Endogenous AND Logic DNA Nanomachine for Highly Specific Cancer Cell Imaging.

Intracellular cancer-related biomarker imaging strategy has been used for specific identification of cancer cells, which was of great importance to accurate cancer clinical diagnosis and prognosis studies. Localized DNA circuits with improved sensitivity showed great potential for intracellular biomarkers imaging. However, the ability of localized DNA circuits to specifically image cancer cells is limited by off-site signal leakage associated with a single-biomarker sensing strategy. Herein, we integrated the endogenous enzyme-powered strategy with logic-responsive and localized signal amplifying capability to construct a self-assembled endogenously AND logic DNA nanomachine (EDN) for highly specific cancer cell imaging. When the EDN encountered a cancer cell, the overexpressed DNA repairing enzyme apurinic/apyrimidinic endonuclease 1 (APE1) and miR-21 could synergistically activate a DNA circuit via cascaded localized toehold-mediated strand displacement (TMSD) reactions, resulting in amplified fluorescence resonance energy transfer (FRET) signal. In this strategy, both endogenous APE1 and miR-21, served as two "keys" to activate the AND logic operation in cancer cells to reduce off-tumor signal leakage. Such a multiplied molecular recognition/activation nanomachine as a powerful toolbox realized specific capture and reliable imaging of biomolecules in living cancer cells.

Logic Signal Amplification System for Sensitive Electrochemiluminescence Detection and Subtype Identification of Cancer Cells.

Achieving sensitive detection and accurate identification of cancer cells is vital for diagnosing and treating the disease. Here, we developed a logic signal amplification system using DNA tetrahedron-mediated three-dimensional (3D) DNA nanonetworks for sensitive electrochemiluminescence (ECL) detection and subtype identification of cancer cells. Specially designed hairpins were integrated into DNA tetrahedral nanostructures (DTNs) to perform a catalytic hairpin assembly (CHA) reaction in the presence of target microRNA, forming hyperbranched 3D nanonetworks. Benefiting from the "spatial confinement effect," the DNA tetrahedron-mediated catalytic hairpin assembly (DTCHA) reaction displayed significantly faster kinetics and greater cycle conversion efficiency than traditional CHA. The resulting 3D nanonetworks could load a large amount of Ru(phen)32+, significantly enhancing its ECL signal, and exhibit detection limits for both miR-21 and miR-141 at the femtomolar level. The biosensor based on modular logic gates facilitated the distinction and quantification of cancer cells and normal cells based on miR-21 levels, combined with miR-141 levels, to further identify different subtypes of breast cancer cells. Overall, this study provides potential applications in miRNA-related clinical diagnostics.

Fucoidan from Ascophyllum nodosum and Undaria pinnatifida attenuate SARS-CoV-2 infection in vitro and in vivo by suppressing ACE2 and alleviating inflammation.

The global healthcare challenge posed by COVID-19 necessitates the continuous exploration for novel antiviral agents. Fucoidans have demonstrated antiviral activity. However, the underlying structure-activity mechanism responsible for the inhibitory activity of fucoidans from Ascophyllum nodosum (FUCA) and Undaria pinnatifida (FUCU) against SARS-CoV-2 remains unclear. FUCA was characterized as a homopolymer with a backbone structure of repeating (1 â†’ 3) and (1 â†’ 4) linked α-l-fucopyranose residues, whereas FUCU was a heteropolysaccharide composed of Fuc1-3Gal1-6 repeats. Furthermore, FUCA demonstrated significantly higher anti-SARS-CoV-2 activity than FUCU (EC50: 48.66 vs 69.52 µg/mL), suggesting the degree of branching rather than sulfate content affected the antiviral activity. Additionally, FUCA exhibited a dose-dependent inhibitory effect on ACE2, surpassing the inhibitory activity of FUCU. In vitro, both FUCA and FUCU treatments downregulated the expression of pro-inflammatory cytokines (IL-6, IFN-α, IFN-γ, and TNF-α) and anti-inflammatory cytokines (IL-10 and IFN-ß) induced by viral infection. In hamsters, FUCA demonstrated greater effectiveness in attenuating lung and gastrointestinal injury and reducing ACE2 expression, compared to FUCU. Analysis of the 16S rRNA gene sequencing revealed that only FUCU partially alleviated the gut microbiota dysbiosis caused by SARS-CoV-2. Consequently, our study provides a scientific basis for considering fucoidans as poteintial prophylactic food components against SARS-CoV-2.

