DETECTION OF BACILLUS-ANTHRACIS SPORES IN SOIL BY PCR

A polymerase chain reaction (PCR)-based protocol was developed for rap id detection of Bacillus anthracis spores in various types of soil. El even soil samples representing miscellaneous vegetation types were mix ed with anthrax spores and incubated overnight. In no case could the s pores of the mixtures be detected directly by PCR. The PCR was based o n primers derived from the capsule-encoding cap genes of the pXO2 plas mid. Therefore, the focus of the study became to develop a suitable me thod for preparation of DNA that would allow rapid detection of the an thrax spews. Various published techniques were investigated, but none enabled detection Of the spores from a majority of the samples. A succ essful Protocol was eventually developed based on a combination of a p ublished preparative technique and a form of nested PCR. First, amplif ication by PCR was performed with a degenerate primer and then samples were diluted and subjected to a second round of amplification using t he B. anthracis-specific cap primers. The protocol is rapid and result ed in DNA of sufficient purity from. all of the 11 investigated soil s amples to permit detection of B. anthracis spores.