Effect of the expression of Escherichia coli fhuA gene in Rhizobium sp. IC3123 and ST1 in planta: Its role in increased nodule occupancy and function in pigeon pea

The inability to utilize a fungal siderophore as source of iron nutrition by most of the rhizobial cultures isolated from pigeon pea, could be considered a negative fitness factor since hydroxamate siderophores are found in significant amounts in natural soils. Thus these cultures were engineered to use ferrichrome a prototype of hydroxamate type siderophore. Pigeon pea Rhizobium spp. IC3123 and ST1 harboring Escherichia coli fhuA gene, responsible for uptake of Fe³⁺-ferrichrome, were obtained by transformation with pGR1, a broad host range plasmid carrying the fhuA gene under the control of the lac promoter of E. coli. Expression of fhuA in transformed rhizobial strains IC3123::pGR1 and ST1::pGR1 was confirmed by the ability of the plasmid-bearing strains to utilize iron bound to ferrichrome. Inoculation of pigeon pea plants with fhuA expressing rhizobial strains in pot experiments showed a significant increase in plant growth as well as nodule density as compared to those inoculated with the parent as well as the empty vector-bearing strain. Inoculation of pigeon pea seedlings with IC3123::pGR1 and ST1::pGR1 led to marked increase in shoot fresh weight, nodule number per plant, chlorophyll content of leaves and effective nodule symbiosis when compared with plants inoculated with the parent strains IC3123 and ST1. The positive effect of IC3123::pGR1 and ST1::pGR1 treatment on plant growth was more significantly observed when ferrichrome producing Ustilago maydis, known to secrete ferrichrome, was co-inoculated along with the transformed rhizobia. The presence of fhuA gene in rhizobial strains also led to an increased survival and root colonization.