High-performance, large-area flexible SERS substrates prepared by reactive ion etching for molecular detection.

In the realm of molecular detection, the surface-enhanced Raman scattering (SERS) technique has garnered increasing attention due to its rapid detection, high sensitivity, and non-destructive characteristics. However, conventional rigid SERS substrates are either costly to fabricate and challenging to prepare over a large area, or they exhibit poor uniformity and repeatability, making them unsuitable for inspecting curved object surfaces. In this work, we present a flexible SERS substrate with high sensitivity as well as good uniformity and repeatability. First, the flexible polydimethylsiloxane (PDMS) substrate is manually formulated and cured. SiO2/Ag layer on the substrate can be obtained in a single process by using ion beam sputtering. Then, reactive ion etching is used to etch the upper SiO2layer of the film, which directly leads to the desired densely packed nanostructure. Finally, a layer of precious metal is deposited on the densely packed nanostructure by thermal evaporation. In our proposed system, the densely packed nanostructure obtained by etching the SiO2layer directly determines the SERS ability of the substrate. The bottom layer of silver mirror can reflect the penetrative incident light, the spacer layer of SiO2and the top layer of silver thin film can further localize the light in the system, which can realize the excellent absorption of Raman laser light, thus enhancing SERS ability. In the tests, the prepared substrates show excellent SERS performance in detecting crystalline violet with a detection limit of 10-11M. The development of this SERS substrate is anticipated to offer a highly effective and convenient method for molecular substance detection.

Evaluation of the Inhibition Potency of Nirmatrelvir against Main Protease Mutants of SARS-CoV-2 Variants.

SARS-CoV-2 continues to pose a threat to public health. Main protease (Mpro) is one of the most lucrative drug targets for developing specific antivirals against SARS-CoV-2 infection. By targeting Mpro, peptidomimetic nirmatrelvir is able to inhibit viral replication of SARS-CoV-2 and reduce the risk for progression to severe COVID-19. However, multiple mutations in the gene encoding Mpro of emerging SARS-CoV-2 variants raise a concern of drug resistance. In the present study, we expressed 16 previously reported SARS-CoV-2 Mpro mutants (G15S, T25I, T45I, S46F, S46P, D48N, M49I, L50F, L89F, K90R, P132H, N142S, V186F, R188K, T190I, and A191V). We evaluated the inhibition potency of nirmatrelvir against these Mpro mutants and solved the crystal structures of representative Mpro mutants of SARS-CoV-2 bound to nirmatrelvir. Enzymatic inhibition assays revealed that these Mpro variants remain susceptible to nirmatrelvir as the wildtype. Detailed analysis and structural comparison provided the inhibition mechanism of Mpro mutants by nirmatrelvir. These results informed the ongoing genomic surveillance of drug resistance of emerging SARS-CoV-2 variants to nirmatrelvir and facilitate the development of next-generation anticoronavirus drugs.

Structural basis of main proteases of HCoV-229E bound to inhibitor PF-07304814 and PF-07321332.

PF-07321332 and PF-07304814, inhibitors against SARS-CoV-2 developed by Pfizer, exhibit broad-spectrum inhibitory activity against the main protease (Mpro) from various coronaviruses. Structures of PF-07321332 or PF-07304814 in complex with Mpros of various coronaviruses reveal their inhibitory mechanisms against different Mpros. However, the structural information on the lower pathogenic coronavirus Mpro with PF-07321332 or PF-07304814 is currently scarce, which hinders our comprehensive understanding of the inhibitory mechanisms of these two inhibitors. Meanwhile, given that some immunocompromised individuals are still affected by low pathogenic coronaviruses, we determined the structures of lower pathogenic coronavirus HCoV-229E Mpro with PF-07321332 and PF-07304814, respectively, and analyzed and defined in detail the structural basis for the inhibition of HCoV-229E Mpro by both inhibitors. Further, we compared the crystal structures of multiple coronavirus Mpro complexes with PF-07321332 or PF-07304814 to illustrate the differences in the interaction of Mpros, and found that the inhibition mechanism of lower pathogenic coronavirus Mpro was more similar to that of moderately pathogenic coronaviruses. Our structural studies provide new insights into drug development for low pathogenic coronavirus Mpro, and provide theoretical basis for further optimization of both inhibitors to contain potential future coronaviruses.

