Transcriptome analysis of long non-coding RNA and mRNA Profiles in VSV-infected BHK-21 Cells.

BACKGROUND: Vesicular stomatitis virus (VSV) is a typical non-segmented negative-sense RNA virus of the genus Vesiculovirus in the family Rhabdoviridae. VSV can infect a wide range of animals, including humans, with oral blister epithelial lesions. VSV is an excellent model virus with a wide range of applications as a molecular tool, a vaccine vector, and an oncolytic vector. To further understand the interaction between VSV and host cells and to provide a theoretical basis for the application prospects of VSV, we analyzed the expression of host differentially expressed genes (DEGs) during VSV infection using RNA-Seq. RESULTS: Our analyses found a total of 1015 differentially expressed mRNAs and 161 differentially expressed LncRNAs in BHK-21 cells infected with VSV for 24 h compared with controls. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment showed that the differentially expressed lncRNAs and their target genes were mainly concentrated in pathways related to apoptosis, cancer, disease, and immune system activation, including the TNF, P53, MAPK, and NF-kappaB signaling pathways. The differentially expressed lncRNA can modulate immune processes by regulating genes involved in these signaling transmissions. Ten randomly selected DEGs, namely, Il12rb2, F2, Masp2, Mcl1, FGF18, Ripk1, Fas, BMF, POLK, and JAG1, were validated using RT-qPCR. As predicted through RNA-Seq analysis, these DEGs underwent either up- or downregulation, suggesting that they may play key regulatory roles in the pathways mentioned previously. CONCLUSIONS: Our study showed that VSV infection alters the host metabolic network and activates immune-related pathways, such as MAPK and TNF. The above findings provide unique insights for further study of the mechanism of VSV-host interactions and, more importantly, provide a theoretical basis for VSV as an excellent vaccine carrier.

Deep Electron Redistributions Induced by Dual Junctions Facilitating Electroreduction of Dilute Nitrate to Ammonia.

The electronic states of metal catalysts can be redistributed by the rectifying contact between metal and semiconductor e.g., N-doped carbon (NC), while the interfacial regulation degree is very limited. Herein, a deep electronic state regulation is achieved by constructing a novel double-heterojunctional Co/Co3O4@NC catalyst containing Co/Co3O4 and Co3O4/NC heterojunctions. When used for dilute electrochemical NO3 - reduction reaction (NO3RR), the as-prepared Co/Co3O4@NC exhibits an outstanding Faradaic efficiency for NH3 formation (FENH3) of 97.9%, -0.4 V versus RHE and significant NH3 yield of 303.5 mmol h-1 gcat -1 at -0.6 V at extremely low nitrate concentrations (100 ppm NO3 --N). Experimental and theoretical results reveal that the dual junctions of Co/Co3O4 and Co3O4/NC drive a unidirectional electron transfer from Co to NC (Co→Co3O4→NC), resulting in electron-deficient Co atoms. The electron-deficient Co promotes NO3 - adsorption, the rate-determining step (RDS) for NO3RR, facilitating the dilute NO3RR to NH3. The design strategy provides a novel reference for unidirectional multistage regulation of metal electronic states boosting electrochemical dilute NO3RR, which opens up an avenue for deep electronic state regulation of electrocatalyst breaking the limitation of the electronic regulation degree by rectifying contact.

Boosting Electrochemical Nitrate Reduction at Low Concentrations Through Simultaneous Electronic States Regulation and Proton Provision.

Electrochemically converting nitrate (NO3 -) into ammonia (NH3) has emerged as an alternative strategy for NH3 production and effluent treatment. Nevertheless, the electroreduction of dilute NO3 - is still challenging due to the competitive adsorption between various aqueous species and NO3 -, and unfavorable water dissociation providing *H. Herein, a new tandem strategy is proposed to boost the electrochemical nitrate reduction reaction (NO3RR) performance of Cu nanoparticles supported on single Fe atoms dispersed N-doped carbon (Cu@Fe1-NC) at dilute NO3 - concentrations (≤100 ppm NO3 --N). The optimized Cu@Fe1-NC presents a FENH3 of 97.7% at -0.4 V versus RHE, and a significant NH3 yield of 1953.9 mmol h-1 gCu -1 at 100 ppm NO3 --N, a record-high activity for dilute NO3RR. The metal/carbon heterojunctions in Cu@Fe1-NC enable a spontaneous electron transfer from Cu to carbon substrate, resulting in electron-deficient Cu. As a result, the electron-deficient Cu facilitates the adsorption of NO3 - compared with the pristine Cu. The adjacent atomic Fe sites efficiently promote water dissociation, providing abundant *H for the hydrogenation of *NOx e at Cu sites. The synergistic effects between Cu and single Fe atom sites simultaneously decrease the energy barrier for NO3 - adsorption and hydrogenation, thereby enhancing the overall activity of NO3 - reduction.

