Simultaneous Stimulation of Soil Respiration and Plant Biomass in Transgenic Bacillus thuringiensis Crop Cultivation

Simultaneous Stimulation of Soil Respiration and Plant Biomass in Transgenic Bacillus thuringiensis Crop Cultivation | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Simultaneous Stimulation of Soil Respiration and Plant Biomass in Transgenic Bacillus thuringiensis Crop Cultivation Lingyan Zhou, Shuxian Jia, Hengshuo Zhang, Kaiyan Zhai, Xuhui Zhou This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6699296/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 16 Dec, 2025 Read the published version in Plant and Soil → Version 1 posted 6 You are reading this latest preprint version Abstract Aims Transgenic Bacillus thuringiensis ( Bt ) crops are cultivated globally to mitigate potential pest crisis and reduce insecticide dependence. However, the response of belowground system to Bt crop cultivation remains controversial, limiting our understanding of Bt crops’ role in agriculture sustainability. The aim of this study was to assess Bt crops’ influence on soil system, including variables of soil respiration, soil carbon and nutrient pools and soil biodiversity. Methods We conducted a global meta-analysis of 88 experimental studies to assess the influence of Bt crop cultivation on soil system. Results Bt crops significantly increased soil respiration by 6.0% (p < 0.05), alongside a 14.3% rise in aboveground biomass and a 4.3% belowground plant biomass. Bt crops reduced rhizospheric soil organic carbon (SOC) by 3.4% but increased bulk soil SOC by 3.1%. SOC, nematode diversity, and fungal diversity accounted for 50%, 34%, and 30% of the variation in soil respiration under Bt crop cultivation, respectively. The decrease of microbial competition for nitrogen under Bt crop cultivation caused a significant reduction in microbial biomass carbon: nitrogen ratio. Conclusions Under Bt crop cultivation, the increased soil respiration might be caused by the stimulated microbe nutrient mining due to plant production-induced system nitrogen deficiency, especially in rhizosphere. Dispite their advantages in biomass accumulation, long-term cultivation of Bt crops may pose ecological risks on soil fertility sustainability and introduce uncertainties in agricultural life cycle assessments. Bt crop soil respiration rhizosphere nutrient ecosystem sustainability Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Introduction As global climate change intensifies, agricultural systems confront mounting pressures from increasingly severe pest infestations, pathogen proliferation, and erratic weather patterns—posing grave risks to worldwide food stability and ecological balance (Singh et al., 2023 , Zhang et al., 2022 ). Transgenic crops, which boast inherent benefits, such as herbicide tolerance, insect resistance, disease resistance, and improved nutritional profiles, have emerged as a key strategy to address these challenges. By 2020, over 500 transgenic events in about 30 different crops had received global approval for cultivation (Kumar et al., 2020 ). Among these, Bacillus thuringiensis ( Bt ) crops stand out as they are genetically engineered to produce endospore (or crystal) toxins of the bacterium, providing them with a natural defense against a range of insect pests, including Lepidoptera, Coleoptera, and Hymenoptera (Abbas, 2018 ). Extensive research has shown that Bt crops, such as those for corn and cotton, consistently outperform their conventional counterparts in terms of yields (Carzoli et al., 2018 , Qaim, 2020 , Shelton et al., 2013 ). The enhanced metabolic efficiency in Bt crops, particularly in pathways such as the tricarboxylic acid cycle and energy metabolism, leads to increased biomass accumulation compared to their non-transgenic counterparts (Zhang et al., 2021 ). This heightened pest suppression capability not only influences carbon allocation at the individual plant level but also alters ecosystem-scale carbon dynamics, shifting the traditional tripartite biotic interactions among pests, plants and soil organisms towards a binary interaction between the latter two (Fig. 1 , Suzuki et al., 2000 ). Consequently, the impact of Bt crop cultivation on soil characteristics, including enzymatic activities and microbial functions, has become a focal point for research (Lee et al., 2017 , Li et al., 2019 , Zhang et al., 2019 ). While Bt crop cultivation is widely perceived as environmentally benign, empirical data suggests that it does not remain without effect on soil processes or biota, especially under long-term cultivation scenarios where the exclusion of pests from the agroecosystem results in increased carbon sequestration within crop biomass (Krogh et al., 2020 , Lee et al., 2017 ). Despite its carbon-sink advantages, the decoupling of pest-mediated carbon-nutrient feedbacks, and long-term shifts in soil enzymatic-microbial dynamics might strain soil nutrient reserves and affect the interactions between crops and soil organisms (Fig. 1 ). The previous interactions among pests, crops and soil organisms in agrosystem would no longer occur due to the unilaterally strategy change in crops (i.e., Bt protein expression by crops), leading to a new Nash equilibrium (a key concept in game theory, in which no player’s expected outcome can be improved by changing one’s own strategy, Fig. 1 , Blaser & Kirschner, 2007 , Cai et al., 2020 , this concept is increasingly applied to model resource competition among organisms in ecosystem). Resisting pest attacks, Bt crops would perform a set of interaction with soil organisms to achieve a new beneficial balance (e.g., enhancing the availability of underground resources), escpecially in rhizosphere, the hotspot for plant - soil organims interactions (Čapek et al., 2018 ). However, the response of soil organism-dominated carbon and nutrient processes (e.g., soil respiration and soil nutrient mineralization) in soil system, as well as soil organism community in rhizosphere or bulk soil under Bt crop cultivation remains largely unexplored. This gap in knowledge constrains our comprehensive understanding of the ecological implications of Bt crop cultivation, particularly in the context of sustainable agricultural systems (Fig. 1 , Mmbando & Ngongolo, 2024 ). Considering coupling relationship between carbon and nutrient cycles within ecosystem, we propose two hypotheses in the new Nash equilibria between Bt crops and soil organisms. Fristly, Bt crops may yield a generally greater biomass and cause a greater demand for nutrients relative to their non-transgenic counterparts. Secondly, the increasing assimilation products in Bt crops would accompany with soil organism regulation of soil carbon and nutrient cycle (e.g., nitrogen and phosphorus), which would be reflected by a change of soil respiration. To test these hypotheses, we employed a meta-analysis to assess responses of soil respiration, soil carbon and nutrient pools, soil biodiversity, and crop biomass to Bt crop cultivation. The diversity of soil organisms and stoichiometry in both rhizosphere and bulk soil were evaluated to provide some underlying explanation for effect of Bt crop cultivation on soil carbon and nutrient cycle. Our fundings indicate that Bt crops cause more carbon sequestrated in both above- and below-ground plant biomass compared to conventional varieties. In the new Nash equilibrium between crops and soil organisms (including bacterias, fungi, and nematodes), the soil respiration significantly enhanced, accompanying with decreases in microbial biomass carbon: nitrogen ratio, and rhizosphere SOC. Materials and Methods Data sources We conducted a comprehensive literature search using the combination “( Bacillus thuringiensis OR Bt ) AND (soil OR carbon OR respiration OR CO 2 OR microbe* OR bacteria* OR fung* OR nitrogen OR ammoni* OR nitrate OR phosphor* OR organism* OR diverisity OR yield OR biomass OR root OR biomass)”. The peer-reviewed journal articles were sought using Web of Science (1990–2024) and China National Knowledge Infrastructure (CNKI, 1992–2024). To avoid bias in publication selection, the studies were selected on the basis of the following considerations: i) The sole differentiating factor between groups of Bt crops and their conventional counterparts was the genetic modification, with all other management practices being consistent across treatments; ii) Experiments were conducted in the field, recording at least one pair of comparative data of conventional non- Bt crop and Bt crop varieties, such as aboveground biomass (AGB), belowground biomass (BGB), soil CO 2 emission (SCE), soil organic carbon (SOC), soil organic nitrogen (SON), soil dissovled organic carbon (DOC), soil dissovled orgnaic nitogen (DON), soil ammonium (soil NH 4 + ), soil nitrate (soil NO 3 − ), soil total phosphorus (STP), soil pH (pH), microbial biomass carbon (MBC), microbial biomass nitrogen (MBN), bacterial, fungal and/or nematode diversity (BD, FD, and/or ND, including Shannon, Richness, InvSimpson (1-Simpson), Chao 1, ACE, and/or operational