The BTLA-HVEM axis restricts CAR T cell efficacy in cancer.

The efficacy of T cell-based immunotherapies is limited by immunosuppressive pressures in the tumor microenvironment. Here we show a predominant role for the interaction between BTLA on effector T cells and HVEM (TNFRSF14) on immunosuppressive tumor microenvironment cells, namely regulatory T cells. High BTLA expression in chimeric antigen receptor (CAR) T cells correlated with poor clinical response to treatment. Therefore, we deleted BTLA in CAR T cells and show improved tumor control and persistence in models of lymphoma and solid malignancies. Mechanistically, BTLA inhibits CAR T cells via recruitment of tyrosine phosphatases SHP-1 and SHP-2, upon trans engagement with HVEM. BTLA knockout thus promotes CAR signaling and subsequently enhances effector function. Overall, these data indicate that the BTLA-HVEM axis is a crucial immune checkpoint in CAR T cell immunotherapy and warrants the use of strategies to overcome this barrier.

Alternative splicing of its 5'-UTR limits CD20 mRNA translation and enables resistance to CD20-directed immunotherapies.

Aberrant skipping of coding exons in CD19 and CD22 compromises the response to immunotherapy in B-cell malignancies. Here, we showed that the MS4A1 gene encoding human CD20 also produces several messenger RNA (mRNA) isoforms with distinct 5' untranslated regions. Four variants (V1-4) were detected using RNA sequencing (RNA-seq) at distinct stages of normal B-cell differentiation and B-lymphoid malignancies, with V1 and V3 being the most abundant. During B-cell activation and Epstein-Barr virus infection, redirection of splicing from V1 to V3 coincided with increased CD20 positivity. Similarly, in diffuse large B-cell lymphoma, only V3, but not V1, correlated with CD20 protein levels, suggesting that V1 might be translation-deficient. Indeed, the longer V1 isoform contained upstream open reading frames and a stem-loop structure, which cooperatively inhibited polysome recruitment. By modulating CD20 isoforms with splice-switching morpholino oligomers, we enhanced CD20 expression and anti-CD20 antibody rituximab-mediated cytotoxicity in a panel of B-cell lines. Furthermore, reconstitution of CD20-knockout cells with V3 mRNA led to the recovery of CD20 positivity, whereas V1-reconstituted cells had undetectable levels of CD20 protein. Surprisingly, in vitro CD20-directed chimeric antigen receptor T cells were able to kill both V3- and V1-expressing cells, but the bispecific T-cell engager mosunetuzumab was only effective against V3-expressing cells. To determine whether CD20 splicing is involved in immunotherapy resistance, we performed RNA-seq on 4 postmosunetuzumab follicular lymphoma relapses and discovered that in 2 of them, the downregulation of CD20 was accompanied by a V3-to-V1 shift. Thus, splicing-mediated mechanisms of epitope loss extend to CD20-directed immunotherapies.

Streamlined measurement of chimeric antigen receptor T-cell concentration, size, viability and two-color phenotyping during manufacturing.

BACKGROUND AIMS: The successful development of CD19-targeted chimeric antigen receptor (CAR) T-cell therapies has led to an exponential increase in the number of patients recieving treatment and the advancement of novel CAR T products. Therefore, there is a strong need to develop streamlined platforms that allow rapid, cost-effective, and accurate measurement of the key characteristics of CAR T cells during manufacturing (i.e., cell number, cell size, viability, and basic phenotype). METHODS: In this study, we compared the novel benchtop cell analyzer Moxi GO II (ORFLO Technologies), which enables simultaneous evaluation of all the aforementioned parameters, with current gold standards in the field: the Multisizer Coulter Counter (cell counter) and the BD LSRFortessa (flow cytometer). RESULTS: Our results demonstrated that the Moxi GO II can accurately measure cell number and cell size (i.e., cell volume) while simultaneously assessing simple two-color flow cytometry parameters, such as CAR T-cell viability and CD4 or CAR expression. CONCLUSIONS: These measurements are comparable with those of gold standard instruments, demonstrating that the Moxi GO II is a promising platform for quickly monitoring CAR T-cell growth and phenotype in research-grade and clinical samples.

