Spatio-temporal microbial regulation of aggregate-associated priming effects under contrasting tillage practices.

Tillage intensity significantly influences the heterogeneous distribution and dynamic changes of soil microorganisms, consequently shaping spatio-temporal patterns of SOC decomposition. However, little is known about the microbial mechanisms by which tillage intensity regulates the priming effect (PE) dynamics in heterogeneous spatial environments such as aggregates. Herein, a microcosm experiment was established by adding 13C-labeled straw residue to three distinct aggregate-size classes (i.e., mega-, macro-, and micro-aggregates) from two long-term contrasting tillage histories (no-till [NT] and conventional plow tillage [CT]) for 160 days to observe the spatio-temporal variations in PE. Metagenomic sequencing and Fourier transform mid-infrared techniques were used to assess the relative importance of C-degrading functional genes, microbial community succession, and SOC chemical composition in the aggregate-associated PE dynamics during straw decomposition. Spatially, straw addition induced a positive PE for all aggregates, with stronger PE occurring in larger aggregates, especially in CT soil compared to NT soil. Larger aggregates have more unique microbial communities enriched in genes for simple C degradation (e.g., E5.1.3.6, E2.4.1.7, pmm-pgm, and KduD in Nitrosospeera and Burkholderia), contributing to the higher short-term PE; however, CT soils harbored more genes for complex C degradation (e.g., TSTA3, fcl, pmm-pgm, and K06871 in Gammaproteobacteria and Phycicoccus), supporting a stronger long-term PE. Temporally, soil aggregates played a significant role in the early-stage PEs (i.e., < 59 days after residue addition) through co-metabolism and nitrogen (N) mining, as evidenced by the increased microbial biomass C and dissolved organic C (DOC) and reduced inorganic N with increasing aggregate-size class. At a later stage, however, the legacy effect of tillage histories controlled the PEs via microbial stoichiometry decomposition, as suggested by the higher DOC-to-inorganic N and DOC-to-available P stoichiometries in CT than NT. Our study underscores the importance of incorporating both spatial and temporal microbial dynamics for a comprehensive understanding of the mechanisms underlying SOC priming, especially in the context of long-term contrasting tillage practices.

In Silico Investigation of Palladium-Catalyzed Chemoselective Monoalkoxycarbonylation of 1,3-diynes for Conjugated Enynes Synthesis.

The palladium-catalyzed monoalkoxycarbonylation of 1,3-diynes provides a chemoselective method for the construction of synthetically useful conjugated enynes. Here, in silico unraveling the detailed mechanism of this reaction and the origin of chemoselectivity were conducted. It is shown that the alkoxycarbonylation reaction preferably proceeds by a NH-Pd pathway, which including three substeps: hydropalladation, CO migratory insertion and methanolysis. The effectiveness of the NH-Pd catalytic system is attributed to the alkynyl-palladium π-back-bonding interaction, C-H⋅⋅⋅π interaction in reactant moiety and d-pπ conjugation between the Pd center and alkenyl group. The hydropalladation step was identified as the rate- and chemoselectivity-determining step, and the first alkoxycarbonylation requires a much lower energy barrier in comparison with the second alkoxycarbonylation, in line with the experimental outcomes that the monoalkoxycarbonylation product was obtained in high yield. Distortion-interaction analysis indicates the more favorable monoalkoxycarbonylation (compared to double alkoxycarbonylation) is caused by steric effect.

Quantitative and modularized CRISPR/dCas9-dCpf1 dual function system in Saccharomyces cerevisiae.

