Comparison of sampling methods to recover germinated Bacillus anthracis and Bacillus thuringiensis endospores from surface coupons.

Aims: In an attempt to devise decontamination methods that are both effective and minimally detrimental to the environment, we evaluated germination induction as an enhancement to strategies for Bacillus anthracis spore decontamination. To determine an optimal method for the recovery of germinating spores from different matrices, it was critical to ensure that the sampling procedures did not negatively impact the viability of the germinating spores possibly confounding the results and downstream analyses of field trial data.
Methods and Results: Therefore, the two main objectives of this study were the following: (i) development of an effective processing protocol capable of recovering the maximum number of viable germinating or germinated spores from different surface materials; and (ii) using a model system of spore contamination, employ this protocol to evaluate the potential applicability of germination induction to wide-area decontamination of B. anthracis spores. We examined parameters affecting the sampling efficiencies of B. anthracis and the surrogate species Bacillus thuringiensis on nonporous and porous materials.
Conclusions: The most efficient extraction from all matrices was observed using PBS with 0·01% Tween 80 extraction buffer. The addition of a sonication and/or extended vortex treatment did not yield significant increases in spore or germinated spore recovery.
Significance and Impact of the Study: Our data demonstrate that previous germination-induction experiments performed in suspension can be reproduced when Bacillus spores are deposited onto reference surfaces materials. Our proof of concept experiment illustrated that a germination pretreatment step significantly improves conventional secondary decontamination strategies and remediation plans.
(Published 2017. This article is a U.S. Government work and is in the public domain in the USA.)