Gamma Irradiation Can Be Used To Inactivate Bacillus anthracis Spores without Compromising the Sensitivity of Diagnostic Assays

The use of Bacillus anthracis as a biological weapon in 2001 heightened awareness of the need for validated methods for the inactivation of B. anthracis spores. This study determined the gamma irradiation dose for inactivating virulent B. anthracis spores in suspension and its effects on real-time PCR and antigen detection assays. Strains representing eight genetic groups of B. anthracis were exposed to gamma radiation, and it was found that subjecting spores at a concentration of 10⁷ CFU/ml to a dose of 2.5 x 10⁶ rads resulted in a 6-log-unit reduction of spore viability. TaqMan real-time PCR analysis of untreated versus irradiated Ames strain (K1694) spores showed that treatment significantly enhanced the detection of B. anthracis chromosomal DNA targets but had no significant effect on the ability to detect targets on the pXO1 and pXO2 plasmids of B. anthracis. When analyzed by an enzyme-linked immunosorbent assay (ELISA), irradiation affected the detection of B. anthracis spores in a direct ELISA but had no effect on the limit of detection in a sandwich ELISA. The results of this study showed that gamma irradiation-inactivated spores can be tested by real-time PCR or sandwich ELISA without decreasing the sensitivity of either type of assay. Furthermore, the results suggest that clinical and public health laboratories which test specimens for B. anthracis could potentially incorporate gamma irradiation into sample processing protocols without compromising the sensitivity of the B. anthracis assays.