A vaccination and challenge model using calcein marked fish

A vaccination and challenge cohabitation model was established and evaluated using Nile tilapia (Oreochromis niloticus), the fluorescent chromophore calcein, and a Streptococcus iniae vaccine. Tilapia were non-invasively calcein marked, sham-vaccinated (CMSV) and cohabited with non-marked sham-vaccinated (NMSV) or non-marked S. iniae vaccinates (NMV) as a single unit. After 30 d, the cohabitants were challenged with a virulent isolate of S. iniae by intraperitoneal (ip) injection and the cumulative mortality was measured over a period of 15 d. Calcein marking did not have a significant effect on S. iniae susceptibility as mortality of CMSV and NMSV was not significantly different (P = 0.6756). Nor did calcein marking have an effect on the vaccination and challenge cohabitation model. The results showed that the cumulative mortality of CMSV (N = 160) was significantly greater (P < 0.0003) than those of NMV (N = 160). The results of the calcein marking trials indicate that the most suitable calcein concentration and exposure time to produce detectable fluorescent marking of tilapia was 500 mg L−1 for 4 h. Furthermore, the calcein marks were readily visible in the calcified skeletal structures of head and fins using a portable handheld UV lamp set at 365 nm wavelength. Calcein appears to be a valuable tool for non-invasive, non-lethal, non-stressful, mass marking of fish to differentiate between sham- and pathogen-vaccinated fish in this cohabitation model. The vaccination and challenge cohabitation model also offers the statistical advantage of using individual fish as the experimental unit maintained in the same aquarium.