Citation

Bacillus anthracis is regarded as a major biological warfare threat. The inhalation form of Bacillus anthracis infection can rapidly lead to a blood infection and kill quickly. Antibiotic treatment can clear the bacterium from the host, but this treatment must be initiated rapidly. By the time clinical symptoms are observed, the toxin, which is rapidly produced, may already be present in lethal amounts. There is a critical need for a rapid, accurate, sensitive and simple assay to determine whether infection has occurred thereby permitting immediate treatment. The presence in the blood of one of the virulence factors, anthrax lethal factor (LF), early in an infection, offers the opportunity for detection prior to catastrophic decline of the patient. This report describes a fast, sensitive, specific and accurate detection method to determine active infection by Bacillus anthracis in plasma. LF is detected using a fluorescently labeled peptide substrate, MAPKKide Plus (US Patent No. US 9,932,570 B2), which is not cleaved by plasma proteases and thus is specific for LF. Three detection strategies have been evaluated. In the simplest method, the substrate is added directly to diluted plasma, and cleavage is monitored by the increase in fluorescence as a function of time. The limit of detection by this method is 25 ng lethal factor/ml of plasma in 15 minutes, 5 ng/ml after 45 minutes, and