Use of enzyme‐linked immunoassays for antibody to types C and D botulinum toxins for investigations of botulism in cattle

Abstract

The development of specific enzyme-linked immunosorbent assays (ELISA) for antibody to types C and D Clostridium botulinum toxins for investigation of botulism in cattle is described. Partially purified type C and D toxins were used as antigens to develop these ELISAs. Specificity of the ELISAs was evaluated on sera from 333 adult beef and dairy cattle from areas with no history or evidence of botulism in animals or water birds. The test was also evaluated on sera from 41 herds that included herds vaccinated against botulism, confirmed botulism cases and herds from areas where the disease is considered endemic. The ELISAs detected the presence of antibody to botulinum toxins in samples from vaccinated cattle and both convalescent and clinically normal animals from unvaccinated herds with outbreaks of botulism. Antibody was also found in unvaccinated animals from herds in which there had been no diagnosed botulism cases in areas where botulism was considered endemic. Sera from some unvaccinated cattle with high ELISA reactivity was shown to be protective for mice in botulinum toxin neutralisation tests. The use of these tests in investigations of botulism in cattle is discussed.

Quantitative Analysis of Levels of Serum Immunoglobulin G against Botulinum Neurotoxin Type D and Association with Protection in Natural Outbreaks of Cattle Botulism

“…Based on the above results, we concluded that the botulism toxoids can confer adequate protection against natural exposure to lethal doses of BoNT/D but that vaccination protocols used on dairy farms were inadequate and should be optimized. However, lower anti-BoNT/D IgG levels in younger animals might be explained by repeated natural exposure to subclinical toxin levels of older animals, as previously demonstrated in Australia (13). The results of our study differ from those of previously reported vaccination studies of cattle (15,34), which demonstrated better protection after fewer inoculations; this difference might be explained by exposure to higher levels of toxin or botulinum strain differences.…”

“…Two days thereafter, a group of pregnant heifers, the biggest and oldest of the VOL. 13,2006 PROTECTIVE LEVELS OF ANTI-BOTULINUM D ANTIBODIES 863 lot, which were in the last feeding group, were found either dead or recumbent (index cases). Within 5 days, 55 of 57 (96%) replacement heifers (Fig.…”

“…ELISA systems, which offer greater flexibility and economy than other immunoassay techniques, were also used for the evaluation of the immune response to various botulinum vaccines in mice (21), primates (19), humans (32), and cattle (3). Previously, the ELISA positive/negative cutoff values used for diagnosis of botulism in cattle were defined as the upper 99% confidence limit of the mean of a population of cattle not exposed to BoNTs plus 3 sample standard deviations (13). Higher ELISA OD values were detected in serum samples from four naturally exposed unvaccinated heifers, which protected mice in a mouse neutralization bioassay (13), and were used for vaccine evaluation in cattle (3).…”

“…Previously, the ELISA positive/negative cutoff values used for diagnosis of botulism in cattle were defined as the upper 99% confidence limit of the mean of a population of cattle not exposed to BoNTs plus 3 sample standard deviations (13). Higher ELISA OD values were detected in serum samples from four naturally exposed unvaccinated heifers, which protected mice in a mouse neutralization bioassay (13), and were used for vaccine evaluation in cattle (3). The threshold of a positive response in vaccinated humans was defined as the mean OD plus 5 standard deviations at a 1:80 dilution of 150 nonvaccinated serum samples (24).…”

Esporos e toxinas de Clostridium botulinum dos tipos C e D em cacimbas no Vale do Araguaia, Goiás

“…No entanto, pode-se deduzir que a ingestão contínua, mesmo de dose subletal, tenha como conseqüência o aumento da susceptibilidade dos animais a uma eventual intoxicação letal. Na realidade, o fato de bovinos criados extensivamente e não vacinados contra o botulismo apresentarem anticorpos contra as toxinas C e D (Gregory et al 1996) revela que este é um evento comum em áreas endêmicas.…”