Ensifer meliloti denitrification is involved in infection effectiveness and N2O emissions from alfalfa root nodules

Abstract

Purpose Alfalfa is one of the most valuable forage crops in temperate climate zones. Ensifer meliloti, the endosymbiont of alfalfa, contains all the denitrification genes but the capacity of alfalfa root nodules to produce nitrous oxide (N2O) is not known. In this work, N2O emissions as well as the influence of bacteroidal denitrification on nodulation competitiveness and N2O release from alfalfa nodules has been investigated. Methods Medicago sativa cv. Victoria plants were inoculated with E. meliloti 1021, a periplasmic nitrate reductase (Nap) defective mutant, a Nap overexpressing strain and a nitrous oxide reductase defective mutant. Plants were grown in the presence of different nitrate and copper treatments and subjected to flooding during one week before harvesting. N2O production by the nodules was analysed by using gas chromatography. Methyl viologen-dependent nitrate reductase (MV+-NR), nitrite reductase (MV+-NIR) and nitrous oxide reductase (N2OR) enzymatic activities were measured in isolated bacteroids. Results Alfalfa root nodules produce N2O in response to nitrate and flooding. Overexpression of Nap improved nodulation competitiveness and induced N2O emissions from nodules. Copper is required for an effective symbiosis as well as triggered a reduction of N2O production due to the induction of the N2OR and a reduction of NIR activities in the bacteroids. Conclusion Alfalfa root nodules emit N2O. Nap is involved in nodulation competitiveness and in N2O emissions by the nodules. Bacteroidal N2OR and NIR activities are modulated by Cu and may be considered as effective targets for the mitigation strategies of N2O emissions derived from alfalfa crops.

Genomic analysis of Sinorhizobium meliloti LPU63, an acid-tolerant and symbiotically efficient alfalfa-nodulating rhizobia

“…Alfalfa root nodules produce N 2 O and Nap has an important role on its release from nodules in response to nitrate and flooding (Pacheco et al, 2023). Recently, a novel strategy to avoid these undesirable metabolic pathways was the screening of S. meliloti strains with spontaneous mutations in napC, which encodes the nitrate reductase structural protein NapC involved in the first step of N 2 O production via the denitrification process.…”

“…It is unknown whether strain LPU63 codes for the complete denitrification pathway. Recently, it has been shown that the overexpression of the nap operon (coding for the enzymes involved in the first step of the denitrification pathway), promotes nodulation when the host plant develops in a hypoxic environment (Pacheco et al, 2023).…”