Rapid & novel microscopy technique to detect germination initiation and specificity in Bacillus spores

Detection and elimination of Bacillus spp. spores is a challenging task being faced by food industry and human life. Spore eradication techniques such as U.V., heat and other stress treatments fails due to dormant and robust nature of spores. Exposure of spores to specific germinants trigger germination and vegetative cells are formed which are metabolically very active and fragile so can be easily eliminated from the site. Mechanistic details for the initiation and specificity of Bacillus spores towards germinants are required. A rapid, simple and inexpensive technique based on fluorescent labeling has been developed for detection of onset and specificity of germination in B. megaterium spores. This technique requires heating the spore suspension followed by addition of germinant and treating it with fluorescent dye diacetate fluorescein which is cell viability stain and is cleaved effectively by germinating spores as compared to dormant spores. Image analysis with EMCCD allows for automated data processing for dormant and germinating spores. A typical increase in fluorescence was observed when spores were germinated with D-Dextrose within minutes. There is a high level of specificity being expressed by spores towards germinants as revealed by their respective florescence pattern. © 2013 Universal Research Publications. All rights reserved