Introduction: Both CRISPR/dCas9 and CRISPR/dCpf1 genome editing systems have shown exciting promises in modulating yeast cell metabolic pathways. However, each system has its deficiencies to overcome. In this study, to achieve a compensatory effect, we successfully constructed a dual functional CRISPR activation/inhibition (CRISPRa/i) system based on Sp-dCas9 and Fn-dCpf1 proteins, along with their corresponding complementary RNAs. Methods: We validated the high orthogonality and precise quantity targeting of selected yeast promoters. Various activating effector proteins (VP64, p65, Rta, and VP64-p65-Rta) and inhibiting effector proteins (KRAB, MeCP2, and KRAB-MeCP2), along with RNA scaffolds of MS2, PP7 and crRNA arrays were implemented in different combinations to investigate quantitative promoter strength. In the CRISPR/dCas9 system, the regulation rate ranged from 81.9% suppression to 627% activation in the mCherry gene reporter system. Studies on crRNA point mutations and crRNA arrays were conducted in the CRISPR/dCpf1 system, with the highest transcriptional inhibitory rate reaching up to 530% higher than the control. Furthermore, the orthogonal CRISPR/dCas9-dCpf1 inhibition system displayed distinct dual functions, simultaneously regulating the mCherry gene by dCas9/gRNA (54.6% efficiency) and eGFP gene by dCpf1/crRNA (62.4% efficiency) without signal crosstalk. Results and discussion: Finally, we established an engineered yeast cell factory for ß-carotene production using the CRISPR/dCas9-dCpf1 bifunctional system to achieve targeted modulation of both heterologous and endogenous metabolic pathways in Saccharomyces cerevisiae. The system includes an activation module of CRISPRa/dCas9 corresponding to a gRNA-protein complex library of 136 plasmids, and an inhibition module of CRISPRi/dCpf1 corresponding to a small crRNA array library. Results show that this CRISPR/dCas9-dCpf1 bifunctional orthogonal system is more quantitatively effective and expandable for simultaneous CRISPRa/i network control compared to single-guide edition, demonstrating higher potential of future application in yeast biotechnology.

Transcriptome analysis used to identify and characterize odorant binding proteins in Agasicles hygrophila (Coleoptera: Chryspmelidae).

The transcriptomes of Agasicles hygrophila eggs and first instar larvae were analyzed to explore the olfactory mechanism of larval behavior. The analysis resulted in 135,359 unigenes and the identification of 38 odorant-binding proteins (OBPs), including 23 Minus-C OBPs, 8 Plus-C OBPs, and 7 Classic OBPs. Further analysis of differentially expressed genes (DEGs) revealed 10 DEG OBPs, with 5 (AhygOBP5, AhygOBP9, AhygOBP12, AhygOBP15 and AhygOBP36) up-regulated in first instar larvae. Verification of expression patterns of these 5 AhygOBPs using qPCR showed that AhygOBP9 and AhygOBP36 were mainly expressed in the adult stage with gradually increasing expression in the larval stage. AhygOBP5, AhygOBP12, and AhygOBP15 were not expressed in eggs and pupae, and their expression in larvae and adults showed no clear pattern. These 5 AhygOBPs may play an olfactory role in larval behavior, providing a basis for further investigation of their specific functions and clarifying the olfactory mechanism of A. hygrophila.

Semiconducting Polymer Nanospherical Nucleic Acid Probe for Transcriptomic Imaging of Cancer Chemo-Immunotherapy.

Real-time in vivo imaging of RNA can enhance the understanding of physio-pathological processes. However, most nucleic acid-based sensors have poor resistance to nucleases and limited photophysical properties, making them suboptimal for this purpose. To address this, a semiconducting polymer nanospherical nucleic acid probe (SENSE) for transcriptomic imaging of cancer immunity in living mice is developed. SENSE comprises a semiconducting polymer (SP) backbone covalently linked with recognition DNA strands, which are complemented by dye-labeled signal DNA strands. Upon detection of targeted T lymphocyte transcript (Gzmb: granzyme B), the signal strands are released, leading to a fluorescence enhancement correlated to transcript levels with superb sensitivity. The always-on fluorescence of the SP core also serves as an internal reference for tracking SENSE uptake in tumors. Thus, SENSE has the dual-signal channel that enables ratiometric imaging of Gzmb transcripts in the tumor of living mice for evaluating chemo-immunotherapy; moreover, it has demonstrated sensitivity and specificity comparable to flow cytometry and quantitative polymerase chain reaction,  yet offering a faster and simpler means of T cell detection in resected tumors. Therefore, SENSE represents a promising tool for in vivo RNA imaging.

