Microbial stimulation of regrowth in ryegrass by the dual‐function ammonia‐oxidizing bacteria strain C5_3

Ammonia‐oxidizing bacteria (AOB) are key drivers of nitrogen cycling and plant growth, yet their roles in cytokinin biosynthesis and regrowth stimulation remain underexplored. This study investigated the multifunctional potential of a novel AOB strain, C5_3, in promoting compensatory regrowth of Lolium multiflorum following clipping. A pot experiment was conducted with six treatments: blank control (BT), nitrification inhibitor (DN), C5_3 inoculation (JI), C5_3 combined with the inhibitor (DJ), cytokinin‐producing strain S2 (JS) and S2 combined with the inhibitor (DS). Measurements were taken on Days 0, 7 and 14 of regrowth, including soil nitrate (NO₃ − –N) content, nitrification rate, zeatin riboside (ZR) content and transport rate, clipped leaf biomass and total plant biomass. The results indicated that inoculation with C5_3 significantly increased nitrification rates, reaching 2.5 times the level observed in the BT treatment, and enhanced C5_3 gene abundance, resulting in higher soil nitrate availability. C5_3 also elevated leaf ZR content and promoted its transport from root to shoot under both light and dark conditions. While S2 primarily enhanced cytokinin levels, C5_3 demonstrated dual functionality by improving both hormonal and nutritional pathways, leading to greater biomass accumulation ( p < .05). The nitrification inhibitor reduced microbial activity and plant regrowth, and its combination with C5_3 only partially mitigated these effects. Overall, these findings demonstrate that C5_3 stimulates compensatory regrowth in L. multiflorum through integrated microbial and hormonal pathways. Its dual functionality highlights its potential as a bioinoculant for sustainable forage production systems requiring rapid recovery after grazing or mowing.