Evaluation of Competitive ELISA for Detection of Antibodies to Brucella Infection in Domestic Animals

Aim

To evaluate competitive enzyme-linked immunosorbent assay (cELISA) for its suitability as an additional serological test for the diagnosis of animal brucellosis.

Methods

cELISA, which was developed at the Veterinary Laboratories Agency, has been evaluated for its accuracy and suitability as an additional serological test for the diagnosis of animal brucellosis. Samples from naturally and experimentally infected animals and those from Brucella-free flocks and herds were tested.

Results

Data obtained since 1991 were analyzed from routine surveillance, animals experimentally infected with Brucella, and stored sera to validate cELISA for the detection of antibodies to Brucella in cows, small ruminants, and pigs. The sensitivity of the test ranged from 92.31% to 100%, in comparison with 77.14% to 100% for the complement fixation test (CFT). Specificities for cELISA, indirect enzyme-linked immunosorbent assay, and CFT were greater than 90%.

Conclusion

cELISA can be used on a variety of animal species, and an added advantage is its suitability for use on poor-quality samples such as those affected by hemolysis.In accordance with EC Directive 91/68/EEC, flocks of sheep and herds of goats in the United Kingdom (UK)are monitored serologically to prove that they are free from Brucella melitensis. In 2006, competitive enzyme-linked immunosorbent assay (cELISA) was introduced to screen these animals as part of a surveillance program in Great Britain (GB), the territory including all of the UK except for Northern Ireland. It replaced the complement fixation test (CFT) because of its much higher specificity and ease of automation. Currently, in excess of 35 000 animals are tested annually.In 2001, a revision to the pig semen directive was introduced by EC Directive 99/608 so that CFT was replaced with the Rose Bengal test (RBT) as the test used for brucellosis on all pigs whose semen is used for artificial insemination. RBT and CFT were run in parallel in addition to cELISA prior to this date in order to assess the effects of changing the testing regime and, at the same time, to validate the use of cELISA for pigs. During 2001, all routine samples that were tested for artificial insemination purposes and were positive by RBT were also tested by cELISA and the results analyzed using different diagnostic thresholds. The aim was to set an appropriate threshold that would provide optimal specificity and sensitivity for cELISA.CFT, RBT, and indirect enzyme linked immunosorbent assay (iELISA) are the conventionally used tests for diagnosis of bovine brucellosis. These tests are described in the Manual for Diagnostic Tests and Vaccines for Terrestrial Animals produced by the World Organisation for Animal Health, previously the Office International des Epizooties (OIE) (1), and this manual gives details of all the diagnostic methods. It also describes the strain of Brucella required for antigen preparation and the procedure for standardization for each test.The cELISA for the detection of antibodies against Brucella spp. was adapted at the Veterinary Laboratories Agency (VLA) from the method described by MacMillan et al in 1990 (2). It was initially developed for the diagnosis of brucellosis in small ruminants and was tested extensively on British sheep and on sheep and goats from France. It has also since been tested on large numbers of cattle and pigs.The aim of this study was to bring together and compare all brucellosis testing results carried out using cELISA, RBT, and iELISA at the VLA since 1991. The samples had been collected and analyzed within the framework of various surveillance screening programs and experimental studies. The present study demonstrates the effectiveness of cELISA compared with other assays currently used as diagnostic tests of brucellosis in domestic animals.