Biomimetic polymeric nanoreactors with photooxidation-initiated therapies and reinvigoration of antigen-dependent and antigen-free immunity.

Immune cell-mediated anticancer modalities usually suffer from immune cell exhaustion and limited efficacy in solid tumors. Herein, the oxygen-carrying biomimetic nanoreactors (BNR2(O2)) have been developed with photooxidation-driven therapies and antigen-dependent/antigen-free immune reinvigoration against xenograft tumors. The BNR2(O2) composes polymeric nanoreactors camouflaged with cancer cell membranes can efficiently target homotypic tumors. It continuously releases O2 to boost intracellular reactive oxygen species (ROS) to oxide diselenide bonds, which controllably releases seleninic acids and anti-folate Pemetrexed compared to hydrogen peroxide and glutathione incubation. The O2-rich microenvironment sensitizes Pemetrexed and blocks programmed cell-death ligand 1 (PD-L1) to reverse T cell immunosuppression. The ROS and Pemetrexed upregulate pro-apoptosis proteins and inhibit folate-related enzymes, which cause significant apoptosis and immunogenic cell death to stimulate dendritic cell maturation for improved secretion of cytokines, expanding antigen-dependent T cell immunity. Furthermore, by regulating the release of seleninic acids, the checkpoint receptor human leukocyte antigen E of tumor cells can be blocked to reinvigorate antigen-free natural killer cell immunity. This work offers an advanced antitumor strategy by bridging biomimetic nanoreactors and modulation of multiple immune cells.

Age Dependent Integration of Cortical Progenitors Transplanted at the Adult CSF-Brain Interface.

There has been renewed interest in neural transplantation of cells and tissues for brain repair. Recent studies have demonstrated the ability of transplanted neural precursor cells and in vitro grown organoids to mature and locally integrate into host brain neural circuitry. Much effort has focused on how the transplant behaves and functions after the procedure, but the extent to which the host brain can properly innervate the transplant, particularly in the context of aging, is largely unexplored. Here we report that transplantation of rat embryonic cortical precursor cells into the cerebrospinal fluid-subventricular zone (CSF-SVZ) of adult rat brains generates a brain-like tissue (BLT) at an ectopic site. This model allows for the assessment of long-range connectivity and cellular interactions between the transplant and the host brain as a function of host age. The transplanted precursor cells initially proliferate, then differentiate, and develop into mature BLTs, which receive supportive cellular components from the host including blood vessels, microglia, astrocytes, and oligodendrocytes. There was integration of the BLT into the host brain which occurred at all ages studied, suggesting that host age does not affect the maturation and integration of the transplant-derived BLT. Long-range axonal projections from the BLT into the host brain were robust throughout the different aged recipients. However, long-distance innervation originating from the host brain into the BLT significantly declined with age. This work demonstrates the feasibility and utility of integrating new neural tissue structures at ectopic sites into adult brain circuits to study host-transplant interactions.

Ultrastructural characterization of hippocampal inhibitory synapses under resting and stimulated conditions.

