Identification of B-cell epitopes located on the surface in the PB2 protein of the H9N2 subtype avian influenza virus.

Avian influenza (AI), caused by H9N2 subtype avian influenza virus (AIV), poses a serious threat to poultry farming and public health due to its transmissibility and pathogenicity. The PB2 protein is a major component of the viral RNA polymerase complex. It is of great importance to identify the antigenic determinants of the PB2 protein to explore the function of the PB2 protein. In this study, the PB2 sequence of H9N2 subtype AIV, from 1090 to 1689 bp, was cloned and expressed. The recombinant PB2 protein with cutting gel was used to immunize BALB/c mice. After cell fusion, the hybridoma cell lines secreting monoclonal antibodies (mAbs) targeting the PB2 protein were screened by indirect ELISA and western blotting, and the antigenic epitopes of mAbs were identified by constructing truncated overlapping fragments in the PB2 protein of H9N2 subtype AIV. The results showed that three hybridoma cell lines (4B7, 4D10, and 5H1) that stably secreted mAbs specific to the PB2 protein were screened; the heavy chain of 4B7 was IgG2α, those of 4D10 and 5H1 were IgG1, and all three mAbs had kappa light chain. Also, the minimum B-cell epitope recognized was 475LRGVRVSK482 and 528TITYSSPMMW537. Homology analysis showed that these two epitopes were conserved among the different subtypes of AIV strains and located on the surface of the PB2 protein. The above findings provide an experimental foundation for further investigation of the function of the PB2 protein and developing monoclonal antibody-based diagnostic kits.

Dual-slab Yb:KGd(WO4)2 regenerative amplifier for spectral shaping and high-power output.

A high-power regenerative amplifier (RA) based on dual-slab Yb:KGd(WO4)2 (Yb:KGW) was demonstrated, which provided a maximum average power of 33.7 W at a repetition rate of 75-200 kHz before compression with a central wavelength of 1039 nm, corresponding to an optical-to-optical conversion efficiency of 51.4%. To the best of our knowledge, this is the highest average power from the Yb:KGW solid-state RA. The compressed pulse duration of 205 fs was realized under the maximum output power. By adjusting the gain of the crystals, respectively, the spectral shaping can be achieved. A combination spectrum with root-mean-square (RMS) bandwidth of 4.5 nm was generated with a central wavelength of 1035 nm at an output power of 20 W, the compressed pulse duration was 159 fs. Meanwhile, effective mitigation of thermal effects by dual-slab configuration guaranteed the nearly diffraction-limited beam quality: M x2 = 1.17 and M y2 = 1.20.

417†W, 2.38†mJ Innoslab amplifier compressible to a high pulse quality of 406†fs.

We demonstrate a 417 W, 175 kHz Innoslab chirped pulse amplification laser compressible to short and clean 406 fs pulse duration. A spectral bandwidth (full width at half maximum, FWHM) of ∼3 nm was maintained at full pump power, and the pulses exhibited good pulse quality in a wide tunable pulse energy range from 1.7 mJ to a maximum of 2.38 mJ. At the maximum output power, the compressed pulses were nearly pedestal free. The comprehensive effects of residual high-order dispersion from the front end, the gain shaping effects of the amplifier, and the slight mismatch of third-order dispersion (TOD) between the stretcher (CFBG) and the gating compressor, along with the small nonlinear phase shift accumulated in the amplifier, could have facilitated the high pulse quality. To the best of our knowledge, this is the shortest pulse duration from the Innoslab amplifiers at hundreds of watts average power in the millijoule energy regime.

Identification of catalytically active domain epitopes in neuraminidase protein of H9N2 subtype of avian influenza virus.

