Forensic application of microbiological culture analysis to identify mail intentionally contaminated with Bacillus anthracis spores

Abstract

The discovery of a letter intentionally filled with dried Bacillus anthracis spores in the office of a United States senator prompted the collection and quarantine of all mail in congressional buildings. This mail was subsequently searched for additional intentionally contaminated letters. A microbiological sampling strategy was used to locate heavy contamination within the 642 separate plastic bags containing the mail. Swab sampling identified 20 bags for manual and visual examination. Air sampling within the 20 bags indicated that one bag was orders of magnitude more contaminated than all the others. This bag contained a letter addressed to Senator Patrick Leahy that had been loaded with dried B. anthracis spores. Microbiological sampling of compartmentalized batches of mail proved to be efficient and relatively safe. Efficiency was increased by inoculating culture media in the hot zone rather than transferring swab samples to a laboratory for inoculation. All mail sampling was complete within 4 days with minimal contamination of the sampling environment or personnel. However, physically handling the intentionally contaminated letter proved to be exceptionally hazardous, as did sorting of cross-contaminated mail, which resulted in generation of hazardous aerosol and extensive contamination of protective clothing. Nearly 8 x 10(6) CFU was removed from the most highly cross-contaminated piece of mail found. Tracking data indicated that this and other heavily contaminated envelopes had been processed through the same mail sorting equipment as, and within 1 s of, two intentionally contaminated letters.

FIG. 1.

Diagram of the negative pressure isolation facility. The workroom dimensions were 25 by 22 by 5 m. Solid rectangles, class II type A biological safety cabinets; solid circles, monitoring of exhaust air using SBA plates during handling of exposed mail (see Materials and Methods for details); open circles, barrels; large open rectangles, work tables; small open rectangles, air handlers with HEPA filters; lines from air handlers, air exhaust or supply hoses (diameter, 30.5 cm) (arrows indicate the direction of airflow); solid squares labeled S1 to S6, slit-to-agar air samplers; large shaded rectangle, loading bay separated by walls from the workroom. The drawing is to scale, and the locations of items (except barrels, which changed positions) were determined by triangulation.

FIG. 2.

Detection of airborne B. anthracis spores during handling of mail. The bars indicate the numbers of colonies discernible on sections of slit-to-agar sample plates representing 100 liters of sampled air (3.3-min exposure) at the time of day indicated. Graphs S1 to S4 show the results for air samplers S1 to S4 (Fig. 1). The x axis begins at 9:00 a.m. and ends at 6:00 p.m. Air monitoring ceased at 5:30 p.m.