Function analysis of choline binding domains (CBDs) of LytA, LytC and CbpD in biofilm formation of Streptococcus pneumoniae.

Biofilm formation is an important strategy for the colonization of Streptococcus pneumoniae, which can increase the capacity to evade antibiotic and host immune stress. Extracellular choline-binding proteins (CBPs) are required for successful biofilm formation, but the function of extracellular CBPs in the process of biofilm formation is not fully understood. In this study, we tend to analyze the functions of LytA, LytC and CbpD in biofilm formation by in vitro studies with their choline-binding domains (CBDs). Biofilm formation of S. pneumoniae was enhanced when cultured in medium supplemented with CBD-C and CBD-D. Parallel assays with ChBp-Is (choline binding repeats with different C-terminal tails) and character analysis of CBDs reveal a higher isoelectric point (pI) is related to promotion of biofilm formation. Phenotype characterization of biofilms revel CBD-C and CBD-D function differently, CBD-C promoting the formation of membrane-like structures and CBD-D promoting the formation of regular reticular structures. Gene expression analysis reveals membrane transport pathways are influenced with the binding of CBDs, among which the phosphate uptake and PTS of galactose pathways are both up-regulated under conditions with CBDs. Further, extracellular substances detection revealed that extracellular proteins increased with CBD-A and CBD-D, exhibiting as increase in extracellular high molecular weight proteins. Extracellular DNA increased under CBD-A but decreased under CBD-C and CBD-D; Extracellular phosphate increased under CBD-C. These support the alterations in membrane transport pathways, and reveal diverse reactions to extracellular protein, DNA and phosphate of these three CBDs. Overall, our results indicated extracellular CBP participate in biofilm formation by affecting surface charge and membrane transport pathways of pneumococcal cells, as well as promoting reactions to extracellular substances.

Dopant-Induced Electronic States Regulation Boosting Electroreduction of Dilute Nitrate to Ammonium.

Electrochemically converting NO3 - into NH3 offers a promising route for water treatment. Nevertheless, electroreduction of dilute NO3 - is still suffering from low activity and/or selectivity. Herein, B as a modifier was introduced to tune electronic states of Cu and further regulate the performance of electrochemical NO3 - reduction reaction (NO3 RR) with dilute NO3 - concentration (≤100 ppm NO3 - -N). Notably, a linear relationship was established by plotting NH3 yield vs. the oxidation state of Cu, indicating that the increase of Cu+ content leads to an enhanced NO3 - -to-NH3 conversion activity. Under a low NO3 - -N concentration of 100 ppm, the optimal Cu(B) catalyst displays a 100 % NO3 - -to-NH3 conversion at -0.55 to -0.6 V vs. RHE, and a record-high NH3 yield of 309 mmol h-1 gcat -1 , which is more than 25 times compared with the pristine Cu nanoparticles (12 mmol h-1 gcat -1 ). This research provides an effective method for conversion of dilute NO3 - to NH3 , which has certain guiding significance for the efficient and green conversion of wastewater in the future.

Development and preliminary testing of a probe-based duplex real-time PCR assay for the detection of African swine fever virus.

An outbreak of African swine fever (ASF) in China in 2018 caused substantial economic losses to the swine industry. To accurately diagnose clinical infection with ASF virus (ASFV), we developed a TaqMan probe-based duplex real-time PCR that simultaneously detected two discontinuous genes in the virus genome, thereby preventing the inaccurate results obtained with only one reaction. Two sets of ASFV gene-specific primers, along with two fluorescent TaqMan probes were designed to target conserved regions of the B646L and B438L genes. This method had high sensitivity and specificity, with a limit of detection of 10 copies/µL, and it did not cross-react with the genomes of other viral pathogens that affect pigs (i.e., CSFV, PRRSV, PEDV, PRV, PPV and PCV2). Overall, 180 clinical samples from ASFV-infected pig farms were used to compare this method with a commercial kit, which yielded excellent consistency (98.3%). This new diagnostic method should greatly improve the efficiency of ASFV surveillance and reduce economic losses, providing benefits for both animal and public health.

