Rastogi Vipin K, Smith Lisa S, Wallace Lalena, Tomasino Stephen F
U.S. Army, Edgewood Chemical Biological Center, BioDefense Team, R & T Directorate, Aberdeen Proving Grounds, MD 21010, USA.
J AOAC Int. 2013 Sep-Oct;96(5):947-50. doi: 10.5740/jaoacint.13-031.
The AOAC Quantitative Three Step Method (TSM; AOAC Official Method SM 2008.05) is validated for testing the efficacy of liquid sporicides against spores of Bacillus subtilis and Bacillus anthracis on selected hard, nonporous, and porous surfaces. The TSM uses 5x5x1 mm inoculated coupons (carriers), which are placed in 400 microL liquid sporicidal agent contained in a microcentrifuge tube. Following exposure of inoculated carriers to the test chemical and subsequent neutralization, viable spores are recovered in three fractions: A (gentle tapping), B (sonication), and C (gentle agitation). The spores in suspension are serially diluted and plated on a recovery medium for enumeration. The plate counts are summed over the three fractions to provide the number of viable spores per carrier, which is log10-transformed to generate a mean log density (LD) value across carriers. As a measure of product efficacy, a log reduction (LR) value is calculated by subtracting the mean LD for treated carriers from the mean LD for control carriers. This paper reports on the comparative evaluation of the current and modified versions of the TSM in order to support a modification to simplify the procedure. The proposed modified TSM (mTSM) consolidates fractions B and C in the same tube. Thus, the sonication (fraction B) and gentle agitation (fraction C) steps are carried out in the same tube, thereby reducing the number of tubes and associated resources and time necessary to complete the test. Glass, steel, pine wood, and ceramic tile carriers were included in the comparative study. Inoculated carriers were evaluated against two preparations of sodium hypochlorite to generate two presumed levels of efficacy (intermediate and high); the control LD and LR values associated with testing each carrier type for the TSM and the mTSM were compared. For control carriers, the mean log densities per carrier (for each carrier material) were not significantly different based on the TSM compared to the mTSM. Furthermore, the treated carrier data showed comparable LR values for the TSM and mTSM. The data provided in this report demonstrate equivalency between the TSM and mTSM and support the proposed procedural modification to consolidate fractions B and C.
美国官方分析化学师协会（AOAC）定量三步法（TSM；AOAC官方方法SM 2008.05）已通过验证，用于测试液体杀孢子剂对选定的坚硬、无孔和多孔表面上枯草芽孢杆菌和炭疽芽孢杆菌孢子的杀灭效果。TSM使用5×5×1毫米的接种试片（载体），将其置于微量离心管中含有的400微升液体杀孢子剂中。接种的载体接触测试化学品并随后进行中和后，存活孢子以三个部分回收：A（轻轻敲击）、B（超声处理）和C（轻轻搅拌）。悬浮液中的孢子进行系列稀释并接种在回收培养基上进行计数。将三个部分的平板计数相加，得出每个载体上存活孢子的数量，将其进行以10为底的对数转换，以生成各载体的平均对数密度（LD）值。作为产品效果的衡量指标，通过从对照载体的平均LD中减去处理后载体的平均LD来计算对数减少（LR）值。本文报告了TSM当前版本和改进版本的比较评估，以支持简化该程序的修改建议。提议的改进TSM（mTSM）将部分B和C合并在同一管中。因此，超声处理（部分B）和轻轻搅拌（部分C）步骤在同一管中进行，从而减少了完成测试所需的管数以及相关资源和时间。比较研究中包括玻璃、钢、松木和瓷砖载体。对接种的载体针对两种次氯酸钠制剂进行评估，以产生两种假定的效果水平（中等和高）；比较了TSM和mTSM测试每种载体类型时的对照LD和LR值。对于对照载体，基于TSM与mTSM相比，每个载体（每种载体材料）的平均对数密度没有显著差异。此外，处理后载体的数据显示TSM和mTSM的LR值相当。本报告提供的数据证明了TSM和mTSM之间的等效性，并支持将部分B和C合并的提议程序修改。
