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Environmental Dose MEOHP Promotes Bladder Cancer Progress through Hybrid EMT Mechanism: Based on the Adverse Outcome Pathway. 环境剂量MEOHP通过混合EMT机制促进膀胱癌进展:基于不良结局途径
IF 11.3 1区 环境科学与生态学 Q1 ENGINEERING, ENVIRONMENTAL Pub Date : 2026-02-06 DOI: 10.1021/acs.est.5c08003
Yuwei Wang, Heng Ni, Sitong Dong, Yahui Shang, Zhenyan Cui, Xiaoyu Zhu, Xinxin Liu, Yu Shi, Dajing Xia, Yihua Wu

Di(2-ethylhexyl) phthalate (DEHP) is one of the major plasticizer pollutants, and numerous studies have reported the harmful effects of DEHP on human health. However, the effects of urinary DEHP metabolites on the progression of bladder cancer remain unclear. Here, we aimed to identify the representative chemical and explore its effect and mechanism on bladder cancer progression with epidemiological and experimental methods based on the adverse outcome pathway (AOP). The quantile-based g-computation (QGC) model showed a positive association between urinary plasticizer metabolites and bladder cancer risks in older men (NHANES 2005-2018), with mono(2-ethyl-5-oxohexyl) phthalate (MEOHP, the secondary-metabolite of DEHP) identified as a main driver factor. We treated human bladder cancer cells with MEOHP at environmentally relevant concentrations (10, 100, and 1000 nM) and found that 100 nM MEOHP exposure activated a hybrid EMT (epithelial-mesenchymal transition) phenotype. Mechanically, we confirmed that the environmental dose of MEOHP increased nuclear transposition of YAP and β-catenin (molecular initiating event, MIE), thereby sustaining the hybrid EMT phenotype of bladder cancer cells through a series of key events. Our study first investigated the effects of plasticizer secondary metabolite on bladder cancer progression, highlighting the potential damage to urinary system health caused by the metabolites of environmental chemicals and providing a new perspective for the toxicity assessment of pollutants in the future.

邻苯二甲酸二(2-乙基己基)酯(DEHP)是主要的增塑剂污染物之一,许多研究报道了DEHP对人体健康的有害影响。然而,尿DEHP代谢物对膀胱癌进展的影响尚不清楚。本研究基于不良预后通路(adverse outcome pathway, AOP),采用流行病学和实验方法鉴定具有代表性的化学物质,探讨其在膀胱癌进展中的作用和机制。基于分位数的g计算(QGC)模型显示,老年男性尿液增塑剂代谢物与膀胱癌风险之间存在正相关(NHANES 2005-2018),邻苯二甲酸单(2-乙基-5-氧己基)酯(MEOHP, DEHP的次级代谢物)被确定为主要驱动因素。我们用环境相关浓度(10,100和1000 nM)的MEOHP处理人膀胱癌细胞,发现100 nM MEOHP暴露激活了混合EMT(上皮-间质转化)表型。机械上,我们证实了MEOHP的环境剂量增加了YAP和β-catenin的核转位(分子启动事件,MIE),从而通过一系列关键事件维持膀胱癌细胞的杂交EMT表型。我们的研究首次探讨了增塑剂次生代谢物对膀胱癌进展的影响,强调了环境化学物质代谢物对泌尿系统健康的潜在损害,为未来污染物的毒性评估提供了新的视角。
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引用次数: 0
The Evolution of Constructed Wetlands from Small-Scale Effluent Polishing Solutions to an Essential Water Treatment Technology. 人工湿地从小规模污水处理方案到基本水处理技术的演变。
IF 11.3 1区 环境科学与生态学 Q1 ENGINEERING, ENVIRONMENTAL Pub Date : 2026-02-05 DOI: 10.1021/acs.est.5c14647
David L Sedlak