Summer heatwaves promote harmful algal blooms in the Fuchunjiang Reservoir, an important drinking water source.

Extensive outbreaks of harmful algal blooms (HABs) occurred in the Fuchunjiang Reservoir in 2022, a crucial urban drinking water source, coinciding with extreme summer heatwaves. We hypothesize that these heatwaves contributed to HABs formation and expansion. Leveraging Landsat 8 and Sentinel-2 data, we employed clustering and machine learning methods to quantify the HABs distribution and area. Concurrent meteorological and water quality data aided in uncovering the effects of heatwave on HABs. When applying different methods to extract HABs from remote sensing images, random forest (RF) analyses indicated accuracies of 99.3% and 99.8% for Landsat 8 and Sentinel-2 data, respectively, while classification and regression tree (CART) analyses indicated 99.1% and 99.7% accuracies, respectively. Support vector machine (SVM) exhibited lower accuracies (83.5% and 97.4%). Thus RF, given its smaller differences between satellites and high accuracy, was selected for further analysis. Both satellites detected extensive HABs in 2022, with Sentinel-2 recording a peak area of 24.13 km2 (44.6% of cloud-free water area) on August 11, 2022. Increasing trends with amplified durations were observed for summer heatwaves in Jiande and Tonglu around the Fuchunjiang Reservoir. Notably, these areas experienced extreme heatwaves for 63 and 58 days in 2022, respectively, more than double the 1980-2022 average. From June 1 to October 8, 2022, water temperature peaks significantly coincided with expansive HABs and elevated chlorophyll a (Chl-a) concentration from 4.8 µg/L to 119.2 µg/L during the summer heatwaves. Our findings indicated that the reservoir became more HAB-prone during heatwave events, escalating the drinking water safety risk. These results emphasize the challenges faced by reservoir managers in dealing with climate-induced extreme heatwaves and underscore the urgency for heightened attention from water source management departments.

Safety and efficacy of a novel anti-CD19 chimeric antigen receptor T cell product targeting a membrane-proximal domain of CD19 with fast on- and off-rates against non-Hodgkin lymphoma: a first-in-human study.

BACKGROUND: Commercial anti-CD19 chimeric antigen receptor T-cell therapies (CART19) are efficacious against advanced B-cell non-Hodgkin lymphoma (NHL); however, most patients ultimately relapse. Several mechanisms contribute to this failure, including CD19-negative escape and CAR T dysfunction. All four commercial CART19 products utilize the FMC63 single-chain variable fragment (scFv) specific to a CD19 membrane-distal epitope and characterized by slow association (on) and dissociation (off) rates. We hypothesized that a novel anti-CD19 scFv that engages an alternative CD19 membrane-proximal epitope independent of FMC63 and that is characterized by faster on- and off-rates could mitigate CART19 failure and improve clinical efficacy. METHODS: We developed an autologous CART19 product with 4-1BB co-stimulation using a novel humanized chicken antibody (h1218). This antibody is specific to a membrane-proximal CD19 epitope and harbors faster on/off rates compared to FMC63. We tested h1218-CART19 in vitro and in vivo using FMC63-CART19-resistant models. We conducted a first-in-human multi-center phase I clinical trial to test AT101 (clinical-grade h1218-CART19) in patients with relapsed or refractory (r/r) NHL. RESULTS: Preclinically, h1218- but not FMC63-CART19 were able to effectively eradicate lymphomas expressing CD19 point mutations (L174V and R163L) or co-expressing FMC63-CAR19 as found in patients relapsing after FMC63-CART19. Furthermore, h1218-CART19 exhibited enhanced killing of B-cell malignancies in vitro and in vivo compared with FMC63-CART19. Mechanistically, we found that h1218-CART19 had reduced activation-induced cell death (AICD) and enhanced expansion compared to FMC63-CART19 owing to faster on- and off-rates. Based on these preclinical results, we performed a phase I dose-escalation trial, testing three dose levels (DL) of AT101 (the GMP version of h1218) using a 3 + 3 design. In 12 treated patients (7 DLBCL, 3 FL, 1 MCL, and 1 MZL), AT101 showed a promising safety profile with 8.3% grade 3 CRS (n = 1) and 8.3% grade 4 ICANS (n = 1). In the whole cohort, the overall response rate was 91.7%, with a complete response rate of 75.0%, which improved to 100% in DL-2 and -3. AT101 expansion correlates with CR and B-cell aplasia. CONCLUSIONS: We developed a novel, safe, and potent CART19 product that recognizes a membrane-proximal domain of CD19 with fast on- and off-rates and showed significant efficacy and promising safety in patients with relapsed B-cell NHL. TRIAL REGISTRATION: NCT05338931; Date: 2022-04-01.

