In silico development of a novel anti-mutation, multi-epitope mRNA vaccine against MPXV variants of emerging lineage and sub-lineages by using immunoinformatics approaches.

Over the past year, an unexpected surge in human monkeypox (hMPX) cases has been observed. This outbreak differs from previous ones, displaying distinct epidemiological characteristics and transmission patterns, believed to be influenced by a newly emerging monkeypox virus (MPXV) lineage. Notably, this emerging MPXV lineage has exhibited several non-synonymous mutations, some of which are linked to immunomodulatory activities and antigenic characteristics that aid in host detection. However, specific treatments or vaccines for human monkeypox are currently lacking. Hence, we aim to develop a multi-epitope mRNA vaccine by using immunoinformatics approaches against the MPXV, particularly its emerging variants. Six proteins (A29L, A35R, B6R, M1R, H3L, and E8L) were chosen for epitope and mutation site identification. Seventeen top-performing epitopes and eight epitopes containing mutation sites were selected and combined with adjuvants, the PADRE sequence, and linkers for vaccine development. The molecular and physical properties of the designed vaccine (WLmpx) were favorable. Immunological characteristics of WLmpx were assessed through molecular docking, molecular dynamics (MD) simulations, and immune simulations. Finally, the vaccine sequence was utilized to formulate an mRNA-based vaccine. The informatics-based predicted results indicated that the designed vaccine exhibits significant potential in eliciting high-level humoral and cellular immune responses, but further validation through in vivo and vitro studies is warranted.Communicated by Ramaswamy H. Sarma.

Short-Term Safety Evaluation of Albumin-Bound Paclitaxel in Intraoperative and Postoperative Hyperthermic Intraperitoneal Chemotherapy for Gastric Cancer.

PURPOSE: To evaluate the short-term safety of albumin-bound paclitaxel in hyperthermic intraperitoneal chemotherapy (HIPEC) during and after gastric cancer (GC) surgery. METHODS: A retrospective analysis of clinical data was conducted for GC surgery patients at Zhongnan Hospital of Wuhan University, from January 2020 to September 2022. The study group (n = 120) received HIPEC and the control group (n = 268) did not receive albumin-bound paclitaxel. Short-term safety indicators including intraoperative complications, hematological toxicity, liver and kidney function, and gastrointestinal function recovery were compared between the two groups. RESULTS: There were no statistically significant differences between the two groups regarding intraoperative complications, hematological toxicity, liver and kidney function, and gastrointestinal function recovery time (P > 0.05 for all). In the study group, patients were further divided into subgroups based on dose and timing. Subgroup analysis revealed no significant differences among the different dose subgroups. However, when focusing on timing subgroups, the postoperative subgroup exhibited significantly higher white blood cell counts and bilirubin levels compared to the intraoperative subgroup, while the intraoperative subgroup had significantly higher bilirubin levels compared to both postoperative and intraoperative plus postoperative subgroups. CONCLUSION: Albumin-bound paclitaxel demonstrates good safety and tolerability in HIPEC during and after GC surgery, without increasing the risk of intraoperative complications.

Ligand-Controlled Cobalt-Catalyzed Regio-, Enantio-, and Diastereoselective Oxyheterocyclic Alkene Hydroalkylation.

Metal-hydride-catalyzed alkene hydroalkylation has been developed as an efficient method for C(sp3)-C(sp3) coupling with broad substrate availability and high functional group compatibility. However, auxiliary groups, a conjugated group or a chelation-directing group, are commonly required to attain high regio- and enantioselectivities. Herein, we reported a ligand-controlled cobalt-hydride-catalyzed regio-, enantio-, and diastereoselective oxyheterocyclic alkene hydroalkylation without chelation-directing groups. This reaction enables the hydroalkylation of conjugated and unconjugated oxyheterocyclic alkenes to deliver C2- or C3-alkylated tetrahydrofuran or tetrahydropyran in uniformly good yields and with high regio- and enantioselectivities. In addition, hydroalkylation of C2-substituted 2,5-dihydrofuran resulted in the simultaneous construction of 1,3-distereocenters, providing convenient access to polysubstituted tetrahydrofuran with multiple enantioenriched C(sp3) centers.