The desire to protect aquatic ecosystems and drinking water supplies from the adverse effects of nutrients, trace organic contaminants, and other constituents in municipal wastewater has led to a need for additional treatment, particularly in watersheds with insufficient dilution. Constructed wetlands have emerged as viable alternatives to advanced wastewater treatment processes for effluent polishing due to their low cost and ancillary benefits. Starting in the late 1980s, experiences gained from several decades of operating wetlands in small communities increased confidence that these nature-based treatment systems could be effective and reliable. As a result, investments were made in larger constructed wetlands, with surface areas greater than a million square meters (1 Mm2) and flows often exceeding 1 m3 s-1, on effluent-dominated rivers and as part of potable water reuse projects. More recently, new wetland designs have improved the constructed wetland system performance by taking advantage of sunlight-mediated processes in the water column and microbial processes on subsurface porous media. Research that provides additional insight into contaminant removal mechanisms, demonstrates long-term viability, and further improves treatment performance could expand the application of constructed wetlands to other difficult-to-solve water quality challenges, including the treatment of municipal water reuse concentrate and the mitigation of nonpoint source pollution.

为了保护水生生态系统和饮用水供应免受营养物质、微量有机污染物和城市废水中其他成分的不利影响,需要进行额外的处理,特别是在稀释度不足的流域。人工湿地由于其低成本和辅助效益,已成为先进废水处理工艺的可行替代方案。从20世纪80年代末开始,从小型社区几十年的湿地操作中获得的经验增加了人们对这些基于自然的处理系统有效和可靠的信心。因此,在以污水为主的河流上以及作为饮用水再利用项目的一部分,对面积超过100万平方米(1平方毫米)、流量通常超过1立方米s-1的大型人工湿地进行了投资。最近,新的湿地设计通过利用水柱中的阳光介导过程和地下多孔介质上的微生物过程来改善人工湿地系统的性能。研究为污染物去除机制提供了更多的见解,证明了长期可行性,并进一步提高了处理性能,可以将人工湿地的应用范围扩大到其他难以解决的水质挑战,包括处理城市水回用浓缩物和减轻非点源污染。
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引用次数: 0
Uncovering Systemic Dynamics through an Integrated WEFE Nexus Index across 21st Century Futures. 通过综合WEFE Nexus指数揭示21世纪期货的系统动力学。
IF 11.3 1区 环境科学与生态学 Q1 ENGINEERING, ENVIRONMENTAL Pub Date : 2026-02-05 DOI: 10.1021/acs.est.5c11740
Zeynep Özcan, Emre Alp

Achieving sustainability under accelerating climate and socioeconomic pressures requires moving beyond siloed sectoral management toward a system-thinking approach. The water-energy-food-ecosystem (WEFE) Nexus offers a holistic lens, yet most applications remain conceptual, short-term, or treat ecosystems as external constraints. This study operationalizes the WEFE Nexus by embedding ecosystems as a coequal, quantified pillar through a hydrologic-regime-based method, since streamflow is a master variable shaping riverine ecosystem health. Long-term foresight is incorporated via dynamically downscaled climate projections and Shared Socioeconomic Pathways within a coupled water and energy systems (WEAP-LEAP) model. Applied to the semiarid Sakarya Basin in Türkiye, the framework evaluates three future periods (2020-2030, 2055-2065, and 2090-2100) across seven subbasins. Results show systemic trade-offs: municipal water security remains high (>90%), but ecosystem integrity and renewable energy goals are consistently compromised. Overall, WEFE Nexus Index values (0.53-0.86) show significant spatial disparities, with arid upstream regions consistently underperforming. Strikingly, SSP2 (business-as-usual) and SSP5 (fossil-fueled growth) yield nearly identical outcomes, underscoring the systemic unsustainability of current trajectories. This framework advances nexus assessment from theory to practice by integrating reproducible metrics, scenario planning, and spatial modeling, creating a practical tool for developing adaptive and resilient sustainability strategies.