Hot Spot Engineering in Hierarchical Plasmonic Nanostructures.

The controllable nanogap structures offer an effective way to obtain strong and tunable localized surface plasmon resonance (LSPR). A novel hierarchical plasmonic nanostructure (HPN) is created by incorporating a rotating coordinate system into colloidal lithography. In this nanostructure, the hot spot density is increased drastically by the long-range ordered morphology with discrete metal islands filled in the structural units. Based on the Volmer-Weber growth theory, the precise HPN growth model is established, which guides the hot spot engineering for improved LSPR tunability and strong field enhancement. The hot spot engineering strategy is examined by the application of HPNs as the surface-enhanced Raman spectroscopy (SERS) substrate. It is universally suitable for various SERS characterization excited at different wavelengths. Based on the HPN and hot spot engineering strategy, single-molecule level detection and long-range mapping can be realized simultaneously. In that sense, it offers a great platform and guides the future design for various LSPR applications like surface-enhanced spectra, biosensing, and photocatalysis.

Structural Basis of the Main Proteases of Coronavirus Bound to Drug Candidate PF-07321332.

The high mutation rate of COVID-19 and the prevalence of multiple variants strongly support the need for pharmacological options to complement vaccine strategies. One region that appears highly conserved among different genera of coronaviruses is the substrate-binding site of the main protease (Mpro or 3CLpro), making it an attractive target for the development of broad-spectrum drugs for multiple coronaviruses. PF-07321332, developed by Pfizer, is the first orally administered inhibitor targeting the main protease of SARS-CoV-2, which also has shown potency against other coronaviruses. Here, we report three crystal structures of the main protease of SARS-CoV-2, SARS-CoV, and Middle East respiratory syndrome (MERS)-CoV bound to the inhibitor PF-07321332. The structures reveal a ligand-binding site that is conserved among SARS-CoV-2, SARS-CoV, and MERS-CoV, providing insights into the mechanism of inhibition of viral replication. The long and narrow cavity in the cleft between domains I and II of the main protease harbors multiple inhibitor-binding sites, where PF-07321332 occupies subsites S1, S2, and S4 and appears more restricted than other inhibitors. A detailed analysis of these structures illuminated key structural determinants essential for inhibition and elucidated the binding mode of action of the main proteases from different coronaviruses. Given the importance of the main protease for the treatment of SARS-CoV-2 infection, insights derived from this study should accelerate the design of safer and more effective antivirals. IMPORTANCE The current pandemic of multiple variants has created an urgent need for effective inhibitors of SARS-CoV-2 to complement vaccine strategies. PF-07321332, developed by Pfizer, is the first orally administered coronavirus-specific main protease inhibitor approved by the FDA. We solved the crystal structures of the main protease of SARS-CoV-2, SARS-CoV, and MERS-CoV that bound to the PF-07321332, suggesting PF-07321332 is a broad-spectrum inhibitor for coronaviruses. Structures of the main protease inhibitor complexes present an opportunity to discover safer and more effective inhibitors for COVID-19.

Quantum Complementarity Approach to Device-Independent Security.

Complementarity is an essential feature of quantum mechanics. The preparation of an eigenstate of one observable implies complete randomness in its complementary observable. In quantum cryptography, complementarity allows us to formulate security analyses in terms of phase-error correction. However, the concept becomes much subtler in the device-independent regime that offers security without device characterization. Security proofs of device-independent quantum cryptography tasks are often complex and quite different from those of their more standard device-dependent cousins. The existing proofs pose huge challenges to experiments, among which large data-size requirement is a crux. Here, we show the complementarity security origin of the device-independent tasks. By linking complementarity with quantum nonlocality, we recast the device-independent scheme into a quantum error correction protocol. Going beyond the identical-and-independent-distribution case, we consider the most general attack. We generalize the sample entropy in classical Shannon theory for the finite-size analysis. Our method exhibits good finite-size performance and brings the device-independent scheme to a more practical regime. Applying it to the data in a recent ion-trap-based device-independent quantum key distribution experiment, one could reduce the requirement on data size to less than a third. Furthermore, the operational meaning of complementarity naturally extends device-independent scenarios to advantage key distillation, easing experiments by tolerating higher loss and lower transmittance.