Simulation-Assisted DNA Nanodevice Serve as a General Optical Platform for Multiplexed Analysis of Micrornas.

Small frame nucleic acids (FNAs) serve as excellent carrier materials for various functional nucleic acid molecules, showcasing extensive potential applications in biomedicine development. The carrier module and function module combination is crucial for probe design, where an improper combination can significantly impede the functionality of sensing platforms. This study explores the effect of various combinations on the sensing performance of nanodevices through simulations and experimental approaches. Variances in response velocities, sensitivities, and cell uptake efficiencies across different structures are observed. Factors such as the number of functional molecules loaded, loading positions, and intermodular distances affect the rigidity and stability of the nanostructure. The findings reveal that the structures with full loads and moderate distances between modules have the lowest potential energy. Based on these insights, a multisignal detection platform that offers optimal sensitivity and response speed is developed. This research offers valuable insights for designing FNAs-based probes and presents a streamlined method for the conceptualization and optimization of DNA nanodevices.

DNAzyme-Mediated Cascade Nanoreactor for Cuproptosis-Promoted Pancreatic Cancer Synergistic Therapy.

Cuproptosis, a kind of newly recognized cell death modality, shows enormous prospect in cancer treatment. The inducer of cuproptosis has more advantages in tumor therapy, especially that can trigger cuproptosis and chemodynamic therapy (CDT) simultaneously. However, cuproptosis is restricted to the deficiency of intracellular copper ions and the nonspecific delivery of copper-based ionophores. Therefore, high level delivery, responsive release, and utilizing synergistic-function of inducer become the key on cuproptosis-based oncotherapy. In this work, a cascade nanosystem is constructed for enhanced cuproptosis and CDT. In the weak acidic environment of tumor cells, DNA, zinc ions, and Cu+ can release from the nanosystem. Since Cu+ having superior performance in mediating both Fenton-like reaction and cuproptosis, the released Cu+ induces cuproptosis and CDT efficiently, accompanied by Cu2+ generation. Then Cu2+ can be converted into Cu+ partially by glutathione (GSH) to from a Cu+ supply loop and ensure the synergistic action. Meanwhile, the consumption of GSH also contributes to cuproptosis and CDT in return. Finally, DNA and Zn2+ form DNAzyme to shear catalase-related RNA, resulting in the accumulation of hydrogen peroxide and further enhancing combination therapy. These results provide a promising nanotherapeutic platform and may inspire the design for potential cancer treatment based on cuproptosis.

Intracellular activated logic nanomachines based on framework nucleic acids for low background detection of microRNAs in living cells.

DNA molecular machines based on DNA logic circuits show unparalleled potential in precision medicine. However, delivering DNA nanomachines into real biological systems and ensuring that they perform functions specifically, quickly and logically remain a challenge. Here, we developed an efficient DNA molecular machine integrating transfer-sensor-computation-output functions to achieve high fidelity detection of intracellular biomolecules. The introduction of pH nanoswitches enabled the nanomachines to be activated after entering the cell, and the spatial-confinement effect of the DNA triangular prism (TP) enables the molecular machine to process complex information at the nanoscale, with higher sensitivity and shorter response time than diffuse-dominated logic circuits. Such cascaded activation molecular machines follow the logic of AND to achieve specific capture and detection of biomolecules in living cells through a multi-hierarchical response, providing a new insight into the construction of efficient DNA molecular machines.

An intelligent DNA nanomachine for amplified MicroRNA imaging and MicroRNA-Guided efficient gene silencing.