Urbanization shifts long-term phenology and severity of phytoplankton blooms in an urban lake through different pathways.

Climate change can induce phytoplankton blooms (PBs) in eutrophic lakes worldwide, and these blooms severely threaten lake ecosystems and human health. However, it is unclear how urbanization and its interaction with climate impact PBs, which has implications for the management of lakes. Here, we used multi-source remote sensing data and integrated the Virtual-Baseline Floating macroAlgae Height (VB-FAH) index and OTSU threshold automatic segmentation algorithm to extract the area of PBs in Lake Dianchi, China, which has been subjected to frequent PBs and rapid urbanization in its vicinity. We further explored long-term (2000-2021) trends in the phenological and severity metrics of PBs and quantified the contributions from urbanization, climate change, and also nutrient levels to these trends. When comparing data from 2011-2021 to 2000-2010, we found significantly advanced initiation of PBs (28.6 days) and noticeably longer duration (51.9 days) but an insignificant trend in time of disappearance. The enhancement of algal nutrient use efficiency, likely induced by increased water temperature and reduced nutrient concentrations, presumably contributed to an earlier initiation and longer duration of PBs, while there was a negative correlation between spring wind speed and the initiation of PBs. Fortunately, we found that both the area of the PBs and the frequency of severe blooms (covering more than 19.8 km2 ) demonstrated downward trends, which could be attributed to increased wind speed and/or reduced nutrient levels. Moreover, the enhanced land surface temperature caused by urbanization altered the thermodynamic characteristics between the land and the lake, which, in turn, possibly caused an increase in local wind speed and water temperature, suggesting that urbanization can differently regulate the phenology and severity of PBs. Our findings have significant implications for the understanding of the impacts of urbanization on PB dynamics and for improving lake management practices to promote sustainable urban development under global change.

Urbanization in developing countries overrides catchment productivity in fueling inland water CO2 emissions.

We compiled a nationwide dataset of carbon dioxide (CO2 ) efflux from 1405 measurements, and found that lakes, reservoirs, and rivers emit a total of 61.9 ± 55.3 TgC as CO2 each year, corresponding to ~6.3% of the annual total national CO2 emission in 2020. Our analysis showed that the presence of anthropogenic disturbances in catchments strongly influences the emission of CO2 from these waters in the non-pristine areas, masking the catchment productivity effect on the emission of CO2 . Our results highlight the need for adjusting climate change models for taking into account anthropogenic effects on CO2 emissions from inland waters.

Hydrological Controls on Dissolved Organic Matter Composition throughout the Aquatic Continuum of the Watershed of Selin Co, the Largest Lake on the Tibetan Plateau.

Alpine river and lake systems on the Tibetan Plateau are highly sensitive indicators and amplifiers of global climate change and important components of the carbon cycle. Dissolved organic matter (DOM) encompasses organic carbon in aquatic systems, yet knowledge about DOM variation throughout the river-lake aquatic continuum within alpine regions is limited. We used optical spectroscopy, ultrahigh-resolution mass spectrometry (Fourier transform ion cyclotron resonance mass spectrometry), and stable water isotopic measurements to evaluate linkages between DOM composition and hydrological connection. We investigated glacial influences on DOM composition throughout the watershed of Selin Co, including upstream glacier-fed rivers and downstream-linked lakes. We found that the dissolved organic carbon concentration increased, whereas specific ultraviolet absorbance (SUVA254) decreased along the river-lake continuum. Relative to rivers, the downstream lakes had low relative abundances of polyphenolic and condensed aromatic compounds and humic-like substances but increased relative abundances of aliphatics and protein-like compounds. SUVA254 decreased while protein-like components increased with enriched stable water isotope δ2H-H2O, indicating that DOM aromaticity declined while autochthonous production increased along the flow paths. Glacier meltwater contributed to elevated relative abundances of aliphatic and protein-like compounds in headwater streams, while increased relative abundances of aromatics and humic-like DOM were found in glacier-fed lakes than downstream lakes. We conclude that changes in hydrological conditions, including glacier melt driven by a warming climate, will significantly alter DOM composition and potentially their biogeochemical function in surface waters on the Tibetan Plateau.