The present study uses electron microscopy to document ultrastructural characteristics of hippocampal GABAergic inhibitory synapses under resting and stimulated conditions in three experimental systems. Synaptic profiles were sampled from stratum pyramidale and radiatum of the CA1 region from (1) perfusion fixed mouse brains, (2) immersion fixed rat organotypic slice cultures, and from (3) rat dissociated hippocampal cultures of mixed cell types. Synapses were stimulated in the brain by a 5 min delay in perfusion fixation to trigger an ischemia-like excitatory condition, and by treating the two culture systems with 90 mM high K+ for 2-3 min to depolarize the neurons. Upon such stimulation conditions, the presynaptic terminals of the inhibitory synapses exhibited similar structural changes to those seen in glutamatergic excitatory synapses, with depletion of synaptic vesicles, increase of clathrin-coated vesicles and appearance of synaptic spinules. However, in contrast to excitatory synapses, no structural differences were detected in the postsynaptic compartment of the inhibitory synapses upon stimulation. There were no changes in the appearance of material associated with the postsynaptic membrane or the length and curvature of the membrane. Also no change was detected in the labeling density of gephyrin, a GABAergic synaptic marker, lining the postsynaptic membrane. Furthermore, virtually all inhibitory synaptic clefts remained rigidly apposed, unlike in the case of excitatory synapses where ~ 20-30% of cleft edges were open upon stimulation, presumably to facilitate the clearance of neurotransmitters from the cleft. The fact that no open clefts were induced in inhibitory synapses upon stimulation suggests that inhibitory input may not need to be toned down under these conditions. On the other hand, similar to excitatory synapse, EGTA (a calcium chelator) induced open clefts in ~ 18% of inhibitory synaptic cleft edges, presumably disrupting similar calcium-dependent trans-synaptic bridges in both types of synapses.

The MYC2 and MYB43 transcription factors cooperate to repress HMA2 and HMA4 expression, altering cadmium tolerance in Arabidopsis thaliana.

Cadmium (Cd) represents a hazardous heavy metal, prevalent in agricultural soil due to industrial and agricultural expansion. Its propensity for being absorbed by edible plants, even at minimal concentrations, and subsequently transferred along the food chain poses significant risks to human health. Accordingly, it is imperative to investigate novel genes and mechanisms that govern Cd tolerance and detoxification in plants. Here, we discovered that the transcription factor MYC2 directly binds to the promoters of HMA2 and HMA4 to repress their expression, thereby altering the distribution of Cd in plant tissues and negatively regulating Cd stress tolerance. Additionally, molecular, biochemical, and genetic analyses revealed that MYC2 interacts and cooperates with MYB43 to negatively regulate the expression of HMA2 and HMA4 and Cd stress tolerance. Notably, under Cd stress conditions, MYC2 undergoes degradation, thereby alleviating its inhibitory effect on HMA2 and HMA4 expression and plant tolerance to Cd stress. Thus, our study highlights the dynamic regulatory role of MYC2, in concert with MYB43, in regulating the expression of HMA2 and HMA4 under both normal and Cd stress conditions. These findings present MYC2 as a promising target for directed breeding efforts aimed at mitigating Cd accumulation in edible plant roots.

End-Member Mixing Model for Methane Carbon Isotope Fractionation During Shale Gas Desorption and Its Application.

Unlike conventional natural gas reservoirs, shale gas development involves systematic changes in methane carbon isotopes that cannot be effectively described by existing isotope fractionation models and mechanisms. Therefore, based on fundamental theories such as Rayleigh fractionation, mass transfer flow, and mass conservation, this study established isotopic fractionation equations for methane in adsorbed and free gas. By considering adsorbed and free gases as two end-members and using an isotope mixing model, a fractionation model for methane carbon isotopes during shale gas desorption was constructed. This model quantifies the isotopic fractionation effects during shale gas desorption and elucidates the mechanism of methane carbon isotope fractionation. Using on-site desorbed gas content and isotope data, parameter fitting and model calculations were conducted to characterize methane carbon isotope variations throughout the process of shale core field desorption. The results show a pattern of "initially negative and then turning positive," consistent with those of physical simulation experiments. It was clarified that differences in mixing the two end-members and isotopic fractionation play key roles in the variation of methane carbon isotopic composition in shale gas. By applying the methane carbon isotope fractionation model, the contribution of adsorbed gas during shale gas production was explored. It was found that in the early stage of development, the adsorbed gas in Well JY 1 was negligible. After nearly seven years of development, the contribution of adsorbed gas in the later stage has only reached nearly 15%, indicating that the production contribution of adsorbed gas is still less than 0.3 million cubic feet per day. The open flow of Well JY 6-2 is more conducive to the production of adsorbed gas, but the production capacity is still mainly contributed by free gas, indicating that the shale gas production capacity in the later stage in the Jiaoshiba gas field is still primarily dominated by free gas.

