The effects of zinc (Zn) speciation on plant growth in Zn-contaminated soil in the presence of bacteria are unknown but are critical to our understanding of metal biodynamics in the rhizosphere where bacteria are abundant. A 6-week pot experiment investigated the effects of two plant growth promoting bacteria (PGPB), Rhizobium leguminosarum and Pseudomonas brassicacearum, on Zn accumulation and speciation in Brassica juncea grown in soil amended with 600 mg kg elemental Zn as three Zn species: soluble ZnSO and nanoparticles of ZnO and ZnS. Measures of plant growth were higher across all Zn treatments inoculated with PGPB compared to uninoculated controls, but Zn species effects were not significant. Transmission electron microscopy identified dense particles in the epidermis and intracellular spaces in roots, suggesting Zn uptake in both dissolved and particulate forms. X-ray absorption near-edge structure (XANES) analysis of roots revealed differences in Zn speciation between treatments. Uninoculated plants exposed to ZnSO contained Zn predominantly in the form of Zn phytate (35%) and Zn polygalacturonate (30%), whereas Zn cysteine (57%) and Zn polygalacturonate (37%) dominated in roots exposed to ZnO nanoparticles. Inoculation with PGPB increased (>50%) the proportion of Zn cysteine under all Zn treatments, suggesting Zn coordination with cysteine as the predominant mechanism of Zn toxicity reduction by PGPB. Using this approach, we show, for the first time, that although speciation is important, the presence of rhizospheric bacteria completely overrides speciation effects such that most of the Zn in plant tissue exists as complexes other than the original form.

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