taxonomic unit (OUT)); iii) The means, standard deviations/errors, and samples sizes of variables could be extracted directly from the text, tables or digitized graphs; iv) The experimental duration were longer than one growing season. Applying these criteria, we compiled a dataset from 88 published papers focusing on the cultivation of Bt genetically modified maize, cotton, or rice (PRISMA in Supplementary Materials). Statistical analysis In this study, we utiliazed the response ratio ( RR ), defined as the natural log of the ratio of the mean value in cultivation of Bt crops ( \(\:\stackrel{-}{{X}_{t}}\) ) to that in control, i.e., cultivation of conventional parent breed ( \(\:\stackrel{-}{{X}_{\text{c}}}\) , Eq. 1 ), to quantify the impact of Bt genetic modification on the concerned variables relative to the conventional cultivation methods (Luo et al., 2006 ). $$\:RR=ln\left(\frac{\stackrel{-}{{X}_{t}}}{\stackrel{-}{{X}_{c}}}\right)={ln}\left(\stackrel{-}{{X}_{t}}\right)-ln\left(\stackrel{-}{{X}_{c}}\right)$$ 1 Subsequently, the weighted average effect sizes ( RR ++ ) and 95% confidence intervals (CI) for each variable was calculated using rma.mv function in metafor package (version 3.4-0, Viechtbauer, 2010 ), based on the numbers of replicates and standard deviations for a given variable under both Bt and conventional crop cultivation. A significant effect of Bt crop cultivation on a variable was considered when the estimated CI did not overlap zero. The percentage change in a concerned variable under Bt crop cultivation was estimated by [exp ( RR ++ ) − 1] × 100%. The Pearson correlation coefficient between response of soil CO 2 emission and that of other variables was analyzed firstly, and then the linear regression was tested using metafor package and drawn with ggplot2 package (version 3.4.4) in R (version 4.3.3, R Core Team, 2024). The 95% CIs of the slopes from two linear regressions were compared for significant differences (p < 0.05). Results Bt crops enhanced the soil respiration with a greater biomass Relative to non- Bt counterparts, Bt crops displayed a significant advantage in biomass accumulation, causing enhancements of 14.3% and 4.3% for aboveground and belowground biomass, respectively (p < 0.05, Fig. 2 a and b). Under Bt crop cultivation, soil respiration enhanced significantly (SR, + 6.0%, p < 0.05, Fig. 3 a), and showed positive correlation with changes of soil organic carbon (SOC, p < 0.05), nematode diversity (ND, p < 0.01), fungal diversity (FD, p < 0.05), microbial biomass nitrogen (MBN, p < 0.05), and dissolved organic carbon (DOC, p < 0.1, Fig. 3 ), respectively. Among these correlated factors, SOC, ND, FD and DOC were increased by Bt crops 2.1%, 2.6%, 0.6%, and 1.6%, respectively (p < 0.05), while MBN was decreased by 9.1% (p < 0.05, Fig. 2 ). The changes of SOC, ND, FD explained 50%, 34%, 30% variation of soil respiration, respectively (Fig. 3 b and c). Different impacts of Bt crops on rhizosphere and bulk soil Cultivation of Bt crops significantly altered soil carbon, nitrogen and phosphorus pools, including SOC (p < 0.001), DOC (p < 0.05), MBC (p < 0.001), DON (p < 0.001), MBN (p < 0.001), soil NH 4 + (p < 0.001), soil NO 3 − (p < 0.001), and STP (p 0.10, Figs. 2 and 4 ). Under the cultivation of Bt crops, rhizospheric soil displayed different response with bulk soil for SOC, DOC, MBC, DON, MBN, NH 4 + , NO 3 − , STP, BD and FD (p < 0.05). Bt crops decreased SOC and STP by 3.4% and 5.1%, in rhizospheric soil, but increased it by 3.1% and 2.0%, in bulk soil, respectively (Fig. 4 ). The variables DOC, DON, NH 4 + , NO 3 − , BD and FD displayed greater positive responses to Bt crops in rhizospheric soil than that in bulk soil, while the trend of MBC was contrary (Fig. 4 ). Rhizospheric soil also displayed a greater decrement of MBN (-19.9%) relative to bulk soil (-9.1%, Fig. 4 ). Bt crops affected soil microbial diversity and stoichiometry Under Bt crop cultivation, the diversity of soil bacterias and nematodes enhanced significantly (Figs. 2 , S1 and S2). The response of soil bacterial diversity in rhizosphere was significantly higher than that in bulk soil (Fig. S1 c). Effects of Bt crops on FD were also different between rhizospheric and bulk soil, with a significant positive influence in the former and a non-significant one in the latter (Fig. S1 d). Bt crop cultivation didn’t affect stoichiometric relationship between DOC and DON in soil, but induced a significant decrement in nitrogen of microbial biomass (MBN) relative to that in carbon (MBC, Fig. 5 ). The slope of MBN responses vs MBC responses (0.3815 ± 0.1679) was significantly lower than that of 1:1 line (p < 0.05, Fig. 5 ). Discussion Enhanced soil respiration under Bt crop cultivation Under Bt crop cultivation, both above- and below-ground biomass accumulation were significantly increased relative to that in their non- Bt counterparts, since the greater growth investment than defensive one due to the advantage of Bt crops in pest and disease resistance (Fig. 2 , Icoz & Stotzky, 2008 , Lee et al., 2017 ). With greater carbon accumulation, Bt crops led to a new Nash equilibria with soil organisms (e.g., soil microbes), specifically, the former derived more organic matter input into soil system (i.e., a rise of SOC), and triggered decompostion acceleration of the latter, and induced a great soil respiration (Figs. 3 a and 4 a, Cai et al., 2020 , Macdonald et al., 2021 , Moore et al., 2020 ). However, the response degree of soil respiration (+ 6.0%, p < 0.05, n = 104) was significantly larger than that of SOC (+ 3.1%, p < 0.05, n = 40, Fig. 6 ). The additional CO 2 efflux from rhizosphere would be the potential reason for the Bt crops’ greater response of soil respiration relative to that of SOC (Fig. 6 , Butterly et al., 2016 , Jackson et al., 2019 ). Relative to their non- Bt counterparts, Bt crops’ advantage in plant growth, especially in root growth, would induce a considerable variation in rhizospheric carbon process, such as root exduation, which might cause a priming effect on rhizospheric microbes for organic matter decomposition (Figs. 2 and 6 , Jackson et al., 2019 , Yin et al., 2020 ). The lower rhizospheric soil organic carbon under Bt crop cultivation than that under non- Bt counterparts, reflected the more organic matter decomposited by microbes in Bt crops’ rhizosphere (Figs. 4 and 6 , López et al., 2023 ). To compete for available nutrients, rhizospheric organisms shifted their strategy and exhibited a heightened respiratory and decomposition activity under Bt crop cultivation, resulting in a disproportionately great increase in soil respiration compared to soil organic carbon (Fig. 5 , Cao et al., 2024 ). Therefore, the ecological impacts of Bt crops could be mitigated through nutrient management strategies, that an appropriate increase in nitrogen fertilization could be a potential strategy to enhance soil carbon sink under Bt crop cultivation (Fig. 6 , Li et al., 2018 , Park et al., 2011 ). Bt crops modulated rhizosphere stoichiometry To support the significant advantage in biomass accumulation (Fig. 2 ), Bt crops are likely to exhibit enhanced nutrient uptake in the rhizophere to fulfill their increased growth demands (Barber, 1979 , Reichardt & Timm, 2020 ). Nutrient uptake at the soil-root interface is anticipated to be modulated in accordance with the growth requirements of plants (Ragland et al., 2024 ). For soil available nitrogen (soil NO 3 − and NH 4 + ) as an example, Bt crops had a significant positive effect in both rhizospheric and bulk soil, but a more pronounced changes in rhizosphere (Fig. 4 ). A great nitrogen need in anabolism for Bt crops (e.g., the Bt gene for nitrogen-based toxin proteins) would trigger an improvement of N uptake, reflected in the significant decrease in aboveground biomass C:N (-18.7 to -27.3%, p < 0.05), indicating a nutritional benefit for both plants and soil from tansgenic crops (Kumar et al., 2020 , Wu et al., 2007 ). However, for microbial nutrients, a substantial shift in microbial stoichiometry (an increase in microbial biomass C:N by 14.1–20.4%, p < 0.05) had been observed under Bt crop cultivation, mediated by rhizo-deposition, crop residues, and gas-water exchanges (Singh & Dubey, 2017 ). The concurrent increase of available NO 3 − and NH 4 + in both rhizospheric and bulk soil suggests a potential decline in relative proportion of microbe-immobilized nitrogen under Bt crop cultivation, indicating a shift in nitrogen competitive interaction between microbes and plants, particularly in the rhizosphere (a geater decline of MBN in rhizosphere compared to the bulk soil, Fig. 6 , Gannett et al., 2024 ). Combined the significant lower regression slope in the response of microbial biomass N vs C compared with that in 1:1 line, and no significant relative change in response of desoilved organic C and N, it was presumed that a reduction in microbial nitrogen competition occurred under Bt cultivation (Fig. 5 , Kuzyakov & Xu, 2013 ). In addition, for Bt crops, the aboveground biomass N:P was decreased by 