Proton therapy for breast cancer: Reducing toxicity.

Radiotherapy is a crucial component in the holistic management of breast cancer, with approximately 60% of individuals diagnosed with breast cancer requiring this treatment. As the survival rate of individuals with breast cancer has significantly increased, there is a growing focus on the long-term well-being of patients. Proton therapy (PT) is a new and rapidly developing radiotherapy method. In comparison with conventional photon therapy, PT offers the benefits of decreased radiation toxicity and increased dosage in the designated region. This can extend patients' lifespan and enhance their overall well-being. The present analysis examines the function of PT in diminishing the harmful effects of radiation in cases of breast cancer, while also providing a brief overview of the future potential and obstacles associated with PT for breast cancer.

Enhanced Energy Storage Properties of Highly Polarized BMT-Based Thin Films through the Multiscale Structure Synergistic Regulation Strategy.

For solving the trade-off relationship of the polarization and breakdown electric field, ferroelectric films with high polarization are playing a critical role in energy storage capacitor applications, especially at moderate/low electric fields. In this work, we propose a multiscale structure (including defect, domain, and grain structures) synergetic optimization strategy to optimize the polarization behavior and energy storage performances of BiMg0.5Ti0.5O3 (BMT) ferroelectric films by introducing Sr0.7La0.2TiO3 (SLT) without compromising the breakdown strength. At a moderate electric field of 2917 kV/cm, a high discharge density of 72.2 J/cm3 has been achieved in 0.9BMT-0.1SLT films, together with good frequency, thermal, and cycle stabilities for energy storage. Importantly, the phase difference Δφ is utilized to quantitatively evaluate the polarization switching mobility of the ferroelectric domain/PNRs at an external electric field stimulation. This research demonstrates that a multiscale structure optimization strategy could effectively regulate the energy storage performance, and ecofriendly BMT-based materials are promising candidates for next-generation energy storage capacitors, especially at moderate/low electric fields.

CDR1, a DUF946 domain containing protein, positively regulates cadmium tolerance in Arabidopsis thaliana by maintaining the stability of OPT3 protein.

Industrial and agricultural production processes lead to the accumulation of cadmium (Cd) in soil, resulting in crops absorb Cd from contaminated soil and then transfer it to human body through the food chain, posing a serious threat to human health. Thus, it is necessary to explore novel genes and mechanisms involved in regulating Cd tolerance and detoxification in plants. Here, we found that CDR1, a DUF946 domain containing protein, localizes to the plasma membrane and positively regulates Cd stress tolerance. The cdr1 mutants exhibited Cd sensitivity, accumulated excessive Cd in the seeds and roots, but decreased in leaves. However, CDR1-OE transgenic plants not only showed Cd tolerance but also significantly reduced Cd in seeds and roots. Additionally, both in vitro and in vivo assays demonstrated an interaction between CDR1 and OPT3. Cell free protein degradation and OPT3 protein level determination assays indicated that CDR1 could maintain the stability of OPT3 protein. Moreover, genetic phenotype analysis and Cd content determination showed that CDR1 regulates Cd stress tolerance and affect the distribution of Cd in plants by maintaining the stability of OPT3 protein. Our discoveries provide a key candidate gene for directional breeding to reduce Cd accumulation in edible seeds of crops.

Polymorphic Localized Heterostructure Design for High-Performance Amorphous/Nanocrystalline Composite Film.

