Biological control of the grapevine dieback fungus Eutypa lata II: Influence of formulation additives and transposon mutagenesis on the antagonistic activity of Bacillus subtilis and Erwinia herbicola

The effect of different supplements on the antagonistic activity of Bacillus subtilis and Erwinia herbicola towards Eutypa lata in vitro and on autoclaved grape wood was investigated. Inhibition zones on grape wood agar were enlarged maximally in B. subtilis by adding peptone (0.25%) and in E. herbicola by adding glucose (0.5%) or high concentrations of peptone (1-2%) to the medium. Nitrate and phosphate salts reduced the antagonistic activity of E. herbicola on glucose-supplemented medium. Glucose as well as manganese ions did not enlarge inhibition zones of B. subtilis. On grape wood, peptone and methylcellulose supplements increased the antagonistic performance and cell densities of B. subtilis B1 alpha , whereas a negative effect was exerted by calcium alginate and carob (88% galactomannane). A combination of 1% peptone and 4% methylcellulose resulted in 100% inhibition of E. lata (70% inhibition without supplements). In E. herbicola JII/E2, cells applied without supplements showed no antagonistic effect. Carob was the only supplement that led to a significant antagonistic effect of suspended cells of E. herbicola (70-100% inhibition). One spontaneous mutant and two transposon mutants of E. herbicola JII/E4 did not inhibit E. lata on tryptic soy czapek dox agar but showed unreduced antagonistic activity on grape wood extract agar amended with glucose or peptone. On grape wood, the transposon mutants inhibited growth of E. lata completely whereas the antagonistic activity of the spontaneous mutant was significantly reduced.