The DNA nanomachines as excellent synthetic biological tools have been widely used for the sensitive detection of intracellular microRNA (miRNA) and DNAzyme-involved gene silencing. However, intelligent DNA nanomachines which have the ability to sense intracellular specific biomolecules and respond to external information in complex environments still remain challenging. Herein, we develop a miRNA-responsive DNAzyme cascaded catalytic (MDCC) nanomachine to perform multilayer cascade reactions, enabling the amplified intracellular miRNA imaging and miRNA-guided efficient gene silencing. The intelligent MDCC nanomachine is designed based on multiple DNAzyme subunit-encoded catalyzed hairpin assembly (CHA) reactants sustained by the pH-responsive Zeolitic imidazolate framework-8 (ZIF-8) nanoparticles. After cellular uptake, the MDCC nanomachine degrades in acidic endosome and releases three hairpin DNA reactants and Zn2+, and the latter can act as an effective cofactor for DNAzyme. In the presence of miRNA-21, a catalytic hairpin assembly (CHA) reaction is triggered, which produces a large number of Y-shaped fluorescent DNA constructs containing three DNAzyme modules for gene silencing. The construction of Y-shaped DNA modified with multisite fluorescence and the circular reaction realizes ultrasensitive miRNA-21 imaging of cancer cells. Moreover, miRNA-guided gene silencing inhibits the cancer cell proliferation through the DNAzyme-specific recognition and cleavage of target EGR-1 (Early Growth Response-1) mRNA, which is one key tumor-involved mRNA. The strategy may provide a promising platform for highly sensitive determination of biomolecules and accurate gene therapy of cancer cells.

Visual analysis of Alzheimer disease biomarker via low-potential driven bipolar electrode.

Developing a simple, economical, and accurate diagnostic method has positive practical significance for the early prevention and intervention of Alzheimer's disease (AD). Herein, combining a closed bipolar electrode (BPE) chip with multicolor electrochemiluminescence (ECL) imaging technology, we constructed a low-voltage driven portable visualized ECL device for the early screening of AD. By introducing parallel resistance, the total resistance of the circuit was greatly reduced. A classical mixture of Ir(ppy)3 and Ru(bpy)32+ was used as multicolor emitters of the anode with TPrA as the co-reactant. Capture of amyloid-ß (Aß) through antigen-antibody recognition, and signal amplification by electroactive covalent organic frameworks (COF) probe at the cathode of BPE caused the significantly increased faradaic current. The electrical balance of the BPE system resulted in the change of the emission color from green to red at the anode. The ECL-BPE sensor shows good reproducibility and high sensitivity with detection limit of 1 pM by naked eye. The driving voltage is 3.0 V, which means the chip could be driven by two fifth batteries. The visualized ECL-BPE sensor provides a promising point-of-care testing (POCT) tool for the screening of Alzheimer's-related diseases in the early stage.

Boron-enriched rice-like homologous carbon nanoclusters with a 51.5% photoluminescent quantum yield for highly sensitive determination of endogenous hydroxyl radicals in living cells.

Exploring the ultrahigh quantum efficiency of a carbon-based probe via a green and simple technique, and utilisation of its sensing ability for highly bioactive molecule detection is still highly challenging. Herein, we prepared a novel boron-enriched rice-like homologous carbon nanoclusters (BRCNs) with an ultrahigh quantum efficiency of ∼51.5% by introduction of a conjugated structure attached to the CN bond and an electron-withdrawing boron active centre. Unexpectedly, the BRCNs obtained showed a stable dispersion of rice-like carbon nanograins, composed of small carbon dot assembled nanoclusters with an average diameter size of ∼30 nm, and containing boron units of ∼24.68 at%. What's exciting is that the BRCNs obtained exhibited an "on-off-on" three-state emission with the addition of an hydroxyl radical (OH˙) and its antioxidants. Thus, two distinctive fluorescent responses for OH˙ and antioxidants based on the BRCN probe had been developed, and the mechanism has been determined using TEM, XPS, FT-IR, FL, UV-vis spectrophotometry, UPS and fluorescent lifetimes. The OH˙, generated from the Fenton's reagent, preferentially attack the electron-deficient vacancy p orbit of the boron atom in the surface of the BRCNs, which results in the boron atom being easily substituted/attacked by OH˙, and leading to spontaneous aggregation induced quenching (AIQ) due to the existence of a strong intermolecular hydrogen bond between denatured BRCNs. Furthermore, the proposed method was also successfully applied to monitor endogenous OH˙ generation in HeLa cells by confocal imaging, which could be used for elucidating OH˙-induced oxidative damage to biological tissues and proteins.