Analogous to linear dielectric, amorphous perovskite dielectrics characterized of high breakdown strength and low remanent polarization possess in-depth application in the sea, land, and air fields. Amorphous engineering is a common approach to balance the inverse relationship between polarization and breakdown strength in dielectric ceramic capacitor, however, the low polarization is the major barrier limiting the improvement of energy storage density. To address this concern, the polymorphic localized heterostructure confirmed by high-resolution transmission electron microscope (HR-TEM) and HADDF images is constructed in BaTiO3-Bi(Ni0.5Zr0.5)O3 amorphous/nanocrystalline composite film with SiO2 addition (BT-BNZ-xS, x = 3, 5, 7, 10 mol%). The stability of nanocrystalline region achieved by Si-rich transition region and the enhancive ultra-short-range ordering in the amorphous region synergistically result in large breakdown strength and nonhysteretic polarized response. This polymorphic localized heterostructure optimizes the thermal stability in a wide temperature range and contributes ultrahigh energy storage density of 149.9 J cm-3 with markedly enhanced efficiency of 79.0%. This study provides a universal strategy to design the polarization behavior in other amorphous perovskite-based dielectrics.

YZL-51N functions as a selective inhibitor of SIRT7 by NAD+ competition to impede DNA damage repair.

The NAD+-dependent deacetylase SIRT7 is a pivotal regulator of DNA damage response (DDR) and a promising drug target for developing cancer therapeutics. However, limited progress has been made in SIRT7 modulator discovery. Here, we applied peptide-based deacetylase platforms for SIRT7 enzymatic evaluation and successfully identified a potent SIRT7 inhibitor YZL-51N. We initially isolated bioactive YZL-51N from cockroach (Periplaneta americana) extracts and then developed the de novo synthesis of this compound. Further investigation revealed that YZL-51N impaired SIRT7 enzymatic activities through occupation of the NAD+ binding pocket. YZL-51N attenuated DNA damage repair induced by ionizing radiation (IR) in colorectal cancer cells and exhibited a synergistic anticancer effect when used in combination with etoposide. Overall, our study not only identified YZL-51N as a selective SIRT7 inhibitor from insect resources, but also confirmed its potential use in combined chemo-radiotherapy by interfering in the DNA damage repair process.

Deep Learning-Based Automated Imaging Classification of ADPKD.

Introduction: The Mayo imaging classification model (MICM) requires a prestep qualitative assessment to determine whether a patient is in class 1 (typical) or class 2 (atypical), where patients assigned to class 2 are excluded from the MICM application. Methods: We developed a deep learning-based method to automatically classify class 1 and 2 from magnetic resonance (MR) images and provide classification confidence utilizing abdominal T 2 -weighted MR images from 486 subjects, where transfer learning was applied. In addition, the explainable artificial intelligence (XAI) method was illustrated to enhance the explainability of the automated classification results. For performance evaluations, confusion matrices were generated, and receiver operating characteristic curves were drawn to measure the area under the curve. Results: The proposed method showed excellent performance for the classification of class 1 (97.7%) and 2 (100%), where the combined test accuracy was 98.01%. The precision and recall for predicting class 1 were 1.00 and 0.98, respectively, with F 1 -score of 0.99; whereas those for predicting class 2 were 0.87 and 1.00, respectively, with F 1 -score of 0.93. The weighted averages of precision and recall were 0.98 and 0.98, respectively, showing the classification confidence scores whereas the XAI method well-highlighted contributing regions for the classification. Conclusion: The proposed automated method can classify class 1 and 2 cases as accurately as the level of a human expert. This method may be a useful tool to facilitate clinical trials investigating different types of kidney morphology and for clinical management of patients with autosomal dominant polycystic kidney disease (ADPKD).

Novel prognostic impact and cell specific role of endocan in patients with coronary artery disease.

