<italic>Bacillus anthracis</italic> endospore germination: An analysis of purine co-germinants, the germination operon <italic>gerH</italic>, and the micro-environments in which <italic>B. anthracis</italic> endospores germinate.

Abstract

The work presented in this dissertation examines <italic>B. anthracis </italic> endospore germination by characterizing the roles played by purine nucleosides and <italic>gerH</italic> in germination, and by using the <italic> gerH</italic> null strain as a tool to further dissect the germination response pathways present in M&phis; conditioned solutions, serum and soil. Furthermore, the mechanisms by which germinants become available to endospores was investigated, as were the different micro-environments in which <italic>B. anthracis</italic> can germinate. The tri-cistronic operon <italic>gerH</italic> was identified, with high homology to <italic>gerI</italic>, an ortholog necessary for inosine-dependent <italic> B. cereus</italic> endospore germination. <italic>B. anthracis</italic> endospores did not germinate in inosine alone, but germination was triggered by inosine in binary combination with amino acid co-germinants grouped into two distinct germination response pathways: inosine-His and purine-Ala. Purines can be substituted for inosine in the inosine-Ala pathway, which includes inosine-Ser, but not in the inosine-His pathway, which requires <italic>gerH</italic>. Unlike <italic>B. anthracis</italic> Sterne endospores, <italic>gerH</italic> null endospores do not germinate in the presence of macrophages and in macrophage-conditioned media. Germination assays performed in alluvial soil extracts demonstrate for the first time that <italic>B. anthracis</italic> endospores can germinate in a natural environment outside of a host, and that <italic>ex vivo</italic> germination utilizes, in part, <italic>gerH</italic>. This finding re-energizes the discussion of the different ecological niches which may support the germination and growth of <italic>B. anthracis</italic>.