include a measurement of potency which in accordance

Corresponding author. Tel.: +44-1483-232441;

fax: +44-1483-232448.

E-mail address: [email protected] (P.V. Barnett).

with the European Pharmacopoiea (EP) requires that vaccine

batches be tested in groups of at least five cattle inoculated

with reduced dose volumes of vaccine so that potency can

be expressed in terms of 50% protective doses [1] (PD50, de-

fined as the factor by which the concentrate may be diluted

such that 50% of vaccinated animals are protected). This

method has practical and logistical problems, and disadvan-

tages from the perspective of animal welfare. For example, in

any extinction point test, approximately 50% of the animals

that are not protected will suffer the painful clinical man-

ifestations of the disease and even some protected animals

may show primary lesions at the site of challenge. Also, only

one valency can be tested in any given trial. Clinically in-

fected animals represent a disease security hazard requiring

expensive high security housing. Finally, given the financial

commitment of purchasing these animals and maintaining

them during the trial period the smallest permitted group

sizes are often used leading to lack of statistical power and

imprecision [2].

However, the European Pharmacopoiea has supported the

use of alternative testing methods provided a correlation

0264-410X/03/$ see front matter 2003 Elsevier Science Ltd. All rights reserved.

doi:10.1016/S0264-410X(03)00219-6


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P.V. Barnett et al. / Vaccine 21 (2003) 32403248 3241

can be established between them and the challenge test.

Serologically-based methods have certain advantages, not

least the relatively insignificant discomfort caused to the

animals as a result of vaccination and subsequent blood sam-

pling. They allow several FMD vaccines to be tested simul-

taneously and/or a number of serotypes to be examined in a

single polyvalent vaccine and can significantly improve theaccuracy of the result. For this reason many research work-

ers developed and evaluated a variety of serological tests and

assessed the correlation of the results with protection. These

have included the virus neutralisation test, plaque reduction

test, the metabolic inhibition test, mouse protection test and

different forms of the enzyme linked immunosorbent assay

(ELISA).

Although some disadvantages have been noted with these

tests, it is generally accepted by FMD research workers that

there is an excellent correlation between the virus neutral-

isation antibody titres of primo-vaccinated cattle and their

protection from virus challenge at 21 days post-vaccination

[36]. Stellman [7] developed this approach further by show-ing how such antibody titres could be used to statistically

assess vaccine and Pay and Parker [8] described a method

for relating log virus neutralisation titres of cattle sera to the

potency of a vaccine in antigen PD50 units. A correlation

was also established between the antigen dose in the vac-

cine, the virus neutralising antibody response and the level

of induced protection [9].

Certainly, there are strong arguments for replacing the

challenge test by in vitro serological assays, however, any

form of test would have to meet certain general criteria. The

test system must be consistent and reproducible, it must be

standardised and include standard reference reagents and pa-rameterised using substantial data sets from which the mod-

els relating titre to protection can be reliably established.

Two methods have primarily been used to calculate the de-

gree of protection in a group of vaccinated animals. Pay

and Hingley [2] and Ahl et al. [10] used mean serum anti-

body titres to calculate a PD50 or a percentage protection.

Bengelsdorff[11] and van Maanen and Terpstra [12] used a

second method in which an individual titre or index was as-

signed to protection and non-protection and the assessment

of a group was calculated from the passed and not-passed

individuals.

In this paper, we have accrued data from two laborato-

ries: The Institute for Animal Health, based at Pirbright in

the United Kingdom and the ARC-Onderstepoort Veterinary

Institute in South Africa, in which cattle challenge tests and

serology have been performed to establish the potency of

many different batches of FMD vaccine. At the Pirbright

Laboratory these tests were performed to either substanti-

ate that a given antigen could be formulated into a high

potency (10 PD50) vaccine for acceptance into the Inter-

national FMD Vaccine Bank[13], or to show stability of the

antigen during ultra-low temperature storage. Similarly, by

using a truncated version of that specified in the OIE man-

ual [14], involving just a single vaccine dilution group, the

laboratory at Onderstepoort performed cattle challenge tests

to ensure that their vaccines had potency values of 8 PD50.