Translocation of Specific DNA Nanocarrier through an Ultrasmall Nanopipette: Toward Single-Protein-Molecule Detection with Superior Signal-to-Noise Ratio.

The use of functional DNA nanostructures as carriers to ship proteins through solid-state nanopores has recently seen substantial growth in single-protein-molecule detection (SPMD), driven by the potential of this methodology and implementations that it may enable. Ultrasmall nanopores have exhibited obvious advantages in spatiotemporal biological detection due to the appropriate nanoconfined spaces and unique properties. Herein, a 6.8 nm DNA tetrahedron (TDN) with a target-specific DNA aptamer (TDN-apt) was engineered to carry the representative target of acetylcholinesterase (AChE) through an ultrasmall nanopipet with a 30 nm orifice, underpinning the advanced SPMD of AChE with good performance in terms of high selectivity, low detection limit (0.1 fM), and especially superior signal-to-noise ratio (SNR). The kinetic interaction between TDN-apt and AChE was studied and the practical applicability of the as-developed SPMD toward real samples was validated using serum samples from patients with Alzheimer's disease. This work not only presented a feasible SPMD solution toward low-abundance proteins in complex samples and but also was envisioned to inspire more interest in the design and implementation of synergized DNA nanostructure-ultrasmall nanopore systems for future SPMD development.

Near-infrared photothermally activated DNA nanotweezers for imaging ATP in living cells.

By assembling nanotweezers with ATP-splitting aptamers on gold nanorods (AuT123L), we constructed a near-infrared-activated ATP sensing device that could time-controllably image ATP levels in living cells. By replacing the aptamers on the nanotweezers, the nanoplatform can be applied to other important biomolecules, opening up more possibilities for the study of time controllable nanodevices.

[Effects of Agasicles hygrophila herbivory on the clonal integration of Alternanthera philoxeroides and A. sessilis]. / èŽ²è‰ç›´èƒ¸è·³ç”²å–é£Ÿå¯¹ç©ºå¿ƒèŽ²å­è‰å’ŒèŽ²å­è‰å ‹éš†æ•´åˆçš„å½±å“.

Many alien invasive plants were clonal species. Examining the relationship between clonal integration characteristics and invasiveness of alien clonal plants is important for clarifying their ecological adaptability and invasion mechanisms. Here, with the invasive plant species Alternanthera philoxeroides and its native congener A. sessilis as the studying objects, we compared the effects of clonal integration on the growth and the biomass allocation of the apical ramets, basal ramets, and the whole fragment of both species under herbivory by the biocontrol beetle Agasicles hygrophila. The results showed that under herbivory by A. hygrophila, leaf number, stolon length, and ramet number of the apical ramets as well as the ground diameter of the whole fragment of A. philoxeroides were significantly higher under clonal integration treatment compared to that without clonal integration, whereas belowground biomass and total biomass of the basal ramets and the whole fragment of A. philoxeroides were conversely decreased by 78.2%, 60.9 % and 48.7%, 37.2%, respectively, under clonal integration treatment compared to that without clonal integration. Ground diameter of the apical ramets and leaf number of the whole fragment of A. sessilis were significantly higher, but the number of basal ramets was 21.7% lower under clonal integration treatment compared to that without clonal integration. The biomass of the apical ramets, basal ramets, and whole fragment of A. sessilis did not significantly differ between clonal integration and without clonal integration treatments. The results of cost-benefit analysis showed that the ramet number and biomass of the apical ramets of A. philoxeroides as well as the ramet number of the apical ramets of A. sessilis were significantly benefited from clonal integration, but the cost-benefit of the ramet number and biomass of the basal ramets of both species were not affected by clonal integration treatment. These results suggested that clonal integration could partly alleviate herbivory pressure by A. hygrophila on the apical ramets of both species, and that the clonal integration ability of A. philoxeroides was stronger than A. sessilis. However, both species seemed not able to gain significant benefits from cloning integration at the whole fragment level.