在不断加剧的气候和社会经济压力下实现可持续发展,需要超越孤立的部门管理,转向系统思维方法。水-能量-食物-生态系统(WEFE)联系提供了一个整体视角,然而大多数应用仍然是概念性的、短期的,或者将生态系统视为外部约束。由于河流流量是影响河流生态系统健康的主要变量,因此本研究通过基于水文制度的方法将生态系统嵌入为一个等量的量化支柱,从而实现了WEFE Nexus的运作。通过在水和能源系统耦合模型(weapon - leap)中动态缩小尺度的气候预测和共享的社会经济路径,将长期预见纳入其中。应用于 rkiye省半干旱的Sakarya盆地,该框架评估了七个子盆地的三个未来时期(2020-2030、2055-2065和2090-2100)。结果显示了系统性的权衡:城市水安全仍然很高(bbb90 %),但生态系统完整性和可再生能源目标不断受到损害。总体而言,WEFE Nexus指数(0.53 ~ 0.86)呈现出显著的空间差异,上游干旱地区表现不佳。引人注目的是,SSP2(一切照旧)和SSP5(化石燃料驱动的增长)产生了几乎相同的结果,强调了当前轨迹的系统性不可持续性。该框架通过整合可重复的度量、情景规划和空间建模,将联系评估从理论推进到实践,为制定适应性和弹性可持续发展战略创造了实用工具。
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引用次数: 0
The Globalized Automotive Industries Have Failed to Improve the Overall Carbon Productivity but Exhibit a Polarized Distribution. 全球化的汽车产业并没有提高整体的碳生产率,而是呈现出两极分化的分布。
IF 11.3 1区 环境科学与生态学 Q1 ENGINEERING, ENVIRONMENTAL Pub Date : 2026-02-05 DOI: 10.1021/acs.est.5c12491
Ailin Kang, Xin Tian, Zhifu Mi, Kailan Tian, Yiling Xiong, Andrea Appolloni, Ludi Liu, Xin Sun, Yiqing Huang, Songyan Wang, Pingdan Zhang

Improving carbon productivity is essential for simultaneously achieving sustainable development goals and tackling climate change. While the Global Value Chains (GVCs) division enhances productive efficiency, its impact on carbon productivity remains elusive. Here we integrated the GVCs theory with the Environmental Expanded Input-output model to investigate the highly globalized automotive manufacturing industry. We found that CO2 emissions intensity, the inverse of carbon productivity, fluctuated between 0.37 and 0.47 kg/USD in automotive manufacturing GVCs during 2001-2021. Notably, developing economies nearly doubled their CO2 emissions intensity during this period, whereas developed economies almost halved theirs. The global distribution of CO2 emissions and value added is becoming increasingly unequal in industrial production and service segments. Lower production levels and energy efficiency in developing economies, coupled with their upstream roles in GVCs (raw materials and industrial parts suppliers), exacerbate these disparities. Our findings indicate that merely global labor division is insufficient to create low-carbon automotive manufacturing GVCs. Formulating emission reduction targets that consider the diverse roles of economies within GVCs, and supporting developing economies in boosting energy productivity, labor value added efficiency, and skill can help narrow the distribution gaps and enhance the carbon productivity of the entire automotive manufacturing GVCs.

提高碳生产率对于同时实现可持续发展目标和应对气候变化至关重要。虽然全球价值链(GVCs)部门提高了生产效率,但其对碳生产率的影响仍然难以捉摸。本文将全球价值链理论与环境扩展投入产出模型相结合,对高度全球化的汽车制造业进行了研究。研究发现,2001-2021年期间,汽车制造业全球价值链的二氧化碳排放强度(与碳生产率相反)在0.37 ~ 0.47 kg/USD之间波动。值得注意的是,在此期间,发展中经济体的二氧化碳排放强度几乎翻了一番,而发达经济体的二氧化碳排放强度几乎减半。在工业生产和服务部门,二氧化碳排放和增加值的全球分布正变得越来越不平等。发展中经济体较低的生产水平和能源效率,加上它们在全球价值链中的上游角色(原材料和工业零部件供应商),加剧了这些差距。研究结果表明,仅靠全球分工不足以形成低碳汽车制造业的全球价值链。制定考虑经济体在全球价值链中不同角色的减排目标,支持发展中经济体提高能源生产率、劳动增值效率和技能,有助于缩小分配差距,提高整个汽车制造业全球价值链的碳生产率。
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引用次数: 0
Tailoring Cu+-Ov-Ti Ensembles with Electrophilic O- Species for Enhanced Catalytic Toluene Oxidation. 用亲电O-物种裁剪Cu+- ov - ti系系以增强甲苯的催化氧化。
IF 11.3 1区 环境科学与生态学 Q1 ENGINEERING, ENVIRONMENTAL Pub Date : 2026-02-05 DOI: 10.1021/acs.est.5c14437
Yarong Fang, Shiqi Ma, Zhixin Yu, Liangwei Li, Yiqing Zeng, Jiahao Chen, Shipeng Wan, Ji Yang, Yanbing Guo, Zhaoxiang Zhong