13.2–22.3% relative to the parent breed in this study (p < 0.05), implying Bt crops absorbed, translocated and utilized more rhizospheric phosphorus compared to their non- Bt counterparts (He et al., 2019 ). The significant reduction of soil total phosphorus (STP) in rhizosphere vs the significant increment in bulk soil under Bt crop cultivation, suggesting Bt crops caused significant limitation of phosphorus resources in rhizosphere but no significant depression in phosphorus returned from crop litter (Veneklaas, 2022 ). Bt -crop cultivation affected rhizospheric biodiversity Trangenic crops, compared to conventional ones, have an advantage in avoiding various biotic disturbances, which are known to significantly affect belowground biotic processes (Zhang et al., 2015 ). As a hotspot of belowground biotic processes, the rhizosphere of Bt crops exhibited a distinct microbial community composition compared to conventional crops (e.g., differences in bacterial and fungal diversity, Figs. S1 and 6, Ahamd et al., 2017 ). Under Bt crop cultivation, the positive response of bacterial diversity in rhizosphere was significantly great than that in the bulk soil (Fig. S1 ), which also triggered more available nitrogen release from organic matter in rhizosphere through potential increase in bacterial function diversity (Fig. 4 , van Wyk et al., 2017 ). In addition, the diveristy of both nematodes and rhizospheric fungi under Bt crop cultivation was also slightly higher than that in conventional crops, potentially due to the increased organic matter exudation by Bt crop roots (Fig. 6 , Zeng et al., 2019 ). Furthermore, the diversity of both fungi and nematodes under Bt crop cultivation correlated positively with soil respiration, especially for changes in nematode diversity, which explained for 34% of the variation in soil respiration (Fig. 3 ). Soil nematodes with higher diversity under Bt crop cultivation, which might enhance the sequestration and redistribution of carbon and energy from the roots or aboveground litter through the food web, and regulate spatial characteristics of soil organic carbon and soil respiration (Fig. 6 , Men et al., 2003 , Young & Unc, 2023 ). This study provided a valuable insight into the ecological impacts associated with the cultivation of Bt crops. Specifically, relative to convential agriculture, Bt crops promoted decomposition of soil organic matter and soil respiration, and caused a considerable impact for soil fertility due to the nutrient mining by rhizospheric microbes (Fig. 6 , Abbas, 2018 , Ahamd et al. , 017). However, it is imperative to acknowledge the limitations inherent in our research. Our investigation focused on the theoretical Nash equilibrium dynamics between crops and soil organisms (Blaser & Kirschner, 2007 ), with a particular emphasis on the belowground processes. Due to the relative insufficience for simutaneous records of plants and soil organisms, the potential relationship between these belowground variables and underlyine paths cannot be explored on the basis of limited data (Čapek et al., 2018 ). It is essential to focus more on the changes in plant-soil organism interactions triggered by genetically modified crops in the future, providing a better understanding of ecological impacts for such crops and informing sustainable agricultural practices (Cao et al., 2024 ) . Conclusion Bt crops, engineered for desired traits, contribute to meeting the needs of a growing global population. This study reveals that Bt crops have a significant advantage in biomass accumulation and carbon sequestration compared to their parent breed, creating a Nash equilibria between crops and soil organisms. The effects of Bt- crop growth on the stoichiometry of crop-soil-microbe system, as well as the diversity of bacterias, fungi and nematodes, particularly in rhizospheric soil, collectively pormote the stimulated soil respiration. For sustainable agriculture, it is thus crucial to dynamically match soil nutrient supply with crop requirements ( Bt crops or parent breed), preventing excessive nutrient mining by microbes from complex soil organic compounds. Therefore, an optimized fertilization practices are necessary to improve rhizospheric nutrient conditions, especially for enhancing life cycle carbon sink potential in transgenic crop cultivation. Declarations Conflict of Interest The authors declare that they have no conflict of interest. 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Pedosphere 29:114–122 Supplementary Files SupplementaryMaterials.docx Cite Share Download PDF Status: Published Journal Publication published 16 Dec, 2025 Read the published version in Plant and Soil → Version 1 posted Editorial decision: Major revisions 25 Aug, 2025 Reviewers agreed at journal 19 Jul, 2025 Reviewers invited by journal 16 Jul, 2025 Editor invited by journal 21 May, 2025 Editor assigned by journal 21 May, 2025 First submitted to journal 20 May, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6699296","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":486415829,"identity":"71e293ef-a1fe-4d92-b3f0-4ab1b13e3f67","order_by":0,"name":"Lingyan Zhou","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA50lEQVRIie3RsQqCQBjA8e8QzsWyUYl6BsOxqFe5Q8jFoak5CK4lcbW3qM0t46BJchVcnJwaapSWLIm2C7eG+w93xx0/vuEAZLI/rAuA622CVt87Iia4IfP2hLchhlsWd5YOAj1GRRVx0FXPgioSEeKOdiy3dyFRRn7CwdxeLeQnQnLud1hO9xngPmIcrMyzFMREhLKaXOgxjdXHi8x+EwfXJKZ7IFh5TzF+Ea3EZnhx7DCja9NnrmYk5eLkC4iuuqVxW04HQcBPt4qNh/rGORSVgECPfE7N12ivJRaAeoz4WSaTyWQATwVITLbvuxP2AAAAAElFTkSuQmCC","orcid":"https://orcid.org/0000-0002-5138-5188","institution":"East China Normal University","correspondingAuthor":true,"prefix":"","firstName":"Lingyan","middleName":"","lastName":"Zhou","suffix":""},{"id":486415830,"identity":"63f43637-bcfa-42c1-b605-b8be4257472a","order_by":1,"name":"Shuxian Jia","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Shuxian","middleName":"","lastName":"Jia","suffix":""},{"id":486415831,"identity":"86ae9951-5920-4a00-ac46-3f226755a80d","order_by":2,"name":"Hengshuo Zhang","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Hengshuo","middleName":"","lastName":"Zhang","suffix":""},{"id":486415832,"identity":"041948fe-3e25-4f6b-8fbd-1e59fe701acf","order_by":3,"name":"Kaiyan Zhai","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Kaiyan","middleName":"","lastName":"Zhai","suffix":""},{"id":486415833,"identity":"470048a4-86a8-4015-9840-dd298a64872d","order_by":4,"name":"Xuhui Zhou","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Xuhui","middleName":"","lastName":"Zhou","suffix":""}],"badges":[],"createdAt":"2025-05-19 13:10:51","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6699296/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6699296/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1007/s11104-025-08189-6","type":"published","date":"2025-12-16T15:58:17+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":87174156,"identity":"22129cd8-1b7c-4417-82c1-aa709f953159","added_by":"auto","created_at":"2025-07-21 08:12:12","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":337023,"visible":true,"origin":"","legend":"\u003cp\u003eSchematic diagram for triple equilibria (pests-crops-soil organisms, \u003cstrong\u003ea\u003c/strong\u003e) and mutual equilibria (\u003cem\u003eBt crop\u003c/em\u003es- soil organisms, \u003cstrong\u003eb\u003c/strong\u003e) in conventional crop and transgenic \u003cem\u003eBt\u003c/em\u003e crop cultivation, respectively.\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-6699296/v1/bd7c37aa09a95851a89368bf.png"},{"id":87174157,"identity":"da99cec9-a8a1-4b81-aa4b-8b71cafdedb0","added_by":"auto","created_at":"2025-07-21 08:12:12","extension":"jpeg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":424604,"visible":true,"origin":"","legend":"\u003cp\u003eEffect of transgenic \u003cem\u003eBt \u003c/em\u003ecrops on above-(\u003cstrong\u003ea\u003c/strong\u003e), below-ground biomass (\u003cstrong\u003eb\u003c/strong\u003e) and other variables in soil (\u003cstrong\u003ec\u003c/strong\u003e). The center of grey circle was the response ratio of individual study, the red circle was the total weighted response ratio (mean ± se) in plot \u003cstrong\u003ea\u003c/strong\u003e and \u003cstrong\u003eb\u003c/strong\u003e. The center of circles in plot \u003cstrong\u003ec \u003c/strong\u003ewas the weighted response ratio, and the error bars indicated 95% confidence interval (CI). If the 95% CI of weighted response ratio did not overlap with zero, the effect of transgenic \u003cem\u003eBt\u003c/em\u003e crops was considered to be statistically significant, and the red or blue symbol * indicated statistical significance of increase and decrease at \u003cem\u003ep\u003c/em\u003e\u0026lt;0.05, respectively, based on two-tailed tests. The number below the blue and orange circles were the sample size. The circles with different letters had statistical difference at \u003cem\u003ep\u003c/em\u003e\u0026lt;0.05, based two-tailed tests. AGB, aboveground biomass; BGB, belowground biomass; MBN, microbial biomass nitrogen; STP, soil total phosphorus; SON, soil organic nitrogen; FD, fungal diversity; ND, nematode diversity; DOC, dissolved organic carbon; SOC, soil organic carbon; SCE, soil CO\u003csub\u003e2\u003c/sub\u003e emission; BD, bacterial diversity; DON, dissolved organic nitrogen; Soil NH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e, soil ammonium; MBC, microbial biomass carbon; Soil NO\u003csub\u003e3\u003c/sub\u003e\u003csup\u003e-\u003c/sup\u003e, soil nitrate.