Impact of processing method on recovery of bacteria from wipes used in biological surface sampling

Autumn S. Downey, Sandra M. Da Silva, Nathan D. Olson, James J. Filliben and Jayne B. Morrow


Environmental sampling for microbiological contaminants is a key component of hygiene monitoring and risk characterization practices used to ensure public health and product integrity.  The reliability of traditional surface sampling methods came under intense scrutiny in 2001 following the widespread contamination of multiple large building complexes with spores identified as Bacillus anthracis.  The qualitative results primarily reported during that event were found to be insufficient for building characterization to assess source and extent of contamination as well as decontamination effectiveness.  In addition to biodefense-related surface sampling needs, emerging hygiene requirements in food production and clinical settings are also fueling demand for improved reliability and efficiency of traditional surface sampling methods.  As a result, over the last decade needs for quantitative assessment capabilities have driven research on the relative effectiveness of different sampling processes as well as the development of validated surface sampling and sample processing methods.  However, confidence in surface sampling results, both in the field and in controlled laboratory studies, has been undermined by large variation in sampling performance results.  Sources of variation include controlled parameters such as sampling materials and processing methods, which often differ among studies, as well as random and systematic error; however, the relative contributions of these factors remain unclear.  The objective of this study was to determine the relative impacts of sample processing methods, including extraction solution and physical dissociation method (vortexing and sonication), on recovery of Gram-positive (Bacillus cereus) and Gram-negative (Burkholderia thailandensis and Escherichia coli) bacteria from directly inoculated wipes.  This work showed that target organism had the largest impact on extraction efficiency and recovery precision, as measured by traditional colony counts.  Physical dissociation method (PDM) had negligible impact, while the effect of the extraction solution was organism dependent.  Overall however, extraction of wipes in PBS with 0.04% Tween 80 (PBST) resulted in highest mean recovery across all three organisms.  The observed differences in recovery between extraction methods and microorganisms could not be attributed to loss of cell viability.  The results from this study contribute to a better understanding of the factors that influence sampling performance, which is prerequisite to the intelligent design and optimization of robust methods that meet multidisciplinary surface sampling needs.