Effects of hydroxy methionine zinc on growth performance, immune response, antioxidant capacity, and intestinal microbiota of red claw crayfish (Procambarus clarkii).

This study aimed to evaluate the effects of varying zinc (Zn) levels on the growth performance, non-specific immune response, antioxidant capacity, and intestinal microbiota of red claw crayfish (Procambarus clarkii (P. clarkii)). Adopting hydroxy methionine zinc (Zn-MHA) as the Zn source, 180 healthy crayfish with an initial body mass of 6.50 ± 0.05 g were randomly divided into the following five groups: X1 (control group) and groups X2, X3, X4, and X5, which were fed the basal feed supplemented with Zn-MHA with 0, 15, 30, 60, and 90 mg kg-1, respectively. The results indicated that following the addition of various concentrations of Zn-MHA to the diet, the following was observed: Specific growth rate (SGR), weight gain rate (WGR), total protein (TP), total cholesterol (TC), the activities of alkaline phosphatase (AKP), phenoloxidase (PO), total antioxidant capacity (T-AOC), total superoxide dismutase (T-SOD) and catalase (CAT), the expression of CTL, GPX, and CuZn-SOD genes demonstrated a trend of rising and then declining-with a maximum value in group X4-which was significantly higher than that in group X1 (P < 0.05). Zn deposition in the intestine and hepatopancreas, the activity of GSH-PX, and the expression of GSH-PX were increased, exhibiting the highest value in group X5. The malonaldehyde (MDA) content was significantly reduced, with the lowest value in group X4, and the MDA content of the Zn-MHA addition groups were significantly lower than the control group (P < 0.05). In the analysis of the intestinal microbiota of P. clarkii, the number of operational taxonomic units in group X4 was the highest, and the richness and diversity indexes of groups X3 and X4 were significantly higher than those in group X1 (P < 0.05). Meanwhile, the dietary addition of Zn-MHA decreased and increased the relative abundance of Proteobacteria and Tenericutes, respectively. These findings indicate that supplementation of dietary Zn-MHA at an optimum dose of 60 mg kg-1 may effectively improve growth performance, immune response, antioxidant capacity, and intestinal microbiota richness and species diversity in crayfish.

Chemical characterization and source identification of PM2.5 in Luoyang after the clean air actions.

Luoyang is a typical heavy industrial city in China, with a coal-dominated energy structure and serious air pollution. Following the implementation of the clean air actions, the physicochemical characteristics and sources of PM2.5 have changed. A comprehensive study of PM2.5 was conducted from October 16, 2019 to January 23, 2020 to evaluate the effectiveness of previous control measures and further to provide theory basis for more effective policies in the future. Results showed that the aerosol pollution in Luoyang in autumn and winter is still serious with the average concentration of 91.1 µg/m3, although a large reduction (46.9%) since 2014. With the contribution of nitrate increased from 12.5% to 25.1% and sulfate decreased from 16.7% to 11.2%, aerosol pollution has changed from sulfate-dominate to nitrate-dominate. High NO3-/SO42- ratio and the increasing of NO3-/SO42- ratio with the aggravation of pollution indicating vehicle exhaust playing an increasingly important role in PM2.5 pollution in Luoyang, especially in the haze processes. Secondary inorganic ions contributed significantly to the enhancement of PM2.5 during the pollution period. The high value of Cl-/Na+ and EC concentration indicate coal combustion in Luoyang is still serious. The top three contributor sources were secondary inorganic aerosols (33.3%), coal combustion (13.6%), and industrial emissions (13.4%). Close-range transport from the western and northeastern directions were more important factors in air pollution in Luoyang during the sampling period. It is necessary to strengthen the control of coal combustion and reduce vehicle emissions in future policies.

Exploring the variation of black and brown carbon during COVID-19 lockdown in megacity Wuhan and its surrounding cities, China.