BACKGROUND: Both the clinical and mechanistic impacts of endocan were not well elucidated especially in coronary artery disease (CAD). OBJECTIVE: This study aimed to investigate the prognostic and potential pathological role of endocan for cardiovascular (CV) events in stable CAD patients. METHODS: A total of 1,071 stable CAD patients with previous percutaneous coronary intervention (PCI) were enrolled prospectively in a nationwide Biosignature study. Another cohort of 76 CAD patients with or without PCI were enrolled for validation. Baseline biomarkers including endocan level was measured and total CV events especially hard CV events (including CV mortality, non-fatal myocardial infection and stroke) during follow-up were identified. Circulating endothelial progenitor cells (EPCs) as an in vivo biological contributor to vascular repairment from CAD patients were used for the in vitro functional study. RESULTS: After 24 months, there were 42 patients (3.92%) with hard CV events and 207 (19.3%) with total CV events in the study group. The incidence of both events was increased with the tertiles of baseline endocan level (hard events: 1.7%,3.4%, and 6.7% in 1st,2nd, and 3rd tertile respectively, p = 0.002; total events: 13.8%vs.16.2%vs.28.0%, p < 0.0001). Multivariate regression analysis revealed the independent association of endocan level with total and hard CV events. These findings were validated in another cohort with a 5-year follow-up. Furthermore, in vitro inhibition of endocan improved cell migration and tube formation capacities, and reduced cell adhesiveness of EPCs from CAD patients. CONCLUSIONS: Endocan might be a novel prognostic indicator, mechanistic mediator, and potential therapeutic target for clinical CAD.

Novel artesunate and isatin hybrid CT3-1 suppresses collagen-induced arthritis through abrogating dendritic cell chemotaxis-induced by CCR5.

BACKGROUND: Chemotaxis and trafficking of dendritic cells (DCs) induced by cytokine receptors are crucial steps in rheumatoid arthritis (RA) pathogenesis. C-C chemokine receptor type 5 (CCR5) plays a key role in DC movement and has been implicated in multitudinous inflammatory and immunology diseases. Thus, targeting CCR5 to suppress DC chemotaxis is considered as a potential strategy for the management of RA. METHODS: Herein, we first synthesized a new hybrid named CT3-1 which based on artesunate and isatin. Besides, we studied the regulating effectiveness of CT3-1 on bone marrow-derived DCs (BMDCs) and on collagen-induced arthritis (CIA) through RNA-seq analysis, cell function experiments in vitro and mice model in vivo. RESULTS: The results shown that CT3-1 mainly reduced CCR5 expression of immature BMDCs and importantly inhibited immature BMDC migration induced by CCR5 in vitro, with no or minor influence on other functions of DCs, such as phagocytosis and maturation. In the mouse model, CT3-1 relieved arthritis severity and inhibited CIA development. Furthermore, CT3-1 intervention decreased the expression of CCR5 in DCs and reduced the proportion of DCs in the peripheral blood of CIA mice. CONCLUSIONS: Our findings suggest that CCR5-induced chemotaxis and trafficking of immature DCs are important in RA. Targeting CCR5 and inhibiting immature DC chemotaxis may provide a novel choice for the treatment of RA and other similar autoimmune diseases. Moreover, we synthesized a new hybrid compound CT3-1 that could inhibit immature DC trafficking and effectively relieve RA by directly reducing the CCR5 expression of immature DCs.

Structurally Diverse Alkaloids with Anti-Renal-Fibrosis Activity from the Centipede Scolopendra subspinipes mutilans.

Twelve new alkaloids, scolopenolines A-L (1-7, 9-11, 13, 14), along with six known analogues, were isolated from Scolopendra subspinipes mutilans, identified by analysis of spectroscopic data and quantum chemical and computational methods. Scolopenoline A (1), a unique guanidyl-containing C14 quinoline alkaloid, features a 6/6/5 ring backbone. Scolopenoline B (2) is a novel sulfonyl-containing heterodimer comprising quinoline and tyramine moieties. Scolopenoline G (7) presents a rare C12 quinoline skeleton with a 6/6/5 ring system. Alkaloids 1, 8, 10, and 15-18 display anti-inflammatory activity, while 10 and 16-18 also exhibit anti-renal-fibrosis activity. Drug affinity responsive target stability and RNA-interference assays show that Lamp2 might be a potentially important target protein of 16 for anti-renal-fibrosis activity.