A Tandem Strategy for Enhancing Electrochemical CO2 Reduction Activity of Single-Atom Cu-S1 N3 Catalysts via Integration with Cu Nanoclusters.

We developed a tandem electrocatalyst for CO2 -to-CO conversion comprising the single Cu site co-coordinated with N and S anchored carbon matrix (Cu-S1 N3 ) and atomically dispersed Cu clusters (Cux ), denoted as Cu-S1 N3 /Cux . The as-prepared Cu-S1 N3 /Cux composite presents a 100 % Faradaic efficiency towards CO generation (FECO ) at -0.65 V vs. RHE and high FECO over 90 % from -0.55 to -0.75 V, outperforming the analogues with Cu-N4 (FECO only 54 % at -0.7 V) and Cu-S1 N3 (FECO 70 % at -0.7 V) configurations. The unsymmetrical Cu-S1 N3 atomic interface in the carbon basal plane possesses an optimized binding energy for the key intermediate *COOH compared with Cu-N4 site. At the same time, the adjacent Cux effectively promotes the protonation of *CO2 - by accelerating water dissociation and offering *H to the Cu-S1 N3 active sites. This work provides a tandem strategy for facilitating proton-coupled electron transfer over the atomic-level catalytic sites.

Occurrence and genomic characterization of ESBL-producing Escherichia coli ST29 strains from swine with abundant virulence genes.

Food-production animals were considered to be a major reservoir of antimicrobial-resistant bacteria and clinically relevant pathogens. The potential of commensal Escherichia coli from pigs as a source of opportunistic pathogens associated with extraintestinal infections in humans needs to be assessed. In this study, 13 E. coli isolates from an intensive pig farm in China were analyzed using whole genome sequencing followed by in-depth in silico analysis. Genomic analysis showed comprehensive antimicrobial resistance profiles, with each isolate carrying between 4 and 22 antimicrobial resistance genes. Although these E. coli isolates were assigned to low-virulence phylogroup A and B1, 31 different virulence genes were detected at least once in the 13 sequenced isolates. Extraintestinal pathogenic E. coli-associated virulence genes, including iss, iha, tsh and iroN, were found in commensal E. coli isolates in this study. Of note, a large number of virulence genes (n = 22) were identified in ESBL-producing E. coli sequence type (ST) 29 isolates. Our study revealed the presence of comprehensive antimicrobial resistance and virulence gene profiles in commensal E. coli isolates of pigs. The emerged ESBL-producing E. coli ST 29 isolates harboring a high abundance of VAGs highlighted that this new clonal linage may pose a threat to public health.

Co-carrying of KPC-2, NDM-5, CTX-M-3 and CTX-M-65 in three plasmids with serotype O89: H10 Escherichia coli strain belonging to the ST2 clone in China.

Carbapenem-resistant Enterobacteriaceae strains as a new serious threat for the public health have been increasingly reported worldwide. In this study, one multi-resistant Escherichia coli strain ZSH6 which co-carried blaKPC-2, blaNDM-5 and blaCTX-M, was isolated from human blood sample. By using plasmid conjugation experiments, ZSH6 was found to harbor three plasmids carrying the blaNDM-5 gene, the blaKPC-2 and blaCTX-M gene, respectively. Whole-genome sequencing of ZSH6 yielded 122 scaffolds of chromosomal DNA and three circular plasmids including pZSH6-blaKPC-2 (46,319 bp), pZSH6-blaNDM-5 (46,161bp) and pZSH6-blaCTX-M (184,723). The isolate was classified to Sequence Type 2 and to the O89: H10 serotype. The results of genome analyses revealed that ZSH6 carried three virulence factors (capU, gad and iss) and twenty resistance genes [blaKPC-2blaNDM-5, blaCTX-M-3, blaCTX-M-65, blaTEM-1, floR, tet(A), tet(B), dfrA17, aadA5, sul1, mdf(A), mph(A), erm(B), aph(3')-Ia, aph(3')-Ib, aph(4)-Ia, aph(6)-Id, aac(3)-Iva, aac(3)-IId]. Therefore, the co-existence of such a large number of resistance genes in multiple plasmids making ZSH6 highly resistant to almost all kinds of commonly used antibiotics, and brings a serious challenge for resistance control and clinical treatment of infections caused by this bacterium.

