Influence of indoleacetic-acid-producing Bacillus isolates on the nodulation of Phaseolus vulgaris by Rhizobium etli under gnotobiotic conditions

Abstract

Twenty-two Bacillus spp. isolates from the rhizosphere of Phaseolus vulgaris 'Contender' were identified using Biolog™, gas chromatographic fatty acid methyl ester, and 23S rDNA analyses. Some of the Bacillus isolates produced significant amounts of the phytohormone indoleacetic acid (IAA) when grown in a liquid culture medium supplemented with 100 μg L-tryptophan/L; less IAA was produced in culture medium not supplemented with L-tryptophan. Thin-layer chromatography, high-performance liquid chromatography, gas chromatography – mass spectrometry, and the avena coleoptile bioassay were used to identify and quantify IAA produced by Bacillus isolates. Significant differences were observed in the amounts of IAA produced by different strains of Bacillus, with amounts varying from 0.40 to 4.88 μg/mL. α-Methyltryptophan-resistant mutants of Bacillus exhibited altered IAA production and excreted tryptophan into the growing medium. The IAA-producing Bacillus isolates promoted root growth and (or) nodulation when coinoculated with Rhizobium etli (TAL 182) on Phaseolus vulgaris 'Contender' under gnotobiotic conditions in growth chambers. Coinoculation resulted in increased nodule number, nodule fresh weight, nitrogenase activity, leghemoglobin content, and total soluble protein content in the root nodules of Phaseolus vulgaris. In contrast, coinoculation with α-methyltryptophan mutants resulted in decreased nodulation, indicating that Bacillus isolates have a direct effect on either the Rhizobium or the plant and the effect may not be singularly attributed to their ability to produce IAA in vitro.Key words: Bacillus, indoleacetic acid production, nodulation enhancement.