Encompassing six different serotypes, the objective of this

analysis was to:

determine the relationship between virus neutralising titre

and protection;

quantify any variation in this relationship between labora-tories or as a result of the use of different vaccine strains

or serotypes;

provide a model that could be used to calculate the prob-

ability that vaccines achieve potency levels at or in excess

of a pre-specified requirement.

2. Methods

2.1. Data

The data comprised the results of vaccine potency tri-

als conducted in the two laboratories (Pirbright and Onder-

stepoort) on six different serotypes (O, A, Asia-1, SAT-1,

SAT-2, and SAT-3). The data is summarised in Table 1 and

Fig. 1. Animals were vaccinated (except control individuals)

with various dilutions of vaccine of varying valency, and the

neutralising antibody titre (log 10 SN50/100 TCID50) to the

vaccine virus strains estimated from serum samples taken at

21 days post-vaccination using the virus neutralisation test

(VNT). At this time animals were challenged with homolo-

gous virus to the vaccine strain by intradermal inoculation of

10,000 ID50 into the tongue. Animals were observed closely

for the subsequent reading period, normally 810 days, and

the occurrence of generalised lesions on any one or morefeet taken as evidence that the animal was not protected by

the vaccination.

2.2. Analysis

The data for each vaccinated animal thus constitutes: lab-

oratory, serotype, strain, log antibody titre from the VNT,

and protected or unprotected status. Logistic regression im-

plemented within MINITAB was used to determine the sig-

nificance of differences between strains within serotypes,

and between laboratories on the relationship between log an-

tibody titre and protection. The significance of all two-way

interactions was examined and those revealed to be insignif-

icant at the 5% level were omitted from subsequent analysis.

When no significant differences between strains or serotypes

were revealed the data were combined and parameters for a

single model estimated. Appropriate minimal models were

determined, and characterised by their intercepts and slopes

(with accompanying standard errors (S.E.)), the covariance

between the estimated slope and intercept, T50 (defined as

the titre at which animals are protected with probability

50%), the 95% confidence intervals (95% CI) on T50 as de-

termined using Fiellers theorem [15] and T95 (defined as the

titre at which animals are protected with probability 95%).


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3242 P.V. Barnett et al. / Vaccine 21 (2003) 32403248

Table 1

Summary of the data (P: Pirbright, O: Onderstepoort)

Serotype Challenge strains No. of animals challenged No. of animals protected (%) Average neutralising Ab titre

of vaccinated animals

Asia-1 P: India 46 37 (80) 2.44 (40)

O P: Manisa 36 31 (86) 2.34 (32)

P: Lausanne 26 19 (73) 2.36 (24)

A P: Cruzeiro 102 53 (52) 1.67 (92)

P: Thailand 36 32 (88) 2.25 (32)

P: Iraq 26 22 (85) 2.29 (24)

SAT-1 O: Sar/9/81 77 49 (64) 1.68 (55)

O: Bot/1/77 12 7 (58) 1.79 (10)

SAT-2 O: Zim/7/83 77 44 (57) 1.79 (55)

O: KNP/19/89 7 1 (14) 1.72 (5)

SAT-3 O: Bec/1/65 14 2 (14) 0.66 (10)

O: KNP/10/90 33 22 (67) 1.70 (25)

The number of animals challenged includes control unvaccinated animals.

The 95% confidence regions (95% CR) for adopted modelswere also determined using methods described in Sokal and

Rolf [16].

The expected probability of protection (i) for an individ-

ual animal with titre ti is given by:

i =exp(a0 + a1ti)

1 + exp(a0 + a1ti)(1)

where a0 and a1 are the intercept and slope of the associated

logistic regression model relating protection to titre. The ex-

pected proportion of protected cattle, , in a group compris-

ing n individuals, with log titres t1, t2, t3, . . . t n, is given by:

= 1/nn

i=1i. An estimate of whether or not this differssignificantly from 50% of the group can be approximated

by inspecting the quantity p =n/2

i=0

n

i

i(1 )(ni)

which constitutes a (one-tailed) test of the hypothesis that

0.5. If n cattle in a potency trial are vaccinated with

vaccine diluted X-fold and the hypothesis that 0.5 is

Table 2

Parameters corresponding to logistic regression models fitted to subsets of the data in which no significant heterogeneity could be detected at the 5% level