Physicochemical changes in microplastics and formation of DBPs under ozonation.

Microplastics (MPs) are substances that pose a risk to both human life and the environment. Their types and production are increasing year on year, and their potential to cause environmental pollution is a worldwide concern. Conventional water treatment processes, particularly coagulation and sedimentation, are not effective at removing all MPs. It is therefore important to assess the morphological changes in the MPs, i.e., the thermoplastic polyurethane (TPU) and polyethylene (PE), during ozonation and the dissolved organic carbon leaching as well as chloroform formation in the subsequent chlorination. The results show that the appearance and surface chemistry of the MPs changed during the ozonation process, most notably for TPU. The trichloromethane (CHCl3) generation during chlorination was 0.168 and 0.152 µmol/L for TPU and PE, respectively, and the ozone pretreatment significantly increased the CHCl3 yield of TPU, while it had a weak effect on PE. Additional disinfection byproducts (DBPs), including CHCl2Br, CHClBr2, and CHBr3, were produced in the presence of bromide ions in the water column, and the total amount of DBPs produced by PE, PE-O, TPU, and TPU-O was significantly increased to 0.787, 0.814, 0.931, and 1.391 µmol/L, respectively. The study provides useful information for the environmental risk assessment of two representative MPs, i.e., TPU and MPs, in disinfection procedures for drinking water.

The impact of chlorination on the tetracycline sorption behavior of microplastics in aqueous solution.

Considering the large volumes of treated water and incomplete elimination of pollutants, wastewater treatment plants (WWTPs) remain a considerable source of microplastics (MPs). Chlorine, the most frequently used disinfectant in WWTPs, has a strong oxidizing impact on MPs. However, little is documented, to date, about the impact of chlorination on the transformation of MPs and the subsequent environmental behaviors of the chlorinated MPs when released into the aquatic environment. This study explored the response of the physicochemical properties of specific thermoplastics, namely polyurethane (TPU) MPs and polystyrene (PS) MPs, to chlorination and their emerging pollutant [tetracycline (TC)] adsorption behavior in aqueous solution. The results indicated that the O/C ratio of the MP surface did not significantly change, and that there were increases in the O-containing functional groups of the TPU and PS MPs, after chlorination. The surface area of the chlorinated TPU MPs increased by 45 %, and that of the chlorinated PS increased by 21 %, compared with the pristine ones, which contributed to the TC adsorption. The adsorption isotherm fitting parameters suggested that the chlorinated TPU fitted the multilayer adsorption, and the chlorinated PS was inclined to the monolayer adsorption. The relative abundance of the O-containing functional groups, on the TPU surface, led to the release of CHCl3 molecules, and the clear surface irregularities and fissures occurred after chlorine treatment. No fissures were found on the surface of the chlorinated PS MPs. The hydrophobicity and electrostatic adsorption were proved to be the major impacts on the TC adsorption of the chlorinated MPs, and the subsequently formed hydrogen bonds led to the stronger adsorption capacity of the chlorinated TPU than the chlorinated PS MPs.

Autonomous Bionanorobots via a Cage-Shaped Silsesquioxane Vehicle for In Vivo Heavy Metal Detoxification.

Nanorobots hold great promise for integrated drug delivery systems that are responsive to molecular triggers. Herein, we successfully developed an automatic smart bionanorobot that has transport capability and recognizes and removes zinc ions from poisoned cells based on nanoscale polyhedral oligomeric silsesquioxane molecules. This intelligent bionanorobot can easily move inside and outside the cell and find zinc ions owing to its highly selective recognition to zinc ions and high cell permeability, especially the well-combined high penetration and strong binding energy. More importantly, it was also found that this intelligent bionanorobot can restore round HeLa cells to a normal fusiform cell morphology following high-concentration zinc treatment and does not interfere with cell proliferation and division. It was also shown by in vivo experiments that the bionanorobot can inhibit persistent enlargement of the liver caused by zinc ion poisoning.

