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. 2020 Jan;128(1):124-137.
doi: 10.1111/jam.14474. Epub 2019 Nov 3.

Combining spore germination and heat inactivation to decontaminate materials contaminated with Bacillus anthracis spores

T L Buhr  1 , Z A Minter  1 , N L Kennihan  1 , A A Young  1 , E L Borgers-Klonkowski  1 , E B Osborn  1 , M D Bohmke  1 , S M Hamilton  1 , M B Kimani  1 , C T Miller  1 , R S Mackie  1 , J M Innocenti  1 , M D Bensman  1 , S D Lilly  1
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Combining spore germination and heat inactivation to decontaminate materials contaminated with Bacillus anthracis spores

T L Buhr et al. J Appl Microbiol. 2020 Jan.

Abstract

Aims: To add a spore germination step in order to reduce decontamination temperature and time requirements compared to the current hot, humid air decontamination parameters, which are 75-80°C, ≥72 h, 70-90% RH, down to ≤60°C and ≤24 h total decontamination time.

Methods and results: Bacillus anthracis spore germination with l-alanine+inosine+calcium dipicolinate (CaDPA) was quantified at 0-40°C, several time points and spore concentrations of 5-9 log10 per ml. Germination efficiency at 0-40°C was >99% at <8 log10 spores per ml. The temperature optimum was 20°C. Germination efficiency was significantly higher but slower at 0°C compared to ≥30°C at ≥8 log10 spores per ml. A single germinant application followed by 60°C, 1-h treatment consistently inactivated >2 log10 (>99%) of spores. However, a repeat application of germinant was needed to achieve the objective of ≥6 log10 spore inactivation out of a 7 log10 challenge (≥99·9999%) for ≤24 h total decontamination time for nylon and aircraft performance coating.

Conclusions: l-alanine+inosine+CaDPA stimulated germination across wide temperature and spore concentration ranges.

Significance and impact of the study: Germination expands the scope of spore decontamination to include materials from any industry sector that can be sprayed with an aqueous germinant solution.

Keywords: Bacillus anthracis; alanine racemase; decontamination; germination; germination inhibition; hot humid air; inosine hydrolase; persister; spore; surrogate.

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