Absorbing carbonaceous aerosols, i.e. black and brown carbon (BC and BrC), affected heavily on climate change, regional air quality and human health. The nationwide lockdown measures in 2020 were performed to against the COVID-19 outbreak, which could provide an important opportunity to understand their variations on light absorption, concentrations, sources and formation mechanism of carbonaceous aerosols. The BC concentration in Wuhan megacity (WH) was 1.9 µg m-3 during lockdown, which was 24% lower than those in the medium-sized cities and 26% higher than those in small city; in addition, 39% and 16-23% reductions occurred compared with the same periods in 2019 in WH and other cities, respectively. Fossil fuels from vehicles and industries were the major contributors to BC; and compared with other periods, minimum contribution (64-86%) mainly from fossil fuel to BC occurred during the lockdown in all cities. Secondary BrC (BrCsec) played a major role in the BrC light absorption, accounting for 65-77% in WH during different periods. BrCsec was promoted under high humidity, and decreased through the photobleaching of chromophores under higher Ox. Generally, the lockdown measures reduced the BC concentrations significantly; however, the variation of BrCsec was slight.

Emodin resists to Cyprinid herpesvirus 3 replication via the pathways of Nrf2/Keap1-ARE and NF-κB in the ornamental koi carp (Cyprinus carpio haematopterus).

Cyprinid herpesvirus 3 (CyHV-3) causes high mortality in carp. Emodin has been shown of the effects of antioxidant, anti-inflammatory and antiviral. In present study, we investigated the preventive effects and mechanism of emodin on CyHV-3 infection. The ornamental koi carp (Cyprinus carpio haematopterus) were intraperitoneally injected with emodin (10 mg/kg, 20 mg/kg, or 40 mg/kg). 72 h later, an intraperitoneal injection of CyHV-3 was administered, and collected the samples one week later to detect the antioxidant parameters, antioxidant genes, inflammatory genes and to perform histopathology assays. The results showed that emodin significantly suppressed CyHV-3 replication (P < 0.05), improved the koi survival rate and slowed the damage caused by CyHV-3. Emodin treatment increased the antioxidant activity and decreased the lipid peroxidation level of the koi. Compared to the CyHV-3 group, emodin treatment resulted in the same antioxidant parameters after CyHV-3 infection. Emodin treatment activated the Nuclear factorery throid 2-related factor 2/Kelch-like ECH-associated protein 1-antioxidatant response element (Nrf2/Keap1-ARE) pathway and upregulated the expression of heme oxygenase 1 (HO-1), superoxide dismutase (SOD), and catalase (CAT) in the hepatopancreas after CyHV-3 infection. Emodin activated the nuclear factor kappa-B (NF-κB) pathway and decreased the expression of interleukin-6 (IL-6), interleukin-8 (IL-8), and tumour necrosis factor-α (TNF-α) in the koi induced by CyHV-3. In conclusion, emodin treatment can suppress CyHV-3 replication and reduce the mortality of koi caused by CyHV-3. Emodin improves antioxidant function, relieves oxidative stress and inflammation cytokines via Nrf2/Keap1-ARE and NF-κB pathways, and protects against the adverse effects induced by CyHV-3.

The Co-regulation of Ethylene Biosynthesis and Ascorbate-Glutathione Cycle by Methy Jasmonate Contributes to Aroma Formation of Tomato Fruit during Postharvest Ripening.

Currently, many fruits are always harvested at the early ripening stage to reduce postharvest losses followed by 1-methylcyclopropene (1-MCP) or ethephon treatment. However, harvesting at the early ripening stage adversely affects fruit quality, especially for the aroma. Methyl jasmonate (MeJA) treatment could induce the biosynthesis of bioactive compounds and maintain postharvest fruit quality. In the present work, the contributions of MeJA to tomato fruit quality during postharvest ripening were studied. The results showed that MeJA treatment significantly promoted the accumulation of volatile organic components (VOCs) by inducing the activities of enzymes related to lipoxygenase pathway and ethylene biosynthesis, whereas 1-MCP treatment largely inhibited the accumulation of VOCs by inhibiting activities of those enzymes. Although the application of ethephon also induced activities of the above enzymes in comparison with control, no significant differences were observed between the VOCs contents of the control and ethephon-treated fruit. Further study revealed that the ethephon treatment resulted in the enhancement of electrical conductivity and malondialdehyde content. Conversely, MeJA treatment inhibited the superoxide anion radical and hydrogen peroxide by regulating the ascorbate-glutathione cycle and further inhibited the enhancement of electrical conductivity and malondialdehyde content, which might be one of the most important reasons why the VOCs contents in fruit treated with ethephon were lower than those in MeJA-treated fruit. Thus, it is considered that MeJA treatment may be an effective and promising strategy to regulate postharvest tomato fruit quality, especially for the aroma, by regulating the ascorbate-glutathione cycle and ethylene biosynthesis.