\u003c/p\u003e","description":"","filename":"floatimage2.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-6699296/v1/8114e4b8d28cd435eeb0bf1b.jpeg"},{"id":87174326,"identity":"1610bc15-cd90-46f0-b699-bf8125aa127a","added_by":"auto","created_at":"2025-07-21 08:20:12","extension":"jpeg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":438331,"visible":true,"origin":"","legend":"\u003cp\u003eThe response of soil CO\u003csub\u003e2\u003c/sub\u003e emission to transgenic \u003cem\u003eBt \u003c/em\u003ecrops, and its relationship with responses of other variables. \u003cstrong\u003ea\u003c/strong\u003e The response ratio and weighted response ratio of soil CO\u003csub\u003e2\u003c/sub\u003e emission;\u003cstrong\u003e b\u003c/strong\u003e-\u003cstrong\u003ed\u003c/strong\u003e The response relationship between soil respiration with fungal diversity (FD), nematode diversity (ND), dissolved organic carbon (DOC), soil organic carbon (SOC), and microbial biomass nitrogen (MBN); \u003cstrong\u003ee\u003c/strong\u003e The Pearson correlation coefficient of response ratio (\u003cem\u003eRR\u003c/em\u003e) of soil CO\u003csub\u003e2\u003c/sub\u003e emission with that of other variables, the symbols *and ** indicated significant correlation coefficient at \u003cem\u003ep\u003c/em\u003e\u0026lt;0. 05 and 0.01, respectively; the symbol ξ indicated the correlation was significant at\u003cem\u003e p\u003c/em\u003e\u0026lt;0.1. The center of grey circle in plot \u003cstrong\u003ea\u003c/strong\u003e was the response ratio of individual study, the red circle was the total weighted response ratio (mean ± se). The grey error bands in plot \u003cstrong\u003eb-d\u003c/strong\u003e represented the upper and lower 95% confidence intervals. The \u003cem\u003ep\u003c/em\u003e-values were calculated from two-tailed tests. BGB, belowground biomass; Soil NH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e, soil ammoniacal nitrogen; BD, bacterial diversity; DON, dissolved organic nitrogen; MBC, microbial biomass carbon; Soil NO\u003csub\u003e3\u003c/sub\u003e\u003csup\u003e-\u003c/sup\u003e, soil nitrate nitrogen.\u003c/p\u003e","description":"","filename":"floatimage3.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-6699296/v1/ef36451b38409602f74b3933.jpeg"},{"id":87174165,"identity":"7cfce3a2-6b0e-413c-a84a-08d792cf1181","added_by":"auto","created_at":"2025-07-21 08:12:13","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":380845,"visible":true,"origin":"","legend":"\u003cp\u003eThe response of soil carbon (\u003cstrong\u003ea-c\u003c/strong\u003e) and nitrogen (\u003cstrong\u003ed-f\u003c/strong\u003e) to transgenic \u003cem\u003eBt \u003c/em\u003ecrops. The center of grey circle was the response ratio of individual study, the red circle was the total weighted response ratio (mean ± se), while the blue and orange circle was the weighted response ratio in rhizosphere and bulk soil, respectively. The error bars of blue and orange circles indicated 95% confidence interval (CI). If the 95% CI of weighted response ratio did not overlap with zero, the effect of transgenic \u003cem\u003eBt \u003c/em\u003ecrops was considered to be statistically significant, and the red or blue symbol * indicated statistical significance of increase and decease at \u003cem\u003ep\u003c/em\u003e\u0026lt;0.05, respectively, based on two-tailed tests. The number beside of the blue and orange circles were the sample size. SOC, soil organic carbon; DOC, dissolved organic carbon; MBC, microbial biomass carbon; SON, soil organic nitrogen; DON, dissolved organic nitrogen; MBN, microbial biomass nitrogen.\u003c/p\u003e","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-6699296/v1/16ec484cc06c47da12b61d50.png"},{"id":87174334,"identity":"61452bed-1d34-481f-b563-94a7b092e2a0","added_by":"auto","created_at":"2025-07-21 08:20:13","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":184972,"visible":true,"origin":"","legend":"\u003cp\u003eThe response relationship between microbial biomass carbon (MBC) vs nitrogen (MBN, \u003cstrong\u003ea\u003c/strong\u003e), and between dissolved organic carbon (DOC) vs nitrogen (DON, \u003cstrong\u003eb\u003c/strong\u003e) in transgenic \u003cem\u003eBt \u003c/em\u003ecrop\u003cem\u003e \u003c/em\u003ecultivation. In plot \u003cstrong\u003ea\u003c/strong\u003e, the red arrow indicated the significant difference between the slope of the regression line and 1 (\u003cem\u003ep\u003c/em\u003e\u0026lt;0.05), based on two-tailed test.\u003c/p\u003e","description":"","filename":"floatimage5.png","url":"https://assets-eu.researchsquare.com/files/rs-6699296/v1/9ecc6d6fda57bd4e4fa7b3a3.png"},{"id":87174162,"identity":"cbc5b065-6da7-4dcf-8410-ae2eca7c522b","added_by":"auto","created_at":"2025-07-21 08:12:13","extension":"jpeg","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":1150396,"visible":true,"origin":"","legend":"\u003cp\u003eIllustration of our finding of transgenic \u003cem\u003eBt \u003c/em\u003ecrop cultivation’s effect on above-, blow-ground plant biomass (AGB and BGB), soil organic carbon (SOC) and soil CO\u003csub\u003e2\u003c/sub\u003e emission (SCE) in comparison with conventional non-\u003cem\u003eBt\u003c/em\u003e crops. The green, blue, yellow and red squares are variables related to carbon, nitrogen, phosphorus and biodiversity. The red frames are variable significantly correlated with soil respiration (Figure 2). The number in parentheses in each square was the sample size for each variable in rhizosphere or/and bulk soil. The black up- and down-ward arrow indicated significant increase and decrease of variables at \u003cem\u003ep\u003c/em\u003e\u0026lt;0.05, respectively, whereas “ns” represented non-significant changes of variables in responding to transgenic \u003cem\u003eBt\u003c/em\u003e crop cultivation.\u003c/p\u003e","description":"","filename":"floatimage6.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-6699296/v1/ceac9ba72ecc2af86a72e0ff.jpeg"},{"id":98815074,"identity":"4b5f65a6-7cb0-46e5-90e6-df518d6aade8","added_by":"auto","created_at":"2025-12-22 16:13:28","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":3633347,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6699296/v1/a51bd0d7-b66a-4f41-a3e4-d6901df0abd9.pdf"},{"id":87174332,"identity":"6691c6ff-a8ac-4538-9c63-0db525e7ec70","added_by":"auto","created_at":"2025-07-21 08:20:13","extension":"docx","order_by":5,"title":"","display":"","copyAsset":false,"role":"supplement","size":304079,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryMaterials.docx","url":"https://assets-eu.researchsquare.com/files/rs-6699296/v1/41c64140e69f33700927c669.docx"}],"financialInterests":"","formattedTitle":"Simultaneous Stimulation of Soil Respiration and Plant Biomass in Transgenic Bacillus thuringiensis Crop Cultivation","fulltext":[{"header":"Introduction","content":"\u003cp\u003eAs global climate change intensifies, agricultural systems confront mounting pressures from increasingly severe pest infestations, pathogen proliferation, and erratic weather patterns\u0026mdash;posing grave risks to worldwide food stability and ecological balance (Singh et al., \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2023\u003c/span\u003e, Zhang et al., \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Transgenic crops, which boast inherent benefits, such as herbicide tolerance, insect resistance, disease resistance, and improved nutritional profiles, have emerged as a key strategy to address these challenges. By 2020, over 500 transgenic events in about 30 different crops had received global approval for cultivation (Kumar et al., \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Among these, \u003cem\u003eBacillus thuringiensis\u003c/em\u003e (\u003cem\u003eBt\u003c/em\u003e) crops stand out as they are genetically engineered to produce endospore (or crystal) toxins of the bacterium, providing them with a natural defense against a range of insect pests, including Lepidoptera, Coleoptera, and Hymenoptera (Abbas, \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Extensive research has shown that \u003cem\u003eBt\u003c/em\u003e crops, such as those for corn and cotton, consistently outperform their conventional counterparts in terms of yields (Carzoli et al., \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2018\u003c/span\u003e, Qaim, \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2020\u003c/span\u003e, Shelton et al., \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2013\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eThe enhanced metabolic efficiency in \u003cem\u003eBt\u003c/em\u003e crops, particularly