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Conservation of tunicamycin biosynthetic gene clusters across Rathayibacter species

Matthew Tancos: USDA ARS FDWSRU

<div>Tunicamycins are a novel class of toxigenic nucleoside antibiotics with broad biological activity, inhibiting both bacterial cell wall biosynthesis and eukaryotic <em>N</em>-linked glycosylation. Tunicamycins are produced by several soil-associated <em>Actinomycetes</em>, including the grass-associated select agent <em>Rathayibacter toxicus. </em>Currently endemic to Australia, <em>R. toxicus</em> causes bacterial toxicosis in livestock when high concentrations of tunicamycin are consumed from infected pastures. <em>R. toxicus </em>is currently the only known phytobacterium that produces tunicamycin, but similar livestock toxicoses have been reported outside Australia. Genomic analyses of available grass-associated <em>Rathayibacter </em>spp. identified two novel tunicamycin gene clusters (TGCs) in <em>R. iranicus </em>and an undescribed South African <em>Rathayibacter </em>sp. termed ‘EV’. The uncharacterized TGCs are similarly sized to the TGC of <em>R. toxicus</em>, but differ in gene orientation, order, and count. <em>R. toxicus</em> possesses a putative TGC that spans 14 genes, of which 12 genes (<em>tunA-tunL</em>) appear essential for toxin production. In contrast, the <em>R. iranicus</em> TGC has 15 genes, but lacks the conserved gene <em>tunG. Rathayibacter </em>sp. EV possesses 14 genes, including all 12 essential genes, but differs greatly in gene order and orientation from <em>R. toxicus</em>. Preliminary bioassays indicate toxin production for both <em>R. iranicus</em> and <em>R</em>. sp. EV, but chemical analyses are ongoing. Conservation of the TGC within species was also investigated.</div>