Mechanically Enhanced Detoxification of Chemical Warfare Agent Simulants by a Two-Dimensional Piezoresponsive Metal-Organic Framework.

Chemical warfare agents (CWAs) refer to toxic chemical substances used in warfare. Recently, CWAs have been a critical threat for public safety due to their high toxicity. Metal-organic frameworks have exhibited great potential in protecting against CWAs due to their high crystallinity, stable structure, large specific surface area, high porosity, and adjustable structure. However, the metal clusters of most reported MOFs might be highly consumed when applied in CWA hydrolysis. Herein, we fabricated a two-dimensional piezoresponsive UiO-66-F4 and subjected it to CWA simulant dimethyl-4-nitrophenyl phosphate (DMNP) detoxification under sonic conditions. The results show that sonication can effectively enhance the removal performance under optimal conditions; the reaction rate constant k was upgraded 45% by sonication. Moreover, the first-principle calculation revealed that the band gap could be further widened with the application of mechanical stress, which was beneficial for the generation of 1O2, thus further upgrading the detoxification performance toward DMNP. This work demonstrated that mechanical vibration could be introduced to CWA protection, but promising applications are rarely reported.

Zinc peroxide-based nanotheranostic platform with endogenous hydrogen peroxide/oxygen generation for enhanced photodynamic-chemo therapy of tumors.

Nanotheranostic platforms, which can respond to tumor microenvironments (TME, such as low pH and hypoxia), are immensely appealing for photodynamic therapy (PDT). However, hypoxia in solid tumors harms the treatment outcome of PDT which depends on oxygen molecules to generate cytotoxic singlet oxygen (1O2). Herein, we report the design of TME-responsive smart nanotheranostic platform (DOX/ZnO2@Zr-Ce6/Pt/PEG) which can generate endogenously hydrogen peroxide (H2O2) and oxygen (O2) to alleviate hypoxia for improving photodynamic-chemo combination therapy of tumors. DOX/ZnO2@Zr-Ce6/Pt/PEG nanocomposite was prepared by the synthesis of ZnO2 nanoparticles, in-situ assembly of Zr-Ce6 as typical metal-organic framework (MOF) on ZnO2 surface, in-situ reduction of Pt nanozymes, amphiphilic lipids surface coating and then doxorubicin (DOX) loading. DOX/ZnO2@Zr-Ce6/Pt/PEG nanocomposite exhibits average sizes of ∼78 nm and possesses a good loading capacity (48.8 %) for DOX. When DOX/ZnO2@Zr-Ce6/Pt/PEG dispersions are intratumorally injected into mice, the weak acidic TEM induces the decomposition of ZnO2 core to generate endogenously H2O2, then Pt nanozymes catalyze H2O2 to produce O2 for alleviating tumor hypoxia. Upon laser (630 nm) irradiation, the Zr-Ce6 component in DOX/ZnO2@Zr-Ce6/Pt/PEG can produce cytotoxic 1O2, and 1O2 generation rate can be enhanced by 2.94 times due to the cascaded generation of endogenous H2O2/O2. Furthermore, the generated O2 can suppress the expression of hypoxia-inducible factor α, and further enable tumor cells to become more sensitive to chemotherapy, thereby leading to an increased effectiveness of chemotherapy treatment. The photodynamic-chemo combination therapy from DOX/ZnO2@Zr-Ce6/Pt/PEG nanoplatform exhibits remarkable tumor growth inhibition compared to chemotherapy or PDT. Thus, the present study is a good demonstration of a TME-responsive nanoplatform in a multimodal approach for cancer therapy.

Correction and Removal of Expression of Concern: Highly stereoselective gram scale synthesis of all the four diastereoisomers of Boc-protected 4-methylproline carboxylates.

[This corrects the article DOI: 10.1039/C9RA06827A.].