Dual Recognition DNA Triangular Prism Nanoprobe: Toward the Relationship between K+ and pH in Lysosomes.

Lysosomal acidification is essential for its degradative function, and the flux of H+ correlated with that of K+ in lysosomes. However, there is little research on their correlation due to the lack of probes that can simultaneously image these two ions. To deeply understand the role of K+ in lysosomal acidification, here, we designed and fabricated a nanodevice using a K+-aptamer and two pH-triggered nanoswitches incorporated into a DNA triangular prism (DTP) as a dual signal response platform to simultaneously visualize K+ and pH in lysosomes by a fluorescence method. This strategy could conveniently integrate two signal recognition modules into one probe, so as to achieve the goal of sensitive detection of two kinds of signals in the same time and space, which is suitable for the detection of various signals with the correlation of concentration. By co-imaging both K+ and H+ in lysosomes, we found that the efflux of K+ was accompanied by a decrease of pH, which is of great value in understanding lysosomal acidification. Moreover, this strategy also has broad prospects as a versatile optical sensing platform for multiplexed analysis of other biomolecules in living cells.

[Effects of above- and below-ground herbivore interactions on interspecific relationship between the invasive plant Alternanthera philoxeroides and its native congener Alternanthera sessilis]. / 地上-åœ°ä¸‹æ¤é£Ÿæ€§å¤©æ•Œå¯¹å ¥ä¾µæ¤ç‰©ç©ºå¿ƒèŽ²å­è‰ä¸Žæœ¬åœ°ç§èŽ²å­è‰ç§é—´å ³ç³»çš„å½±å“.

Biological invasion is a major threat to global biodiversity. The relative interspecific competition abilities of invasive species compared to those native species determine their invasion success. In this study, we examined the effects of the specialist leaf beetle Agasicles hygrophila and the nematode Meloidogyne incognita on the growth and interspecific relationship between the invasive plant Alternanthera philoxeroides and its native congener A. sessilis. Compared without herbivory, nematode herbivory alone significantly reduced shoot height of A. sessilis by 28.1%, but conversely significantly increased the shoot height of A. philoxeroides by 52.8% and aboveground biomass of A. sessilis by 63.7%. Beetle herbivory alone significantly reduced shoot height of A. sessilis by 40.7%, but did not affect that of A. philoxeroides. The combination of beetle and nematode herbivory significantly reduced shoot height of A. sessilis by 35.3% as well as the belowground biomass of A. philoxeroides by 62.2%, but significantly increased the aboveground biomass of A. sessilis by 69.1%. Herbivore stress did not affect stem diameter, branch number, and root length of both species. The relative neighbor effect index (RNE) of the two species without herbivory were positive, and the RNE value of A. philoxeroides was 21.3% higher than that of A. sessilis. However, the RNE values of A. philoxeroides were negative under all above- and below-ground herbivory treatments. The RNE values of A. sessilis were positive under the beetle or the nematode herbivory alone and negative under the beetle + nematode herbivory combination. These results indicated that above- and below-ground herbivore interactions could change the interspecific relationship between the two species, and in turn might accelerate the invasion of A. philoxeroides.

Chlorogenic acid enhances autophagy by upregulating lysosomal function to protect against SH-SY5Y cell injury induced by H2O2.