H9N2 subtype of avian influenza virus (AIV) is primarily a bird virus, which is widespread in clinical avian disease, and reported in cases of human infection. As one of the surface proteins of AIV, the neuraminidase (NA) protein plays an important role mainly in viral budding. However, vaccine development and detection methods for NA of H9N2 AIVs are in urgent clinical need. In this study, a truncated NA gene (205-900 bp) was cloned from the NA sequence of H9N2 strain, and then expressed using pET-28a (+) vector. This purified recombinant NA protein was used to immunize BALB/c mice, and the monoclonal antibodies were screened through the indirect enzyme-linked immunosorbent assay (ELISA). Next, eight prokaryotic expression vectors were constructed for epitope identification. After cell fusion, three hybridoma cell lines producing the antibodies special to NA protein were screened by ELISA, western blotting, and indirect immunofluorescence; these were named 1B10, 2B6, and 5B2, respectively. Epitope scanning techniques were used to identify three B-cell epitopes recognized by these three monoclonal antibodies, 196KNATASIIYDGMLVD210, 210DSIGSWSKNIL220 and 221RTQESECVCI230. The subsequent homology analysis revealed the three epitopes were highly conserved in H9N2 AIV strains. The structural predictions of the antigenic epitopes indicated that all three epitopes were located in the catalytic region of NA. These results provide a basis for studying the function of the NA protein of H9N2 AIV and technical support for the development of a universal detection method based on anti-NA monoclonal antibodies.

Investigation into the Rheological Properties and Microstructure of Silt/Crumb Rubber Compound-Modified Asphalt.

Near the coast of China, a large amount of sediment is produced during construction work. In order to mitigate the environmental damage caused by sediment and enhance the performance of rubber-modified asphalt effectively, solidified silt material and waste rubber were prepared to modify asphalt, and its macroscopic properties, such as viscosity and chemical composition, were determined via a routine physical test, DSR, Fourier Transform Infrared Spectroscopy (FTIR), and Fluorescence Microscopy (FM). The results show that, with the increase in powder particles and the addition of a certain amount of hardened mud, the mixing and compaction temperature of modified asphalt can be significantly increased-still reaching the design standard. In addition, the high thermal stability and fatigue resistance of the modified asphalt were clearly better than those of the ordinary asphalt. From the FTIR analysis, rubber particles and hardened silt only exhibited mechanical agitation with the asphalt. Considering that excessive silt might result in the aggregation of matrix asphalt, the addition of an appropriate amount of hardened solidified silt material can eliminate the aggregation. Therefore, the performance of modified asphalt was optimum when solidified silt was added. Our research can provide an effective theoretical basis and reference values for the practical application of compound-modified asphalt. Therefore, 6%HCS(6:4)-CRMA have better performance. Compared to ordinary rubber-modified asphalt, the composite-modified asphalt binder has better physical properties and a more suitable construction temperature. The composite-modified asphalt uses discarded rubber and silt as raw materials, which can effectively protect the environment. Meanwhile, the modified asphalt has excellent rheological properties and fatigue resistance.

Single-pulse characterization of the focal spot size of X-ray free-electron lasers using coherent diffraction imaging.

The characterization of X-ray focal spots is of great significance for the diagnosis and performance optimization of focusing systems. X-ray free-electron lasers (XFELs) are the latest generation of X-ray sources with ultrahigh brilliance, ultrashort pulse duration and nearly full transverse coherence. Because each XFEL pulse is unique and has an ultrahigh peak intensity, it is difficult to characterize its focal spot size individually with full power. Herein, a method for characterizing the spot size at the focus position is proposed based on coherent diffraction imaging. A numerical simulation was conducted to verify the feasibility of the proposed method. The focal spot size of the Coherent Scattering and Imaging endstation at the Shanghai Soft X-ray Free Electron Laser Facility was characterized using the method. The full width at half-maxima of the focal spot intensity and spot size in the horizontal and vertical directions were calculated to be 2.10 ± 0.24 µm and 2.00 ± 0.20 µm, respectively. An ablation imprint on the silicon frame was used to validate the results of the proposed method.

