Relationship between the germination and spore wall proteins in Nosema bombycis

To study the composition of spore wall proteins(SWPs)in Nosema bombycis and the relevance with spore germination,we firstly developed a method called GDGC,which is spores germination in vitro activated by 0.1 mol/L K_2CO_3,combined with Density Gradient Centrifugation to purify the germinated spore coats.Using this method,we obtained the spore coats,and then comparatively analyzed the composition of SWPs among the supernatant after spore germination,the purified spore coats and normal spores.Results showed that the highly pure spore coats were acquired with GDGC method,and their density is 1.113 g/cm3.The major spore wall proteins including SWP32,SWP30 and SWP25 can be extracted from the purified spore coats.However,the concentration of SWP32 and SWP25 was decreased.In addition,the SWP32 was also detected by the electrophoresis analysis of spore wall proteins from the germinated supernatant.Moreover,LC-MS/MS data also displayed that three major spore wall proteins were existed in the germinated supernatant.To confirm the SWP32 and SWP25 are relevant to spore germination,the frozen spores were treated with 0.1 mol/l K_2CO_3,and no spores were germinated.Besides,the results of the electrophoresis analysis showed that only the SWP30 protein appeared in 0.1 mol/l K_2CO_3 solution.These data suggested that the solution of SWP32 and SWP25 from the spore coat could be not related to alkali liquor but concerned with the spore germination.