Model no. Virus/lab Vaccine/challenge strain n a0 S.E. a0 a1 S.E. a1 Covariance

(a0, a1)

T50 T50 (95% CI) T95

Onderstepoort

1 SAT-1 Sar/9/81 77 5.155 1.18 4.726 1.054 1.190 1.091 0 .932 1 .258 1.7142 SAT-1 Bot/1/77 12 15.200 12.96 9.780 8.008 102.020 1.554 Insufficient data 1.855

3 SAT-1 Sar/9/81 + Bot/1/77 89 4.780 1.02 4.157 0.840 0.820 1.150 0 .988 1 .307 1.858

4 SAT-2 Zim/7/83 77 6.002 1.30 4.482 0.934 1.162 1.339 1 .164 1 .517 1.996

5 SAT-2 KNP/19/89 7 Insufficient data

6 SAT-2 Zim/7/83 + KNP/19/89 84 5.760 1.18 4.065 0.805 0.912 1.417 1 .243 1 .594 2.141

7 SAT-3 Bec/1/65 + KNP/10/90 47 5.000 1.41 4.310 1.287 1.740 1.160 0 .965 1 .469 1.843

Pirbright

8 A Cruzeiro + Thailand + Iraq 164 6.920 1.14 4.773 0.728 0.807 1.450 1 .326 1 .560 2.067

9 O Manisa + Lausanne 62 8.889 3.43 5.673 1.967 6.668 1.567 1 .116 1 .749 2.086

10 Asia-1 India 46 10.123 5.31 5.803 2.634 13.860 1.744 0.409 1.977 2.252

11 All 3

combined

Cruzeiro + Thailand + Iraq

+ Manisa + Lausanne

+ India

272 6.923 0.957 4.658 0.581 0.540 1.486 1 .377 1 .581 2.118

rejected in favour of the hypothesis that > 0.5 then it canbe concluded that the PD50 of the vaccine is greater than X.

The test is conservative because the variance of a mixed bi-

nomial process (in which each trial succeeds or fails with

a different probability) isn

i=1i(1 i), which is always

greater than the variance for a homogenous process with the

same mean (in which each trial succeeds or fails with the

same probability)which is n(1 ) [17]. Thus, assum-

ing the process to be homogenous when it is in fact mixed

should lead to an overestimate of p.

We can, in principle, use a bootstrap analysis to inves-

tigate the relationship between the number of animals in

such challenge-free trials, and the probability that signifi-cantly more than half the animals in such trials would be

protected. We can randomly assign all vaccinated individu-

als in a trial of n animals a titre from the set of observed

titres for that serotype, and use the model combined over

all strains for that serotype reported in Table 2 to calculate

the mean and variance of the expected number of animals


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Fig. 1. Histograms showing the frequency distribution of titres induced by vaccines of different FMD serotypes.

protected, and the P-value indicative of whether significantly

more than half the animals were protected. Repeating this

process 500 times each for a range of different possible

sizes of trials, allows us to compute the probability that sig-

nificantly more than half the animals in a trial would be

protected as a function of the number of animals in the

trial.

3. Results

3.1. The relationship between virus neutralising titre

and protection

No significant interactions between neutralising anti-

body titre and serotype, or antibody titre and strain were


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detected in any of the analyses described below. Analysis

indicated a highly significant effect of laboratory on the

relationship between titre and protection, we therefore pro-

ceeded to analyse results from the different laboratories

separately. The parameter estimates, with standard errors,

covariances and T50 and T95 values are summarised in

Table 2.