The Synergism of 1-Methylcyclopropene and Ethephon Preserves Quality of "Laiyang" Pears With Recovery of Aroma Formation After Long-Term Cold Storage.

A "Laiyang" pear is a climacteric fruit with a special taste and nutritional value but is prone to a post-harvest aroma compound loss and a loss in fruit quality. In this study, pears were pretreated with 0.5 µl L-1 1-methylcyclopropene (1-MCP) at 20°C for 12 h and then stored at 0 ± 1°C for 150 days to evaluate the influence of 1-MCP on fruit quality and the changes in components of volatile aromas. In addition, pears were further treated with 2 mmol L-1 ethephon. The effects of ethephon on the recovery of aroma production were investigated during the 150 day storage at 0 ± 1°C and the subsequent 7 day shelf life at 20 ± 1°C. Treatment with 1-MCP inhibited firmness loss, increased electrical conductivity, reduced respiration and ethylene production rates as well as the contents of soluble solids, and maintained the storage quality of the fruits. However, 1-MCP treatment inhibited the emission of volatile aromas in pear fruits by decreasing the activities of various enzymes, such as lipoxygenase (LOX), hydroperoxide lyase (HPL), alcohol dehydrogenase (ADH), pyruvate carboxylase (PDC), and alcohol acetyltransferase (AAT). During the shelf-life, activities of the above mentioned enzymes were significantly enhanced, and a higher content of volatile aromas were found in fruits treated with 1-MCP + ethephon, while other qualities were not compromised. These results showed that 1-MCP treatment could effectively maintain the quality of the "Laiyang" pear during cold storage, and the additional application of ethephon on fruits during shelf-life may be a promising way to restore volatile aromas in pear fruits after long-term storage.

SlARG2 contributes to MeJA-induced defense responses to Botrytis cinerea in tomato fruit.

BACKGROUND: Arginase plays key roles in methyl jasmonate (MeJA)-mediated quality maintenance in vegetables and fruits. MeJA treatment induced the Arginase 2 (SlARG2) expression, which is one of the most important encoding genes of arginase. In addition, the treatment with MeJA induced resistance to pathogenic infection in many plants. However, the functions of SlARG2 in MeJA-induced defense to Botrytis cinerea are unclear. In our work, control and SlARG2-silenced tomato fruits (Solanum lycopersicum) were treated with 0.05 mmoL L-1 MeJA before storage to assay the roles of SlARG2 in MeJA-induced defense responses to Botrytis cinerea. RESULTS: Our results indicated that MeJA treatment induced both pathogenesis-related gene expression (PR1, PR2a, PR2b and PR3b), and the activity of defense-related enzymes, as well as upregulated arginine metabolism. Compared to control fruits, the treatment with MeJA also induced the activity of arginase, arginine decarboxylase (ADC) and ornithine aminotransferase (OAT), and expression of SlARG2, SlADC, ornithine decarboxylase (SlODC) and SlOAT, and consequently increased the accumulation of arginine, proline, glutamate, putrescine and spermine. However, the induction effects by MeJA were significantly reduced in fruits in which SlARG2 was silenced and severe disease symptoms were observed. CONCLUSION: MeJA fumigation could inhibit disease development by inducing pathogenesis-related gene expression (PR1, PR2a, PR2b and PR3b) and defense-related enzymes activity, as well as upregulated arginine metabolism. In addition, SlARG2 silencing could inhibit the functions of MeJA in inducing the accumulation of the above substances. Overall, our study provided strong evidence that SlARG2 was essential for MeJA-induced tomato fruit defense responses to Botrytis cinerea. © 2020 Society of Chemical Industry.