Efficient high-power orthogonal dual-slab Yb:KGd(WO4)2 laser oscillator with a TEM00 mode.

We demonstrated a TEM00 mode orthogonal dual-slab Yb:KG(WO4)2(Yb:KGW) laser oscillator with high average power. Polarization anisotropy of thermal lenses was investigated and alleviating the astigmatism based on orthogonal dual-slab. In addition, the laser polarization was directly controlled by adjusting the net gain of the two crystals. The maximum output power was highly enhanced compared with single crystal due to effective thermal distribution. For an absorbed pump power of 52.4 W, this oscillator delivered an average power of 26.5 W, corresponding to an optical-to-optical conversion efficiency of 50.6%. Meanwhile, the ellipticity of the output laser was optimized to 0.940. Nearly diffraction-limited beam quality was measured to be M x2 = 1.19 and M y2=1.18.

Effect of astragaloside IV and salvianolic acid B on antioxidant stress and vascular endothelial protection in the treatment of atherosclerosis based on metabonomics.

Vascular endothelial cells and oxidation reduction system play an important role in the pathogenesis of atherosclerosis (AS). If these conditions are disordered, it will inevitably lead to plaque formation and even rupture. Astragaloside IV (AsIV) and salvianolic acid B (Sal B) are the main active ingredients of Astragalus membranaceus and Salvia miltiorrhiza, respectively, and found to ameliorate vascular endothelial dysfunction and protect against oxidative stress in recent studies. However, it is still unknown if the combination of AsIV and Sal B (AsIV + Sal B) can inhibit the development of plaque through amplifying the protective effect of vascular endothelial cells and anti-oxidative stress effect. To clarify the role of AsIV + Sal B in AS, we observed the efficacy of each group (Control, Model, AsIV, Sal B, and AsIV + Sal B) by biomolecular assays, such as observing the pathological morphology of the aorta by oil red O staining, evaluating the level of oxidative stress and endothelial cells in the serum by the Elisa test, and analyzing the changes of all small molecule metabolites in liver tissue by UPLC-QTOF-MS. Results showed that AsIV, Sal B and AsIV + Sal B decreased the deposition of lipid in the arterial wall, so as to exert the effect of anti-oxidant stress and vascular endothelial protection, where the inhibitory effect of AsIV + Sal B was the most obvious. Metabonomics analysis showed that Sal B regulated the metabolic pathways of arginine and proline. AsIV regulated glycerol metabolism and saturated fatty acid biosynthesis metabolism. AsIV + Sal B is mainly related to the regulation of the citrate cycle (TCA cycle), alanine, aspartic acid, and glutamate metabolism, cysteine, and methionine metabolism. Succinic acid and methionine are synergistic metabolites that exert an enhancing effect when AsIV and Sal B were used in combination. In conclusion, we demonstrated that AsIV acompanied with Sal B can be successfully used for anti-oxidative stress and vascular endothelial protection of AS, and succinic acid and methionine are the synergistic metabolites.

Piezo1 mediates endothelial atherogenic inflammatory responses via regulation of YAP/TAZ activation.