Experimental Mode-Pairing Measurement-Device-Independent Quantum Key Distribution without Global Phase Locking.

In the past two decades, quantum key distribution networks based on telecom fibers have been implemented on metropolitan and intercity scales. One of the bottlenecks lies in the exponential decay of the key rate with respect to the transmission distance. Recently proposed schemes mainly focus on achieving longer distances by creating a long-arm single-photon interferometer over two communication parties. Despite their advantageous performance over long communication distances, the requirement of phase locking between two remote lasers is technically challenging. By adopting the recently proposed mode-pairing idea, we realize high-performance quantum key distribution without global phase locking. Using two independent off-the-shelf lasers, we show a quadratic key-rate improvement over the conventional measurement-device-independent schemes in the regime of metropolitan and intercity distances. For longer distances, we also boost the key rate performance by 3 orders of magnitude via 304 km commercial fiber and 407 km ultralow-loss fiber. We expect this ready-to-implement high-performance scheme to be widely used in future intercity quantum communication networks.

Crystal structures of main proteases of SARS-CoV-2 variants bound to a benzothiazole-based inhibitor.

Main protease (M pro) serves as an indispensable factor in the life cycle of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) as well as its constantly emerging variants and is therefore considered an attractive target for antiviral drug development. Benzothiazole-based inhibitors targeting M pro have recently been investigated by several groups and proven to be promising leads for coronaviral drug development. In the present study, we determine the crystal structures of a benzothiazole-based inhibitor, YH-53, bound to M pro mutants from SARS-CoV-2 variants of concern (VOCs) or variants of interest (VOIs), including K90R (Beta, B.1.351), G15S (Lambda, C.37), Y54C (Delta, AY.4), M49I (Omicron, BA.5) and P132H (Omicron, B.1.1.529). The structures show that the benzothiazole group in YH-53 forms a C-S covalent bond with the sulfur atom of catalytic residue Cys145 in SARS-CoV-2 M pro mutants. Structural analysis reveals the key molecular determinants necessary for interaction and illustrates the binding mode of YH-53 to these mutant M pros. In conclusion, structural insights from this study offer more information to develop benzothiazole-based drugs that are broader spectrum, more effective and safer.

SERS detection of foodborne pathogens in beverage with Au nanostars.

The aim of this study is to develop a simple but rapid method for the determination of foodborne pathogens in complex matrices (beverages) by surface enhanced Raman spectroscopy (SERS) combined with Au nanostar solid-phase substrates. The star-shaped singlet Au nanostructure was formed on the surface of a stainless steel sheet by chemical replacement reaction. Rhodamine 6G verified the sensitivity and reproducibility of this substrate, and the relative standard deviations of the SERS intensity at 1312 cm-1, 1364 cm-1, and 1510 cm-1 displacements were 3.40%, 5.64%, and 3.48%, respectively. By detecting four pathogens in beverage samples on Au nanostar substrates, the utility of the SERS assay was demonstrated, while the combination of principal component analysis (PCA) and hierarchical cluster analysis (HCA) further enabled the isolation and identification of pathogens. The results of spiked beverages were validated in conventional culture identification and Vitek 2 Compact biochemical identification system experiments. Thus, this research demonstrated that Au nanostar substrates can be effectively utilized for the recognition of pathogenic bacteria and have immense promise to advance the progress of quick detection of foodborne pathogens and food safety.

Lack of social touch alters anxiety-like and social behaviors in male mice.

