Submitted to: ASM Biodefense Research Meeting
Publication Type: Abstract only
Publication Acceptance Date: 2/8/2005
Publication Date: 3/20/2005
Citation: Hang, J., Sundaram, A., Shelton, D.R., Karns, J.S., Zhu, P., Amstutz, P., Tang, C. 2005. Rapid Germination and Growth of Bacillus Anthracis. ASM Biodefense Research Meeting, March 20-23, 2005, Baltimore, MD. p.60. Interpretive Summary:
Technical Abstract: Bacillus anthracis spore is a major threat as an agent of bioterrorism, due to its lethal potency and ease with which spores can be aerosolized. Creatv MicroTech is currently developing biosensor instruments and assays for rapid detection of B. anthracis spores that include a culture step, in order to verify viability and to increase the number of genetic copies for subsequent PCR identification. Various heat shock temperatures and time durations were tested and germinants evaluated. Different combinations of culture broths and supplements were evaluated at three growth temperatures for the most rapid growth of vegetative cells. Microscopic observation of plated cells in early growth revealed that cells tend to remain attached, forming long chains. Consequently, a quantitative real-time PCR technique was developed to provide more accurate estimates of growth rates than is possible with conventional colony counting methods, as described in a separate poster entitled 'A Biosensor Platform for Genetic Identification and Quantitation of Bacillus anthracis.' In solutions containing selected germinants, almost 100% of spores germinated within ten minutes. Brain Heart Infusion (BHI) at 37°C was selected as the preferred medium, because it allowed the germination and growth steps to be conducted concomitantly in one procedure. Three supplements, 3-amino-L-tyrosine, luminol and bicarbonate, significantly accelerated cell growth in TSB; however, they only provided minor enhancement in BHI. The effects of heat shock on spore germination and cell growth were also tested. No significant difference was observed in the germination or growth rates for spores that were heat-treated, as compared to spores that were not heat-treated, other than that heating at 80°C for longer than 5 minutes did inhibit growth. For the culture step of Creatv's biosensor system, BHI at 37°C will be used for germination and growth without heat shock.