Composite Sampling Approaches for Bacillus anthracis Surrogate Extracted from Soil.

Brian France,Trudy Scholten,Emily Chang,William Bell,Raymond Schuch

doi:10.1371/journal.pone.0145799

Abstract

Any release of anthrax spores in the U.S. would require action to decontaminate the site and restore its use and operations as rapidly as possible. The remediation activity would require environmental sampling, both initially to determine the extent of contamination (hazard mapping) and post-decon to determine that the site is free of contamination (clearance sampling). Whether the spore contamination is within a building or outdoors, collecting and analyzing what could be thousands of samples can become the factor that limits the pace of restoring operations. To address this sampling and analysis bottleneck and decrease the time needed to recover from an anthrax contamination event, this study investigates the use of composite sampling. Pooling or compositing of samples is an established technique to reduce the number of analyses required, and its use for anthrax spore sampling has recently been investigated. However, use of composite sampling in an anthrax spore remediation event will require well-documented and accepted methods. In particular, previous composite sampling studies have focused on sampling from hard surfaces; data on soil sampling are required to extend the procedure to outdoor use. Further, we must consider whether combining liquid samples, thus increasing the volume, lowers the sensitivity of detection and produces false negatives. In this study, methods to composite bacterial spore samples from soil are demonstrated. B. subtilis spore suspensions were used as a surrogate for anthrax spores. Two soils (Arizona Test Dust and sterilized potting soil) were contaminated and spore recovery with composites was shown to match individual sample performance. Results show that dilution can be overcome by concentrating bacterial spores using standard filtration methods. This study shows that composite sampling can be a viable method of pooling samples to reduce the number of analysis that must be performed during anthrax spore remediation.

Highlights

If an airport or seaport is shut down by biological agent contamination, the economic loss for each missed day would be enormous; it is absolutely essential to restore operations as rapidly as possible
Potting soil is a level of sophistication towards actual environmental sampling. In this test we reduced the number of spores added to each individual sample, from 2.1x105 colony forming units (CFU) used on the Arizona test dust samples to under 2000 CFUs
In the event of widespread anthrax spore contamination, the time required to obtain and analyze the samples needed to identify the location of contamination and verify decontamination constitute a bottleneck in the recovery process

Summary

Introduction

If an airport or seaport is shut down by biological agent contamination, the economic loss for each missed day would be enormous; it is absolutely essential to restore operations as rapidly as possible Improved decon methods such as an electrochemical decon system (eClO2) produces 100% kill of anthrax spores in less than one minute [1]. If a pool tests positive, the individuals in that pool will be retested so that the infected individuals can be identified; if the pool is negative, a large amount of time is saved because only one test has to be run, rather than testing all samples in the pool (10, 100 or whatever pool size is selected) This pooling or composite sampling procedure can greatly reduce the analysis time and costs with no loss in accuracy [5]. In a wide area biological agent restoration event we could follow this procedure if desirable, but to save valuable time, the more likely course is to re-treat the area covered by the group that tests positive until decontamination is achieved

Methods

Results

Conclusion