3.2. ARC-Onderstepoort Veterinary Institute

Of the two strains of SAT-1 examined, significant differ-

ences in model intercept were established between Sar/9/81

and Bot/1/77 (P < 0.05): a significant relationship existed

between titre and probability of protection against Sar/9/81

(P < 0.001) but not against Bot/1/77 (presumably a con-

sequence of the small number of potency results available

for this strain). Similarly, a significant relationship between

titre and protection existed for the SAT-2 strain Zim/7/83

(P < 0.001) but insufficient data existed to establish such a

relationship for the other SAT-2 strain, KNP/19/89. No sig-

nificant differences between titre and protection could be es-

tablished for the SAT-3 strains Bec/1/65 and KNP/10/90 so

these were combined and a significant relationship between

titre and protection was apparent (P < 0.001). When strains

Sar/9/81, Zim/7/83, and (KNP/10/90 + Bec/1/65) represent-

ing the three SAT strains are compared, the Zim/7/83 was

found to be modelled by a lower intercept to the SAT-1 and

SAT-3 strains (which were not distinguishable) indicating

that this SAT-2 strain requires a higher titre to achieve the

same level of protection compared to Sar/9/81, KNP/10/90

and Bec/1/65. This is reflected in the T95 values for SAT-1,

SAT-2, and SAT-3 vaccines which were 1.858, 2.141, and1.843, respectively (for models with strains combined within

serotypes).

Fig. 2. The best fitting models to four different subsets of the data.

3.3. Institute for Animal Health, Pirbright

Significant relationships existed between antibody titre

and probability of protection for all strains for which data

were available for serotypes Asia-1, O, and A. In addition,

no significant differences were revealed in the intercepts of

logistic models fitted to the three strains of the A serotype,namely, A15 Thailand, A22 Iraq and A24 Cruzeiro, or the

two strains, Manisa and Lausanne, of the O strain. Analysis

of all three serotypes combined also revealed no significant

differences in intercepts between serotypes. T95 values for

serotypes A, O, and Asia-1 vaccines were 2.067, 2.086, and

2.252, respectively (once again for models with strains com-

bined within serotypes). An observation previously reported

using independent data from tests involving other O, A, and

C vaccine strains [18].

The probabilities of protection with different titres of anti-

body for different subsets of the data sets that are statistically

indistinguishable are indicated in Fig. 2. The estimates of

slope and intercept for the logistic regressions exhibit nega-

tive covariance (see Table 2). This means that uncertainty in

the estimated value of the intercept is negatively correlated

with uncertainty in the estimate of the slope. Therefore, the

confidence regions in parameter space which encapsulate the

true parameters governing the relationship between titre

and protection with 95% probability, are angled ellipses (see

Fig. 3a and b).

3.4. Determining the probability that a particular test

vaccine has a specified/required PD50

Suppose that a vaccine concentrate of serotype Asia-1from Pirbright was diluted X-fold and used to vaccinate eight

test animals. At 21 days post-vaccination, these animals were


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Fig. 3. The confidence regions for the models fitted to the data from vaccine potency trials for serotypes from the two laboratories. The confidence

regions are for models 3, 6, and 7 and 8, 9, and 10 (see Table 2). The angle in the orientation of these ellipses arises from the negative covariance

between estimates of slope and intercepts reported in Table 2.

bled, and log antibody titre determined. Suppose the eight

log titres, ti, were: 2.70, 2.15, 2.85, 2.85, 2.25, 3.15, 3.15, and

1.65. Using these titres and the estimated model (a0 and a1

model number 10) this laboratory from Table 2, we would

estimate that the expected proportion of animals protected

would be = (1/8)8

i=1(exp(a0 + a1ti))/(1 + exp(a0 +

a1ti)) = 0.90260 (in reality seven of eight (87.5%) of these

animals with these titres were protected from challenge).

The quantityn/2

i=0

n

i

i(1 )(ni) yields a P-value of

0.0046. If the PD50 of the test vaccine was Xthen we would

expect 50% of the animals to be protected, but from these

results we anticipate that over 90% would be protected, so

the probability that the PD50 really is Xor less is in this case

very small and we could be 99.54% confident that the PD50of the stored vaccine concentrate was greater than X.

As a second example suppose that 10 test animals had

been vaccinated with a SAT-1 vaccine from Onderstepoort,

diluted Y-fold. Suppose the 21-day log titres were: 1.80,

1.90, 1.40, 1.70, 2.10, 2.40, 2.10, 1.50, 1.30, and 1.60. Us-

ing these titres and the estimated model (number 3) for the

combined SAT-1 results from this laboratory from Table 2,

we would estimate that the expected proportion of animals

protected would be = (1/10)10

i=1(exp(a0 + a1ti))/(1 +

exp(a0 + a1ti)) = 0.882 (in reality 7 of the 10 (70%)

animals with these titres were protected from challenge).