Bloom-induced internal release controlling phosphorus dynamics in large shallow eutrophic Lake Taihu, China.

High phosphorus (P) concentrations are commonly observed in lakes during algal blooms despite massive efforts on external nutrient reduction. However, the knowledge about the relative contribution of internal P loading linked with algal blooms on lake phosphorus (P) dynamics remains limited. To quantify the effect of internal loading on P dynamics, we conducted extensive spatial and multi-frequency nutrient monitoring from 2016 to 2021 in Lake Taihu, a large shallow eutrophic lake in China, and its tributaries (2017-2021). The in-lake P stores (ILSP) and external loading were estimated and then internal P loading was quantified from the mass balance equation. The results showed that the in-lake total P stores (ILSTP) ranged from 398.5 to 1530.2 tons (t), and exhibited a dramatic intra- and inter-annual variability. The annual internal TP loading released from sediment ranged from 1054.3 to 1508.4 t, which was equivalent to 115.6% (TP loading) of the external inputs on average, and responsible for the fluctuations of ILSTP on a weekly scale. High-frequency observations exemplified that ILSTP increased by 136.4% during algal blooms in 2017, while by only 47.2% as a result of external loading after heavy precipitation in 2020. Our study demonstrated that both bloom-induced internal loading and storm-induced external loading are likely to run counter significantly to watershed nutrient reduction efforts in large shallow lakes. More importantly, bloom-induced internal loading is higher than storm-induced external loading over the short term. Given the positive feedback loop between internal P loadings and algal bloom in eutrophic lakes, which explains the significant fluctuation of P concentration while nitrogen concentration decreased. It is emphasized that internal loading and ecosystem restoration are unignorable in shallow lakes, particularly in the algal-dominated region.

Sunlight irradiation promotes both the chemodiversity of terrestrial DOM and the biodiversity of bacterial community in a subalpine lake.

Alpine lake habitats are evolving into subalpine lakes under the scenario of climate change, where the vegetation are promoted due to increasing temperature and precipitation. The abundant terrestrial dissolved organic matter (TDOM) leached from watershed soil into subalpine lakes would undergo strong photochemical reaction due to the high altitude, with the potential to alter DOM composition and affect the bacterial communities. To reveal the transformation of TDOM by both photochemical and microbial processes in a typical subalpine lake, Lake Tiancai (located 200 m below the tree line) was chosen. TDOM was extracted from the surrounding soil of Lake Tiancai and then subjected to the photo/micro-processing for 107 days. The transformation of TDOM was analyzed by Fourier-transform ion cyclotron resonance mass spectrometry (FT-ICR MS) and fluorescence spectroscopy, and the shift of bacterial communities was analyzed using 16s rRNA gene sequencing technology. Dissolved organic carbon and light-absorbing components (a350) decay accounted for approximately 40% and 80% of the original, respectively, in the sunlight process, but both less than 20% in the microbial process for 107 days. The photochemical process promoted the chemodiversity as there were ∼7000 molecules after sunlight irradiation, compared to ∼3000 molecules in the original TDOM. Light promoted the production of highly unsaturated molecules and aliphatics, which were significantly associated with Bacteroidota, suggesting that light may influence bacterial communities by regulating the DOM molecules. Carboxylic-rich alicyclic molecules were generated in both photochemical and biological processes, suggesting TDOM was converted to a stable pool over time. Our finding on the transformation of terrestrial DOM and the alternation of bacterial community under the simultaneously photochemical and microbial processes will help to reveal the response of the carbon cycle and lake system structure to climate change for high-altitude lakes.

Unraveling the Role of Anthropogenic and Natural Drivers in Shaping the Molecular Composition and Biolability of Dissolved Organic Matter in Non-pristine Lakes.

