<i>Bacillus</i> sp. G2112 Detoxifies Phenazine-1-carboxylic Acid by <i>N</i>5 Glucosylation

Abstract

Microbial symbionts of plants constitute promising sources of biocontrol organisms to fight plant pathogens. Bacillus sp. G2112 and Pseudomonas sp. G124 isolated from cucumber (Cucumis sativus) leaves inhibited the plant pathogens Erwinia and Fusarium. When Bacillus sp. G2112 and Pseudomonas sp. G124 were co-cultivated, a red halo appeared around Bacillus sp. G2112 colonies. Metabolite profiling using liquid chromatography coupled to UV and mass spectrometry revealed that the antibiotic phenazine-1-carboxylic acid (PCA) released by Pseudomonas sp. G124 was transformed by Bacillus sp. G2112 to red pigments. In the presence of PCA (>40 µg/mL), Bacillus sp. G2112 could not grow. However, already-grown Bacillus sp. G2112 (OD600 > 1.0) survived PCA treatment, converting it to red pigments. These pigments were purified by reverse-phase chromatography, and identified by high-resolution mass spectrometry, NMR, and chemical degradation as unprecedented 5N-glucosylated phenazine derivatives: 7-imino-5N-(1′β-D-glucopyranosyl)-5,7-dihydrophenazine-1-carboxylic acid and 3-imino-5N-(1′β-D-glucopyranosyl)-3,5-dihydrophenazine-1-carboxylic acid. 3-imino-5N-(1′β-D-glucopyranosyl)-3,5-dihydrophenazine-1-carboxylic acid did not inhibit Bacillus sp. G2112, proving that the observed modification constitutes a resistance mechanism. The coexistence of microorganisms—especially under natural/field conditions—calls for such adaptations, such as PCA inactivation, but these can weaken the potential of the producing organism against pathogens and should be considered during the development of biocontrol strategies.

Keywords

• antibiotic
• biocontrol
• co-cultivation
• epiphyte
• glycosylation
• mass spectrometry