Development, Validation, and Utilization of a Competitive Enzyme-Linked Immunosorbent Assay for the Detection of Antibodies against Brucella Species in Marine Mammals

Abstract

Abstract. A competitive enzyme-linked immunosorbent assay (cELISA) was developed by using a wholecell antigen from a marine Brucella sp. isolated from a harbor seal (Phoca vitulina). The assay was designed to screen sera from multiple marine mammal species for the presence of antibodies against marine-origin Brucella. Based on comparisons with culture-confirmed cases, specificity and sensitivity for cetacean samples tested were 73% and 100%, respectively. For pinniped samples, specificity and sensitivity values were 77% and 67%, respectively. Hawaiian monk seal (Monachus schauinslandi; n 5 28) and bottlenose dolphin (Tursiops truncatus; n 5 48) serum samples were tested, and the results were compared with several other assays designed to detect Brucella abortus antibodies. The comparison testing revealed the marine-origin cELISA to be more sensitive than the B. abortus tests by the detection of additional positive serum samples. The newly developed cELISA is an effective serologic method for detection of the presence of antibodies against marine-origin Brucella sp. in marine mammals.

Assessment of Clinical Pathology and Pathogen Exposure in Sea Otters (Enhydra Lutris) Bordering the Threatened Population in Alaska

“…The increased prevalence could also be due to crossreactivity with antibodies to other Gramnegative bacteria. Although not a part of this study, evaluation of cross-reactivity of the marine Brucella-based cELISA with other Gram-negative bacteria (Meegan et al, 2010) found that it did not occur with Yersinia enterocolitica O:9 antibody-positive sera; however, cross-reactivity was found with B. abortus-and B. melitensis-positive sera. Thus, because the ma-rine Brucella cELISA does cross-react with other terrestrial Brucella spp., it is unclear whether exposure to a marine or terrestrial strain was detected.…”

“…Samples with .30% inhibition were considered positive. This test has not been validated in sea otters; however, based on comparisons with culture-confirmed cases, specificity and sensitivity for cetacean samples were 73% and 100%, respectively, and for pinnipeds 77% and 67%, respectively (Meegan et al, 2010). Additionally comparison testing revealed the marine-origin cELISA was more sensitive for detecting anti-Brucella antibodies in marine mammal samples than was the B. abortus tests available.…”

“…was measured using a competitive enzyme-linked immunosorbent assay (cELISA) at the Mystic Aquarium and Institute for Exploration, Mystic, Connecticut, USA (Meegan et al, 2010). The assay was developed using a whole-cell antigen of a strain of marine Brucella species isolated from a harbor seal (Phoca vitulina), characterized and available commercially through NVSL (case 04-0281, accession 311585).…”

A multiplex real-time polymerase chain reaction assay with two internal controls for the detection of Brucella species in tissues, blood, and feces from marine mammals

“…19,26,30 Clinical manifestations of brucellosis in marine mammals are so far described only in cetaceans, with abortion and placentitis, orchitis, visceral and blubber abscesses, musculoskeletal lesions, and meningoencephalitis reported. 12,20,27 High levels of seropositivity are frequently reported in pinnipeds, and microbiologic isolation is reported in many pinniped species.…”

Detection of Francisella tularensis-Specific Antibodies in Patients with Tularemia by a Novel Competitive Enzyme-Linked Immunosorbent Assay

“…Our cELISA is based on the measurement of competition between the test sample and MAb for LPS, the major antigen of F. tularensis (14,24,25). Similar assays using MAb against LPS have been described for Brucella species (26,27). The typical ladder-like bands observed on WB demonstrated that both MAb and F. tularensis-infected human and mice serum contained antibodies predominantly recognizing the LPS antigen (Fig.…”