Lakes receive and actively process terrestrial dissolved organic matter (DOM) and play an important role in the global carbon cycle. Urbanization results in elevated inputs of nonpoint-source DOM to headwater streams. Retention of water in lakes allows time for alteration and transformation of the chemical composition of DOM by microbes and UV radiation. Yet, it remains unclear how anthropogenic and natural drivers impact the composition and biolability of DOM in non-pristine lakes. We used optical spectroscopy, Fourier transform ion cyclotron mass spectrometry, stable isotopic measurements, and laboratory bioincubations to investigate the chemical composition and biolability of DOM across two large data sets of lakes associated with a large gradient of urbanization in lowland Eastern China, encompassing a total of 99 lakes. We found that increased urban land use, gross domestic products, and population density in the catchment were associated with an elevated trophic level index, higher chlorophyll-a, higher bacterial abundance, and a higher amount of organic carbon with proportionally higher contribution of aliphatic and peptide-like DOM fractions, which can be highly biolabile. Catchment areas, water depth, lake area: catchment area, gross primary productivity, δ18O-H2O, and bacterial abundance, however, had comparatively little linkage with DOM composition and biolability. Urban land use is currently intensifying in many developing countries, and our results anticipate an increase in the level of biolabile aliphatic DOM from nonpoint sources and accelerated carbon cycling in lake ecosystems in such regions.

Changes in Water Chemistry Associated with Rainstorm Events Increase Carbon Emissions from the Inflowing River Mouth of a Major Drinking Water Reservoir.

Large reservoirs are hotspots for carbon emissions, and the continued input and decomposition of terrestrial dissolved organic matter (DOM) from upstream catchments is an important source of carbon emissions. Rainstorm events can cause a surge in DOM input; however, periodic sampling often fails to fully capture the impact of these discrete rainstorm events on carbon emissions. We conducted a set of frequent observations prior to and following a rainstorm event in a major reservoir Lake Qiandao (China; 580 km2) from June to July 2021 to investigate how rainstorms alter water chemistry and CO2 and CH4 emissions. We found that the mean CO2 efflux (FCO2) (13.2 ± 9.3 mmol m-2 d-1) and CH4 efflux (FCH4) (0.12 ± 0.02 mmol m-2 d-1) in the postrainstorm campaign were significantly higher than those in the prerainstorm campaign (-3.8 ± 3.0 and +0.06 ± 0.02 mmol m-2 d-1, respectively). FCO2 and FCH4 increased with increasing nitrogen and phosphorus levels, elevated DOM absorption (a350), specific UV absorbance SUVA254, and terrestrial humic-like fluorescence. Furthermore, FCO2 and FCH4 decreased with increasing chlorophyll-a (Chl-a), dissolved oxygen (DO), and pH. A five-day laboratory anoxic bioincubation experiment further revealed a depletion of terrestrial-DOM concurrent with increased CO2 and CH4 production. We conclude that rainstorms boost the emission of CO2 and CH4 fueled by the surge and decomposition of fresh terrestrially derived biolabile DOM in this and likely many other reservoir's major inflowing river mouths.

High-resolution temporal detection of cyanobacterial blooms in a deep and oligotrophic lake by high-frequency buoy data.

Cyanobacterial blooms are increasing in magnitude, frequency, and duration worldwide. However, our knowledge of cyanobacterial blooms dynamics and driving mechanisms is still limited due to their high spatiotemporal variability. To determine the potential driving mechanisms of cyanobacterial blooms in oligotrophic lakes, we collected a high-frequency depth profile of chlorophyll fluorescence (ChlF) and synchronous water quality, hydrometeorological data in early spring 2016 in oligotrophic Lake Qiandaohu. The vertical distribution of ChlF exhibited two patterns, "aggregated" and "discrete", using Morisita's index, and the aggregated ChlF presented subsurface chlorophyll maxima during the thermal stratification period. The ChlF concentration was positively correlated with water temperature and negatively correlated with turbidity. Significantly linear relationships were observed between ChlF vertical structure parameters (e.g., Morisita's index, subsurface chlorophyll maxima depth and thickness) and thermal stratification parameters (e.g., mixing layer depth and relative water column stability). After rainstorm floods, the ChlF pattern suddenly change from "aggregated" to "discrete" and a ChlF concentration <1 µg/L was observed for 7-11 days with a significant increase in the mixing depth layer and turbidity. The results suggest that cyanobacterial blooms are robustly associated with thermal stratification and rainstorm floods in the deep and oligotrophic lake. Thermal stratification boosts surface phytoplankton accumulation by increasing water temperature, enhancing light availability and restricting phytoplankton vertical distribution. Rainstorm floods interrupt the accumulation by disrupting thermal stratification and decreasing the available light. Furthermore, wind speed and air temperature both regulate the phytoplankton dynamics by affecting thermal stratification. Our research quantifies the cyanobacterial bloom dynamics and their relationship between environmental factors, improving our knowledge of the driving mechanisms of cyanobacterial bloom for the protection of drinking water safety and aquatic organism health in lakes.

