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Rose Bengal antigen and smooth lipopolysaccharide (s-LPS) were produced from a field strain of Brucella ceti ("homologous" antigens) and from the reference strain B. abortus S99 ("heterologous" antigens); they are currently used for the diagnosis of brucellosis in cattle, water buffaloes, sheep, goats, and pigs, as recommended in the Manual of Diagnostic Tests and Vaccines for Terrestrial Animals of the World Organization for Animal Health (WOAH). "Homologous" and "heterologous" antigens were used in a rapid serum agglutination test (Rose Bengal test, RBT) and a competitive ELISA assay (c-ELISA) to test a panel of sera, blood, and other body fluids (cerebrospinal fluid, pericardial fluid, tracheal fluid, and aqueous humor) collected from 71 individuals belonging to five cetacean species (Stenella coeruleoalba; Tursiops truncatus; Grampus griseus; Globicephala melas; and Ziphius cavirostris), which were found stranded on the Italian coastline. Six animals were positive for Brucella spp. for bacterial isolation and/or PCR, and 55 animals were negative; for the remaining 10 animals, no PCR/isolation data were available. A total of 90 samples were tested. Results obtained from the two tests were compared in order to identify the most suitable antigen for the serological diagnosis of Brucella infection in cetaceans. The RBT performed with the "homologous" antigen showed the best results in comparison with the "heterologous" antigen: diagnostic sensitivity, specificity, and accuracy were 80.0%, 44.1%, and 46.9% for the "homologous" antigen and 80.0%, 17.0%, and 21.9% for the "heterologous" antigen. For the c-ELISA, "homologous" and "heterologous" s-LPS showed similar results (diagnostic sensitivity 66.7%, diagnostic specificity 97.3%, and diagnostic accuracy 95.0%). Therefore, the RBT using the "homologous" antigen and c-ELISA with "homologous" or "heterologous" s-LPS could be used in parallel for the detection of antibodies against Brucella spp. in cetaceans. [ABSTRACT FROM AUTHOR]

In Italy, veterinary autogenous vaccines manufacturing is regulated by the legislative decree of the Ministry of Health, March 17th, 1994, n. 287. The production is performed by the network of the 'Istituti Zooprofilattici Sperimentali' (IZSs), public health institutes scattered all over the Italian territory. The aim of this research was to evaluate the feasibility of an in vitro method to test the abnormal toxicity of autogenous bacterial vaccines as an alternative to animal models routinely employed. For this purpose, the Istituto Zooprofilattico Sperimentale dell'Abruzzo e del Molise (IZSAM) in partnership with the Istituto Zooprofilattico Sperimentale della Lombardia e dell'Emilia Romagna (IZSLER), evaluated the toxicity of 49 batches of autogenous bacterial vaccines, previously shown to be safe in guinea pigs and mice, on animal model, by means of the methyl tetrazolium (MTT) assay. All vaccines showed cytotoxic effects when tested 1:2 diluted and undiluted; overall, all vaccines lost toxicity at 1:128 dilution. As expected, these findings suggest a different susceptibility of this assay  compared to the laboratory animal model. On the other hand, these results do not clarify which components of the vaccines are responsible for the cytotoxic effect. Overall, more experiments are warranted in order to standardize the MTT assay which could be coupled with the trials in laboratory animals.