The importance of social interactions has been reported in a variety of animal species. In human and rodent models, social isolation is known to alter social behaviors and change anxiety or depression levels. During the coronavirus pandemic, although people could communicate with each other through other sensory cues, social touch was mostly prohibited under different levels of physical distancing policies. These social restrictions inspired us to explore the necessity of physical contact, which has rarely been investigated in previous studies on mouse social interactions. We first conducted a long-term observation to show that pair-housed mice in a standard laboratory cage spent nearly half the day in direct physical contact with each other. Furthermore, we designed a split-housing condition to demonstrate that even with free access to visual, auditory, and olfactory social signals, the lack of social touch significantly increased anxiety-like behaviors and changed social behaviors. There were correspondingly higher levels of the pro-inflammatory cytokine interleukin-6 in the hippocampus in mice with no access to physical contact. Our study demonstrated the necessity of social touch for the maintenance of mental health in mice and could have important implications for human social interactions.

Suspended 3D metallic dimers with sub-10 nm gap for high-sensitive SERS detection.

The suspended metallic nanostructures with tiny gaps have certain advantages in surface-enhanced Raman scattering (SERS) due to the coaction of the tiny metallic nanogaps and the substrate-decoupled electromagnetism resonant modes. In this study, we used the lithographic HSQ/PMMA electron-beam bilayer resist exposure combined with a deposition-induced nanogap-narrowing process to define elevated suspended metallic nanodimers with tiny gaps for surface-enhanced Raman spectroscopy detection. By adjusting the deposited metal thickness, the metallic dimers with sub-10 nm gaps can be reliably obtained. These dimers with tunable nanogaps successfully served as excellent SERS substrates, exhibiting remarkable high-sensitivity detection ability for crystal violet molecules. Systematic experiments and simulations were conducted to explain the origin of the improved SERS performance. The results showed that the 3D elevated suspended metallic dimers could achieve a higher SERS enhancement factor than the metallic dimers on HSQ pillars and a common Si substrate, demonstrating that this kind of suspended metallic dimer is a promising route for high-sensitive SERS detection and other plasmonic applications.

Biologically Safe, Versatile, and Smart Bismuthene Functionalized with a Drug Delivery System Based on Red Phosphorus Quantum Dots for Cancer Theranostics.

Two-dimensional nanomaterials are attracting attention for cancer therapy. However, high toxicity, insensitivity to external stimuli and single therapeutic modality are still key issues hindering their clinical application. Therefore, the construction of a safe, intelligent and versatile nanocomposite is needed to meet clinical expectations. Herein, we developed a nanocomposite of Bi@RP-PEG-DOX with 2D bismuthene loaded with 0D red phosphorus quantum dots and DOX. The nanocomposite with DOX loading capacity (ca. 250 %) and photothermal conversion efficiency (ca. 54 %) showed both photothermal and photodynamic effects and a sensitive response of drug release to the acidic tumor microenvironment or NIR II laser irradiation. The nanocomposite exhibits good biosafety. Through the X-ray attenuation properties of bismuth, the nanocomposite serves as an excellent CT contrast agent, providing potential to perform CT-guided therapy.

Npa3 interacts with Gpn3 and assembly factor Rba50 for RNA polymerase II biogenesis.

RNA polymerase II is one of the most vital macromolecular complexes in eukaryotes and the assembly of such complete enzyme requires many factors. Three members of GPN-loop GTPase family Npa3/Gpn1, Gpn2, and Gpn3 participate in the biogenesis of RNA polymerase II with nonredundant roles. We show here that rapid degradation of each GPN protein in yeast leads to cytoplasmic accumulation of Rpb1 and defects in the assembly of RNA polymerase II, suggesting conserved functions of GPN paralogs for RNA polymerase II biogenesis as in humans. Taking advantage of a multicopy genetic screening, we identified GPN3 and assembly factor RBA50 among others as strong suppressors of npa3ts mutants. We further demonstrated that Npa3 interacts with Gpn3 and Rba50, similarly human Gpn1 physically interacts with Gpn3 and RPAP1 (human analog of Rba50). Moreover, a mutual dependency of protein levels of Npa3 and Gpn3 was also clearly presented in yeast using an auxin-inducible degron (AID) system. Interestingly, Rpb2, the second largest subunit of RNA polymerase II was determined to be the subunit that interacts with both Gpn1 and Rba50, indicating a close association of Npa3 and Rba50 in Rpb2 subcomplex assembly. Based on these results, we conclude that Npa3 interacts with Gpn3 and Rba50, for RNA polymerase II biogenesis. We therefore propose that multiple factors may coordinate through conserved regulatory mechanisms in the assembly of RNA polymerase complex.