Autophagy serves an important role in amyloid-ß (Aß) metabolism and τ processing and clearance in Alzheimer's disease. The progression of Aß plaque accumulation and hyperphosphorylation of τ proteins are enhanced by oxidative stress. A hydrogen peroxide (H2O2) injury cell model was established using SH-SY5Y cells. Cells were randomly divided into normal, H2O2 and chlorogenic acid (5-caffeoylquinic acid; CGA) groups. The influence of CGA on cell viability was evaluated using a Cell Counting Kit-8 assay and cell death was assessed using Hoechst 33342 nuclear staining. Autophagy induction and fusion of autophagic vacuoles assays were performed using monodansylcadaverine staining. Additionally, SH-SY5Y cells expressing Ad-mCherry-green fluorescent protein-LC3B were established to detect autophagic flow. LysoTracker Red staining was used to evaluate lysosome function and LysoSensor™ Green staining assays were used to assess lysosomal acidification. The results demonstrated that CGA decreased the apoptosis rate, increased cell viability and improved cell morphology in H2O2-treated SH-SY5Y cells. Furthermore, CGA alleviated the accumulation of autophagic vacuoles, reduced the LC3BII/I ratio and decreased P62 levels, resulting in increased autophagic flux. Additionally, CGA upregulated lysosome acidity and increased the expression levels of cathepsin D. Importantly, these effects of CGA on H2O2-treated SH-SY5Y cells were mediated via the mTOR-transcription factor EB signaling pathway. These results indicated that CGA protected cells against H2O2-induced oxidative damage via the upregulation of autophagosomes, which promoted autophagocytic degradation and increased autophagic flux.

Proportions of Th17 cells and Th17-related cytokines in neuromyelitis optica spectrum disorders patients: A meta-analysis.

BACKGROUND: T helper (Th17) cells play an important role in many autoimmune diseases. In this meta-analysis, we aimed to specify the proportion of Th17 cells and the levels of Th17-related cytokines in neuromyelitis optica spectrum disorders (NMOSD) patients, we did this meta-analysis. METHODS: Using previously reported data from PubMed, EMBASE, and Web of Science and Cochrane, we explored the proportion of Th17 cells in CD4+ T cells in peripheral blood (PB) and the level of Th17-related cytokines, such as interleukin (IL)1ß, IL6, IL17, IL21, IL22, IL23 and transforming growth factor -beta (TGFß), in cerebrospinal fluid (CSF), plasma, and serum in NMOSD patients compared to control group and multiple sclerosis (MS) patients. RESULTS: In total, 38 trials were included for our analysis. Results showed that the proportion of Th17 cells was higher in NMOSD patients than in the control and MS groups. The levels of IL1ß, IL6, IL17 and IL21 in CSF and plasma, and IL6, IL21, IL22, and IL23 in the serum were higher in NMOSD patients than in the control group. The levels of IL6 in CSF and serum and IL17 in plasma and serum were higher in NMOSD patients than in MS patients. CONCLUSION: The proportion of Th17 cells and the levels of Th17-related cytokines was increased in NMOSD patients compared with the control group and MS patients. The results of this meta-analysis indicated that Th17 cells and Th17-associated cytokines may play an essential role in the pathogenesis of NMOSD. PROSPERO registration: CRD42019128785.

Association between bone mineral density and benign paroxysmal positional vertigo: a meta-analysis.

BACKGROUND: The association between bone mineral density (BMD) and benign paroxysmal positional vertigo (BPPV) has been investigated by multiple studies, but the conclusions are controversial. This meta-analysis was conducted to evaluate whether the bone mineral density is associated with BPPV. METHODS: The relevant studies were identified by searching PubMed, EMBASE, Cochrane Library, ScienceDirect, Web of Science database up to June 2018. Statas14.0 software was used for meta-analysis. We used the pooled odds ratio (OR) and 95% confidence interval (CI) to assess the incidence of osteoporosis and osteopenia in patients with BPPV and controls (free of BPPV disease). The standardized mean difference (SMD) and 95% confidence interval (CI) were used to assess the T score in BPPV patients and controls. This meta-analysis has been registered at International Prospective Register of Systematic Reviews (PROSPERO) (number CRD42018082271). RESULTS: A total of 11 studies were eligible for meta-analysis, including 1982 subjects. When compared with the controls, the total incidence of osteoporosis and osteopenia was significantly higher in BPPV patients (OR 3.27, 95% CI 2.66-4.03, p < 0.0001). Further analysis was conducted by separate discussion about the incidence of osteoporosis and osteopenia in BPPV patients, the result of which shows that both the incidence of osteoporosis (OR 3.48, 95% CI 1.86-6.51, p < 0.0001) and the incidence of osteopenia (OR 1.75, 95% CI 1.01-3.04, p < 0.0001) were higher in BPPV patients than that in controls. There was an significant reduction in T scores of BPPV patients (SMD - 0.82, 95% CI -1.18 to - 0.46, p < 0.0001). Publication bias for each analysis was evaluated by Egger's test and Begg's indicating that no publication bias existed. Sensitivity analysis was conducted for each analysis demonstrating that the results were robust. CONCLUSIONS: Our meta-analysis provided stronger evidence that patients with BPPV were associated with a lower T score and a higher risk of osteoporosis and osteopenia. The results demonstrated that lower bone mineral density may be a risk factor for BPPV. However, large-scare, multicenter clinical studies need to be carried out to explore the precise risk of osteoporosis and osteopenia in patients with BPPV in future.

