Diseases of the respiratory tract are significant in cetaceans. In fact, bacterial pneumonia is responsible for more deaths in captive dolphins than any other disease. Infections of the respiratory tract are usually associated with predisposing diseases. Conditions may be acute or chronic, depending on the bacterial organism and the immune status of the animal. Once the clinical signs of malaise, anorexia, increased respiratory rate, coughing and foul smelling breath are demonstrated, successful therapy is more difficult. Therefore, it is important to find ways to diagnose respiratory tract disease early in its course so that therapy can be instituted in the early phase of the disease. Specimens of respiratory exudate to be analyzed for cytological evidence of disease may be collected by behavioral training of the dolphin to exhale forcefully into a canister placed above the blowhole. Respiratory exudates from four dolphins over a period of eight months were examined as described below.

Material and Methods

Blowhole samples were collected from four dolphins which were behaviorally trained to exhale forcefully into a canister on a presentation of a signal. The samples were collected at least once every two weeks, or more often if necessary, over a period of eight months. To the sputum (blowhole sample collected in canster), 200 microliters of new methylene blue were added and the mixture was thoroughly mixed. Fifty to 100 microliters of the mixture was placed on a slide with cover slip and examined under a microscope, using 10X for eye pieces and 10X for the objective, giving a final magnification of 100. A total number of 200 cells (epithelial and white blood cells) were counted and a ratio of white blood cells (WBC) to epithelial cells was established.

Results

A total number of 98 samples were collected over a period of eight months. White blood cells were observed in seven out of 26, six out of 20, 10 out of 25, and 14 out of 21 samples from dolphins nos. 1, 2, 3 and 4, respectively. The ratio of WBC to epithelial cells ranged from 1:12 to 1:19 in dolphin no. 1; 1:4 to 1:99 in dolphin no. 2; 1:1 to 1:99 in dolphin no. 3; and 100:6 to 1:99 in dolphin no. 4. Cytology specimens with a WBC/epithelial cell ratio of greater than 1:1 are indicative of an active inflammatory process. In the case of dolphin no. 4, complete blood count was unremarkable. Vibrio parahemolyticus was isolated on culture. Serum iron was significantly reduced and blood sedimentation rate was moderately elevated. Antibiotic treatment was instituted.

Discussion

Training dolphins to exhale forcefully on a presentation of a signal into a canister was relatively simple and a minimally stressful method to collect respiratory exudate for analysis of inflammatory cells in the sputum. It proved to be a sensitive method for diagnosing respiratory infection and allowed early intervention with therapy. Most of the time either no or very few white blood cells were seen in the samples. However, in animal no. 4 there was a period when the number of WBC to epithelial cells was greater than 1:2, clearly indicating an infection. After the antibiotic treatment, the WBC to the epithelial cell ratio returned to normal. Our observations are in agreement with those of Sweeny, (4) where he has suggested that a ratio of 2:1 would be indicative of respiratory tract inflammation or infection.

Table 1 Blowhole samples from Dolphins nos. 1 through 4, showing the ratio of epithelial cells to white blood cells.