Isolation and characterization of hydrogen oxidizing bacteria in hydrogen treated soil and soil adjacent to Hup(-) soybean nodules

Previous studies have shown that the hydrogen (H2) gas evolved from legume nodules inoculated by hydrogen releasing (Hup -) rhizobium strains contributes to growth promotion of subsequent crop plants by altering the populations of soil H2 oxidizing bacteria; this may be responsible for the major beneficial effect of legumes in crop rotation. To further study the plant growth promoting bacteria in H2 treated soil and soil adjacent to the Hup- legume nodules, the H2 oxidizing bacteria were isolated from H2 treated soil and soil adjacent to Hup- soybean nodules grown in greenhouse and field. A much higher bacterial density was found in soils with a higher H2 uptake rate. Twenty isolates of H2 oxidizing bacteria were obtained. They are slow growing bacteria on mineral salt medium incubated under H2-enriched air. All H2 oxidizing bacterial strains exhibited one high Km for H2 at 1000ppm. The isolates of H2 oxidizing bacteria were identified as Variovorax paradoxus, Flavobacterium johnsonae and Burkholderia sordidicola by conventional microbiological tests and 16S rRNA gene sequence analysis. All isolated H2 oxidizing bacteria increased the root elongation of spring wheat seedlings ranging from 21% to 254% compared to controls. Seventeen of them had a significantly positive effect on root elongation (p<0.005). The strains of V. paradoxus found in both H 2 treated soil and soil adjacent to Hup- soybean nodules had the highest direct plant growth effect. The results confirm the hypothesis that the microorganisms responsible for the oxidation of H2 released by Hup- legume nodules are bacteria and they act as plant growth promoting rhizobacteria (PGPR).