Figure 4.

MccJ25^I13T demonstrates improved specificity

For pre-purified activity quantification MccJ25 mutants were first grown and induced in liquid culture, then the supernatants were isolated and sterilized by heating to 98 ºC for 15 min. and then plated on agar containing target organisms yielding a spectrum of responses (A, left: STen, middle: PUTI 53, right: FVEC 964). The specific activity, assessed via dilution plating, of wild-type MccJ25 (B, light bars) and MccJ25^I13T (B, dark bars), as well as the specific normalized activity (C) are shown against 18 commensal E. coli human isolates (grey bars) as well as well as two strains of Salmonella (blue bars). Data use Salmonella enterica serovar enteritidis (SE) as a reference point. These data demonstrate significant reduction of specific normalized activity for 15 of the 18 E. coli isolates (p<0.05). (D) To differentiate bactericidal from only bacteriostatic response, a subset of commensal E. coli and both Salmonella were incubated for one hour in exponential phase growth with MccJ25 or MccJ25^I13T and colony forming units (cfu) determined. A value below unity (10⁰) is indicative of bactericidal activity.