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Fig. 4. The probability of being able to show that the PD50 is significantly greater than that used in these vaccine trials as a function of the number

of individuals used in the trial. Each point is the proportion of 500 re-sampled sets of titre values (from the indicated serotype) in which significantly

more than 50% of individuals in the set would be protected. The number of individuals (the number of re-sampled titres) in each trial is indicated on

the x-axis. Parameter values are those for the combined strains model for each serotype (numbers 3, 610).

If the PD50 of the test vaccine was Y then we would ex-

pect 50% of the animals to be protected, but from these re-

sults we anticipate that 88.2% would be protected, so the

probability that the PD50 really is Y or less is in this casen/2i=0

n

i

i(1 )(ni) = 0.0034 and in this case, we

could be 99.66% confident that the PD50 of the vaccinetested was greater than Y.

The number of animals required in a trial to establish with

95% confidence that the vaccine protected 50% of vacci-

nated animals obviously depends on the data used but some

examples using data analysed in this study are shown in

Fig. 4. If the titres from each serotype analysed here were

representative generally of those obtainable from vaccines

prepared from these serotypes then only modest numbers

of animals (as few as six) are required to establish that the

most effective and consistent vaccines (those of serotype O

and Asia-1) protect half the animals when diluted as they

were in these trials. For the more variable titres obtained

from SAT-1 and SAT-3 vaccines, even using as many as 18

animals would not necessarily indicate 50% protection with

any degree of confidence.

4. Discussion

This analysis reveals that the relationships documented

between vaccinally induced antibody titre and protection

against clinical foot-and-mouth disease differs between two

different laboratories. Whether this is because of method-

ological differences in the adopted procedures, or because

different relationships hold for the different serotypes stud-

ied in each of the different laboratories cannot be determined

from the data available. However, previous studies have also

documented differences in the test results between different

laboratories [19]. In general, it appears that so long as sam-

ple sizes were not too small, results from different strains

of the same serotype usually conformed to a single logis-tic model (this was the case for three serotypes represented

by more than one strain: A, O, and SAT-3). The parame-

terisation of these models for each serotype enables the use

of challenge-free trials to estimate the effectiveness of these

vaccines. We developed a simple statistical framework for

analysis of challenge-free trials for which existing data sug-

gests that for some serotypes, six to eight animals might

provide sufficient statistical power to confidently establish

required PD50 values. This is fewer animals than that cur-

rently stipulated by the European Pharmacopoiea.

Usually potency testing of inactivated vaccines is per-

formed on the target species. This takes into account not

only the active substance, i.e. antigen, but other compo-

nents such as the adjuvants which are incorporated to fur-

ther enhance the immunity of the vaccine. This is true for

foot-and-mouth disease vaccine as this vaccine always in-

corporates either aluminium hydroxide/saponin or mineral

oil-based adjuvants. This in vivo potency assessment usually

involves cattle, which makes the test very costly and involves

the examination of virus challenged animals to establish the

protective ability of the vaccines. The extremely infectious

nature of FMDV means that high level containment facilities

are required to perform such tests, which further limits ac-

cessibility and adds to the overall cost. The results are then


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based on whether generalisation of disease has occurred at

sites other than that of challenge. This of course means that

animals not protected will suffer the painful clinical man-

ifestations of FMD and even some protected animals may

undergo some pain due to the development of primary le-

sions at the site of challenge. Results are, therefore, either

black or white, with no intermediary level in which an an-imal could be perceived as being partially protected. These

results are then extrapolated into percentages of protected

animals within the vaccine dose groups and converted to a

PD50 value by the Krber method [20] or some similar pro-

cedure, to establish the lowest dilution of vaccine that would

protect 50% of the animals. Such tests are often based on

relatively small numbers of animals. For example, the Eu-

ropean Pharmacopoeia monograph for foot-and-mouth dis-

ease vaccines requires three vaccine dilution groups of five

animals per group. This test has been viewed as both statis-

tically weak and inaccurate with a 90% confidence interval

for the PD50 value lying anywhere between 45 and 220% of

the potency [21]. A further important consideration is thatoften FMD vaccines can be multivalent containing three or

four different strains of virus and it is not possible to under-

take a challenge test with more than a single strain of virus

at any one time.

