The Enzyme Glucose‐1‐Phosphate Thymidylyltransferase <scp>RmlA</scp> Plays a Crucial Role in the Pathogenesis of <i>Pectobacterium actinidiae</i> <scp>GX1</scp>

Pectobacterium actinidiae is one of the primary pathogens that causes summer canker disease in kiwifruit, yet its pathogenic mechanisms remain unknown. The exopolysaccharide PCAP-1a, isolated from the fermentation broth of P. actinidiae strain GX1, exhibits notable cytotoxicity and acts as a virulence factor facilitating host infection. Genome-wide analysis revealed a 21-gene cluster responsible for the biosynthesis of exopolysaccharides in GX1. Homologous recombination was used to systematically knock out these genes, which led to the identification of RmlA as a key protein in the synthesis of the PCAP-1a precursor. The deletion of the rmlA gene significantly affected the yield of PCAP-1a and resulted in a direct reduction in GX1 pathogenicity. Further studies revealed that mutations in the substrate binding site of RmlA weakened its capacity to bind G-1-P and dTTP, which led to markedly reduced pathogenicity in the corresponding complemented strains. This study indicates that the exopolysaccharide PCAP-1a serves as a virulence factor in the pathogenesis of GX1, and its biosynthesis depends on the polysaccharide synthesis gene rmlA and the substrate binding activity of its encoded protein.