Abstract

Knowledge about the colonization behaviour of microbial inoculants is a prerequisite to their effective use in agriculture. In this study, we used GUS reporter gene to observe the behaviour of symbiotic nitrogen-fixing and plant growth-promoting bacteria in alfalfa, white clover, and soyabean. The gusA gene from Escherichia coli S17-1 lambda-pir with plasmid mTn5SSgusA20 was inserted into the genome of the recipient strains by triparental mating. There was no significant difference in the acetylene reduction activity (ARA) and nodulation of alfalfa when inoculated with either gusA-marked Rhizobium meliloti or parent strain, indicating that GUS marking did not affect the nitrogen-fixing properties of the transconjugants. Co-inoculation of Rhizobium leguminosarum bv. trifolii and gusA-marked Azospirillum lipoferum increased the ARA and nodulation of white clover by 2-3 times from 5 to 20 days after inoculation (DAI) and ARA by 2.3-2.7 times at 20 DAI. The abundant colonization of A. lipoferum on the roots, root hairs, and sites near or on the nodules suggests that the formation of additional infection site by A. lipoferum may be the mechanism that enhances the ARA and nodulation of white clover. Co-inoculation of Bradyrhizobium japonicum and gusA-marked Pseudomonas fluorescens also increased the ARA and nodule number of soyabean at 10 and 20 DAI and stimulated the growth and colonization of B. japonicum on soyabean roots. The results indicate that the enhanced ARA and nodulation of soyabean are due to the high colonization of P. fluorescens on soyabean roots and the production of substances that stimulate the growth of B. japonicum.