Temporal dependence of chlorophyll a-nutrient relationships in Lake Taihu: Drivers and management implications.

Eutrophication and its associated algal blooms are principal environmental challenges confronting lakes worldwide. The empirical relationships between nutrient (total nitrogen, TN; total phosphorus, TP) and chlorophyll a (Chla) level are widely used as a theoretical basis for lake eutrophication management. Here, seasonal environmental variables and Chla from 2005 to 2020 in Chinese shallow eutrophic Lake Taihu were examined and Chla-nutrient equations in the entire period and annually from 2005 to 2020 were explored using 95% quantile regression model. The results showed robust linear relationships of logChla-logTN and logChla-logTP in the vast majority of cases. Based on Chla-nutrient equations in the entire study period, 0.69 mg/L TN and 52 µg/L TP are recommended as nutrient threshold in Lake Taihu. Furthermore, the results revealed increasing Chla sensitivity to nutrient for each study month (i.e. February, May, August, and November) from 2005 to 2020, whose drivers included increase in water temperature and water level, decrease in wind speed, mass ratio of nitrogen to phosphorus, and grazing effect. It is noteworthy that atmospheric stilling is likely to be the key climatic factor promoting annual peak Chla in Lake Taihu. For one, the deviations of the sub-index of Trophic State Index indicated that light is a critical limiting factor of summer Chla in Lake Taihu. For another, calmer water mainly due to atmospheric stilling decreased near 40% non-algal turbidity and a substantially increased buoyant cyanobacteria during the study period, improving phytoplankton "light niche". Thus, increasing algal sensitivity to nutrient occurred until the additional algal-turbidity induce further light limitations or the exhaustion of TN (or TP) cause nutrient limitation. Given atmospheric stilling is a global phenomenon, this study would affect future algal bloom mitigation efforts in shallow lakes as temperature is always the focus in the recent literatures on global climate change.

Monitoring water transparency, total suspended matter and the beam attenuation coefficient in inland water using innovative ground-based proximal sensing technology.

Long-term and high-frequency observations are vital to reveal water quality dynamics and responses to climate change and human activities. However, the datasets collected from traditional in situ and satellite observations may miss the rapid dynamics of water quality in the short term due to low temporal-spatial monitoring frequency and cloudy or rainy weather. To address this shortage, innovative ground-based proximal sensing (GBPS) technology was proposed to monitor water quality and identify emergencies with a wavelength range of 400-1000 nm, a spectral resolution of 1 nm and a minimal observation interval of 30 s. The GBPS was equipped with a hyperspectral imager placed 4-5 m above the water surface to minimize the impacts of the atmosphere and clouds. In this study, combined with 583 water samples obtained from four field samplings, GBPS datasets were first applied to estimate the total suspended matter (TSM), Secchi disk depth (SDD) and beam attenuation coefficient at 550 nm (C(550)) in Taihu Lake (TL), Liangxi River (LR) and Funchunjiang Reservoir (FR). The results demonstrated good performance with the TSM (R2 = 0.83, RMSE = 8.35 mg/L, MAPE = 24.0%), SDD (R2 = 0.88, RMSE = 0.09 m, MAPE = 14.7%), and C(550) (R2 = 0.79, RMSE = 3.55 m-1, MAPE = 35.8%). The time series of TSM and C(550) at the second-minute level showed consistent changes, but they were opposite to those of SDD. Taking TSM as an example, the datasets captured two mutations in TL with an 853.6% increase in 65 min and a rapid change from 40.3 mg/L to 256.9 mg/L and then to 51.0 mg/L in 224 min on November 1 and 3, respectively. Meanwhile, a significant decreasing trend (r = -0.83, p < 0.01) in LR from November 7 to 9 and a periodic diurnal increasing trend of TSM in FR during November 11 to 13 (0.46 ≤ R2 ≤ 0.70, p < 0.01) were observed. GBPS, with the advantages of high-frequency observations and the applicability of complex weather conditions, compensates for the in situ, aircraft and satellite observation deficiencies. Therefore, GBPS allows us to capture more detailed water quality information and episodic events, which is an important part of an integrated air-space-ground monitoring system in the future.

