In Pursuit of Protein Targets: Proteomic Characterization of Bacterial Spore Outer Layers

Bacillus cereus, responsible for food poisoning, and Clostridium difficile, the causative agent of Clostridium difficile-associated diarrhea (CDAD), are both spore-forming pathogens involved in food spoilage, food intoxication, and other infections in humans and animals. The proteinaceous coat and the exosporium layers from spores are important for their resistance and pathogenicity characteristics. The exosporium additionally provides an ability to adhere to surfaces eventually leading to spore survival in food. Thus, studying these layers and identifying suitable protein targets for rapid detection and removal of spores is of the utmost importance. In this study, we identified 100 proteins from B. cereus spore coat, exosporium and 54 proteins from the C. difficile coat insoluble protein fraction. In an attempt to define a universal set of spore outer layer proteins, we identified 11 superfamily domains common to the identified proteins from two Bacilli and one Clostridium species. The evaluated orthologue relationships of identified proteins across different spore formers resulted in a set of 13 coat proteins conserved across the spore formers and 12 exosporium proteins conserved in the B. cereus group, which could be tested for quick and easy detection or targeted in strategies aimed at removal of spores from surfaces.蜡样芽孢杆菌（Bacillus cereus）可引发食物中毒，艰难梭菌（Clostridium difficile）则是艰难梭菌相关性腹泻（Clostridium difficile-associated diarrhea, CDAD）的致病菌，二者均为产芽孢病原菌，可导致食品腐败、食物中毒以及人和动物的其他感染性疾病。芽孢的蛋白质衣壳（proteinaceous coat）与芽胞外膜（exosporium）层对于其抗性与致病特性至关重要。芽胞外膜还可赋予芽孢黏附于物体表面的能力，最终使其得以在食品环境中存活。因此，研究这些结构层并筛选适用于芽孢快速检测与清除的蛋白靶点具有极高的研究价值。本研究从蜡样芽孢杆菌的芽孢衣壳、芽胞外膜中鉴定出100种蛋白质，同时从艰难梭菌芽孢衣壳的不溶性蛋白组分中鉴定出54种蛋白质。为了确定一套通用的芽孢外层蛋白集合，研究人员在这两种芽孢杆菌与一种梭菌的已鉴定蛋白中，找到了11个共有的超家族结构域。通过对不同产芽孢菌的已鉴定蛋白进行直系同源（orthologue）关系分析，最终得到了13种在所有产芽孢菌中保守的芽孢衣壳蛋白，以及12种在蜡样芽孢杆菌群中保守的芽胞外膜蛋白——此类蛋白均可用于开发便捷快速的检测手段，或是作为靶向策略清除物体表面的芽孢。