Fabrication of single-nanometer metallic gaps via spontaneous nanoscale dewetting.

Ultrasmall metallic nanogaps are of great significance for wide applications in various nanodevices. However, it is challenging to fabricate ultrasmall metallic nanogaps by using common lithographic methods due to the limited resolution. In this work, we establish an effective approach for successful formation of ultrasmall metallic nanogaps based on the spontaneous nanoscale dewetting effect during metal deposition. By varying the initial opening size of the exposed resist template, the influence of dewetting behavior could be adjusted and tiny metallic nanogaps can be obtained. We demonstrate that this method is effective to fabricate diverse sub-10 nm gaps in silver nanostructures. Based on this fabrication concept, even sub-5 nm metallic gaps were obtained. SERS measurements were performed to show the molecular detection capability of the fabricated Ag nanogaps. This approach is a promising candidate for sub-10 nm metallic gaps fabrication, thus possessing potential applications in nanoelectronics, nanoplasmonics, and nano-optoelectronics.

Single-Copies Estimation of Entanglement Negativity.

Entanglement plays a central role in quantum information processing and quantum physics. However, few effective ways are known to detect the amount of entanglement of an unknown quantum state. Here, we propose a scheme to estimate the entanglement negativity for any bipartition of a composite system. The proposed scheme is based on the random unitary evolution and local measurements on a single-copy quantum state, which is more practical compared to former methods based on collective measurements on many copies of the identical state. Meanwhile, we generalize the scheme to quantify the total correlation. We demonstrate the efficiency of the scheme with statistical analyses and numerical simulations. Our scheme is quite suitable for state-of-the-art quantum platforms, which can serve as a useful benchmarking tool to advance quantum technologies and a probe to study fundamental quantum physics like entanglement dynamics.

Lower levels of plasma NT-proBNP are associated with higher prevalence of NASH in patients with biopsy-proven NAFLD.

BACKGROUND AND AIMS: Emerging evidence suggests that plasma N-terminal pro-brain natriuretic peptide (NT-proBNP) levels are decreased in patients with imaging-defined nonalcoholic fatty liver disease (NAFLD), but no data are currently available on the association between plasma NT-proBNP levels and the histological severity of NAFLD. METHODS AND RESULTS: We enrolled 351 (73.5% men) consecutive adult patients with biopsy-proven NAFLD without a prior history of cardiovascular disease (CVD). Plasma NT-proBNP levels were measured using a commercially available immunochemical system (VITROS® 5600, Johnson, New Jersey). Fifty-three percent of these subjects had nonalcoholic steatohepatitis (NASH). After stratification of patients by plasma NT-proBNP tertiles; compared to those in the 1st tertile (NT-proBNP ≤16 pg/ml), the odds ratio for NASH was 0.52 (95% CI 0.29-0.95) in patients in the 2nd tertile (NT-proBNP of 17-33 pg/ml) and 0.49 (95% CI 0.26-0.93) in those in the 3rd tertile (NT-proBNP ≥34 pg/ml) of plasma NT-proBNP levels, even after adjustment for age, sex, body mass index, homeostasis model assessment (HOMA)-estimated insulin resistance, pre-existing diabetes, hypertension, and dyslipidemia. CONCLUSION: In subjects with biopsy-proven NAFLD without known CVD, this cross-sectional study shows for the first time, that lower plasma NT-proBNP levels are strongly associated with a higher prevalence of NASH.

An optimized yeast display strategy for efficient scFv antibody selection using ribosomal skipping system and thermo resistant yeast.

