Effects of inactivation methods on the analysis of Bacillus atrophaeus endospores using real-time PCR and MALDI-TOF-MS

A wide range of analytical methods are available for the detection and identification of biological warfare agents. These technologies are often hampered in their performance when the inactivated samples are analyzed. To work with pathogens outside of biosafety level 3 laboratories, a complete inactivation is mandatory when appropriate protection equipment is unavailable. When methods of inactivation are used, the detection of bacteria becomes more difficult. In contrast to measuring viable organisms, inactivation steps can have a massive impact on the intrinsic cellular information. This study examined the effects of autoclaving and chemical inactivation methods on Bacillus spores using biological warfare detection setups like real-time PCR and MALDI-TOF-MS. Here, the inactivation of Bacillus atrophaeus spores with formaldehyde, which is a suggested model for biological warfare spore agents, was compared with other inactivation reagents like Wofasteril (R) E400, a commercially available decontaminant based on peroxyacetic acid. With Wofasteril (R) E400 the critical factor of inactivation time was reduced to about 15 min and a limit of detection of 8500 spores by PCR was still measurable using five-times-washed spores. It has also been shown that MALDI-TOF-MS peak information can be hampered by inactivation methods.