The vascular endothelium plays a key role in the pathobiology of atherosclerotic cardiovascular disease. Endothelial cell Piezo1 mediates blood vessel formation, angiogenesis and regulation of blood pressure. However, changes of Piezo1 expression in atherosclerosis (AS) and the role of Piezo1 in the progression of atherosclerotic diseases remains obscure. Thus, the current study is to elucidate the role and mechanism of which Piezo1 mediates vascular inflammation in atherosclerotic mice and vascular endothelial inflammation induced by oxidized low density lipoprotein (ox-LDL) in vitro. Here, we have shown that the expression of Piezo1 was significantly increased in the stenotic carotid artery of ApoE-/- mice fed by high-fat diet (HFD). Pharmacological inhibition of Piezo1 (GsMTx-4) attenuated plaque formation, decreased the level of inflammation related factors (JNK, TNF-α, NF-κB, VCAM-1) of carotid plaque in atherosclerotic mice. Meanwhile, ox-LDL also upregulates Piezo1 and inflammation proteins (NF-κB, JNK and TNF-α) in endothelium cells (ECs). YAP/TAZ is activated accompanied by the enhanced Piezo1 activity in ECs induced by ox-LDL. Interference by siRNA of Piezo1 abolished the expression of YAP/TAZ and inflammation proteins (JNK, NF-κB and TNF-α). In addition, Ca2+ influx in ECs induced by ox-LDL was increased than control group, Piezo1 siRNA can reduce the calcium content. Piezo1 agonist Yoda1 increased Ca2+ influx and promote YAP nucleus translocation in ECs, genetic deletion of Piezo1 reversed it. Our results indicate that Piezo1 could mediate endothelial atherogenic inflammatory responses via regulation of YAP/TAZ activation and nuclear localization. Piezo1 may be a potential therapeutic target for atherosclerotic diseases in the future.

Mitogenomic characterization and phylogeny of Scelimena melli Günther (Orthoptera: Tetrigoidea: Scelimeninae).

The mitochondrial genome (mitogenome) of Scelimena melli, which belongs to Orthoptera, Tetrigoidea, Tetrigidae, Scelimeninae was determined. The mitogenome has a length of 14,598 bp and consists of 37 genes including 13 protein-coding genes (PCGs), two rRNA genes, and 22 tRNA genes. Phylogenetic analysis using 37 mitochondrial genes with other 22 Tetrigoidea species revealed that S. melli had a closer relationship with Paragavialidium sichuanense, but the monophyly of Scelimeninae was not recovered. The mitogenome data of S. melli would provide useful resources for further evolutionary studies of Scelimeninae and Tetrigoidea species.

Author Correction: Pretreatment-Integration for Milk Protein Removal and Device-Facilitated Immunochromatographic Assay for 17 Items.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Salvianolic acid B ameliorates atherosclerosis via inhibiting YAP/TAZ/JNK signaling pathway in endothelial cells and pericytes.

Atherosclerosis (AS) is a chronic disease of the arterial wall where both innate and adaptive immunoinflammatory mechanisms are involved. Inflammation plays an important role in the pathological process of atherosclerosis at various stages. Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ, also known as WWTR1) behave as a novel drug target against atherosclerosis. Therefore, the mechanism relationship of YAP/TAZ, inflammation and AS was explored in this study. Experiments demonstrated that serine dephosphorylation and nuclear translocation of YAP was increased in ECs and pericytes induced by oxidative low-density lipoprotein (ox-LDL), while the inhibition of YAP degraded the expression of downstream inflammatory factors. The expression of YAP/TAZ and inflammation proteins (JNK, NF-κB and TNF-α) in ECs and pericytes was suppressed through the application of Sal-B. Besides, Sal-B protects ECs and pericytes from oxidative stress and apoptosis. In vivo, Sal-B reduced en face and aortic root sinus lesions size, and decreased the expression of inflammation related factors (IL-6, IL-1ß, TNF-α) and ox-LDL in serum sample of ApoE-/- mice fed a high fat diet. Therefore, our work provides a potential therapeutic strategy of using Sal-B to attenuate the development of atherosclerosis, the anti-atherosclerosis effects of Sal-B is related to regulate YAP/TAZ/JNK signaling pathway.

An anti-BSA antibody-based immunochromatographic assay for chloramphenicol and aflatoxin M1 by using carboxy-modified CdSe/ZnS core-shell nanoparticles as label.

