Identification and application in plant of plant growth-promoting endophytic bacteria

Abstract

Seventy-eight endophytic bacteria isolated from 15 plant samples in Thailand including 50 isolates from sugarcane root samples obtained from 6 Provinces and 28 isolates from stems and leaves of healthy plants in Kanchanaburi Province, were identified and evaluated for plant growth promoting capability using rice as a plant model. They were identified as Gluconacetobacter (37 isolates), Pantoea (14 isolates), Burkholderia (2 isolates), Pseudomonas (4 isolates), Priestia (4 isolates), Enterobacter (2 isolates), Acinetobacter (2 isolates), Novosphingobium (2 isolates), Curtobacterium (2 isolates), and each of Nguyenibacter, Aureimonas, Bacillus, Peribacillus, Sphingobium, Staphylococcus, Brevibacillus, Aneurinibacillus, and Pseudarthrobacter based on their phenotypic characteristics and 16S rRNA gene analyzes. According to the 16S rRNA gene phylogeny, strain Sx8-5T was a member of genus Novosphingobium and shared the highest sequence similarity to Novosphingobium barchaimii LL02T of 99.4% and other Novosphingobium species (<99.4%). The ANIb and ANIm values of whole genome comparison between strain Sx8-5T and seven closely related type strains were 72.33-82.14%, 83.82-87.38%, respectively, and the digital DNA-DNA hybridization (dDDH) values ranged from 21.0% to 28.6% when compared to the type strains of the member of Novosphingobium. Based on the results obtained, demonstrated that strain Sx8-5T represents a novel species of the genus Novosphingobium, the name Novosphingobium kaempferiae sp. nov. is proposed. Sixty-one strains were positive in fixing nitrogen through nitrogenase activity by converting of N2 in the air to ammonia, which was supported by nif genes found in the genome of G. dulcium PS25. Sixty-nine strains were able to solubilize tricalcium phosphate (SI=1.18-4.4) and 72 strains were able to solubilize zinc oxide (SI=1.59-5.6). Sixteen strains could produce IAA (6.13 to 202.25 µg/mL) in NF medium supplemented with 0.01% L-tryptophan, a main precursor for IAA biosynthesis. The maximum IAA produced by the Sphingobium sp. Sx8-8 increased to 232.1 µg/mL at optimized conditions: pH 7, 30oC, 0.5% L-tryptophan and 48 h cultivation. In in vitro 2 rice varieties germination, the Sphingobium sp. Sx8-8 and Pantoea sp. S5-1 increased root length (4.83, 3.24 cm), shoot length (18.7, 13.75 cm) and number of lateral roots of rice ‘RD6’ seedlings more than control and standard IAA, similar to Aureimonas sp. SK2, G. dulcium PS25 and G. liquefaciens LSG1 significantly increased root length (3.09-3.49 cm), shoot length (10.14-11.21 cm), number of lateral roots, and biomass of ‘Khao Dawk Mali 105’ rice seedlings. This study indicated that endophytic bacteria could potentially promote plant growth and be utilized as bioinoculant or biofertilizers in agriculture.