FIG 2.

Secondary screening by TonB-dependent siderophore nutrition assays and colicin killing tests. (A) We tested the 20 selected primary hits for interference with siderophore nutrition assays with FeEnt and Fc; the negative and positive controls were untreated bacteria and bacteria treated with CCCP, respectively. We observed several types of inhibition by the compounds: larger halos (that imply a lower rate of iron uptake [10, 44, 45]; illustrated by the control, CCCP, at 15 and 10 μM), distorted, aberrant halos, and the complete absence of a halo. Inhibition was identified by a loss of halo or an increase in the diameter of the halo around the ferric siderophore disc (Table 2). (B) We performed ColB and ColIa killing tests in the presence of the 20 selected compounds, with the same controls. The graph depicts the percent survival of bacteria in the presence of ColB or ColIa in the absence and presence of inhibitory compounds (see also Table 2). We performed each experiment 2 or 3 times and averaged the percent survival of colicin killing in the absence and presence of each compound. Error bars represent the standard deviations of the mean values. Compound abbreviations: CCCP, carbonyl cyanide m-chlorophenylhydrazone; Bai, baicalein; CA, cadmium acetate; CD, carbidopa; CP, carboplatin; CHL, p-chloranil; DS, dideoxyscleroin; DC, dequalinium chloride; EB, ebselen; EA, ellagic acid; MC, methylcatechol; THM, thimerosal; ZP, zinc pyrithione; 120304, lead compound from reference 71. See Fig. 1 and Table 2 for more information about the enumerated compounds.