False Negative Rates of a Macrofoam-Swab Sampling Method with Low Surface Concentrations of Two Bacillus anthracis Surrogates via Real-Time PCR

Hutchison, Janine R.¹  Piepel, Gregory F.¹  Amidan, Brett G.¹  Sydor, Michael A.¹  Kaiser, Brooke L.D.¹ 

[1]Pacific Northwest National Lab. (PNNL), Richland, WA (United States)

关键词: Bacillus anthracis;    Macrofoam swab sampling;    Polymerase chain reaction;    False negative rate;    Limit of detection;   

DOI  :  10.2172/1260869
RP-ID  :  PNNL--24204 Rev. 1
PID  :  OSTI ID: 1260869
Others  :  Other: 400904120

学科分类：生物科学（综合）

美国|英语

来源: SciTech Connect

【 摘 要 】

Surface sampling for Bacillus anthracis spores has traditionally relied on detection via bacterial cultivation methods. Although effective, this approach does not provide the level of organism specificity that can be gained through molecular techniques. False negative rates (FNR) and limits of detection (LOD) were determined for two B. anthracis surrogates with modified rapid viability-polymerase chain reaction (mRV-PCR) following macrofoam-swab sampling. This study was conducted in parallel with a previously reported study that analyzed spores using a plate-culture method. B. anthracis Sterne (BAS) or B. atrophaeus Nakamura (BG) spores were deposited onto four surface materials (glass, stainless steel, vinyl tile, and plastic) at nine target concentrations (2 to 500 spores/coupon; 0.078 to 19.375 colony-forming units [CFU] per cm2). Mean FNR values for mRV-PCR analysis ranged from 0 to 0.917 for BAS and 0 to 0.875 for BG and increased as spore concentration decreased (over the concentrations investigated) for each surface material. FNRs based on mRV-PCR data were not statistically different for BAS and BG, but were significantly lower for glass than for vinyl tile. FNRs also tended to be lower for the mRV-PCR method compared to the culture method. The mRV-PCR LOD95 was lowest for glass (0.429 CFU/cm2 with BAS and 0.341 CFU/cm2 with BG) and highest for vinyl tile (0.919 CFU/cm2 with BAS and 0.917 CFU/cm2 with BG). These mRV-PCR LOD95 values were lower than the culture values (BAS: 0.678 to 1.023 CFU/cm2 and BG: 0.820 to 1.489 CFU/cm2). The FNR and LOD95 values reported in this work provide guidance for environmental sampling of Bacillus spores at low concentrations.