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A two-dimensional waveguide is the building block in integrated photonic circuits. In this work, the 5.0 MeV oxygen ion implantation at a dose of [Formula: see text] ions/cm2 was carried out on the fused silica glass for the formation of the planar waveguide. Then, a femtosecond laser was employed to ablate the ion-implanted surface at a speed of 200 [Formula: see text]m/s for the construction of the ridge waveguide. The energy depositions of the oxygen ion implantation into the fused silica glass were simulated by the SRIM 2013. The microscopic morphology of the ridge waveguide was photographed by a Nikon microscope. The near-field intensity distributions of the ridge waveguide at 532 nm and 976 nm were measured by using the end-face coupling system. The ion-implanted and femtosecond-ablated ridge waveguide on the fused silica glass has the potential as a photonic device for integrated optical systems at the visible and near-infrared bands.

Becker's nevus (BN) manifests as a hyperpigmented, sometimes hypertrichotic plaque/patch over the chest and shoulder, and it is in the category of benign cutaneous hamartomas. BN has elongation and fusion of the rete ridge, keratotic plugging, sebaceous hyperplasia, smooth muscle hyperplasia, and hyperpigmentation of the basal/suprabasal layer histologically. This article highlights all issues involved in pathogenesis and treatment options of BN. According to current research, postzygotic ACTB mutations induce BN and Becker's nevus syndrome (BNS). Although several therapy strategies were utilized to treat the pigmentary and hypertrichotic aspects of BN, no definitive standard treatment was identified to far, and further research is needed to better educate BN care.

Giant ragweed (Ambrosia trifida) pollen is closely associated with respiratory allergy in late summer and autumn, and the prevalence of giant ragweed pollen allergy progressively increases. Compared with short ragweed (Ambrosia artemisiifolia), allergenic components from giant ragweed pollen are poorly investigated. To promote component-resolved diagnosis and treatment for giant ragweed pollen allergy, it becomes necessary to identify and characterize unknown allergens from giant ragweed pollen. In the present study, we identified and characterized a new cysteine-protease (CP) allergen from giant ragweed pollen, named as Amb t CP. The cloned Amb t CP gene encoded 387 amino acids. Recombinant Amb t CP (rAmb t CP) and natural Amb t CP (nAmb t CP) were purified by high-affinity Ni

To investigate the indentification method of samples mistyped as O phenotype and to explore the precision transfusion strategy.The blood samples from donors and patients admitted in our center from 2018 to 2019 was collected. The samples with O phenotype suspected subtypes were further determined by tube test, adsorption-elution test, etc. Molecular testing was used to sequence the related blood type genes of the subjects.Among 14 subjects misjudged as O, 11 different genotypes were identified, in which 3 blood donors were Ael02/O02, Bel03/O02, and one para-Bombay with B101/O02 (FUT1: h3h3; FUT2: SeThe phenotypes of Ael, Bel, Aw and para-Bombay subtypes are easily misjudged as type O. Molecular technology is helpful to identify the genotype of subtypes, and the corresponding transfusion strategies could be reasonably performed.误判O型的疑难血型鉴定及安全输血策略的研究.综合探讨多种误判O型的疑难血型鉴定及安全输血策略.收集2018-2019年本中心接收的无偿献血者和临床患者疑难血型复检标本，对表型为O型但疑似亚型标本使用试管法、吸收放散法等进一步验证；并采用分子生物学方法对受检者的相关血型基因进行检测.14例表型被误判为O型的受检者中，共鉴定出11种基因型，其中无偿献血者3例，ABO血型基因型分别为Ael02/O02、Bel03/O02及1例类孟买B101/O02（FUT1: h3h3; FUT2: SeAel、Bel、Aw、类孟买等多种亚型的表型易被误判为O型。分子生物学手段有助于鉴定亚型基因型，合理安排相应的输血策略.

In recent years, the outbreaks of porcine epidemic diarrhea (PED) caused by the highly virulent porcine epidemic diarrhea virus (PEDV) variants occurred frequently in China, resulting in severe economic impacts to the pork industry. In this study, we selected and analyzed the genetic evolution of 15 PEDV representative strains that were identified in fecal samples of diarrheic piglets in 10 provinces and cities during 2011–2017. The phylogenetic analysis indicated that all the 15 PEDV isolates clustered into G2 genotype associated with the current circulating strains. Compared with the genome of the prototype strain CV777, these strains had 103–120 amino acid mutations in their S proteins, most of which were in the N terminal domain of S1 (S1-NTD). We also found 37 common mutations in all these 15 strains, although these strains shared 96.9–99.7% nucleotide homology and 96.3–99.8% amino acid homology in the S protein compared with the other original pandemic strains. Computational analysis showed that these mutations may lead to remarkable changes in the conformational structure and asparagine (N)-linked glycosylation sites of S1-NTD, which may be associated with the altered pathogenicity of these variant PEDV strains. We evaluated the pathogenicity of the PEDV strain FJzz1 in piglets through oral and intramuscular infection routes. Compared with oral infection, intramuscular infection could also cause typical clinical signs but with a slightly delayed onset, confirming that the variant PEDV isolate FJzz1 was highly pathogenic to suckling piglets. In conclusion, we analyzed the genetic variation and pathogenicity of the emerging PEDV isolates of China, indicating that G2 variant PEDV strains as the main prevalent strains that may mutate continually. This study shows the necessity of monitoring the molecular epidemiology and the etiological characteristics of the epidemic PEDV isolates, which may help better control the PED outbreaks., Highlights • G2 variant PEDV strains as the main prevalent strains continue to mutate along with time. • The isolated strains were prone to mutate in the S protein with many common amino acid mutations. • These mutations may lead to remarkable changes in the conformational structure of S1-NTD and the pathogenicity of PEDV. • Both oral and intramuscular routes caused typical clinical signs to piglets.

