Serological Investigation for Brucella ceti in Cetaceans from the Northwestern Mediterranean Sea

Brucella ceti is a marine bacterium that causes neurological, reproductive and skeletal disease in free-ranging cetaceans. Its zoonotic potential and importance for wild animals has prompted, over the years, the search for a reliable diagnostic method to detect antibodies and infer the level of infection. In this work, we perform an exploratory serological study on cetaceans stranded in the Northwestern Mediterranean Sea. Antibody levels were higher in animals with confirmed Brucella disease and infection in juveniles and in animals with chronic morbilliviral infection. This provides the first seroprevalence estimation in this area and reaffirms the active circulation of Brucella in wild cetaceans. Neurobrucellosis in cetaceans, caused by Brucella ceti, is a relevant cause of death in striped dolphins (Stenella coeruleoalba) from the Mediterranean Sea. Serological tests are not used as a routinary technique for the diagnosis of this infection. We briefly describe the pathological findings of nine free-ranging stranded cetaceans diagnosed with Brucella disease or infection in our veterinary necropsy service from 2012 to 2022. The findings included focal diskospondylitis and non-suppurative meningitis, choroiditis and radiculitis. Additionally, an exploratory serological study was conducted in sixty-six frozen sera collected in the period 2012-2022 from fifty-seven striped dolphins, five Risso's dolphins (Grampus griseus), two common bottlenose dolphins (Tursiops truncatus), one common dolphin (Delphinus delphis) and one pilot whale (Globicephala melas) to compare antibody levels in Brucella -infected (n = 8) and non-infected (n = 58) animals, classified by the cause of death, sex, age class and cetacean morbillivirus (CeMV) infection status. The authors hypothesized that active infection in cases of neurobrucellosis would elicit a stronger, detectable humoral response compared to subclinical infections. We performed a commercial competition ELISA (cELISA) using serial serum dilutions for each sample, considering a percentage of inhibition (PI) of ≥40% as positive. A titer of 1:160 was arbitrarily determined as the seropositivity threshold. Seropositive species included striped dolphins and Risso's dolphins. Seroprevalence was higher in animals with neurobrucellosis (87.5%) compared to the overall seroprevalence (31.8%) and to other causes of death, indicating, likely, a high sensitivity but low specificity for neurobrucellosis. Animals with chronic CeMV seemed to have higher seroprevalences, as well as juveniles, which also had a higher disease prevalence. These results indicate, as in other studies, that antibodies are not decisive against clinical brucellosis, although they may indicate a carrier state, and that CeMV may influence Brucella epidemiology. More research is required to elucidate the epidemiology and pathogenesis and to resolve the complicated host-pathogen interaction in Brucella species.