A test approach that relies on large numbers of defined

sera that have an established correlation with potency and

protection in the target species therefore seems an attractive

alternative. By far the most explored area of investigation

has been the correlation between protection and serological

parameters such as neutralising antibody [2,7,2224]. The

data of this sort collected at Onderstepoort and Pirbright per-

mits a feasibility study of this approach. In order to developlogistic models it is important that the data span a wide range

of titres. Data for serotype C from Pirbright could not be

used in this analysis because no records of vaccinated but

unprotected animals existedthe vaccine always protected

vaccinates, and thus a logistic model could not be fitted.

The bootstrap analysis of the number of animals required

to conclude with reasonable confidence that significantly

more than half the animals in a group are protected (Fig. 4)

is only intended for illustratory purposes. Specifically, we

do not claim that the titres included in the analysis here are

representative of those that would be expected from field use

of market ready product. Data for some strains and serotypes

(very usefully) includes vaccine used at higher dilutions, and

thus induce a wider range of titres than would be expected

from non-experimental use. However, it does show, first, that

its very straightforward to use this kind of data to suggest the

number of animals required in such trials, and second, that

if a vaccine regularly induces high levels of protection then

this number could be quite modestpossibly as low as 68.

In summary, statistical analysis of serological and protec-

tion data accrued from challenge/potency tests performed

at two independent research laboratories and encompassing

vaccine strains representing six of the seven serotypes of

FMD indicates a significant effect of laboratory on the rela-

tionship between neutralising antibody titre and protection.

However, individual analysis of the data from each labora-

tory showed that, provided reasonable sample sizes are avail-

able, a significant relationship can be established between

antibody titre and probability of protection. Furthermore, no

significant differences were observed in the parameters of

logistic models fitted to combined strains within serotypesA, O, and SAT-3, or when the strains from serotypes A, O,

and Asia-1 or SAT-1 and SAT-3 are combined. These mod-

els permit the development of a truncated test, that avoids

the use of virus challenge and requires only a single group

of cattle administered with a vaccine diluted to a level equal

to or greater than the required potency value (PD50) potency,

providing an alternative method for evaluating the vaccine

potency required.

Acknowledgements

Daniel T. Haydon was supported by the Wellcome Trust.We thank Victoria Edge for helpful discussions.

References

[1] European Pharmacopoeia 1997. Foot-and-Mouth Disease (Rumin-

ants) Vaccine 1997;63:8756.

[2] Pay TWF, Hingley PJ. The use of serum neutralising antibody assay

for the determination of the potency of foot-and-mouth disease

vaccines in cattle. Develop Biol Stand 1986;64:15361.

[3] van Bekkum JG. Neutraliserende antistoffen in sera van regen

mond and Klauwzeer geente runderen. Thesis. University of Utrecht,

Utrecht, 1959.

[4] Hecke F, Uhlmann W, Lorenz R. Bericht uber die ergebnisseder grossversuchs 19571958 zur wirkaamkeitsprufung der Maul-

und klavenseuche-koncentratvakzine nach pyl unf der origina-

lvakzine nach waldmann und kobe. IV. Beziehung zwischen

serumneutralisation und immunitat. Mh Tierheildk 1960;12:111.[5] Mackowiak C, Lang R, Fontaine J, Camand R, Petermann MB.

Relation entre titre danticorps neutralisants et protection des animaux

aprs vaccination antiaphteuse. Ann Ins Pasteur 1962;103:25261.

[6] Bercan A, Muntiu N, Dohotaru U, Tomescu A. Sur la corrlation

entre le test de sro-neutralization sur sourceux et immunisation

anti-aphtheuse crosee chez les bovins. Bull Off Int Epizoot

1969;71:38192.