Microbiomes of China's Space Station During Assembly, Integration, and Test Operations.

Sufficient evidence indicates that orbiting space stations contain diverse microbial populations, which may threaten astronaut health and equipment reliability. Understanding the composition of microbial communities in space stations will facilitate further development of targeted biological safety prevention and maintenance practices. Therefore, this study systematically investigated the microbial community of China's Space Station (CSS). Air and surface samples from 46 sites on the CSS and Assembly Integration and Test (AIT) center were collected, from which 40 bacteria strains were isolated and identified. Most isolates were cold- and desiccation-resistant and adapted to oligotrophic conditions. Bacillus was the dominant bacterial genus detected by both cultivation-based and Illumina MiSeq amplicon sequencing methods. Microbial contamination on the CSS was correlated with encapsulation staff activities. Analysis by spread plate and qPCR revealed that the CSS surface contained 2.24 × 103-5.47 × 103 CFU/100 cm2 culturable bacteria and 9.32 × 105-5.64 × 106 16S rRNA gene copies/100cm2; BacLight™ analysis revealed that the viable/total bacterial cell ratio was 1.98-13.28%. This is the first study to provide important systematic insights into the microbiome of the CSS during assembly that describes the pre-launch microbial diversity of the space station. Our findings revealed the following. (1) Bacillus strains and staff activities should be considered major concerns for future biological safety. (2) Autotrophic and multi-resistant microbial communities were widespread in the AIT environment. Although harsh cleaning methods reduced the number of microorganisms, stress-resistant strains were not completely removed. (3) Sampling, storage and analytical methods for the space station were thoroughly optimized, and are expected to be applicable to low-biomass environments in general. Microbiology-related future works will follow up to comprehensively understand the changing characteristics of microbial communities in CSS.

[Canopy vertical structure and understory plant regeneration of an evergreen broadleaved forest in Damingshan, Guangxi, China.] / å¤§æ˜Žå±±å¸¸ç»¿é˜”å¶æž—å† å±‚åž‚ç›´ç»“æž„ä¸Žæž—ä¸‹æ¤ç‰©æ›´æ–°.

In order to reveal the dynamics of canopy vertical structure and its effects on understory regeneration, we built 24 permanent plots (20 m×20 m) on the upslope, midslopeand downslope, respectively, in a typical evergreen broadleaved forest in Damingshan, Guangxi, China. We measured the crown area of each tree with diameter at breast height (DBH)≥1.0 cm, and surveyed the understory regeneration in growing season from 2009 to 2011. The results showed that the total canopy cover significantly increased from 54.0% in 2009 to 67.4% in 2011 after the frozen disaster in 2008. A significant difference existed in the cover and increment of different canopy layers. The canopy cover in the upper layers was markedly higher than that in the middle and lower layers. The increment of canopy coverage in the middle and lower layers was significantly higher than that in the upper layer. There were 55 regenerated woody plant species, and the dominant families and species of regenerated plants were in accord with those in the evergreen broadleaved forest. Biodiversity index of regenerated plants in the same slope position was significantly different among different years, and no significant difference was observed among different slope positions in the same year. The correlation between the coverage at different canopy layers and the species richness and abundance of regenerated plants was not significant. Total canopy cover and canopy coverage at the middle and lower layers were significantly negatively correlated with the Shannon index, Simpson index, and Pielou evenness index of the understory regenerated plants. It indicated that canopy coverage had a significant influence on the regeneration of understory, and the middle and lower layers had a stronger influence on the biodiversity of regenerated plants.