in pathways such as the tricarboxylic acid cycle and energy metabolism, leads to increased biomass accumulation compared to their non-transgenic counterparts (Zhang et al., \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). This heightened pest suppression capability not only influences carbon allocation at the individual plant level but also alters ecosystem-scale carbon dynamics, shifting the traditional tripartite biotic interactions among pests, plants and soil organisms towards a binary interaction between the latter two (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e1\u003c/span\u003e, Suzuki et al., \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2000\u003c/span\u003e). Consequently, the impact of \u003cem\u003eBt\u003c/em\u003e crop cultivation on soil characteristics, including enzymatic activities and microbial functions, has become a focal point for research (Lee et al., \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2017\u003c/span\u003e, Li et al., \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2019\u003c/span\u003e, Zhang et al., \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). While \u003cem\u003eBt\u003c/em\u003e crop cultivation is widely perceived as environmentally benign, empirical data suggests that it does not remain without effect on soil processes or biota, especially under long-term cultivation scenarios where the exclusion of pests from the agroecosystem results in increased carbon sequestration within crop biomass (Krogh et al., \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2020\u003c/span\u003e, Lee et al., \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Despite its carbon-sink advantages, the decoupling of pest-mediated carbon-nutrient feedbacks, and long-term shifts in soil enzymatic-microbial dynamics might strain soil nutrient reserves and affect the interactions between crops and soil organisms (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eThe previous interactions among pests, crops and soil organisms in agrosystem would no longer occur due to the unilaterally strategy change in crops (i.e., \u003cem\u003eBt\u003c/em\u003e protein expression by crops), leading to a new Nash equilibrium (a key concept in game theory, in which no player\u0026rsquo;s expected outcome can be improved by changing one\u0026rsquo;s own strategy, Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e1\u003c/span\u003e, Blaser \u0026amp; Kirschner, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2007\u003c/span\u003e, Cai et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2020\u003c/span\u003e, this concept is increasingly applied to model resource competition among organisms in ecosystem). Resisting pest attacks, \u003cem\u003eBt\u003c/em\u003e crops would perform a set of interaction with soil organisms to achieve a new beneficial balance (e.g., enhancing the availability of underground resources), escpecially in rhizosphere, the hotspot for plant - soil organims interactions (Čapek et al., \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). However, the response of soil organism-dominated carbon and nutrient processes (e.g., soil respiration and soil nutrient mineralization) in soil system, as well as soil organism community in rhizosphere or bulk soil under \u003cem\u003eBt\u003c/em\u003e crop cultivation remains largely unexplored. This gap in knowledge constrains our comprehensive understanding of the ecological implications of \u003cem\u003eBt\u003c/em\u003e crop cultivation, particularly in the context of sustainable agricultural systems (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e1\u003c/span\u003e, Mmbando \u0026amp; Ngongolo, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2024\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eConsidering coupling relationship between carbon and nutrient cycles within ecosystem, we propose two hypotheses in the new Nash equilibria between \u003cem\u003eBt\u003c/em\u003e crops and soil organisms. Fristly, \u003cem\u003eBt\u003c/em\u003e crops may yield a generally greater biomass and cause a greater demand for nutrients relative to their non-transgenic counterparts. Secondly, the increasing assimilation products in \u003cem\u003eBt\u003c/em\u003e crops would accompany with soil organism regulation of soil carbon and nutrient cycle (e.g., nitrogen and phosphorus), which would be reflected by a change of soil respiration. To test these hypotheses, we employed a meta-analysis to assess responses of soil respiration, soil carbon and nutrient pools, soil biodiversity, and crop biomass to \u003cem\u003eBt\u003c/em\u003e crop cultivation. The diversity of soil organisms and stoichiometry in both rhizosphere and bulk soil were evaluated to provide some underlying explanation for effect of \u003cem\u003eBt\u003c/em\u003e crop cultivation on soil carbon and nutrient cycle. Our fundings indicate that \u003cem\u003eBt\u003c/em\u003e crops cause more carbon sequestrated in both above- and below-ground plant biomass compared to conventional varieties. In the new Nash equilibrium between crops and soil organisms (including bacterias, fungi, and nematodes), the soil respiration significantly enhanced, accompanying with decreases in microbial biomass carbon: nitrogen ratio, and rhizosphere SOC.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003eData sources\u003c/h2\u003e\u003cp\u003eWe conducted a comprehensive literature search using the combination \u0026ldquo;(\u003cem\u003eBacillus thuringiensis\u003c/em\u003e OR \u003cem\u003eBt\u003c/em\u003e) AND (soil OR carbon OR respiration OR CO\u003csub\u003e2\u003c/sub\u003e OR microbe* OR bacteria* OR fung* OR nitrogen OR ammoni* OR nitrate OR phosphor* OR organism* OR diverisity OR yield OR biomass OR root OR biomass)\u0026rdquo;. The peer-reviewed journal articles were sought using Web of Science (1990\u0026ndash;2024) and China National Knowledge Infrastructure (CNKI, 1992\u0026ndash;2024). To avoid bias in publication selection, the studies were selected on the basis of the following considerations: i) The sole differentiating factor between groups of \u003cem\u003eBt\u003c/em\u003e crops and their conventional counterparts was the genetic modification, with all other management practices being consistent across treatments; ii) Experiments were conducted in the field, recording at least one pair of comparative data of conventional non-\u003cem\u003eBt\u003c/em\u003e crop and \u003cem\u003eBt\u003c/em\u003e crop varieties, such as aboveground biomass (AGB), belowground biomass (BGB), soil CO\u003csub\u003e2\u003c/sub\u003e emission (SCE), soil organic carbon (SOC), soil organic nitrogen (SON), soil dissovled organic carbon (DOC), soil dissovled orgnaic nitogen (DON), soil ammonium (soil NH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e), soil nitrate (soil NO\u003csub\u003e3\u003c/sub\u003e\u003csup\u003e\u0026minus;\u003c/sup\u003e), soil total phosphorus (STP), soil pH (pH), microbial biomass carbon (MBC), microbial biomass nitrogen (MBN), bacterial, fungal and/or nematode diversity (BD, FD, and/or ND, including Shannon, Richness, InvSimpson (1-Simpson), Chao 1, ACE, and/or operational taxonomic unit (OUT)); iii) The means, standard deviations/errors, and samples sizes of variables could be extracted directly from the text, tables or digitized graphs; iv) The experimental duration were longer than one growing season. Applying these criteria, we compiled a dataset from 88 published papers focusing on the cultivation of \u003cem\u003eBt\u003c/em\u003e genetically modified maize, cotton, or rice (PRISMA in Supplementary Materials).\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec4\" class=\"Section2\"\u003e\u003ch2\u003eStatistical analysis\u003c/h2\u003e\u003cp\u003eIn this study, we utiliazed the response ratio (\u003cem\u003eRR\u003c/em\u003e), defined as the natural log of the ratio of the mean value in cultivation of \u003cem\u003eBt\u003c/em\u003e crops (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\stackrel{-}{{X}_{t}}\\)\u003c/span\u003e\u003c/span\u003e) to that in control, i.e., cultivation of conventional parent breed (\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\stackrel{-}{{X}_{\\text{c}}}\\)\u003c/span\u003e\u003c/span\u003e, Eq.\u0026nbsp;\u003cspan refid=\"Equ1\" class=\"InternalRef\"\u003e1\u003c/span\u003e), to quantify the impact of \u003cem\u003eBt\u003c/em\u003e genetic modification on the concerned variables relative to the conventional cultivation methods (Luo et al., \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2006\u003c/span\u003e).