Elucidating phytoplankton limiting factors in lakes and reservoirs of the Chinese Eastern Plains ecoregion.

Knowledge of phytoplankton limiting factors is essential for cost-efficient lake eutrophication management. Herein, we propose a statistical framework to explore site-specific phytoplankton limiting factors and their dependence on water depth (WD) in 54 lakes in the Chinese Eastern Plains ecoregion. First, the maximal chlorophyll a (Chla) response to total N (TN) or P (TP), representing a region-specific "standard" model where phytoplankton were primarily N- or P-limited, was quantified using a 95% quantile regression. Second, site-specific limiting factors were identified using analogical residual analysis. N- or P-limitation was inferred if FractionTN (i.e. fraction of Chla observed and predicted by the "standard" model for a given TN) > 0.95 or FractionTP >0.95; if both FractionTN and FractionTP <0.95 in a specific environmental condition (e.g. high non-algal turbidity), light limitation was suggested. As a result, 5%, 7%, 4%, 36%, 16%, 2%, and 30% of the sampling sites were limited by N, P, N+P, light availability, rapid flushing, abundant macrophytes, and unmeasured factors, respectively. Bloom control suggestions in the short run are proposed considering these actual limiting factors. Furthermore, the maximal FractionTN or FractionTP response to WD was explored, reflecting the effect of WD on FractionTN (or FractionTP) without significant confounders. The results indicated that phytoplankton in the studied freshwaters would be potentially light-limited, N-limited, N+P-co-limited, or P-limited depending on WD (<1.8, 1.8-2.1, 2.1-5.2, or >5.2 m, respectively), because N will gradually become surplus with increasing WD, while at very shallow depths, strong sediment re-suspension induces light limitation. This finding implies that long-term nutrient management strategies in the studied freshwaters that have WDs of 0-2.1, 2.1-5.2, and >5.2 m can entail control of N, N+P, and P, respectively. This study provides essential information for formulating context-dependent bloom control for lakes in our study area and serves as a valuable reference for developing a cost-efficient eutrophication management framework for other regions.

Computer-designed surgical templates improve the extraction of impacted supernumerary teeth in the hard palate.

OBJECTIVES: The surgical procedure of bony impacted supernumerary teeth (SNT) in hard palate is commonly done with poor visualization and uncomfortable posture. This study aims to introduce our primary practice of presurgical evaluation and guiding exodontia of bony impacted supernumerary teeth (SNT) in the hard palate to reduce surgical trauma, duration and uncertainty. STUDY DESIGN: Twelve patients with impacted supernumerary teeth in hard palate were included. Intraoral scan and the three-dimensional (3D) reconstruction of the cone beam computed tomography (CBCT) file was superimposed, and virtual simulation of flap elevation and osteotomy was conducted on the rebuilt 3D model. A couple of surgical templates were designed with surgical planning software Mimics, fabricated by a 3D printer and were used to guide the extraction of the impacted SNT. RESULTS: The surgical templates fitted well to the teeth and operation site. All the impacted SNTs were accurately located and extracted without damaging the adjacent vital anatomical structures. All patients had an uneventful postoperative recovery without infection or sensory disturbance. CONCLUSIONS: The application of 3D printed surgical templates reduced trauma and increased the accuracy and predictability of surgical extraction of bony impacted SNT in hard palate. The results of this study increased the accuracy and predictability of surgical extraction of bony impacted SNT in hard palate, and reduced the surgeon's embarrassment and surgical trauma because of location difficulty.