[7] Stellman C, Bornarel P, Lang R, Terre J. Controle quantitatif du

vaccin antiaphtheux. Analyse statistique de la relation liant les titres

danticorps neutralisant au pourcentage de protection bovine. Rec

Med Vet (Alfort) 1968;144:32551.[8] Pay TWF, Parker NJ. Some statistical and experimental design

problems in the assessment of FMDV vaccine potency. Develop Biol

Stand 1977;35:36983.

[9] Pay TWF, Hingley PJ. Correlation of 140S antigen dose with the

serum neutralizing antibody response and the level of protection

induced in cattle by foot-and-mouth disease vaccines. Vaccine

1987;5:604.[10] Ahl R, Haas B, Lorenz RJ, Wittman G. Assessment of potency of

foot-and-mouth disease vaccines by means of antibody assays with

sera from vaccinated cattle. Report of the Meeting of the Research

Group of the Standing Technical Committee of the European

Commission for the Control of Foot-and-Mouth Disease. Prague,

Czechoslovakia, FAO, Rome, Appendix 17, 2023 September 1988.

p. 99111.


9/9


[11] Bengelsdorff HJ. Testing the efficacy of FMD vaccines: relationships

between test infection results and corresponding neutralization

titres of vaccinated cattle. Berliner und Munchener Tierarztliche

Wochenschrift 1989;102:1938.

[12] van Maanen C, Terpstra C. Comparison of the liquid phase blocking

sandwich ELISA and the serum neutralisation test to evaluate

immunity in potency tests for foot-and-mouth disease vaccines.

Session of the Research Group of the European Commission for the

Control of Foot-and-Mouth Disease. Prague, Czechoslovakia, FAO,

Rome, 2023 September 1989. p. 918.

[13] Barnett PV, Carabin H. A review of emergency foot-and-mouth

disease (FMD) vaccines. Vaccine 2002;20:150514.

[14] Anon. Manual of Standards for Diagnostic Tests and Vaccines. Office

International des Epizooties. 4th ed. Part 2. Section 2.1. OIE Listed

Diseases. Foot-and-Mouth Disease, 2000, Chapter 2.1.1. p. 77

92.

[15] Finney DJ. Probit analysis. 3rd ed. London: Cambridge University

Press; 1971.

[16] Sokal RR, Rolf FJ. Biometry. 3rd ed. New York: Freeman; 1995.

[17] Feller W. An introduction to probability theory and its applications.

New York: Wiley; 1968.

[18] Ahl R, Wittmann G. Further studies on the assessment of potency

of FMD vaccines by means of antibody assays with sera fromvaccinated cattle. Report of the Session of the Research Group of

the Standing Technical Committee of the European Commission for

the Control of Foot-and-Mouth Disease. Rio de Janeiro, Brazil, FAO,

Rome, Appendix 6, 1518 October 1985. p. 459.

[19] Sutmoller P, Vieira A. The relationship of neutralizing antibody

titres for FMDV and the protection of cattle. Boletin del Centro

Panamericano de Fiebre Aftosa 1980;3940:5762.

[20] Karber G. Beitrag zur kollekiven Behandlung pharmakologischer

Reihenversuche. Arche Exp Pathol Pharmakol 1931;162:480.

[21] Hendriksen CFM. Laboratory animals in vaccine production and

control: replacement, reduction and refinement. Dordretch: Kluwer

Academic Publishers; 1988.

[22] Sutmller P, Gomes I, Astudillo VM. Potency estimation of FMD

vaccines according to antibody assay results. Boletin del Centro

Panamericano Fiebre Aftosa 1984;4550:314.

[23] Ahl R, Haas B, Lorenz RJ, Wittman G. Alternative potency test of

FMD vaccines and results of comparative antibody assays in different

cell systems and ELISA. Report of the Meeting of the Research

Group of the Standing Technical Committee of the European

Commission for the Control of Foot-and-Mouth Disease. Lindholm,

Denmark; FAO, Rome, June 1990. p. 5160.

[24] Pay TWF, Hingley PJ. Foot-and-mouth disease vaccine potency

tests in cattle: the interrelationship of antigen dose, serum

neutralizing antibody response and protection from challenge.Vaccine 1992;10:699706.

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