\u003cdiv id=\"Equ1\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equ1\" name=\"EquationSource\"\u003e\n$$\\:RR=ln\\left(\\frac{\\stackrel{-}{{X}_{t}}}{\\stackrel{-}{{X}_{c}}}\\right)={ln}\\left(\\stackrel{-}{{X}_{t}}\\right)-ln\\left(\\stackrel{-}{{X}_{c}}\\right)$$\u003c/div\u003e\u003cdiv class=\"EquationNumber\"\u003e1\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003eSubsequently, the weighted average effect sizes (\u003cem\u003eRR\u003c/em\u003e\u003csub\u003e\u003cem\u003e++\u003c/em\u003e\u003c/sub\u003e) and 95% confidence intervals (CI) for each variable was calculated using \u003cem\u003erma.mv\u003c/em\u003e function in \u003cem\u003emetafor\u003c/em\u003e package (version 3.4-0, Viechtbauer, \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2010\u003c/span\u003e), based on the numbers of replicates and standard deviations for a given variable under both \u003cem\u003eBt\u003c/em\u003e and conventional crop cultivation. A significant effect of \u003cem\u003eBt\u003c/em\u003e crop cultivation on a variable was considered when the estimated CI did not overlap zero. The percentage change in a concerned variable under \u003cem\u003eBt\u003c/em\u003e crop cultivation was estimated by [exp (\u003cem\u003eRR\u003c/em\u003e\u003csub\u003e\u003cem\u003e++\u003c/em\u003e\u003c/sub\u003e)\u0026thinsp;\u0026minus;\u0026thinsp;1] \u0026times; 100%. The Pearson correlation coefficient between response of soil CO\u003csub\u003e2\u003c/sub\u003e emission and that of other variables was analyzed firstly, and then the linear regression was tested using \u003cem\u003emetafor\u003c/em\u003e package and drawn with \u003cem\u003eggplot2\u003c/em\u003e package (version 3.4.4) in R (version 4.3.3, R Core Team, 2024). The 95% CIs of the slopes from two linear regressions were compared for significant differences (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05).\u003c/p\u003e\u003c/div\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cb\u003eBt\u003c/b\u003e \u003cb\u003ecrops enhanced the soil respiration with a greater biomass\u003c/b\u003e\u003c/p\u003e\u003cp\u003eRelative to non-\u003cem\u003eBt\u003c/em\u003e counterparts, \u003cem\u003eBt\u003c/em\u003e crops displayed a significant advantage in biomass accumulation, causing enhancements of 14.3% and 4.3% for aboveground and belowground biomass, respectively (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05, Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e2\u003c/span\u003ea and b). Under \u003cem\u003eBt\u003c/em\u003e crop cultivation, soil respiration enhanced significantly (SR, +\u0026thinsp;6.0%, p\u0026thinsp;\u0026lt;\u0026thinsp;0.05, Fig.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e3\u003c/span\u003ea), and showed positive correlation with changes of soil organic carbon (SOC, p\u0026thinsp;\u0026lt;\u0026thinsp;0.05), nematode diversity (ND, p\u0026thinsp;\u0026lt;\u0026thinsp;0.01), fungal diversity (FD, p\u0026thinsp;\u0026lt;\u0026thinsp;0.05), microbial biomass nitrogen (MBN, p\u0026thinsp;\u0026lt;\u0026thinsp;0.05), and dissolved organic carbon (DOC, p\u0026thinsp;\u0026lt;\u0026thinsp;0.1, Fig.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e3\u003c/span\u003e), respectively. Among these correlated factors, SOC, ND, FD and DOC were increased by \u003cem\u003eBt\u003c/em\u003e crops 2.1%, 2.6%, 0.6%, and 1.6%, respectively (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05), while MBN was decreased by 9.1% (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05, Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e2\u003c/span\u003e). The changes of SOC, ND, FD explained 50%, 34%, 30% variation of soil respiration, respectively (Fig.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e3\u003c/span\u003eb and c).\u003c/p\u003e\u003cp\u003e\u003cb\u003eDifferent impacts of\u003c/b\u003e \u003cb\u003eBt\u003c/b\u003e \u003cb\u003ecrops on rhizosphere and bulk soil\u003c/b\u003e\u003c/p\u003e\u003cp\u003eCultivation of \u003cem\u003eBt\u003c/em\u003e crops significantly altered soil carbon, nitrogen and phosphorus pools, including SOC (p\u0026thinsp;\u0026lt;\u0026thinsp;0.001), DOC (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05), MBC (p\u0026thinsp;\u0026lt;\u0026thinsp;0.001), DON (p\u0026thinsp;\u0026lt;\u0026thinsp;0.001), MBN (p\u0026thinsp;\u0026lt;\u0026thinsp;0.001), soil NH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e (p\u0026thinsp;\u0026lt;\u0026thinsp;0.001), soil NO\u003csub\u003e3\u003c/sub\u003e\u003csup\u003e\u0026minus;\u003c/sup\u003e (p\u0026thinsp;\u0026lt;\u0026thinsp;0.001), and STP (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05), excepting for SON (p\u0026thinsp;\u0026gt;\u0026thinsp;0.10, Figs.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e2\u003c/span\u003e and \u003cspan refid=\"Fig10\" class=\"InternalRef\"\u003e4\u003c/span\u003e). Under the cultivation of \u003cem\u003eBt\u003c/em\u003e crops, rhizospheric soil displayed different response with bulk soil for SOC, DOC, MBC, DON, MBN, NH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e, NO\u003csub\u003e3\u003c/sub\u003e\u003csup\u003e\u0026minus;\u003c/sup\u003e, STP, BD and FD (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). \u003cem\u003eBt\u003c/em\u003e crops decreased SOC and STP by 3.4% and 5.1%, in rhizospheric soil, but increased it by 3.1% and 2.0%, in bulk soil, respectively (Fig.\u0026nbsp;\u003cspan refid=\"Fig10\" class=\"InternalRef\"\u003e4\u003c/span\u003e). The variables DOC, DON, NH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e, NO\u003csub\u003e3\u003c/sub\u003e\u003csup\u003e\u0026minus;\u003c/sup\u003e, BD and FD displayed greater positive responses to \u003cem\u003eBt\u003c/em\u003e crops in rhizospheric soil than that in bulk soil, while the trend of MBC was contrary (Fig.\u0026nbsp;\u003cspan refid=\"Fig10\" class=\"InternalRef\"\u003e4\u003c/span\u003e). Rhizospheric soil also displayed a greater decrement of MBN (-19.9%) relative to bulk soil (-9.1%, Fig.\u0026nbsp;\u003cspan refid=\"Fig10\" class=\"InternalRef\"\u003e4\u003c/span\u003e).\u003c/p\u003e\u003cp\u003e\u003cb\u003eBt\u003c/b\u003e \u003cb\u003ecrops affected soil microbial diversity and stoichiometry\u003c/b\u003e\u003c/p\u003e\u003cp\u003eUnder \u003cem\u003eBt\u003c/em\u003e crop cultivation, the diversity of soil bacterias and nematodes enhanced significantly (Figs.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e2\u003c/span\u003e, S1 and S2). The response of soil bacterial diversity in rhizosphere was significantly higher than that in bulk soil (Fig. \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003ec). Effects of \u003cem\u003eBt\u003c/em\u003e crops on FD were also different between rhizospheric and bulk soil, with a significant positive influence in the former and a non-significant one in the latter (Fig. \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003ed). \u003cem\u003eBt\u003c/em\u003e crop cultivation didn\u0026rsquo;t affect stoichiometric relationship between DOC and DON in soil, but induced a significant decrement in nitrogen of microbial biomass (MBN) relative to that in carbon (MBC, Fig.\u0026nbsp;\u003cspan refid=\"Fig11\" class=\"InternalRef\"\u003e5\u003c/span\u003e). The slope of MBN responses \u003cem\u003evs\u003c/em\u003e MBC responses (0.3815\u0026thinsp;\u0026plusmn;\u0026thinsp;0.1679) was significantly lower than that of 1:1 line (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05, Fig.\u0026nbsp;\u003cspan refid=\"Fig11\" class=\"InternalRef\"\u003e5\u003c/span\u003e).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003e\u003cb\u003eEnhanced soil respiration under\u003c/b\u003e \u003cb\u003eBt\u003c/b\u003e \u003cb\u003ecrop cultivation\u003c/b\u003e\u003c/p\u003e\u003cp\u003eUnder \u003cem\u003eBt\u003c/em\u003e crop cultivation, both above- and below-ground biomass accumulation were significantly increased relative to that in their non-\u003cem\u003eBt\u003c/em\u003e counterparts, since the greater growth investment than defensive one due to the advantage of \u003cem\u003eBt\u003c/em\u003e crops in pest and disease resistance (Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e2\u003c/span\u003e, Icoz \u0026amp; Stotzky, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2008\u003c/span\u003e, Lee et al., \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). With greater carbon accumulation, \u003cem\u003eBt\u003c/em\u003e crops led to a new Nash equilibria with soil organisms (e.g., soil microbes), specifically, the former derived more organic matter input into soil system (i.e., a rise of SOC), and triggered decompostion acceleration of the latter, and induced a great soil respiration (Figs.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e3\u003c/span\u003ea and \u003cspan refid=\"Fig10\" class=\"InternalRef\"\u003e4\u003c/span\u003ea, Cai et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2020\u003c/span\u003e, Macdonald et al., \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2021\u003c/span\u003e, Moore et al., \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). However, the response degree of soil respiration (+\u0026thinsp;6.0%, p\u0026thinsp;\u0026lt;\u0026thinsp;0.05, n\u0026thinsp;=\u0026thinsp;104) was significantly larger than that of SOC (+\u0026thinsp;3.1%, p\u0026thinsp;\u0026lt;\u0026thinsp;0.05, n\u0026thinsp;=\u0026thinsp;40, Fig.\u0026nbsp;\u003cspan refid=\"Fig12\" class=\"InternalRef\"\u003e6\u003c/span\u003e). The additional CO\u003csub\u003e2\u003c/sub\u003e efflux from rhizosphere would be the potential reason for the \u003cem\u003eBt\u003c/em\u003e crops\u0026rsquo; greater response of soil respiration relative to that of SOC (Fig.