Monitoring Wound Healing with Topically Applied Optical NanoFlare mRNA Nanosensors.

An effective wound management strategy needs accurate assessment of wound status throughout the whole healing process. This can be achieved by examining molecular biomarkers including proteins, DNAs, and RNAs. However, existing methods for quantifying these biomarkers such as immunohistochemistry and quantitative polymerase chain reaction are usually laborious, resource-intensive, and disruptive. This article reports the development and utilization of mRNA nanosensors (i.e., NanoFlare) that are topically applied on cutaneous wounds to reveal the healing status through targeted and semi-quantitative examination of the mRNA biomarkers in skin cells. In 2D and 3D in vitro models, the efficacy and efficiency of these nanosensors are demonstrated in revealing the dynamic changes of mRNA biomarkers for different stages of wound development. In mouse models, this platform permits the tracking and identification of wound healing stages and a normal and diabetic wound healing process by wound healing index in real time.

Transdermal delivery of Chinese herbal medicine extract using dissolvable microneedles for hypertrophic scar treatment.

Hypertrophic scars are unfavorable skin diseases characterized by excessive collagen deposition. Although systemic treatments exist in clinic to manage hypertrophic scars, they pose significant side effects and tend to lose efficacy over prolonged applications. Traditional Chinese medicine (TCM) offers as a promising candidate to treat pathological scars. A large number of TCMs have been studied to show anti-scarring effect, however, the natural barrier of the skin impedes their penetration, lowering its therapeutic efficacy. Herein, we reported the use of dissolvable hyaluronic acid (HA) microneedles (MNs) as a vehicle to aid the transdermal delivery of therapeutic agent, a model TCM called shikonin for the treatment of hypertrophic scars. Here, shikonin was mixed with HA to make MNs with adequate mechanical strength for skin penetration, making its dosage controllable during the fabrication process. The therapeutic effect of the shikonin HA MNs was studied in vitro using HSFs and then further verified with quantitative reverse transcriptase polymerase chain reaction. Our data suggest that the shikonin HA MNs significantly reduce the viability and proliferation of the HSFs and downregulate the fibrotic-related genes (i.e., TGFß1, FAP-α and COL1A1). Furthermore, we observed a localized therapeutic effect of the shikonin HA MNs that is beneficial for site-specific treatment.

Enhanced Drug Permeation into Human Keloid Tissues by Sonophoresis-Assisted Microneedling.

Keloids are disfiguring pathological scars that could cause pain and pruritus. The conventional treatments, such as bolus injection of drugs or surgery, are invasive and require a personal visit to clinic. Microneedle (MN) technology has great potential to offer a self-administered and minimally invasive treatment of keloids. However, drugs delivered using MNs suffer from limited penetration in keloid tissue. This study demonstrates enhanced drug penetration in human keloid scar tissue by combining MN and sonophoresis.

Factor analysis of the relationship between PANSS score and family burden of patients with schizophrenia.

OBJECTIVE: This study aims to investigate the burden of family caregivers of patients with schizophrenia, and its influencing factors METHODS: A total of 105 patients with schizophrenia and their caregivers were investigated using the positive and negative symptom scale (PANSS) and family burden scale of disease (FBS) RESULTS: There was a strong correlation between the patient's recovery and family burden, especially between positive and negative symptoms and family financial burden, family daily activities, family recreational activities, and family relationship CONCLUSION: There is a strong correlation between the patient's recovery and family burden, and this is especially correlated to family economic burden, family daily activities, family recreational activities, and family relationship. Medical staff should pay attention to the psychological characteristics of patients and fully understand and avoid the adverse effects of family burden on the rehabilitation of patients.

Cryomicroneedles for transdermal cell delivery.

