Canine distemper virus antibodies in the Asian elephant (Elaphas maximus)

FREE-RANGING domestic dogs are frequently suspected of acting as disease reservoirs for wildlife species. Few community dogs in Thailand are vaccinated (Mitmoonpitak and others 1997) and they may therefore provide a source of infection for other species (Cleaveland and others 2000). Canine distemper virus (CDV) is capable of infecting many wildlife species, including members of the Felidae, Procyonidae, Mustelidae, Ursidae, Ailuridae, Phocidae and Tayassuidae (Appel and others 1994, Anderson 1995, Mamaev and others 1996, Kennedy and others 2000, Noon and others 2003). Some species, such as the Suidae, can mount an antibody response to the virus without developing clinical disease (Appel and Summers 1995). It is likely that this is also the case in the Asian elephant (Elaphas maximus), as no clinical signs of distemper have previously been reported in this species. This short communication describes a serological survey for antibodies to CDV in Asian elephants in Thailand. Serum samples were collected in June and July 2004 from Asian elephants in seven different tourist camps: 75 from Kanchanaburi province, and 69 from Chiang Mai and Lampang provinces. Questionnaires were used at the time of sampling to assess the presence of dogs within each camp and their vaccination status, although no records were made to determine the prevalence of clinical disease. A protein G ELISA was used to detect anti-CDV immunoglobulin G (IgG) in the elephant sera, using a modified version of the method described by Takikawa and others (1996). Positive antigen was prepared using CDV vaccine (Caniffa DH Bivirovax; Merial) inoculated into Vero cells. Virus-infected cells were scraped into Dulbecco’s modification of Eagle’s medium (DMEM) containing 5 per cent fetal calf serum (Gibco). Test serum was diluted to 1:50, and viral and control antigens were diluted to 1:80. Optical densities (ODs) were read using an ELISA plate reader (Lucy 2; Anthos) at a wavelength of 650 nm. The ELISA value for each sample was taken as the OD of the viral antigen minus the OD of the control antigen. Of the 144 serum samples, 25 (17 per cent) had a final ELISA OD value greater than 0·1. The virus neutralisation (VN) test (OIE 1996) was applied to 10 of these samples to establish a cut-off value. All samples were negative for neutralising antibodies, with the exception of two samples with an OD value greater than or equal to 0·2. Endpoint serum titres were determined as 1:20 and 1:320. There are no previous reports of testing for CDV antibodies in the Asian elephant, and thus there are no published cut-off values for infection. The VN test is considered a reference standard for determining titre levels of CDV antibodies (Bohm and others 2004), and was used to determine a cutoff value for positive samples in this study. The two positive samples had an OD cut-off value greater than 0·45 for CDV antibodies. It is interesting to note that the two VN-positive samples were collected from elephants in camps in Chiang Mai province, where there were no dogs present, so it is difficult to be certain about the source of the CDV. However, many of the elephants in tourist camps usually pass through different areas before purchase, and these elephants may have come into contact with dogs at any time. One sample taken from Kanchanaburi province had an OD value greater than 0·45, yet it was VN negative. The reason for this is unclear. Two samples had an OD control value that was higher than the OD virus value. This may indicate some degree of reaction to non-CDV proteins, for example, in the Vero cell supernatant, or ELISA plate contamination. Alternatively, there may have been cross-reaction with another morbillivirus, such as measles virus transmitted from the mahouts (elephant trainers) or their families (Ariyasriwatana and Kalayanarooj 2004). The measles virus is antigenically related to CDV (Orvell and Norrby 1980, Sheshberadaran and others 1986) and may provide limited immunity to distemper (Carlstrom 1956, Millian and others 1960). Thus, the results of serological testing of Asian elephants may not necessarily reflect actual exposure and infection. Further studies such as virus isolation are needed to confirm the presence of CDV and investigate the nature of the antigen-antibody reaction in the elephants.