Effects of berberine on spore germination and outgrowth.

(A) Spores (~300) were directly spread on LB plates containing various berberine concentrations, and with or without 20 μg/mL INF55, and incubated at 37°C for 12–18 h. (B) Spores (~107/mL) were first germinated with L-valine for 1 hr at 37°C as described in Methods plus various berberine concentrations, and with or without 20 μg/mL INF55. The germinated spores (~300) were then spread on LB plates without berberine and INF55, plates incubated at 37°C for 12 -18h, and growth was determined as described in Methods. (C) Germination of individual spores germinated at 37°C with L-valine and without berberine or with 200 μg/mL berberine. The kinetic germination was determined by the data for > 390 individual spores. (D) Germination and outgrowth of PS533 spores in LB medium plus valine, and without berberine or with 200 μg/mL berberine. Heat-activated spores were added at zero time, and the OD600 falls as DPA is released and the cortex is hydrolyzed. In the absence of berberine, the germinated spores outgrow and ultimately grow vegetatively leading to an increase in OD600; however, with berberine there is no increase in OD600 and thus no outgrowth.