The acceleration of industrialization has driven the increased emission of volatile organic compounds (VOCs), posing significant threats to both the ecological environment and public health. The deficiency of reactive oxygen species fundamentally restricts the low-temperature catalytic toluene combustion in transition-metal oxide catalysts. Herein, we report a strategy for intelligently designing active Cu+-Ov-Ti ensembles by coupling isolated Cu with adjacent oxygen vacancy, which can synergistically activate chemisorbed O2 into reactive superoxide species (O2-). The defective Cu/TiO2-x catalyst exhibited remarkable catalytic performance for toluene oxidation, achieving a T90 of 225 °C, significantly 100 °C lower than that of the pristine Cu/TiO2 catalyst. The low coordination geometry and electron transfer within Cu+-Ov-Ti ensembles synergistically activated O2 to form the Cu-(O-O)ad-Ti bridged superoxide O2- intermediate with an elongated O═O bond. In addition, the distinctive Cu-(O-O)ad-Ti bridging structure with localized electrons facilitated the chemisorbed O2 dissociation into electrophilic monatomic O- species, which subsequently nucleophilically attack the methyl C-H of toluene. These benzyl alcohol-derived Ph-CH2-O- intermediates can be readily and flexibly converted into reactive benzaldehyde and benzoic acid species, which were available for subsequent aromatic ring-opening reactions. This study not only advances mechanistic insights into the Cu+-Ov-Ti ensembles and electrophilic O- species in toluene catalytic oxidation but also establishes a design Cu+-Ov-Ti principle for engineering efficient VOC elimination catalysts.

随着工业化进程的加快,挥发性有机化合物(VOCs)的排放不断增加,对生态环境和公众健康构成重大威胁。活性氧缺乏从根本上制约了过渡金属氧化物催化剂中低温催化甲苯燃烧的发展。在此,我们报告了一种智能设计活性Cu+- ov - ti系综的策略,通过将分离的Cu与邻近的氧空位偶联,可以协同激活化学吸附的O2成为活性超氧化物(O2-)。缺陷Cu/TiO2-x催化剂对甲苯的氧化表现出显著的催化性能,T90为225℃,比原始Cu/TiO2催化剂显著降低100℃。Cu+- ov - ti系综中的低配位几何和电子转移协同激活O2,形成具有延长O = O键的Cu-(O-O)ad-Ti桥接的超氧化物O2-中间体。此外,独特的Cu-(O-O)和ti桥接结构与局部电子促进化学吸收的O2解离成亲电的单原子O-物种,随后亲核攻击甲苯的甲基C-H。这些苯甲醇衍生的Ph-CH2-O中间体可以很容易和灵活地转化为活性苯甲醛和苯甲酸,可用于随后的芳开环反应。本研究不仅对甲苯催化氧化过程中Cu+- ov - ti系群和亲电性O-组分的机理有了深入的了解,而且还建立了Cu+- ov - ti的设计原理,用于设计高效的VOC消除催化剂。
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引用次数: 0
Contrasting Responses of Wind and Solar Energy Potential to Forestation-Induced Climate Feedbacks. 风能和太阳能潜力对森林气候反馈的响应对比。
IF 11.3 1区 环境科学与生态学 Q1 ENGINEERING, ENVIRONMENTAL Pub Date : 2026-02-05 DOI: 10.1021/acs.est.5c12835
Zhaorong Zhu, Pengyi Zhang, Xiaoting Zhong, Yuquan Qu, Mengyang Xu, Mengyu Wang, Binbin Liu, Jiasheng Li, Chao Yue

