Isolation of anthrax spores from the environment and ecological investigation of Bacillus anthracis in endemic regions of northern Canada

Bacteriological studies of anthrax endemic regions in northern Canada were initiated in order to determine the extent of anthrax spore contamination and to examine the ecology of Bacillus anthracis spores. Efforts to develop a field immunoassay to detect viable anthrax spores proved unsuccessful and experiments were undertaken to investigate ways to improve the sensitivity of selective polymyxin, lysozyme, ethylenediaminetetraacetic acid, thallium acetate (PLET) medium In laboratory tests with seeded sterile soil, solutions of high specific gravity sucrose plus nonionic detergent extracted significantly more spores than the standard water extraction used with PLET medium.; PLET medium was evaluated for recovery of spores from a number of environmental and clinical strains of Bacillus species. While inhibitory to most non-anthracis Bacillus, the medium supported the growth of B. anthracis ATCC 4229, B. mycoides MU 711/84, B. thuringiensis QC 12093, B. subtilis 1A289, and environmental strains of B. pumilus and B. circulans. Not all B. anthracis ATCC 4229 spores germinated when applied to PLET indicating it is not an optimal medium for anthrax spore recovery.; Using a 1.22 g/ml sucrose plus 0.5% (v/v) Triton X-100 extraction solution, ethanol purification, PLET medium and confirmatory assays, B. anthracis spores were detected in 11 of 588 (1.9%) environmental specimens collected from anthrax endemic regions in northern Canada. All positive samples were associated with disposal sites of bison carcasses from previous anthrax epizootics. Viable anthrax spores were found at three of six carcass burial sites surveyed in the Parson's Lake Road region and at four of eight carcass cremation sites examined in the Falaise Lake region. Anthrax spore concentrations of >500 spores/g were observed in a red fox scat and specimens collected from a distinct bone bed found within cremation sites. The positive fox scat is the first bacteriological evidence that carnivorous mammals are capable of disseminating anthrax spores in northern Canada. Within the cremation bone beds, mats of bison hair and maggot casings were also found with anthrax spores, indicating the bed had been protected from the heat of the blaze by the bulk of the carcass.