\u0026nbsp;\u003cspan refid=\"Fig12\" class=\"InternalRef\"\u003e6\u003c/span\u003e, Butterly et al., \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2016\u003c/span\u003e, Jackson et al., \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2019\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eRelative to their non-\u003cem\u003eBt\u003c/em\u003e counterparts, \u003cem\u003eBt\u003c/em\u003e crops\u0026rsquo; advantage in plant growth, especially in root growth, would induce a considerable variation in rhizospheric carbon process, such as root exduation, which might cause a priming effect on rhizospheric microbes for organic matter decomposition (Figs.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e2\u003c/span\u003e and \u003cspan refid=\"Fig12\" class=\"InternalRef\"\u003e6\u003c/span\u003e, Jackson et al., \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2019\u003c/span\u003e, Yin et al., \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). The lower rhizospheric soil organic carbon under \u003cem\u003eBt\u003c/em\u003e crop cultivation than that under non-\u003cem\u003eBt\u003c/em\u003e counterparts, reflected the more organic matter decomposited by microbes in \u003cem\u003eBt\u003c/em\u003e crops\u0026rsquo; rhizosphere (Figs.\u0026nbsp;\u003cspan refid=\"Fig10\" class=\"InternalRef\"\u003e4\u003c/span\u003e and \u003cspan refid=\"Fig12\" class=\"InternalRef\"\u003e6\u003c/span\u003e, L\u0026oacute;pez et al., \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). To compete for available nutrients, rhizospheric organisms shifted their strategy and exhibited a heightened respiratory and decomposition activity under \u003cem\u003eBt\u003c/em\u003e crop cultivation, resulting in a disproportionately great increase in soil respiration compared to soil organic carbon (Fig.\u0026nbsp;\u003cspan refid=\"Fig11\" class=\"InternalRef\"\u003e5\u003c/span\u003e, Cao et al., \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). Therefore, the ecological impacts of \u003cem\u003eBt\u003c/em\u003e crops could be mitigated through nutrient management strategies, that an appropriate increase in nitrogen fertilization could be a potential strategy to enhance soil carbon sink under \u003cem\u003eBt\u003c/em\u003e crop cultivation (Fig.\u0026nbsp;\u003cspan refid=\"Fig12\" class=\"InternalRef\"\u003e6\u003c/span\u003e, Li et al., \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2018\u003c/span\u003e, Park et al., \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2011\u003c/span\u003e).\u003c/p\u003e\u003cp\u003e\u003cb\u003eBt\u003c/b\u003e \u003cb\u003ecrops modulated rhizosphere stoichiometry\u003c/b\u003e\u003c/p\u003e\u003cp\u003eTo support the significant advantage in biomass accumulation (Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e2\u003c/span\u003e), \u003cem\u003eBt\u003c/em\u003e crops are likely to exhibit enhanced nutrient uptake in the rhizophere to fulfill their increased growth demands (Barber, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e1979\u003c/span\u003e, Reichardt \u0026amp; Timm, \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Nutrient uptake at the soil-root interface is anticipated to be modulated in accordance with the growth requirements of plants (Ragland et al., \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2024\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eFor soil available nitrogen (soil NO\u003csub\u003e3\u003c/sub\u003e\u003csup\u003e\u0026minus;\u003c/sup\u003e and NH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e) as an example, \u003cem\u003eBt\u003c/em\u003e crops had a significant positive effect in both rhizospheric and bulk soil, but a more pronounced changes in rhizosphere (Fig.\u0026nbsp;\u003cspan refid=\"Fig10\" class=\"InternalRef\"\u003e4\u003c/span\u003e). A great nitrogen need in anabolism for \u003cem\u003eBt\u003c/em\u003e crops (e.g., the \u003cem\u003eBt\u003c/em\u003e gene for nitrogen-based toxin proteins) would trigger an improvement of N uptake, reflected in the significant decrease in aboveground biomass C:N (-18.7 to -27.3%, p\u0026thinsp;\u0026lt;\u0026thinsp;0.05), indicating a nutritional benefit for both plants and soil from tansgenic crops (Kumar et al., \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2020\u003c/span\u003e, Wu et al., \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). However, for microbial nutrients, a substantial shift in microbial stoichiometry (an increase in microbial biomass C:N by 14.1\u0026ndash;20.4%, p\u0026thinsp;\u0026lt;\u0026thinsp;0.05) had been observed under \u003cem\u003eBt\u003c/em\u003e crop cultivation, mediated by rhizo-deposition, crop residues, and gas-water exchanges (Singh \u0026amp; Dubey, \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). The concurrent increase of available NO\u003csub\u003e3\u003c/sub\u003e\u003csup\u003e\u0026minus;\u003c/sup\u003e and NH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e in both rhizospheric and bulk soil suggests a potential decline in relative proportion of microbe-immobilized nitrogen under \u003cem\u003eBt\u003c/em\u003e crop cultivation, indicating a shift in nitrogen competitive interaction between microbes and plants, particularly in the rhizosphere (a geater decline of MBN in rhizosphere compared to the bulk soil, Fig.\u0026nbsp;\u003cspan refid=\"Fig12\" class=\"InternalRef\"\u003e6\u003c/span\u003e, Gannett et al., \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). Combined the significant lower regression slope in the response of microbial biomass N \u003cem\u003evs\u003c/em\u003e C compared with that in 1:1 line, and no significant relative change in response of desoilved organic C and N, it was presumed that a reduction in microbial nitrogen competition occurred under \u003cem\u003eBt\u003c/em\u003e cultivation (Fig.\u0026nbsp;\u003cspan refid=\"Fig11\" class=\"InternalRef\"\u003e5\u003c/span\u003e, Kuzyakov \u0026amp; Xu, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2013\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eIn addition, for \u003cem\u003eBt\u003c/em\u003e crops, the aboveground biomass N:P was decreased by 13.2\u0026ndash;22.3% relative to the parent breed in this study (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05), implying \u003cem\u003eBt\u003c/em\u003e crops absorbed, translocated and utilized more rhizospheric phosphorus compared to their non-\u003cem\u003eBt\u003c/em\u003e counterparts (He et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). The significant reduction of soil total phosphorus (STP) in rhizosphere \u003cem\u003evs\u003c/em\u003e the significant increment in bulk soil under \u003cem\u003eBt\u003c/em\u003e crop cultivation, suggesting \u003cem\u003eBt\u003c/em\u003e crops caused significant limitation of phosphorus resources in rhizosphere but no significant depression in phosphorus returned from crop litter (Veneklaas, \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2022\u003c/span\u003e).\u003c/p\u003e\u003cp\u003e\u003cb\u003eBt\u003c/b\u003e\u003cb\u003e-crop cultivation affected rhizospheric biodiversity\u003c/b\u003e\u003c/p\u003e\u003cp\u003eTrangenic crops, compared to conventional ones, have an advantage in avoiding various biotic disturbances, which are known to significantly affect belowground biotic processes (Zhang et al., \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). As a hotspot of belowground biotic processes, the rhizosphere of \u003cem\u003eBt\u003c/em\u003e crops exhibited a distinct microbial community composition compared to conventional crops (e.g., differences in bacterial and fungal diversity, Figs. S1 and 6, Ahamd et al., \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Under \u003cem\u003eBt\u003c/em\u003e crop cultivation, the positive response of bacterial diversity in rhizosphere was significantly great than that in the bulk soil (Fig. \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e), which also triggered more available nitrogen release from organic matter in rhizosphere through potential increase in bacterial function diversity (Fig.\u0026nbsp;\u003cspan refid=\"Fig10\" class=\"InternalRef\"\u003e4\u003c/span\u003e, van Wyk et al., \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2017\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eIn addition, the diveristy of both nematodes and rhizospheric fungi under \u003cem\u003eBt\u003c/em\u003e crop cultivation was also slightly higher than that in conventional crops, potentially due to the increased organic matter exudation by \u003cem\u003eBt\u003c/em\u003e crop roots (Fig.\u0026nbsp;\u003cspan refid=\"Fig12\" class=\"InternalRef\"\u003e6\u003c/span\u003e, Zeng et al., \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Furthermore, the diversity of both fungi and nematodes under \u003cem\u003eBt\u003c/em\u003e crop cultivation correlated positively with soil respiration, especially for changes in nematode diversity, which explained for 34% of the variation in soil respiration (Fig.