A lateral-flow immunochromatographic assay with excellent sensitivity and wide application potential is described. The bovine serum albumin (BSA) antibody was immobilized in the test line for universality, and preincubation was introduced for high method sensitivity. Carboxy-modified CdSe/ZnS core-shell nanoparticles were used as label, and the fluorescence peaking at 605 nm was detected. The fluorescence in the test line was negative against the relevant analyte content. The chloramphenicol (CAP) and the aflatoxin M1 (AFM1) in milk were detected using the same strip to validate the universality. After optimization, the detection limit for CAP is 10 pg·mL-1, which is three times less that of a conventional assay (30 pg·mL-1). The detection limit for AFM1 was 6 pg·mL-1, which was 13 times less than that of a conventional assay (8 pg·mL-1). The method was applied in the analysis of spiked milk samples. The performance was compared with that of the commercial ELISA kit, and good agreement was observed. Graphical abstractSchematic representation of the universal and sensitive combined immunochromatographic assay (USICA) and conventional immunochromatographic assay (TICA) of chloramphenicol (CAP) and aflatoxin M1.

Ovalbumin antibody-based fluorometric immunochromatographic lateral flow assay using CdSe/ZnS quantum dot beads as label for determination of T-2 toxin.

This work describes an anti-ovalbumin antibody-based lateral flow immunoassay (LFI) for T-2 toxin. The antibody uses a coating antigen as a bifunctional element for universality and introduces preincubation to improve the detection limits of the method. T-2 toxin and ovalbumin-modified T-2 toxin competitively binds on the anti-T-2 toxin monoclonal antibody modified on CdSe/ZnS quantum dot beads during preincubation. The modified T-2 toxin acts as a bifunctional element that forms immuno complexes during preincubation and combines with anti-ovalbumin antibody coated in the test line through the ovalbumin terminal. Fluorescence is detected at 610 nm on the test zone following photoexcitation at 365 nm. It has a reverse dose-effect relationship with the amount of T-2 toxin. The calibration plot is linear in the 20-110 fg mL-1 T-2 toxin concentration range, and the limit of detection (LOD) is 10 fg mL-1, which is lower by 8-fold than that of the traditional LFI system (LOD 80 fg mL-1) and one order of magnitude than those of LFIs with labels of colloidal gold nanoparticles (LOD 150 fg mL-1) or fluorophores (LOD 190 ng mL-1). Universality was verified through aflatoxin B1 detection using the established ovalbumin antibody-based LFI system (LOD 10 fg mL-1). The performance of the method was compared with that of established systems and a commercial ELISA kit (LOD 360 fg mL-1). Graphical abstractSchematic representation of ovalbumin antibody-based immunochromatographic lateral flow assay for T-2 toxin. Preincubation is introduced for high sensitivity. T-2- anti-ovalbumin acts as a bi-functional element for universality. CdSe/ZnS quantum dot beads act as label. Fluorometric signal is detected at 610 nm.

Pretreatment-Integration for Milk Protein Removal and Device-Facilitated Immunochromatographic Assay for 17 Items.

Accurate and comprehensive immunochromatographic assay (ICA) data are urgently required in the daily supervision of plants, schools, testing institutions, and law-enforcing departments. Through pretreatment-integration and device-facilitated operation, a quantitative ICA with high sensitivity and throughput was realized on the basis of a commercialized semi-quantitative ICA strip. Three pretreatment methods, namely, acid base, heavy metal salt, and organic solvent methods, have less than three steps. The pretreatment was established for protein removal. A total of 17 pretreated ICA items in milk were considered for the identification of the most suitable pretreatment method. The items are composed of six items pretreated by the acid-base method, six by the heavy salt method, and five by the organic solvent method. Then, the ICA results with pretreatment were compared with those without pretreatment. After pretreatment, the signal intensity increased by 39%, the detection limit decreased to 12%, the half maximal inhibitory concentration decreased to 18%, and the detection range increased fourfold. A device with mixing and centrifugation functions was designed for the pretreatment-related operations. A pre-incubation sampling device was used to facilitate incubation in batch and high-throughput detection. An ICA reader was used. The detection throughput reached 8 samples per batch or 32 samples per hour. The designed devices were printed through 3D printing and rapid prototyping.

