Elisa for Detection of Brucellosis in Bottlenose Dolphins (Tursiops truncatus)
IAAAM Archive
W. George Miller1; Tracy Romano2; Roberta Pugh2; Garry Adams2; Sam H. Ridgway3
1US Navy Marine Mammal Program, San Diego, CA; 2Texas A & M University, College Station, TX;3University of California-San Diego, Department of Pathology, Center for Veterinary Medicine, San Diego, CA


Two bottlenose dolphins (Tursiops truncatus)aborted fetuses that died as a result of Brucella placentitisinfection.1,4 In a third case, an identical organism was cultured from lung necropsy tissue of an adult female T. truncatus.1 The Brucella isolate recovered in all three cases proved to be identical by microbiology results and polymerase chain reaction results.1 Microbiology, specific polymerase chain reaction, and pulsed-field gel electrophoresis results support an additional genomic group(s),1,2,3,4,5 that Miller et al.1 has designated Brucella delphini, for isolates adapted to T. truncatus.

Current serologic diagnostic tests reliable for known terrestrial Brucella species are unreliable in detecting dolphin brucellosis.1 Infected placenta, vaginal, uterine, lactational and seminal fluids may transmit Brucella delphini to other cetaceans. Our findings, together with previous reports, suggest that brucellosis in marine mammals is a naturally occurring disease.1,2,3,4,5,6 Brucella infection in cetaceans can adversely impact reproduction.1

An enzyme labeled immunosorbent assay (ELISA) test was developed to detect antibody titers to Brucella delphini. A fixed amount of purified whole Brucella delphini antigen was coated onto the surface of a 96-well microplate. Dilutions of dolphin serum (both test and control) were added to the plate, followed by a rabbit anti-dolphin Ig secondary antibody conjugated with horseradish peroxidase. The addition of an enzyme substrate allowed for direct visualization and measurement of the chromagen reaction at 405nm.

This assay can be used as a diagnostic tool for Brucella exposure in cetaceans and in managing the breeding and reproduction of cetaceans at marine facilities.


1.  Miller WG; Adams LG; Ficht TA; Cheville NF; Payeur JP; Harley DR; House C; Ridgway SH 1999. Brucella-induced abortions and infection in bottlenose dolphins (Tursiops truncatus). Space and Naval Warfare Systems Center, San Diego, California 92152-6266, USA. J Zoo Wildl Med 30(1):100-110.

2.  Bricker B J, Ewalt DR, MacMillan AP, Foster G, Brew S. characterization of Brucella strains isolated from marine mammals. 38(3):1258-1262. J Clin Microbiol.

3.  Jensen AE, Cheville NF, Thoen CO, MacMillan AP, Miller WG. 1999. Genomic fingerprinting and development of a dendrogram for Brucella spp. isolated from seals, porpoises, and dolphins. J Vet Diagn Invest. 11(2):152-157.

4.  Ewalt DR, Payeur JB, Martin BM, Cummins DR, Miller WG. 1994. Characteristics of a Brucella species from a bottlenose dolphin (Tursiops truncatus). J Vet Diagn Invest. 6(4):448-452.

5.  Cloeckaert A, Grayon M, Grepinet O. 2000. An IS711 element downstream of the bp26 gene is a specific marker of brucella spp. Isolated from marine mammals. Clin Diagn Lab Immunol. 7(5):835-839.

6.  Garner MM; Lambourn DM; Jeffries SJ; Hall PB; Rhyan JC; Ewalt DR; Polzin LM; Cheville NF. 1997. Evidence of Brucella infection in Parafilaroides lungworms in a Pacific harbor seal (Phoca vitulina richardsi). J Vet Diagn Invest. 9(3):298-303.

Speaker Information
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W. George Miller, DVM, PhD
San Diego, CA, USA

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