This study was to determine the median lethal concentration (LC50) of copper to Javenese carp, Puntius gonionotus (Bleeker), and the immune response after the fish were exposed to sublethal levels of copper and challenged with formalin killed Aeromonas hydrophila. The LC50 of copper on P. gonionotus at 24, 48, 72, 96 and 120 h were estimated as 2.17, 0.91, 0.57, 0.53 and 0.42 mg l(-1), respectively. To determine the effect of copper on the immune system, fish were exposed for 66 days to 0.05, 0.10 and 0.15 mg Cu l(-1). After 56 days of initial exposure to copper, fish were challenged with 0.1 ml of 4.5 x 10(5) cfu ml(-1) formalin killed A. hydrophila and maintained in the same concentration of copper. After the challenge, the immune response was monitored for 2 weeks using haematological and serological assays. During the initial phase of exposure to copper, significant changes were noted in the white blood cell, lysozyme, potential killing activity, total plasma protein, total immunoglobulin and haematocrit levels between the control and treated fish. One week after challenge with A. hydrophila, there was a significant increase in the values of white blood cells, total protein and total immunoglobulin compared to the values before the challenge. However, these values were not significantly different (P>0.05) between the control and the treated fish. In contrast, NBT and lysozyme assays exhibited a significant difference (P<0.05) in fish exposed to 0.10 mg Cu l(-1) (0.525 +/- 0.17; 24.42 +/- 3.35 x 10(2) micromg ml(-1)) and 0.15 mg Cu 1(-1) (0.536 +/- 0.19; 21.78 +/- 1.29 x 10(2) micromg ml(-1)) compared to the control (0.746 +/- 0.31; 30.73 +/- 5.42 x 10(2) micromg ml(-1)) after the bacterial challenge (day 61). There was however no significant difference (P>0.05) in NBT and lysozyme levels in fish exposed to lower level of copper (0.05 mg Cu l(-1)), suggesting the absence of immunosuppressive effects at lower level of exposure.
The production of tilapia Oreochromis spp. is rapidly growing throughout the world, but atypical motile aeromonad septicemia (MAS) is a current threat to the tilapia farming industry. The etiological agent of this disease is usually Aeromonas hydrophila. Mortality rates due to MAS are frequently high, resulting in a devastating negative impact on this industry worldwide; therefore, proper control measures regarding both prevention and treatment are necessary. Although vaccines against MAS for tilapia are available, their effectiveness is entirely dependent on the specific strain of problematic bacteria. Until now, whole-cell inactivated A. hydrophila vaccines for tilapia have exhibited the highest level of protection over live attenuated and recombinant vaccines. Among the various vaccine administration systems, only intraperitoneal (i.p.) injections of the A. hydrophila vaccine into tilapia were found to provide prominent immune protection. Vaccine efficacy was primarily measured by using the i.p. injection challenge model and estimating the relative percent survival of the immunized tilapia. Freund's incomplete adjuvant showed to be the most effective for tilapia MAS vaccines. In this review, multiple factors that directly or indirectly influence the efficacy of MAS vaccines for tilapia (adjuvants, challenge models, immunization doses and duration, and size of vaccinated fish) are discussed.