Pantoea agglomerans strain EH318 produces two antibiotics that inhibit Erwinia amylovora in vitro

Pantoea agglomerans (synonym: Erwinia herbicola) strain Eh318 produces thro ugh antibiosis a complex zone of inhibited growth in an overlay seeded with Erwinia amylovora, the causal agent of fire blight, This zone is caused by two antibiotics, named pantocin A and B. Using a genomic library of Eh318, two cosmids, pCPP702 and pCPP704, were identified that conferred on Escher ichia coli the ability to inhibit growth of E. amylovora. The two cosmids c onferred different antibiotic activities on E. coil DH5 alpha. and had dist inct restriction enzyme profiles. A smaller, antibiotic-conferring DNA segm ent from each cosmid was cloned, Each subclone was characterized and mutage nized with transposons to generate clones that were deficient in conferring pantocin A and B production, respectively. Mutated subclones were introduc ed into Eh318 to create three antibiotic-detective marker exchange mutants: strain Eh421 (pantocin A deficient); strain Eh439 (pantocin B deficient), and Eh440 (deficient in both pantocins), Cross-hybridization results, restr iction maps, and spectrum-of-activity data using the subclones and marker e xchange mutants, supported the presence of two distinct antibiotics, pantoc in A and pantocin B, whose biosynthetic genes were present in pCPP702 and p CPP704, respectively. The structure of pantocin A is unknown, whereas that of pantocin B has been determined as (R)-N-[((S)-2-amino-propanoylamino)-me thyl]-2-methanesulfonyl-succinamic acid. The two pantocins mainly affect ot her enteric bacteria, based on limited testing.