Forestation and renewable energy sources are both critical tools for climate mitigation and human sustainable development. However, climate feedback from forestation, such as changes in wind speed and downwelling shortwave radiation following forestation driven by modified land-atmosphere heat, momentum, and moisture exchanges, could potentially compromise the availability of weather-dependent renewable energy. Here, based on coupled land-atmosphere simulations with contrasting forest distributions, our results showed that, in idealized forestation scenarios, pixels of forestation remaining suitable for renewable energy deployment are projected to experience a 27.8 ± 1.1% decline in wind energy potential and a 1.9 ± 0.1% reduction in solar energy potential. For the future emission scenario of SSP1-2.6, which is compatible with the goal of the Paris Agreement that limits global warming below 2 °C, forestation-driven reduction (-9.5 ± 1.9%) in wind energy potential for current onshore wind installations can even surpass that caused by climate change (-2.1 ± 1.5%). In contrast, solar energy change for current onshore solar installations in the SSP1-2.6 scenario is projected to be dominated by climate-driven increases (+3.9 ± 0.7%) rather than forestation-induced change (+0.3 ± 0.2%). Our study highlights the need to account for the trade-off between forestation-based carbon removal and renewable-based emission reduction when formulating climate mitigation pathways.

造林和可再生能源都是减缓气候变化和人类可持续发展的重要工具。然而,来自造林的气候反馈,如由陆地大气热量、动量和水分交换改变驱动的造林后的风速和下坡短波辐射的变化,可能会损害依赖天气的可再生能源的可用性。通过对比森林分布的陆地-大气耦合模拟,我们的研究结果表明,在理想的造林情景下,仍然适合可再生能源部署的造林像素预计将经历27.8±1.1%的风能潜力下降和1.9±0.1%的太阳能潜力下降。在SSP1-2.6的未来排放情景中,符合《巴黎协定》将全球变暖限制在2°C以下的目标,目前陆上风电装置的森林驱动的风能潜力减少(-9.5±1.9%)甚至可以超过气候变化导致的风能潜力减少(-2.1±1.5%)。相比之下,在SSP1-2.6情景下,目前陆上太阳能装置的太阳能变化预计将由气候驱动的增加(+3.9±0.7%)而不是由森林引起的变化(+0.3±0.2%)主导。我们的研究强调,在制定气候减缓途径时,需要考虑基于森林的碳去除和基于可再生能源的减排之间的权衡。
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引用次数: 0
Efficient Depolymerization and Low-Toxicity Leaching of Polyester Microplastics through Alkali-Hydrothermal Treatment of Sewage Sludge. 污泥碱水热处理聚酯微塑料的高效解聚和低毒浸出。
IF 11.3 1区 环境科学与生态学 Q1 ENGINEERING, ENVIRONMENTAL Pub Date : 2026-02-05 DOI: 10.1021/acs.est.5c18091
Xingfeng Yang, Shiyu Niu, Xiang Huang, Yulong Niu, Kailiang Shen, Yue Chen, Seinn Lei Aye, Xiaowei Li

Most microplastics (MPs) in wastewater are retained within the sewage sludge. These MPs enter the soil environment through land application, posing a threat to ecosystems. This study proposes an effective control strategy using alkali pretreatment (pH 10, 5 days) followed by hydrothermal treatment at 180 °C (AHT), achieving a degradation rate of 81.83% for polyethylene terephthalate MPs (PET-MPs) in sludge. AHT promotes the formation and solubilization of key active components in sludge, such as alkalinity, which drives nucleophilic attack, metal ions, which catalyze reactions, and organic matter, which acts as radical donors. These components synergistically disintegrate PET-MPs through hydrolysis and radical oxidation pathways during hydrothermal treatment. Meanwhile, hydrothermal treatment induces polymer chain motion and physical structural disruption, accelerating the penetration and reaction of active components, thereby achieving efficient degradation of PET-MPs. Spectral and high-resolution mass spectrometry analyses reveal that sludge AHT facilitates the transformation of MP-derived dissolved organic matter (MP-DOM) into molecules characterized by low-aromaticity, low-molecular-weight, high-saturation, and high-bioavailability. Concurrently, MP-DOM exhibits low acute toxicity toward aquatic organisms and the immortalized human liver cell line (THLE-2 cells). Therefore, sludge AHT effectively degrades and converts polyester MPs into MP-DOM with low-toxicity, thereby mitigating the risks of sludge-based MPs to ecosystems.