Cell therapies for the treatment of skin disorders could benefit from simple, safe and efficient technology for the transdermal delivery of therapeutic cells. Conventional cell delivery by hypodermic-needle injection is associated with poor patient compliance, requires trained personnel, generates waste and has non-negligible risks of injury and infection. Here, we report the design and proof-of-concept application of cryogenic microneedle patches for the transdermal delivery of living cells. The microneedles are fabricated by stepwise cryogenic micromoulding of cryogenic medium with pre-suspended cells, and can be easily inserted into porcine skin and dissolve after deployment of the cells. In mice, cells delivered by the cryomicroneedles retained their viability and proliferative capability. In mice with subcutaneous melanoma tumours, the delivery of ovalbumin-pulsed dendritic cells via the cryomicroneedles elicited higher antigen-specific immune responses and led to slower tumour growth than intravenous and subcutaneous injections of the cells. Biocompatible cryomicroneedles may facilitate minimally invasive cell delivery for a range of cell therapies.

Impact assessment of odor nuisance, health risk and variation originating from the landfill surface.

Many researchers are concerned that municipal solid waste (MSW) threatens public health, causing them to increasingly focus on odor pollution. In this study, the odor nuisance and health risk impacts of landfill surface gas on eight sensitive receptors were assessed. The emission rates of odor and 145 volatile organic compounds (VOCs) were acquired by considering various landfilling operations, including high-density polyethylene (HDPE) membrane removal (MR), landfill tipping area (TA), temporary HDPE membrane cover (MC), top of the HDPE membrane (LM) and dumping platform (DP). Furthermore, differences in landfill surface geometry, such as emission height and source area, and variations in residential living floors were considered in odor assessment with the air dispersion model. Based on these uncertain factors, normal-, medial-, and worst-case scenarios were defined to elucidate the odor nuisance effect and health risk impact. Four of the eight sensitive receptors, which were 2.6 km away from the landfill surface, basically experienced odor nuisance and health risk impacts. Dichloromethane exerted an indelible and crucial impact on body health based on a comprehensive investigation of aromatics, halocarbons, and other chemicals. The odor nuisance and health risk impacts were notable near the landfill, and the local environment was remarkably damaged.

Comparison of hypertrophic scarring on a red Duroc pig and a Guangxi Mini Bama pig.

Pigs are the most promising models for the study of wound healing and hypertrophic scarring because they are anatomically and physiologically similar to human beings. The Red Duroc pig and Mini Bama pig are two swine models that have attracted a lot of attention. The aim of the present study was to examine and compare the scarring process in a red Duroc pig and a Mini Bama pig, providing knowledge for researchers and clinicians to enable them to choose the most suitable pig model for studies.

Stress-driven dynamic regulation of multiple tolerance genes improves robustness and productive capacity of Saccharomyces cerevisiae in industrial lignocellulose fermentation.

Yeast productivity in lignocellulosic ethanol fermentation is clearly impeded by stress. Enhancing the robustness of xylose-fermenting yeast is important for improving lignocellulosic ethanol production. In this study, the glutathione biosynthesis pathway and acetic acid degradation pathway were strengthened to enhance yeast tolerance to stress due to elevated reactive oxygen species (ROS) and acetic acid. Dynamic feedback regulation of the anti-stress genetic circuits was achieved using stress-driven promoters discovered from the transcriptome to maintain low intracellular ROS, relieve the metabolic burden, and ultimately improve the robustness and ethanol production of yeast. The cell growth, xylose utilization and ethanol production of the engineered strain were enhanced under both stress and nonstress conditions. The engineered strain showed 49.5% and 17.5% higher ethanol productivity in laboratory media and industrial lignocellulosic media, respectively, at 36 °C compared with the parent strain. This study provides novel insights on the rational design and construction of feedback genetic circuits for dynamically improving yeast robustness.

A Double-Layered Microneedle Platform Fabricated through Frozen Spray-Coating.