Correction and removal of expression of concern: Total synthesis of tubulysin U and N14-desacetoxytubulysin H.

Correction and removal of expression of concern for 'Total synthesis of tubulysin U and N14-desacetoxytubulysin H' by Bohua Long et al., Org. Biomol. Chem., 2020, 18, 5349-5353, https://doi.org/10.1039/D0OB01109F.

A novel bioprospecting strategy via 13C-based high-throughput probing of active methylotrophs inhabiting oil reservoir surface soil.

Methane-oxidizing bacteria (MOB) have long been considered as a microbial indicator for oil and gas prospecting. However, due to the phylogenetically narrow breath of ecophysiologically distinct MOB, classic culture-dependent approaches could not discriminate MOB population at fine resolution, and accurately reflect the abundance of active MOB in the soil above oil and gas reservoirs. Here, we presented a novel microbial anomaly detection (MAD) strategy to quantitatively identify specific indicator methylotrophs in the surface soils for bioprospecting oil and gas reservoirs by using a combination of 13C-DNA stable isotope probing (SIP), high-throughput sequencing (HTS), quantitative PCR (qPCR) and geostatistical analysis. The Chunguang oilfield of the Junggar Basin was selected as a model system in western China, and type I methanotrophic Methylobacter was most active in the topsoil above the productive oil wells, while type II methanotrophic Methylosinus predominated in the dry well soils, exhibiting clear differences between non- and oil reservoir soils. Similar results were observed by quantification of Methylobacter pmoA genes as a specific bioindicator for the prediction of unknown reservoirs by grid sampling. A microbial anomaly distribution map based on geostatistical analysis further showed that the anomalous zones were highly consistent with petroleum, geological and seismic data, and validated by subsequent drilling. Over seven years, a total of 24 wells have been designed and drilled into the targeted anomaly, and the success rate via the MAD prospecting strategy was 83 %. Our results suggested that molecular techniques are powerful tools for oil and gas prospecting. This study indicates that the exploration efficiency could be significantly improved by integrating multi-disciplinary information in geophysics and geomicrobiology while reducing the drilling risk to a greater extent.

Identifying Active Rather than Total Methanotrophs Inhabiting Surface Soil Is Essential for the Microbial Prospection of Gas Reservoirs.

Methane-oxidizing bacteria (MOB) have long been recognized as an important bioindicator for oil and gas exploration. However, due to their physiological and ecological diversity, the distribution of MOB in different habitats varies widely, making it challenging to authentically reflect the abundance of active MOB in the soil above oil and gas reservoirs using conventional methods. Here, we selected the Puguang gas field of the Sichuan Basin in Southwest China as a model system to study the ecological characteristics of methanotrophs using culture-independent molecular techniques. Initially, by comparing the abundance of the pmoA genes determined by quantitative PCR (qPCR), no significant difference was found between gas well and non-gas well soils, indicating that the abundance of total MOB may not necessarily reflect the distribution of the underlying gas reservoirs. 13C-DNA stable isotope probing (DNA-SIP) in combination with high-throughput sequencing (HTS) furthermore revealed that type II methanotrophic Methylocystis was the absolutely predominant active MOB in the non-gas-field soils, whereas the niche vacated by Methylocystis was gradually filled with type I RPC-2 (rice paddy cluster-2) and Methylosarcina in the surface soils of gas reservoirs after geoscale acclimation to trace- and continuous-methane supply. The sum of the relative abundance of RPC-2 and Methylosarcina was then used as specific biotic index (BI) in the Puguang gas field. A microbial anomaly distribution map based on the BI values showed that the anomalous zones were highly consistent with geological and geophysical data, and known drilling results. Therefore, the active but not total methanotrophs successfully reflected the microseepage intensity of the underlying active hydrocarbon system, and can be used as an essential quantitative index to determine the existence and distribution of reservoirs. Our results suggest that molecular microbial techniques are powerful tools for oil and gas prospecting.