Universal and Ultrasensitive Immunochromatographic Assay by Using an Antigen as a Bifunctional Element and Antialbumin Antibody on a Test Line.

A universal and ultrasensitive immunochromatographic assay (ICA) was established using antigen as a bifunctional element and antialbumin antibody in a test line. Preincubation was introduced for competitive recognition. After optimization, the linear detection of aflatoxin M1 (AFM1) with quantum dot bead (QB)-based ICA (QB-ICA) sensor ranged from 10 to 52 pg mL-1, with a 50% inhibitory concentration (IC50) of 23 pg mL-1, which was nearly 49.6-fold lower than those of ICA on a traditional structure with traditional pretreatment (IC50 = 1.10 ng mL-1) and 10-fold lower than those of ICA on a traditional structure with acid aid pretreatment (IC50 = 0.25 ng mL-1). The limit of detection (LOD) for AFM1 was 16 pg mL-1 in milk, which was approximately 16.3-fold times higher than those of ICA on a traditional structure with traditional pretreatment and 6.3-fold higher than those of ICA on a traditional structure with acid aid pretreatment. The LOD improved by 20-fold by using the proposed structure compared to that of conventional enzyme-linked immunosorbent assay (ELISA) for AFM1-spiked milk samples (IC10 = 0.12 ng mL-1). The performance and practicability of the established QB-ICA sensor were validated with a commercial ELISA kit. To evaluate universality, we successfully detected chloramphenicol, with IC50 of 0.42 ng mL-1. Given its high sensitivity and universality, the proposed QB-ICA can be used as an alternative for rapid, sensitive, and universal quantitative detection of all small-molecule analytes.

Immunochromatographic assay for T-2 toxin based on luminescent quantum dot beads.

A quantum dot bead based immunochromatographic assay (QB-ICA) system was established for T-2 toxin (T-2), which widely occurs in agriculture and could be used as a potential biological warfare agent. After optimization, the dynamic linear detection range of T-2 calculated from a calibration curve was from 0.12 to 0.67 ng mL-1 and the limit of detection (LOD) was 0.08 ng mL-1, which is lower than those of the ICA based on colloidal gold nanoparticles or a fluorescent material or an antibody-based biochip in other reports. The performance and practicability of the established ICA system were validated with a commercial ELISA kit and the two methods were comparable. The proposed QB-ICA for T-2 could be an alternative for rapid, sensitive, and quantitative on-site detection of this toxin in biosafety monitoring in agriculture and for susceptibility testing of the potential release of this biological warfare agent.

[Prognostic analysis of different treatments for synchronous colorectal liver metastasis].

OBJECTIVE: To evaluate the effects of different treatments on the prognosis of patients with synchronous colorectal liver metastasis(CLM). METHODS: Clinicopathological data of 121 patients with synchronous CLM in The First Affiliated Hospital of Xinjiang Medical University between January 2010 and December 2014 were retrospectively analyzed. According to the metastatic lesions, patients were divided into simple operation group(22 patients, receiving operation only), simple chemotherapy group(43 patients, receiving chemotherapy only), and combination group(56 patients, receiving chemotherapy based on operation). The prognosis of patients in three groups was investigated. Univariate and multivariate analyses were performed through the long-rank test and Cox model to evaluate the related factors affecting prognosis. RESULTS: The median survival time was 10(3-39) months in simple operation group, 7(3-36) months in simple chemotherapy group, and 18 (4-66) months in combination group. The differences among groups were all statistically significant (all P<0.05). Extent of lymph node metastasis, number of liver metastatic lesion, size of liver metastatic lesion, resection edge extent of liver metastatic lesion, and treatment method were independent factors of synchronous CLM(all P<0.05). CONCLUSION: Combination therapy of a variety of treatment can prolong the survival of patients with synchronous CLM.