CRISPR/Cas9-Mediated SlMYC2 Mutagenesis Adverse to Tomato Plant Growth and MeJA-Induced Fruit Resistance to Botrytis cinerea.

Methyl jasmonate (MeJA), a natural phytohormone, played a critical role not only in plant growth but also in plant defense response to biotic and abiotic stresses. MYC2, a basic helix-loop-helix transcription factor, is a master regulator in MeJA signaling pathway. In the present work, slmyc2 mutants were generated by the clustered regularly interspaced short palindromic repeats and associated Cas9 protein (CRISPR/Cas9) system to investigate the role of SlMYC2 in tomato plant growth and fruit disease resistance induced by exogenous MeJA. The results showed that slmyc2 mutants possessed a higher number of flowers and a lower fruit setting rate in comparison with wild-type plants. In addition, the fruit shape of slmyc2 mutant was prolate, while the control fruits were oblate. Knockout of SlMYC2 significantly decreased the activities of disease defensive and antioxidant enzymes, as well as the expression levels of pathogen-related (PR) genes (SlPR-1 and SlPR-STH2) and the key genes related to jasmonic acid (JA) biosynthesis and signaling pathway including allene oxide cyclase (SlAOC), lipoxygenase D (SlLOXD), SlMYC2, and coronatine insensitive 1 (SlCOI1), and consequently aggravated the disease symptoms. By contrast, the disease symptoms were largely reduced in MeJA-treated fruit that possessed higher activities of these enzymes and expression levels of genes. However, the induction effects of MeJA on fruit disease resistance and these enzymes' activities and genes' expressions were significantly attenuated by knockout of SlMYC2. Therefore, the results indicated that SlMYC2 played positive regulatory roles not only in the growth of tomato plants but also in MeJA-induced disease resistance and the antioxidant process in tomato fruits.

SlMYC2 are required for methyl jasmonate-induced tomato fruit resistance to Botrytis cinerea.

The mechanism of SlMYC2, involved in methyl jasmonate (MJ)-induced tomato fruit resistance to pathogens, was investigated. The data indicated that MJ treatment enhanced the accumulation of total phenolics and flavonoids, as well as individual phenolic acids and flavonoids, which might be caused by the increased phenylalanine ammonia-lyase and polyphenol oxidase activities, induced pathogenesis-related gene (PR) expression, ß-1,3-glucanase and chitinase activities, as well as α-tomatine, by inducing GLYCOALKALOID METABOLISM gene expression. These effects, induced by MJ, partly contributed to tomato fruit resistance to Botrytis cinerea. Nevertheless, the induction effects of MJ were almost counteracted by silence of SlMYC2, and the disease incidence and lesion diameter in MJ + SlMYC2-silenced fruit were higher than those in MJ-treated fruit. These observations are the first evidence that SlMYC2 plays vital roles in MJ-induced fruit resistance to Botrytis cinerea, possibly by regulating defence enzyme activities, SlPRs expression, α-tomatine, special phenolic acids and flavonoid compounds.

Generation and functional evaluation of a DNA vaccine co-expressing Cyprinid herpesvirus-3 envelope protein and carp interleukin-1 beta.

Cyprinid herpesvirus 3 (CyHV-3) infection in carp causes a fatal and highly contagious disease that results in huge economic losses in common and koi carp aquaculture worldwide. Thus the development of an effective vaccine to protect carp stocks against the CyHV3 virus is imperative. In this study, we immunized common carps with a DNA vaccine consisting of a plasmid that co-expresses the CyHV-3 envelope protein ORF25 and the carp IL-1ß gene in order to evaluate the adjuvant potential of IL-1ß. Our result shows that antibodies specific to ORF25 can be detected as early as one week after intramuscular injection of the DNA vaccine at low dosage. Moreover, the co-expression of IL-1ß can enhance the potency of the vaccine, as demonstrated by a higher antibody level after the third immunizations. Importantly, the DNA vaccine reduced mortality in carps when they were immunized prior to a CyHV-3 challenge, as compared to negative control groups. However, despite being able to induce higher neutralizing antibody titres, the co-expression of IL-1ß in the DNA vaccine did not significantly improve the overall survival of immunized fish following virus challenge. Furthermore, the DNA vaccine can protect carps from tissue damage and histopathological alteration caused by viral infection. These strongly suggests that the vaccine can efficiently elicit protective immunity against CyHV-3 infection. In conclusion, the DNA vaccine formulated with the pIRES-ORF25-IL-1ß DNA construct can protect carp against CyHV-3 infection and has potential applicability in the aquaculture industry.