废水中的大多数微塑料(MPs)被保留在污水污泥中。这些MPs通过土地利用进入土壤环境,对生态系统构成威胁。本研究提出了一种有效的控制策略,采用碱预处理(pH 10, 5天),然后在180°C (AHT)下进行水热处理,对污泥中的聚对苯二甲酸乙二醇酯MPs (PET-MPs)的降解率达到81.83%。AHT促进污泥中关键活性成分的形成和增溶,如驱动亲核攻击的碱度、催化反应的金属离子和充当自由基供体的有机物。在水热处理过程中,这些成分通过水解和自由基氧化途径协同分解PET-MPs。同时,水热处理引起聚合物链运动和物理结构破坏,加速活性组分的渗透和反应,从而实现PET-MPs的高效降解。光谱和高分辨率质谱分析表明,污泥AHT促进了MP-DOM衍生的溶解有机质(MP-DOM)转化为低芳香性、低分子量、高饱和度和高生物利用度的分子。同时,MP-DOM对水生生物和永生化人肝细胞系(THLE-2细胞)具有低急性毒性。因此,污泥AHT能有效降解聚酯MPs,并将其转化为低毒的MP-DOM,从而降低了污泥基MPs对生态系统的风险。
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引用次数: 0
The Wind Energy Industry Faces Significant Sustainability Challenges. 风能产业面临着重大的可持续性挑战。
IF 11.3 1区 环境科学与生态学 Q1 ENGINEERING, ENVIRONMENTAL Pub Date : 2026-02-05 DOI: 10.1021/acs.est.5c18375
Haonan Wen, Hao Yu, Xuezhi Yang, Qian Liu, Guibin Jiang
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引用次数: 0
Introducing Emission Trading Scheme To Accelerate Low-Carbon Transition in China's Urban Central Heating. 引入排放权交易机制加快中国城市集中供热低碳转型。
IF 11.3 1区 环境科学与生态学 Q1 ENGINEERING, ENVIRONMENTAL Pub Date : 2026-02-05 DOI: 10.1021/acs.est.5c12218
Xiaodan Guo, Bowen Xiao, Ce Jia, Zhuang Liang, Wenjun Meng, Lei Zhu, Pengfei Liu, Shu Tao

Accelerating the low-carbon transition of an urban centralized heating system is a key concern for policymakers. We propose a novel equilibrium analysis framework to assess the feasibility of implementing ETS in China's central heating sector. The results demonstrate that implementing an emission trading scheme (ETS) within the central heating sector can accelerate decarbonization process by shifting the focus of the current Clean Heating Campaign from gas boilers to industrial surplus heat and geothermal energy, aligning with China's goal of reaching 25% nonfossil energy by 2030. It can provide a more cost-effective and sustainable decarbonization pathway in which a 1% annual reduction in carbon quotas leads to emissions peak at 2033, as well as it is expected to generate significant co-benefits by reducing heating-related air pollution. A well-designed subsidy reallocation and phase-out strategy can enable the ETS to drive decarbonization and alleviate fiscal pressures. This combination of ETS and subsidies ensures a smooth transition to low-carbon heating in the short term and is sustainable in the long run through endogenous technological advancement and market self-regulation.

