Agglutination Method for Fast Identification bacteria Aeromonas hydrophila on Freshwater Fish¹⁾

Aeromonas hydrophila are gram-negative bacteria that are very harmful on freshwater aquaculture activity. This bacterium attacks causes the MAS (Motile Aeromonas Septicemia) disease which is fatal. Signs of this disease can be seen from the clinical damage on body of the fish, but this characteristic cannot be the basis, because of many other diseases attack that causes similar characteristics. Further identification conducted by the conventional method (biochemical and morphological test) on pure culture from isolation results. Obstacles of conventional testing are the high degree of difficulty in implementation, and require considerably long time. Moreover, because depending on the skill of each analyst, the results of conventional tests often there difference on reading of final result (because many variables tested).

More accurate method in detecting the presence of bacteria Aeromonas is the agglutination method. This method utilizes a defense mechanism of specific antibody against antigen which is produced by Aeromonas hydrophila bacteria. Because of specific characteristic, antigen from certain bacteria will only react to particular type of antibody as well. The mechanism is connection between the lock with a key. The resulting reaction is clumping (agglutination) which is a form of the system immune response. In the agglutination method, specific antibody reacted with purified isolate of A. hydrophila bacteria was on a glass slide. If those bacteria are A. hydrophila will be formed clots on glass slide, whereas if negative mixture of antibody and bacteria will remain homogeneous. Agglutination reaction that occurs is specific for A. hydrophila bacterial antigens, if other bacteria although both gram negative will not cause clots on glass slide. This characteristic distinguishing and accurately guarantee type of bacteria which is we identify.

A. hydrophila specific antibody obtained from rabbit blood serum isolation that has been injected with weakened A. hydrophila bacteria. Specific antibody production stages starting with debilitating pure isolate of A. hydrophila bacteria with formalin and hot temperatures to produce the antigen substance A. hydrophila (Ag.A). Furthermore antigen diluted with physiological solution until 10⁹ concentrations (standard McFarland). Ag.A injected into rabbit with gradual dose every week for duration 4 weeks (0.5 ml, 1 ml, 2 ml, 3 ml). Rabbit blood serum harvest that containing specific antibody conducted at week V. Storage serum in 0.1% Sodium azide can be used in testing bacteria for duration 1 year.

Keywords; Aeromonas hydrophila, identification, antigen, specific antibody, agglutination