Characterization of Heat Shock Response in Brucella melitensis and Interaction of Heat Shock Protein with Sick and Healthy Sera

Brucella is one of the most important zoonotic diseases and is caused by the members of Brucella genus especially B. melitensis. The bacteria begin to synthesize heat shock proteins(hsp) when facing elevated temperatures. In this investigation, clinical isolates of B. melitensis were subjected to heat shocks and the hsps produced were surveyed by SDS-PAGE electrophoresis. The immunogenicity of hsp-60 was then investigated in both sick and healthy sera by Western blot. In this analytical descriptive study, five B. melitensis isolated from sick people were cultured. The bacterial isolates were subjected to 39, 40, and 42˚C heat shocks and after lysing the cells by lysozome, cell proteins were extracted by SDS(sodium dodecyl sulfate). The extracted proteins were exposed to electrophoresis in SDS-PAGE followed by staining with Coomasie Blue. Finally, antibodies against hsp-60 in control as well as sick sera were surveyed by Western blot. SDS-PAGE gels revealed protein bands mainly in the range of 10-100 KDa. The major protein groups were in the range of 45-75, 20-30, and 14-20 KDa. The amount of 60 KDa protein band(hsp-60) was significantly enhanced following heat shock in comparison to unheated cells. The sera from Brucellosis patients reacted with several of these cell-derived protein bands in Western blots, none of which were reactive with the sera from healthy individuals. These reactive proteins were in the range of 10, 60, and 100 KDa. The 60 KDa band was the most significant one and showed strong reactions with all Brucellosis serum samples. Significant differences in protein bands were detected by the electrophoresis of the cells subjected to 39 and 40˚C heat shocks in comparison to unheated bacteria. The SDS-PAGE results indicated that Brucella melitensis begins to synthesize heat shock proteins when facing elevated temperatures. The Western blot protein bands of the heat shocked bacteria incubated with sera from sick and healthy individuals showed striking differences. This observation points to the immunogenic properties of hsps, especially the overwhelming response to hsp-60. Therefore, hsp-60 can be a good antigen candidate for ELISA test development as well as for engineering subunit vaccine against Brucella.