Engineered Escherichia coli nrfA Reduces Cadmium Accumulation in Rice by Modulating Bacterial-Driven Nitrogen Cycling...

Nitrogen supply in the rhizosphere affects cadmium (Cd) accumulation in rice. To study bacterial nitrogen transformation effects on Cd uptake in rice, Escherichia coli MG1655 overexpressing nrfA, which encodes cytochrome c nitrite reductase involved in dissimilatory nitrite reduction to ammonium (DNRA), was constructed using a Tn5 transposon system. The nrfA genomic copy number in the engineered strain was approximately 3-fold that in wild-type, and RNA-seq analysis confirmed significantly increased nrfA transcript. Transcriptomics showed enrichment of ribosomes, oxidative phosphorylation, and carbon metabolism pathways. In pot experiments, inoculation with the nrfA-overexpressing strain selectively enriched nitrogen-transformation and iron-cycling taxa, including Actinobacteria and Flavobacterium. Organic nitrogen metabolism gene abundances increased following strain inoculation, particularly in the nrfA-overexpressing treatment; in contrast, denitrification gene abundances decreased, with the greatest reduction in the nrfA-overexpressing treatment. Consistent with these results, soil ammonium nitrogen was 79.4% and 34.1% higher in the nrfA-overexpressing treatment than in uninoculated control and wild-type treatment, respectively. The nrfA-overexpressing strain also promoted iron plaque formation on rice root surfaces, enhancing root Cd interception Cd by >20% versus both other groups. Consequently, compared with uninoculated control, inoculation with nrfA-overexpressing E. coli significantly reduced Cd concentration in rice roots, stems, and grains by 36.2%, 61.0%, and 57.4%, respectively. Collectively, nrfA-overexpressing E. coli reshaped rhizosphere nitrogen cycling toward sustained ammonium nitrogen enrichment, potentially suppressing root Cd uptake through cation competition and strengthening root-surface Cd retention, achieving simultaneous nitrogen enrichment and Cd reduction in paddy soil. This study may offer theoretical support for microbial remediation of Cd-contaminated farmland.