Vaccination of plasmid DNA encoding ORF81 gene of CJ strains of KHV provides protection to immunized carp.

In order to construct the recombinant plasmid of pIRES-ORF81, the nucleic acid isolated from Koi herpes virus-CJ (KHV-CJ) strains was used as a template to insert the ORF81 gene fragments amplified by PCR into the pIRES-neo, a kind of eukaryotic expression vector. Using Western blotting analysis, it was verified that ORF81 gene protein can be expressed correctly by pIRES-ORF81, after MFC cells were transfected. The recombinant plasmid pIRES-ORF81 was set into three immunization dose gradients: 1, 10, and 50 µg/carp. Empty plasmid group, PBS group, and blank control group were set simultaneously. Giving intramuscular injections to healthy carps with an average body mass of 246 ± 20 g, indirect ELISA was used to regularly determine antibody levels after three times immunization injection. Neutralizing antibodies were detected by neutralization assay. The results of inoculation tests showed that the pIRES-ORF81 recombinant plasmid can induce the production of carp-specific antibodies. The differences of immune effect between the three different doses of immune gradients were not significant (P > 0.05), but they can induce the production of neutralizing antibodies. After 25 d of inoculation, carp mortality of pIRES-neo empty vector treatment groups was 85%, while the carp mortality of eukaryotic expression recombinant plasmid pIRES-ORF81 injected with three different doses of immune gradients was 20, 17.5, and 12.5%, respectively. Differences in comparison to the control group were highly significant (P < 0.01). However, histopathological section of immunohistochemistry organization revealed no significant changes. It demonstrated that the DNA vaccine pIRES-ORF81 constructed in the experiment displayed a good protective effect against KHV, which had the potential to industrial applications.

The primary culture of mirror carp snout and caudal fin tissues and the isolation of Koi herpesvirus.

The explosive Koi herpesvirus (KHV) epidemic has caused the deaths of a large number of carp and carp variants and has produced serious economic losses. The mirror carp (Cyprinus carpio var. specularis) exhibits strong environmental adaptability and its primary cells can be used to isolate KHV. This study utilized the tissue explant method to systematically investigate primary cell culture conditions for mirror carp snout and caudal fin tissues. We demonstrated that cells from these two tissue types had strong adaptability, and when cultured in Medium 199 (M199) containing 20% serum at 26 to 30°C, the cells from the snout and caudal fin tissues exhibited the fastest egress and proliferation. Inoculation of these two cell types with KHV-infected fish kidney tissues produced typical cytopathic effects; additionally, identification by electron microscopy, and PCR indicated that KHV could be isolated from both cell types.

[Analysis of Chinese syndrome features and combination laws of 2029 patients with coronary heart disease angina].

OBJECTIVE: To explore the element distribution features and syndrome combination laws of coronary heart disease angina. METHODS: The syndrome database of 2 029 patients with coronary heart disease angina was established to study the syndrome elements and syndrome combination laws. RESULTS: (1) The syndrome element distribution of coronary heart disease angina was featured as: blood stasis > qi deficiency > phlegm turbid > yin deficiency > phlegm turbid with more warm property > yang deficiency > stagnant qi > phlegm turbid with more cold > cold coagulation. Of them, qi deficiency and blood stasis were the main two syndrome elements, and phlegm turbid with more warm and yin deficiency also occupied important positions. (2) Syndrome combination laws of coronary heart disease angina: three elements syndrome and two elements syndrome were dominant. The combination of sthenia syndrome element and asthenia syndrome element was the most important combination laws. Qi deficiency and blood stasis was the main combination form. CONCLUSIONS: Qi deficiency, yin deficiency, phlegm turbid, and blood stasis form four key links of its pathogenesis, in which, qi deficiency and blood stasis was the most basic pathogenesis. The syndrome element combination had some laws.