A requirement of TolC1 for effective survival, colonization and pathogenicity of Actinobacillus pleuropneumoniae.

To establish infection in the host, pathogens have evolved sophisticated systems to cope with environmental conditions and to protect cells against host immunity. TolC is the outer membrane channel component of type 1 secretion systems and multidrug efflux pumps that plays critical roles during the infection process in many pathogens. However, little is known about the exact roles of TolC1 in the pathogenicity of A. pleuropneumoniae, an etiological agent of the porcine contagious pleuropneumoniae that causes severe respiratory disease. In this study, deletion of tolC1 causes apparent ultrastructural defects in A. pleuropneumoniae cell examined by transmission electron microscopy. The tolC1 mutant is hypersensitivity to oxidative, osmotic and acid challenges by in vitro stress assays. Analysis on secreted proteins shows that the excretion of ApxIIA and an ApxIVA-like protein, ApxIVA-S, is abolished in the absence of TolC1. This result confirms the essential role of TolC1 in the secretion of Apx toxins and this is the first identification of an ApxIVA-like protein in in vitro culture of A. pleuropneumoniae. Besides, disruption of TolC1 leads to a significant attenuation of virulence in mice by an intraperitoneal route of A. pleuropneumoniae. The basis for the attenuation is further investigated using a mouse intranasal infection model, which reveals an impaired ability to colonize and induce lesions in the lungs for the loss of TolC1 of A. pleuropneumoniae. In conclusion, our findings demonstrate significant roles of TolC1 in facilitating bacterial survival in hostile conditions, maximum colonization as well as pathogenicity during the infection of A. pleuropneumoniae.

Frequency of virulence factors in high biofilm formation blaKPC-2 producing Klebsiella pneumoniae strains from hospitals.

The aim of this study is to determine the frequency of virulence genes in high biofilm formation blaKPC-2 producing Klebsiella pneumoniae strains collected over a period of two years. A total of 43 non-repetitive high biofilm blaKPC-2 producing isolates were screened from 429 strains. The MIC of carbapenems (imipenem and meropenem) ranged from 4 to 32 µg/ml. The OD595 value of the biofilm ranged from 0.56 to 2.56. The K1, K2, K5, K20, K54, K57 genotypes, MLST and virulence factors, including entB, ybtS, mrkD, fimH, rmpA, allS, iutA, kfu, wcaG, aerobaction, fecIRA, shiF, magA and pagO gene, were determined by PCR. The results showed that, among the 43 isolates, 5 of 43 were K1 type, 25 of 43 were K2 type, 4 strains and 2 strains were K5 and K57 respectively. The MLST results showed that 23/43 strains were ST11, followed by ST433(4/43), ST107(4/43), ST690(4/43), ST304(2/43), ST2058(1/43), ST1(1/43), ST146(1/43), ST914(1/43), ST2636(1/43), ST2637(1/43). As to the virulence factors, all 43 strains carried entB, ybtS and mrkD gene, followed by fimH(38/43), rmpA(14/43), allS(34/43), iutA(27/43), kfu(25/43), wcaG(21/43), aerobaction(16/43), fecIRA(15/43), shiF(10/43), magA(5/43) and pagO(5/43). This study demonstrated that high frequency of virulence factors emerging in high biofim blaKPC-2 producing strains. It also suggested that we should continue to focus on the toxicity variation and it's high time to enhance clinical awareness to the infections causing by Klebsiella pneumoniae.

Co-location of the blaKPC-2, blaCTX-M-65, rmtB and virulence relevant factors in an IncFII plasmid from a hypermucoviscous Klebsiella pneumoniae isolate.

