{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,2]],"date-time":"2026-06-02T15:15:38Z","timestamp":1780413338647,"version":"3.54.1"},"reference-count":59,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2026,6,2]],"date-time":"2026-06-02T00:00:00Z","timestamp":1780358400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Shanghai Agricultural Science and Technology Innovation Project","award":["T2024310"],"award-info":[{"award-number":["T2024310"]}]},{"name":"Shanghai Sailing Program","award":["24YF273800"],"award-info":[{"award-number":["24YF273800"]}]},{"name":"Shanghai Agricultural Technology System","award":["202509"],"award-info":[{"award-number":["202509"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Microorganisms"],"abstract":"<jats:p>Spent mushroom substrate (SMS) return is a vital strategy for agricultural waste recycling and soil fertility improvement, yet its ecological impacts of duration remain poorly understood. This study employed metagenomic sequencing to explore soil fertility, microbial dynamics, and nitrogen cycling across different SMS return durations (0, 1, and 3 years) within rice\u2013mushroom crop rotation systems. Soil nutrients (organic matter, total nitrogen, total phosphorus) initially decreased and then increased throughout the rice growth cycle. The one-year return (y1) induced early nutrient depletion, whereas the three-year return (y3) significantly enhanced late-stage nutrient accumulation. With increasing duration, bacterial and archaeal assembly shifted from stochastic toward deterministic processes, while fungal diversity and stochasticity decreased continuously. Co-occurrence network analysis demonstrated that SMS return increased network complexity and intercommunity competition. This transition was accompanied by a functional shift in keystone taxa from those responsive to exogenous organic matter in y1 to those mediating nitrogen fixation, anammox, and sulfur metabolism in y3. Nitrogen cycling in y1 increased potential N2O emission risks through nirS upregulation and nosZ downregulation, whereas y3 mitigated inorganic nitrogen loss by upregulating gene abundances of ammonia assimilation, nitrification, and DNRA genes. Notably, the structure of nitrogen-cycling genes fluctuated in y1 but was resilient to y0 levels in y3. These findings demonstrated that while initial SMS return triggered ecological fluctuations and environmental risks, continuous return (y3) achieved functional stability by reshaping microbial niches. This study highlights the importance of SMS return duration in balancing soil fertility enhancement with environmental risk mitigation in sustainable paddy ecosystems.<\/jats:p>","DOI":"10.3390\/microorganisms14061251","type":"journal-article","created":{"date-parts":[[2026,6,2]],"date-time":"2026-06-02T14:38:42Z","timestamp":1780411122000},"page":"1251","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Duration of Spent Mushroom Substrate Return Affects Microbial Assembly and Nitrogen Metabolism to Promote Functional Stabilization in Rice\u2013Mushroom Crop Rotation Systems"],"prefix":"10.3390","volume":"14","author":[{"given":"Yihong","family":"Yue","sequence":"first","affiliation":[{"name":"National Engineering Research Center of Edible Fungi, Key Laboratory of Edible Fungi Resources and Utilization (South), Ministry of Agriculture, Institute of Edible Fungi, Shanghai Academy of Agricultural Sciences, Shanghai 201403, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yu","family":"Jiang","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Environmental Criteria and Risk Assessment, State Environmental Protection Key Laboratory of Simulation and Control of Groundwater Pollution, Chinese Research Academy of Environmental Sciences, Beijing 100012, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0756-1153","authenticated-orcid":false,"given":"Yuchen","family":"Zhang","sequence":"additional","affiliation":[{"name":"National Engineering Research Center of Edible Fungi, Key Laboratory of Edible Fungi Resources and Utilization (South), Ministry of Agriculture, Institute of Edible Fungi, Shanghai Academy of Agricultural Sciences, Shanghai 201403, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tingting","family":"Xiao","sequence":"additional","affiliation":[{"name":"National Engineering Research Center of Edible Fungi, Key Laboratory of Edible Fungi Resources and Utilization (South), Ministry of Agriculture, Institute of Edible Fungi, Shanghai Academy of Agricultural Sciences, Shanghai 201403, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Haibo","family":"Hao","sequence":"additional","affiliation":[{"name":"National Engineering Research Center of Edible Fungi, Key Laboratory of Edible Fungi Resources and Utilization (South), Ministry of Agriculture, Institute of Edible Fungi, Shanghai Academy of Agricultural Sciences, Shanghai 201403, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Qian","family":"Wang","sequence":"additional","affiliation":[{"name":"National Engineering Research Center of Edible Fungi, Key Laboratory of Edible Fungi Resources and Utilization (South), Ministry of Agriculture, Institute of Edible Fungi, Shanghai Academy of Agricultural Sciences, Shanghai 201403, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1827-2165","authenticated-orcid":false,"given":"Zongjun","family":"Tong","sequence":"additional","affiliation":[{"name":"National Engineering Research Center of Edible Fungi, Key Laboratory of Edible Fungi Resources and Utilization (South), Ministry of Agriculture, Institute of Edible Fungi, Shanghai Academy of Agricultural Sciences, Shanghai 201403, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jinjing","family":"Zhang","sequence":"additional","affiliation":[{"name":"National Engineering Research Center of Edible Fungi, Key Laboratory of Edible Fungi Resources and Utilization (South), Ministry of Agriculture, Institute of Edible Fungi, Shanghai Academy of Agricultural Sciences, Shanghai 201403, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hui","family":"Chen","sequence":"additional","affiliation":[{"name":"National Engineering Research Center of Edible Fungi, Key Laboratory of Edible Fungi Resources and Utilization (South), Ministry of Agriculture, Institute of Edible Fungi, Shanghai Academy of Agricultural Sciences, Shanghai 201403, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,6,2]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"133767","DOI":"10.1016\/j.biortech.2025.133767","article-title":"Waste to wealth: Sustainable approach of alchemizing mushroom SMS into bioenergy","volume":"443","author":"Gupta","year":"2026","journal-title":"Bioresour. 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