A new enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies against foot-and-mouth disease virus. I. Development and method of ELISA.

A liquid-phase blocking sandwich ELISA has been developed for the quantification of antibodies against foot-and-mouth disease virus which may replace the virus neutralisation (VN) test. This test employs the incubation of a constant amount of antigen with a range of test serum dilutions in the liquid-phase before being assayed using a trapping ELISA. Thus it does not rely on the availability or growth of tissue culture cells. The assay is rapid and relatively simple to perform, reagents are used economically and results may be recorded within 24 h. The ELISA is sensitive and results are more specific and more reproducible than those obtained by VN. Results are expressed as reciprocal antibody titres which are analogous and of a similar order to those recorded by VN. Individual titres, therefore, may be easily assessed by workers in the field who are already familiar with VN.

A new enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies against foot-and-mouth disease virus. II. Application.

The liquid-phase blocking sandwich ELISA has been evaluated for the serological study of antibodies against foot-and-mouth disease virus (FMDV). The titres recorded for sera from a population of more than 300 British uninfected, unvaccinated cattle which were examined against each of the seven immunologically distinct FMDV types were less than 1 in 40. A positive correlation between ELISA and VN titres was recorded for sera either vaccinated or involved in outbreaks of FMDV. The overall regression between the ELISA/VN data showed that 1 in 16 by VN was equivalent to 1 in 40 by ELISA. Thus ELISA is sensitive, specific and reproducible and results may be directly correlated to those recorded by VN. Serum titres may be interpreted as positive or negative and the number of sera which require retesting would be considerably less than by VN. It is suggested that this ELISA may be used as an alternative to the existing VN test for the quantification of antibodies against FMD virus.

An enzyme-linked immunosorbent assay (ELISA) for the detection and quantification of antibodies against swine vesicular disease virus (SVDV).

A liquid phase blocking sandwich ELISA has been compared with virus neutralisation for testing pig sera for antibodies against swine vesicular disease (SVD) virus. Highest infectivity titre of SVD virus was obtained using a multiplicity of infection of 30 pfu/cell and harvesting after 21 h. Titres obtained for 300 clinically normal animals were assessed by ELISA and 89% were found to be 1/6 or less. Results were skewed and spread up to 1/45. Comparison of known positive sera resulted in a correlation between the two methods of 0.68 and showed that a virus neutralisation titre of 1/16 was equivalent to 1/40 (log10 1.61) by ELISA. Variation in results obtained by replicate testing using ELISA and virus neutralisation was almost identical. Overlap between positive and negative sera was shown to be reduced to 1-1/2 fold in ELISA. Therefore, the ELISA correlated well with virus neutralisation and has several advantages over the latter.

The duration of the foot-and-mouth disease virus carrier state in African buffalo (i) in the individual animal and (ii) in a free-living herd.

The maintenance of a virus depends on a number of factors, including the duration of infectivity and the size of the available host population. In this work, foot-and-mouth disease virus was shown to persist in individual African buffalo (Syncerus caffer) for up to at least five years; thus, the duration of infectivity is more than adequate to cover the normal periods between calving peaks. In a small isolated free-living population which varied from 30 to 100 buffalo, two immunological types of foot-and-mouth disease virus were maintained for at least 24 years and through several generations.

Experimental infection of eland (Taurotrages oryx), sable antelope (Ozanna grandicomis) and buffalo (Syncerus caffer) with foot-and-mouth disease virus.

The course of experimental infection of a type SAT 1 FMDV strain was studied in buffalo, sable antelope and eland following tongue inoculation and contact and has been compared with that in cattle. All species became infected, although disease was less severe in the game animals and larger amounts of virus were required to infect game animals than cattle. Neutralizing antibody titres were high and were maintained for an extended period in buffalo, sable antelope and eland. The carrier state was demonstrated in buffalo for the longest period. Cattle carried virus for up to 56 days. Virus persistence in sable antelope was transitory and did not occur in eland.

Comparative studies of United Kingdom isolates of swine vesicular disease virus.

The characteristics of four United Kingdom isolates of swine vesicular disease (SVD) virus from 1981 to 1982 have been compared with those of an isolate obtained from the first outbreak of swine vesicular disease diagnosed in the United Kingdom in 1972. When the virus structural proteins were examined by polyacrylamide gel electrophoresis the four isolates from 1981-82 all had the same polypeptide pattern, which was different from that of the 1972 isolate. Immunodiffusion tests with the 1972 isolate and one 1982 isolate did not reveal any antigenic difference between the viruses but minor antigenic differences were shown by cross-neutralisation tests between the 1972 isolate and the four isolates from 1981-82. In experimentally infected pigs the 1972 isolate produced typical SVD lesions whereas the four more recent SVD viruses produced only very mild clinical disease. Clinical lesions scored numerically were four- to 10- and five- to 11-fold higher at seven and 14 days after infection for pigs infected with the 1972 isolate than with the four isolates from 1981-82. The serum of pigs infected with the 1972 isolate contained significantly higher levels of neutralising antibody than those of pigs infected with more recent isolates. The antibody titres of pigs with only primary lesions ranged from log10 1.9 to 2.8 and one clinically normal pig had a titre of log10 2.4 at 14 days after infection. Attention is drawn to the implication of these findings for SVD control policies based only on the recognition and reporting of clinical disease.

Experimental transmission of foot-and-mouth disease virus from carrier African buffalo (Syncerus caffer) to cattle in Zimbabwe.

Four female cattle and three male African buffalo (Syncerus caffer) which were free of foot-and-mouth disease (FMD) virus were held together on an island in Lake Kariba, Zimbabwe. The buffalo were experimentally infected with FMD virus type SAT2, developed generalised disease and became virus carriers. While the buffalo were in the acute phase of the disease the susceptible contact cattle did not show lesions, no virus was recovered from them and they did not develop serum antibodies. However, five months later the cattle developed severe foot-and-mouth disease. Direct nucleotide sequencing of the virus used to infect the buffalo and of the virus from the in-contact cattle showed that the two isolates were almost identical. The results suggest that in nature it is possible for the virus to be transmitted from buffalo to cattle under the influence of factors not yet defined, and that there was very little change in the nucleotide sequence of the virus during the carrier period of five months.