Hypervirulent variants of klebsiella pneumoniae (hvKP), which cause serious infections not only healthy individuals, but also the immunocompromised patients, have been increasingly reported recently. One conjugation of a hypermucoviscous strian SWU01 co-carried the resistance gene blaKPC-2 and virulence gene iroN by the PCR detection from three carbapenem-resistance hvKP. To know the genetic context of this plasmid. The whole genome of this strain was sequenced. We got a 162,552-bp plasmid (pSWU01) which co-carried the resistance gene blaKPC-2 and virulence gene iroN. It is composed of a typical IncFII-type backbone, five resistance genes including blaCTX-M-65, blaKPC-2, blaSHV-12, blaTEM-1 and rmtB, and several virulence relevant factors including iroN, traT and toxin-antitoxin systems. The plasmid pSWU01 co-carrying the multidrug resistance determinants and virulence relevant factors from the hypermucoviscous K. pneumoniae represents a novel therapeutic challenge.

Thermoregulated Phase-Transition Synthesis of Two-Dimensional Carbon Nanoplates Rich in sp2 Carbon and Unimodal Ultramicropores for Kinetic Gas Separation.

The development of highly selective, chemically stable and moisture-resistant adsorbents is a key milestone for gas separation. Porous carbons featured with random orientation and cross-linking of turbostratic nanodomains usually have a wide distribution of micropores. Here we have developed a thermoregulated phase-transition-assisted synthesis of carbon nanoplates with more than 80 % sp2 carbon, unimodal ultramicropore and a controllable thickness. The thin structure allows oriented growth of carbon crystallites, and stacking of crystallites in nearly parallel orientation are responsible for the single size of the micropores. When used for gas separation from CH4 , carbon nanoplates exhibit high uptakes (5.2, 5.3 and 5.1 mmol g-1 ) and selectivities (7, 71 and 386) for CO2 , C2 H6 and C3 H8 under ambient conditions. The dynamic adsorption capacities are close to equilibrium uptakes of single components, further demonstrating superiority of carbon nanoplates in terms of selectivity and sorption kinetics.

HtrA Is Important for Stress Resistance and Virulence in Haemophilus parasuis.

Haemophilus parasuis is an opportunistic pathogen that causes Glässer's disease in swine, with polyserositis, meningitis, and arthritis. The high-temperature requirement A (HtrA)-like protease, which is involved in protein quality control, has been reported to be a virulence factor in many pathogens. In this study, we showed that HtrA of H. parasuis (HpHtrA) exhibited both chaperone and protease activities. Finally, nickel import ATP-binding protein (NikE), periplasmic dipeptide transport protein (DppA), and outer membrane protein A (OmpA) were identified as proteolytic substrates for HpHtrA. The protease activity reached its maximum at 40°C in a time-dependent manner. Disruption of the htrA gene from strain SC1401 affected tolerance to temperature stress and resistance to complement-mediated killing. Furthermore, increased autoagglutination and biofilm formation were detected in the htrA mutant. In addition, the htrA mutant was significantly attenuated in virulence in the murine model of infection. Together, these data demonstrate that HpHtrA plays an important role in the virulence of H. parasuis.

Modulation of natural killer cell antitumor activity by the aryl hydrocarbon receptor.

The aryl hydrocarbon receptor (AhR) has become increasingly recognized for its role in the differentiation and activity of immune cell subsets; however, its role in regulating the activity of natural killer (NK) cells has not been described. Here, we show that AhR expression is induced in murine NK cells upon cytokine stimulation. We show that in the absence of AhR, NK cells have reduced cytolytic activity and reduced capacity to control RMA-S tumor formation in vivo, despite having normal development and maturation markers. Although AhR was first identified to bind the xenobiotic compound dioxin, AhR is now known to bind a variety of natural exogenous (e.g., dietary) and endogenous ligands. We show that activation of AhR with an endogenous tryptophan derivative, 6-formylindolo[3,2-b]carbazole, potentiates NK cell IFN-γ production and cytolytic activity. Further, administration of 6-formylindolo[3,2-b]carbazole in vivo enhances NK cell control of tumors in an NK cell- and AhR-dependent manner. Finally, similar effects on NK cell potency occur with AhR dietary ligands, potentially explaining the numerous associations that have been observed in the past between diet and NK cell function. Our studies introduce AhR as another regulator of NK cell activity in vivo.