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e3\u003c/span\u003e). Soil nematodes with higher diversity under \u003cem\u003eBt\u003c/em\u003e crop cultivation, which might enhance the sequestration and redistribution of carbon and energy from the roots or aboveground litter through the food web, and regulate spatial characteristics of soil organic carbon and soil respiration (Fig.\u0026nbsp;\u003cspan refid=\"Fig12\" class=\"InternalRef\"\u003e6\u003c/span\u003e, Men et al., \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2003\u003c/span\u003e, Young \u0026amp; Unc, \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2023\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eThis study provided a valuable insight into the ecological impacts associated with the cultivation of \u003cem\u003eBt\u003c/em\u003e crops. Specifically, relative to convential agriculture, \u003cem\u003eBt\u003c/em\u003e crops promoted decomposition of soil organic matter and soil respiration, and caused a considerable impact for soil fertility due to the nutrient mining by rhizospheric microbes (Fig.\u0026nbsp;\u003cspan refid=\"Fig12\" class=\"InternalRef\"\u003e6\u003c/span\u003e, Abbas, \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2018\u003c/span\u003e, Ahamd \u003cem\u003eet al.\u003c/em\u003e, 017). However, it is imperative to acknowledge the limitations inherent in our research. Our investigation focused on the theoretical Nash equilibrium dynamics between crops and soil organisms (Blaser \u0026amp; Kirschner, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2007\u003c/span\u003e), with a particular emphasis on the belowground processes. Due to the relative insufficience for simutaneous records of plants and soil organisms, the potential relationship between these belowground variables and underlyine paths cannot be explored on the basis of limited data (Čapek et al., \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). It is essential to focus more on the changes in plant-soil organism interactions triggered by genetically modified crops in the future, providing a better understanding of ecological impacts for such crops and informing sustainable agricultural practices (Cao et al., \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2024\u003c/span\u003e) .\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003e\u003cem\u003eBt\u003c/em\u003e crops, engineered for desired traits, contribute to meeting the needs of a growing global population. This study reveals that \u003cem\u003eBt\u003c/em\u003e crops have a significant advantage in biomass accumulation and carbon sequestration compared to their parent breed, creating a Nash equilibria between crops and soil organisms. The effects of \u003cem\u003eBt-\u003c/em\u003ecrop growth on the stoichiometry of crop-soil-microbe system, as well as the diversity of bacterias, fungi and nematodes, particularly in rhizospheric soil, collectively pormote the stimulated soil respiration. For sustainable agriculture, it is thus crucial to dynamically match soil nutrient supply with crop requirements (\u003cem\u003eBt\u003c/em\u003e crops or parent breed), preventing excessive nutrient mining by microbes from complex soil organic compounds. Therefore, an optimized fertilization practices are necessary to improve rhizospheric nutrient conditions, especially for enhancing life cycle carbon sink potential in transgenic crop cultivation.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003ch2\u003eConflict of Interest\u003c/h2\u003e\u003cp\u003eThe authors declare that they have no conflict of interest.\u003c/p\u003e\u003c/p\u003e\u003ch2\u003eAcknowledgement\u003c/h2\u003e\u003cp\u003eThis research was financially supported by the National Natural Science Foundation of China (Grant No. 32471683, 31600352) and the Anhui Provincial Natural Science Foundation of China (Grant No. 1708085QC53).\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eAbbas MST (2018) Genetically engineered (modified) crops \u003cem\u003e(Bacillus thuringiensis\u003c/em\u003e crops) and the world controversy on their safety. 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Pedosphere 29:114\u0026ndash;122\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"plant-and-soil","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"plso","sideBox":"Learn more about [Plant and Soil](https://www.springer.com/journal/11104)","snPcode":"11104","submissionUrl":"https://submission.nature.com/new-submission/11104/3","title":"Plant and Soil","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Bt crop, soil respiration, rhizosphere, nutrient, ecosystem sustainability","lastPublishedDoi":"10.21203/rs.3.rs-6699296/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6699296/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eAims\u003c/h2\u003e\u003cp\u003eTransgenic \u003cem\u003eBacillus thuringiensis\u003c/em\u003e (\u003cem\u003eBt\u003c/em\u003e) crops are cultivated globally to mitigate potential pest crisis and reduce insecticide dependence. However, the response of belowground system to \u003cem\u003eBt\u003c/em\u003e crop cultivation remains controversial, limiting our understanding of \u003cem\u003eBt\u003c/em\u003e crops\u0026rsquo; role in agriculture sustainability. The aim of this study was to assess \u003cem\u003eBt\u003c/em\u003e crops\u0026rsquo; influence on soil system, including variables of soil respiration, soil carbon and nutrient pools and soil biodiversity.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e\u003cp\u003eWe conducted a global meta-analysis of 88 experimental studies to assess the influence of \u003cem\u003eBt\u003c/em\u003e crop cultivation on soil system.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e\u003cp\u003e\u003cem\u003eBt\u003c/em\u003e crops significantly increased soil respiration by 6.0% (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05), alongside a 14.3% rise in aboveground biomass and a 4.3% belowground plant biomass. \u003cem\u003eBt\u003c/em\u003e crops reduced rhizospheric soil organic carbon (SOC) by 3.4% but increased bulk soil SOC by 3.1%. SOC, nematode diversity, and fungal diversity accounted for 50%, 34%, and 30% of the variation in soil respiration under \u003cem\u003eBt\u003c/em\u003e crop cultivation, respectively. The decrease of microbial competition for nitrogen under \u003cem\u003eBt\u003c/em\u003e crop cultivation caused a significant reduction in microbial biomass carbon: nitrogen ratio.\u003c/p\u003e\u003ch2\u003eConclusions\u003c/h2\u003e\u003cp\u003eUnder \u003cem\u003eBt\u003c/em\u003e crop cultivation, the increased soil respiration might be caused by the stimulated microbe nutrient mining due to plant production-induced system nitrogen deficiency, especially in rhizosphere. Dispite their advantages in biomass accumulation, long-term cultivation of \u003cem\u003eBt\u003c/em\u003e crops may pose ecological risks on soil fertility sustainability and introduce uncertainties in agricultural life cycle assessments.\u003c/p\u003e","manuscriptTitle":"Simultaneous Stimulation of Soil Respiration and Plant Biomass in Transgenic Bacillus thuringiensis Crop Cultivation","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-07-21 08:12:08","doi":"10.21203/rs.3.rs-6699296/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Major revisions","date":"2025-08-25T05:35:16+00:00","index":"","fulltext":""},{"type":"reviewerAgreed","content":"","date":"2025-07-19T04:41:27+00:00","index":0,"fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-07-16T14:53:23+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"Plant and Soil","date":"2025-05-21T22:57:03+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-05-21T11:59:15+00:00","index":"","fulltext":""},{"type":"submitted","content":"Plant and Soil","date":"2025-05-20T09:05:53+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"plant-and-soil","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"plso","sideBox":"Learn more about [Plant and Soil](https://www.springer.com/journal/11104)","snPcode":"11104","submissionUrl":"https://submission.nature.com/new-submission/11104/3","title":"Plant and Soil","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"f9d1162c-ec6e-44cf-afd9-532fe99e5adf","owner":[],"postedDate":"July 21st, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2025-12-22T16:09:24+00:00","versionOfRecord":{"articleIdentity":"rs-6699296","link":"https://doi.org/10.1007/s11104-025-08189-6","journal":{"identity":"plant-and-soil","isVorOnly":false,"title":"Plant and Soil"},"publishedOn":"2025-12-16 15:58:17","publishedOnDateReadable":"December 16th, 2025"},"versionCreatedAt":"2025-07-21 08:12:08","video":"","vorDoi":"10.1007/s11104-025-08189-6","vorDoiUrl":"https://doi.org/10.1007/s11104-025-08189-6","workflowStages":[]},"version":"v1","identity":"rs-6699296","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6699296","identity":"rs-6699296","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}