加快城市集中供暖系统的低碳转型是政策制定者关注的一个关键问题。我们提出了一个新的均衡分析框架来评估在中国集中供热行业实施碳排放交易体系的可行性。研究结果表明,通过将当前清洁供暖运动的重点从燃气锅炉转移到工业余热和地热能,在集中供暖部门实施排放交易计划(ETS)可以加速脱碳进程,这与中国到2030年达到25%非化石能源的目标相一致。它可以提供更具成本效益和可持续的脱碳途径,其中每年减少1%的碳配额将导致2033年的排放量达到峰值,并且有望通过减少与供暖相关的空气污染产生显着的协同效益。精心设计的补贴重新分配和逐步淘汰战略可以使碳排放交易体系推动脱碳并减轻财政压力。碳排放交易体系和补贴的结合确保了短期内向低碳供暖的平稳过渡,并通过内生的技术进步和市场自我调节,确保了长期的可持续发展。
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引用次数: 0
Method Development and Sensitivity Optimization Paradigm for One-Step RPA-CRISPR/Cas12a Rapid Detection of the Hard-to-Amplify Gene blaNDM. 难扩增基因blaNDM的一步法快速检测方法开发及灵敏度优化范例
IF 11.3 1区 环境科学与生态学 Q1 ENGINEERING, ENVIRONMENTAL Pub Date : 2026-02-05 DOI: 10.1021/acs.est.5c10280
Lingtao Sun, Zishu Liu, Chifei Dong, Taolve Chi, Jiang Chen, Dongqing Cheng, Lizhong Zhu, Baolan Hu

The presence and spread of antibiotic resistance genes (ARGs) across various habitats have increased the risks of antibiotic resistance, highlighting the urgent need for effective monitoring methods. One key challenge in method development lies in balancing sensitivity, speed, and portability. To address it, a one-step assay targeting the carbapenem resistance gene blaNDM was developed based on recombinase polymerase amplification (RPA) combined with CRISPR/Cas12a. A sensitivity optimization paradigm─MOSAIC (multistrategy optimized sensitive assay via integrated CRISPR/Cas12a)─was proposed, incorporating component optimization, suboptimal-PAM-mediated CRISPR inhibition, and glycerol-assisted phase separation. The glycerol-assisted strategy exhibited the largest enhancement, followed by the suboptimal-PAM strategy and component optimization. When combined, these strategies demonstrated a synergistic effect, yielding greater improvement (10 000-fold) than a single strategy alone. MOSAIC reached a limit of detection (LOD) of 260 copies/μL, comparable to that of qPCR, and enabled faster quantification of blaNDM at 37 °C within 1 h on a standard plate reader. It achieved 100% diagnostic sensitivity and 95.45% specificity in clinical isolates, and 77.41-99.73% accuracy in environmental matrix-spiked samples, comparable to that of qPCR. It provides a technological foundation for on-site detection of blaNDM and offers an optimization paradigm and new insights for the development of one-step RPA-CRISPR/Cas12a assays targeting various genes.

抗生素耐药基因(ARGs)在各种栖息地的存在和传播增加了抗生素耐药的风险,突出表明迫切需要有效的监测方法。方法开发中的一个关键挑战在于平衡灵敏度、速度和可移植性。为了解决这一问题,我们基于重组酶聚合酶扩增(RPA)和CRISPR/Cas12a技术,建立了靶向碳青霉烯类耐药基因blaNDM的一步检测方法。提出了一种灵敏度优化范式─MOSAIC(通过整合CRISPR/Cas12a的多策略优化敏感试验)─,结合组分优化、次优pam介导的CRISPR抑制和甘油辅助相分离。甘油辅助策略表现出最大的增强,其次是次优pam策略和组分优化。当结合使用时,这些策略显示出协同效应,比单独使用策略产生更大的改善(10 000倍)。MOSAIC的检测限(LOD)为260拷贝/μL,与qPCR相当,并且可以在标准平板读取器上在37°C下在1 h内快速定量blaNDM。该方法对临床分离株的诊断灵敏度为100%,特异性为95.45%,对环境基质加标样品的诊断准确度为77.41 ~ 99.73%,与qPCR相当。为blaNDM的现场检测提供了技术基础,为开发针对多种基因的一步法rna - crispr /Cas12a检测提供了优化范例和新见解。
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