Using hollow carbon nanospheres as a light-induced free radical generator to overcome chemotherapy resistance.

Under evolutionary pressure from chemotherapy, cancer cells develop resistance characteristics such as a low redox state, which eventually leads to treatment failures. An attractive option for combatting resistance is producing a high concentration of produced free radicals in situ. Here, we report the production and use of dispersible hollow carbon nanospheres (HCSs) as a novel platform for delivering the drug doxorubicine (DOX) and generating additional cellular reactive oxygen species using near-infrared laser irradiation. These irradiated HCSs catalyzed sufficiently persistent free radicals to produce a large number of heat shock factor-1 protein homotrimers, thereby suppressing the activation and function of resistance-related genes. Laser irradiation also promoted the release of DOX from lysosomal DOX@HCSs into the cytoplasm so that it could enter cell nuclei. As a result, DOX@HCSs reduced the resistance of human breast cancer cells (MCF-7/ADR) to DOX through the synergy among photothermal effects, increased generation of free radicals, and chemotherapy with the aid of laser irradiation. HCSs can provide a unique and versatile platform for combatting chemotherapy-resistant cancer cells. These findings provide new clinical strategies and insights for the treatment of resistant cancers.

Comparative proteome analysis of the extracellular proteins of two Haemophilus parasuis strains Nagasaki and SW114.

This study used a comparative proteomics approach to distinguish between the two-dimensional electrophoresis profiles of extracellular proteins in Nagasaki and SW114. Protein spots were identified using matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry. The ten proteins unique to Nagasaki were putative adhesin AidA protein, putative extracellular serine protease (autotransporter) (771aa), putative extracellular serine protease (autotransporter) (780aa), protective surface antigen D15, 30S ribosomal protein S2, periplasmic serine protease do/hhoA-like protein, acid phosphatase, membrane protein, protein-disulfide isomerase, and iron ABC transporter substrate-binding protein. Meanwhile, the two proteins unique to SW114 were C4-dicarboxylate ABC transporter substrate-binding protein and peptide ABC transporter substrate-binding protein. Quantitative PCR was used to analyze the mRNA transcript levels of three randomly selected proteins. The afuA, AidA, and ompD15 genes encoding iron ABC transporter substrate-binding protein, putative adhesin AidA protein and protective surface antigen D15 respectively demonstrated significantly higher mRNA transcript levels (39.606, 3.924, and 36.668, respectively) in Nagasaki than in SW114. These observations suggest the levels of differentially expressed proteins were directly proportional to their cellular mRNA levels. Three virulence-related proteins, namely, putative adhesin AidA protein, putative extracellular serine protease (autotransporter) (771aa) and putative extracellular serine protease (autotransporter) (780aa) were identified in Nagasaki.

Tocopherol-deficient rice plants display increased sensitivity to photooxidative stress.

Tocopherols are lipophilic antioxidants that are synthesized exclusively in photosynthetic organisms. Despite extensive in vivo characterization of tocopherol functions in plants, their functions in the monocot model plant, rice, remain to be determined. In this study, transgenic rice plants constitutively silenced for homogentisate phytyltransferase (HPT) and tocopherol cyclase (TC) activity were generated. Silencing of HPT and TC resulted in up to a 98 % reduction in foliar tocopherol content relative to the control plants, which was also confirmed by transcript level analysis. When grown under normal conditions, HPT and TC transgenics showed no distinctive phenotype relative to the control plants, except a slight reduction in plant height and a slight decrease in the first leaf length. However, when exposed to high light at low temperatures, HPT and TC transgenics had a significantly higher leaf yellowing index than the control plants. The tocopherol-deficient plants decreased their total individual chlorophyll levels, their chlorophyll a/b ratio, and the maximum photochemical efficiency of photosystem II, whereas increased lipid peroxidation levels relative to the control plants. Tocopherol deficiency had no effect on ascorbate biosynthesis, but induced glutathione, antheraxanthin, and particularly zeaxanthin biosynthesis for compensation under stressful conditions. However, despite these compensation mechanisms, HPT and TC transgenics still exhibited altered phenotypes under high light at low temperatures. Therefore, it is suggested that tocopherols cannot be replaced and play an indispensable role in photoprotection in rice.