Biochemical and molecular characterization of Bacillus amyloliquefaciens, B. subtilis and B. pumilus isolates with distinct antagonistic potential against Xanthomonas campestris pv. campestris

Fifty-one Bacillus isolates were characterized by fatty acid methyl ester (FAME) analysis; universal primer polymerase chain reaction (UP-PCR) fingerprinting; production of secondary metabolites and antagonistic activity against Xanthomonas campestris pv. campestris (causal agent of black rot in cabbage) in vitro and in vivo . Based on FAME analysis and/or PCR fingerprinting, the isolates were clustered into three different groups, named as Bacillus amyloliquefaciens , B . subtilis and B . pumilus . Seed treatment with Bacillus spp. generally reduced germination of seeds and incidence of black rot, but no relationship was found between the results of in vitro and in vivo experiments. The B .  amyloliquefaciens group contained isolates that were generally the most effective at reducing attack of black rot in vivo . The metabolic profiles of these isolates suggested that they produced surfactin, iturin, bacillomycine and/or azalomycin F. Isolates belonging to the B . subtilis group were mostly able to synthesize surfactin and arthrobactin. Surfactin, amphomycin, arthrobactin and valinomycin were generally found in culture extracts of isolates belonging to the B . pumilus group. No effect on growth of the pathogen was detected when the activity of filtered culture extracts and selected metabolites produced by the three different Bacillus species was tested in vitro against X . c . pv. campestris . However, inhibition was seen when bacterial liquid cultures were used. When the ability to colonize cabbage endophytically was examined for seven selected isolates with different antagonistic potential against black rot, it was found that the ability was related to the species and not to the antagonistic activity of the isolates.