This work reports a frozen spray-coating method for the fabrication of double-layered microneedles (MNs). Taking swellable methacrylated hyaluronic acid (MeHA)-derived MNs as the model, both hydrophobic molecules (Nile red, Cy5) and hydrophilic ones (FITC, FITC-Dextran, Insulin) can be homogeneously coated without impacting the mechanical properties of the original MeHA MNs. The prepared double-layered MNs can execute multiple roles. It is demonstrated that insulin-coated MeHA double-layered MNs allow the effective delivery of the insulin into circulation of mice for controlling the blood glucose level while they also permit the extraction of skin interstitial fluid for the timely analysis of the biomarker (glucose).

Omics Analysis Reveals the Mechanism of Enhanced Recombinant Protein Production Under Simulated Microgravity.

Simulated microgravity (SMG) is regarded as a suitable environment to produce recombinant proteins. This study showed that ß-glucuronidase expressing Escherichia coli had higher productivity of recombinant protein and higher plasmid copy number under SMG compared with the normal gravity condition. The cellular changes were analyzed at both transcriptomic and proteomic levels. The upregulation of a group of ribosome/RNA polymerase genes and a cluster of genes involving energy metabolism at transcriptomic level stood out for improved production of recombinant protein under SMG. The protein folding modulators such as chaperones were upregulated at proteomic level, which could be a result of the increased activity of protein synthesis and can help recombinant protein production. Protein export was also strengthened, which was revealed at both transcriptomic and proteomic levels. The results demonstrated that SMG is a favorable environment for recombinant protein production arousing the upregulation of protein synthesis, protein folding, and protein export.

A comparative study of the effects of microbial agents and anaerobic sludge on microalgal biotransformation into organic fertilizer.

The effects of Lactobacillus bulgaricus, Rhodopseudomonas palustris, Issatchenkia orientalis and anaerobic sludge on anaerobic digestion of microalgae to organic fertilizer were studied. High-throughput sequencing was used to analyze characteristics of microbial community structure during anaerobic digestion of microalgae using different inocula. Lactobacillales and Saccharomycetales were more likely to become dominant bacteria and eukaryotes. The relative abundance of Lactobacillales was 98.15%, 88.61% and 81.73% of total bacteria at the beginning, middle and end of the experiment, respectively. Meanwhile, the relative abundance of Saccharomycetales was 90.91%, 98.41% and 98.8% of eukaryotes at the beginning, middle and end of the experiment, respectively. At the end of digestion, the microcystin content in the reactor inoculated with Issatchenkia orientalis decreased to 0.71 µg/kg, which met drinking water standards. Rhodopseudomonas palustris did not become a dominant microorganism and had the most negative impact on the atmosphere. Volatile organic compounds were 11.92 mg/kg while the odor concentration reached 97,724 ou/m3. The organic matter content in reactors inoculated with specific groups of microbial agents, which was higher than the standard required for bio-organic fertilizer, occupying over 96% dry weight. In addition, the effective microorganism counts of Issatchenkia orientalis and Lactobacillus bulgaricus in fermentation products reached 1.8E+09 colony-forming units (cfu)/g and 1.6E+09 cfu/g, respectively, which are suitable values for microbial fertilizer.

The preparations of novel cellulose/phenylboronic acid composite intelligent bio-hydrogel and its glucose, pH-responsive behaviors.

Novel intelligent cellulose/4-vinyl-phenylboronic acid (VPBA) composite bio-hydrogels with glucose and pH-responsiveness were successfully prepared via electron beam irradiation technology at room temperature. The composites were characterized by Fourier transform infrared spectrum (FT-IR) and X-ray photoelectron spectroscopy (XPS). The electron beam irradiation results in the appearance of carbonyl in the polymerization of 4-ethenyl-phenylboronic acid, grafting and cross linking reaction in composites, and a novel composite hydrogel was formed between the poly-4-ethenyl-phenylboronic acid and cellulose matrix. By means of the incorporation of phenylboronic acid groups, the composite hydrogels with pH and glucose responsive properties was produced, and glucose responsive properties were investigated by the self-regulation of insulin release of composite hydrogel through a serial glucose solution with different concentrations, which is having great potential applications in many fields.