Biodegradable dissolved organic carbon shapes bacterial community structures and co-occurrence patterns in large eutrophic Lake Taihu.

Interactions between dissolved organic matter (DOM) and bacteria are central in the biogeochemical cycles of aquatic ecosystems; however, the relative importance of biodegradable dissolved organic carbon (BDOC) compared with other environmental variables in structuring the bacterial communities needs further investigation. Here, we investigated bacterial communities, chromophoric DOM (CDOM) characteristics and physico-chemical parameters as well as examined BDOC via bioassay incubations in large eutrophic Lake Taihu, China, to explore the importance of BDOC for shaping bacterial community structures and co-occurrence patterns. We found that the proportion of BDOC (%BDOC) correlated significantly and positively with the DOC concentration and the index of the contribution of recent produced autochthonous CDOM (BIX). %BDOC, further correlated positively with the relative abundance of the tryptophan-like component and negatively with CDOM aromaticity, indicating that autochthonous production of protein-like CDOM was an important source of BDOC. The richness of the bacterial communities correlated negatively with %BDOC, indicating an enhanced number of species in the refractory DOC environments. %BDOC was identified as a significant stronger factor than DOC in shaping bacterial community composition and the co-occurrence network, suggesting that substrate biodegradability is more significant than DOC quantity determining the bacterial communities in a eutrophic lake. Environmental factors explained a larger proportion of the variation in the conditionally rare and abundant subcommunity than for the abundant and the rare bacterial subcommunities. Our findings emphasize the importance of considering bacteria with different abundance patterns and DOC biodegradability when studying the interactions between DOM and bacteria in eutrophic lakes.

[Rapid screening of single guide RNA targeting pig genome and the harvesting of monoclonal cells by microarray seal].

The emergence of regular short repetitive palindromic sequence clusters (CRISPR) and CRISPR- associated proteins 9 (Cas9) gene editing technology has greatly promoted the wide application of genetically modified pigs. Efficient single guide RNA (sgRNA) is the key to the success of gene editing using CRISPR/Cas9 technology. For large animals with a long reproductive cycle, such as pigs, it is necessary to screen out efficient sgRNA in vitro to avoid wasting time and resource costs before animal experiments. In addition, how to efficiently obtain positive gene editing monoclonal cells is a difficult problem to be solved. In this study, a rapid sgRNA screening method targeting the pig genome was established and we rapidly obtained Fah gene edited cells, laying a foundation for the subsequent production of Fah knockout pigs as human hepatocyte bioreactor. At the same time, the method of obtaining monoclonal cells using pattern microarray culture technology was explored.

Long-term variation of zooplankton communities in a large, heterogenous lake: Implications for future environmental change scenarios.

In recent decades, freshwater ecosystems have been threatened worldwide by multiple simultaneous stressors, including eutrophication, climate change and competing demands for water sources. However, understanding of the long-term variation of zooplankton communities remains limited because long-term observations are lacking. Here, using a long-term (19 year) monitoring dataset, we demonstrate the spatio-temporal variation of zooplankton communities in Lake Taihu, a large, shallow, heterogenous lake in China. With the development of eutrophication, the abundance and biomass of zooplankton first increased from 1998 to 2004, and then exhibited a decreasing trend thereafter. Specifically, the population of rotifer dramatically declined after 2001, while the abundance of copepod and cladoceran showed an increasing trend even though their biomass decreased significantly after 2008. The dominance of small cladocerans (Bosmina coregoni and Ceriodaphnia cornuta) and copepod (Limnothora sinensis) significantly increased with decreasing rotifer density after 2014. Moreover, the zooplankton community structure exhibited heterogenous spatial population dynamics. Cladoceran and rotifer were predominant in cyanobacteria-dominated regions, while a higher proportion of copepod were found in macrophyte-dominated regions. Analyses revealed that zooplankton communities were strongly affected by climate warming and nutrients. These results reinforce previous work demonstrating that the development of eutrophication and climate warming could change the structure of zooplankton community and increase the dominance of small-bodied crustacean. Our findings address the recognized gap in understanding the variation of the zooplankton community in Lake Taihu, and provide an opportunity to evaluate ongoing changes in the zooplankton community related to future environmental change scenarios.