OBJECTIVES: Establish a method to restrict unexpected fragments including stop codons in scFv library and generate a thermo resistant strain for screening of thermal stable scFv sequences. RESULTS: Here, we have constructed a T2A-Leu2 system for selection of yeast surface display libraries that blocks amplification of "stop codon" plasmids within the library, thereby increasing the quality of the library and efficiency of the selection screen. Also, we generated a temperature-resistant yeast strain, TR1, and validated its combined use with T2A-Leu2 for efficient screening. Thus, we developed a general approach for a fast and efficient screening of scFv libraries using a ribosomal skipping system and thermo-resistant yeast. CONCLUSIONS: The method highlights the utility of the T2A-Leu2-based ribosomal skipping strategy for increasing the quality of the input library for selection, along with an optimized selection protocol based on thermo-resistant yeast cells.

Effect of chronic toxicity of the crystalline forms of TiO2 nanoparticles on the physiological parameters of Daphnia magna with a focus on index correlation analysis.

The widespread use of titanium dioxide nanoparticles (NPs) and their inevitable release into aquatic environments have caused great concerns about their ecotoxicity. However, the chronic toxicity to TiO2 NPs of aquatic organisms has not been fully understood. In particular, research is lacking on the influence of the crystalline forms of TiO2 NPs on their mechanisms of toxicity. This study investigated the chronic toxicity (i.e., 21-day toxicity tests) of 5 types of TiO2 NPs with various percentages of crystalline forms on Daphnia magna. Results revealed that the crystalline form composed of 80% anatase and 20% rutile (i.e., the M1 form) had the highest energy band gap (i.e., Eg, the energy interval between the valence band edge and the conduction band edge) and caused maximal D. magna mortality compared with other crystalline forms. The crystalline form comprising 100% rutile (i.e., the R-S form) had the lowest Eg and exhibited a minimal effect on the physiological parameters of D. magna. Moreover, in a suitable environment without TiO2 NPs, D. magna progenies could recover to a normal physiological level (e.g., the mortalities of D. magna progenies were lower than those of parental D. magna that were exposed to TiO2 NPs at a concentration of 0.5 mg/L). Correlation analysis revealed that the body length, time of first brood, and number of neonates in the first brood of D. magna were negatively correlated with titanium accumulation in vivo. Furthermore, the indices of Ti accumulation and the product of Eg and Ti accumulation (i.e., Eg × Ti accumulation) were positively correlated (p < 0.05) with D. magna mortality, thus indicating that crystalline forms with a high Eg may cause severe toxicity to aquatic organisms at the same TiO2 bioaccumulation level.

Emission of odorous volatile organic compounds from a municipal manure treatment plant and their removal using a biotrickling filter.

Odorous volatile organic compounds (VOCs) from municipal manure treatment facilities are considered as a major nuisance issue for operators and nearby residents. In this study, up to 71 odorous VOCs were detected by gas chromatography-mass spectrometry at the manure treatment plant. These compounds can be classified into five different categories, including alkanes, olefins, aromatics, volatile organosulphur compounds and terpenes. Toluene, dimethyl disulphide, dimethyl sulphide, xylene and ethylbenzene were the five most abundant pollutants. A pilot-scale biotrickling filter (BTF) was employed to treat the complex odorous gases. Correlation analysis showed that the removal efficiency (RE) of the BTF was related with the molecular weight and chemical structure of contaminants. Higher than 85% of REs could be reached for aromatic, terpenes and most alkanes compounds after 180 days of operation. Comparatively, most olefins and partial alkanes compounds with a molecular weight lower than 70 were not removed easily. The REs of these compounds ranged from 0% to 94%, and the average removal efficiency (RE) was only about 33.3%.

[Research status and development trends of the heart valve mechanical properties].

The study of mechanical properties on heart valves can provide an important theoretical basis for doctors to repair heart valves and prosthetic valve materials research. In this paper, we present the current status of the mechanical property study methods of heart valve, expound the methods and special requirements about uniaxial tensile test and biaxial tensile test of the heart valve, and further discuss several establishment methods of heart valve constitutive models. We also discuss the development trend of heart valve mechanics.