We developed a method to synthesize fluorinated 1,4-unsaturated dicarbonyl compounds via photoredox catalyzed radical addition process. Commercially available ethyl bromodifluoroacetate (BrCF2CO2Et) as fluoroalkyl source, the corresponding fluoro-containing dicarbonyl compounds could be obtained in moderate to good yields.

Porcine reproductive and respiratory syndrome (PRRS) is one of the major drivers of economic loss in the swine industry worldwide. In commercial pig production, vaccination is the first option in an attempt to control infectious diseases. Pigs are therefore often immunized with different vaccines, and almost all of them are delivered via the intramuscular (IM) route. However, the IM injection may result in physical damage, stress reactions, and is labor demanding. An alternative route is urgently needed to reduce the disadvantages of conventional vaccination. In this study, a needle-free intradermal (ID) delivery system was evaluated for delivering a live PRRS vaccine as compared with the traditional needle-syringe method. Fifty-two 4-week-old piglets were divided into six groups: piglets in groups A-C were immunized using ID delivery system with 104, 105 and 106 TCID50 of PRRS candidate vaccine strain rHN-NP49, respectively; piglets in group D were immunized IM with 105 TCID50 of rHN-NP49; and group E and F were used as challenge and control groups, respectively. At 28 days post vaccination, piglets in group A to E were challenged with a lethal dose of highly-pathogenic PRRSV. Similar results were found in viremia and antibody response among the ID and IM groups during the immunization stage. After challenge, similar results were found in average body weight gain, viral shedding, serum viral load, and clinical score among the immunization groups, with a higher protection ratio in the ID group compared with IM group with the same immunization dose. These results demonstrated that the ID delivery system could provide similar or even better protection compared with IM route, and could be an effective route for PRRS vaccination.

Geosynthetics-reinforced soil (GRS) structures have been widely used for the prevention of geological hazards. As a recently introduced product, the triaxial geogrid has been confirmed to provide improved performance due to the more stable grid structure. This paper presents an evaluation of the mechanical behavior based on a series of laboratory tests. The unconfined tensile strength of biaxial geogrid and triaxial geogrid in different loading directions relative to the orientation of ribs was investigated. Then, more than 8 pullout tests were conducted on the triaxial geogrid specimens embedded in the compacted sand. The internal displacements along the geogrid length were monitored. The results show that the triaxial geogrid has been shown to provide nearly uniform tensile strength in all loading directions as compared with the biaxial geogrid. The triaxial geogrid deformation is mainly characterized by rib bending and nodal distortion along with an inward squeeze perpendicular to the pullout direction. The interface friction between the soil and the geogrid develops in a progressive mode, and an elasto-plastic-softening characteristic is detected experimentally due to the extensibility of geogrid.

Novel porcine circovirus type 3 (PCV3), first identified in the United States, has been detected in many other countries. Porcine circovirus is associated with postweaning multisystemic wasting syndrome, reproductive failure, congenital tremors, and other clinical symptoms. In this study, we established a double polymerase chain reaction assay for detecting both porcine circovirus type 2 (PCV2) and PCV3. This is the first study to detect and characterize the PCV3 genome in the Tianjin region of North China. We collected a total of 169 tissue samples from seven farms between 2016 and 2018. The PCV3-positive rate of all tissue samples was 37.3% (63/169) and the rate of PCV2 and PCV3 coinfection was 14.8% (25/169). PCV2 and PCV3 coinfections with more serious clinical symptoms were found in only three farms. We sequenced three PCV3 strains selected from tissue samples that were positively identified. The complete genome sequences of the three strains shared 97.6–99.4% nucleotide identities with the PCV3 strains in GenBank. Our results showed the extent of PCV3’s spread in Tianjin, and the need to further study PCV3’s pathobiology, epidemiology, isolation, and coinfection.

The parent HZSM-5 was activated by NaOH solution with different concentration to obtain a series of alkaline activated HZSM-5-m (m denotes the concentration of NaOH). Zn (2 wt%) was supported on HZSM-5 and HZSM-5-m via the impregnation method by using Zn(NO3)2·6H2O as a precursor. The materials were systematically characterized by XRD, FT-IR, N2 adsorption–desorption at low temperature, and NH3-TPD techniques. It was found that the acidity and textural properties of the HZSM-5-m could be adjusted by altering the concentration of NaOH. Moreover, the amount and distribution of acidity and the pore structure of catalyst had a significantly influence on heptane conversion and aromatics content. As a result, an efficient Zn/HZSM-5-m catalyst for aromatization reaction was obtained by changing the concentration of NaOH.

// Feng Wen 1 , Jinyue Guo 2 , Guangzhi Tong 1, 3 , Dingren Bi 2 , Qi Wang 1 , Xiaomin Liu 1 , Shuaiyong Wang 1 , Tonglin Shan 1 , Wu Tong 1 , Yanjun Zhou 1 , Guoxin Li 1 and Hai Yu 1, 3 1 Division of Swine Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China 2 College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, China 3 Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou 225009, China Correspondence to: Jinyue Guo, email: guojinyue12@163.com Hai Yu, email: haiyu@shvri.ac.cn Keywords: influenza A, H7N9, H5N1, meta-analysis, extracellular matrix Received: April 04, 2017 Accepted: June 16, 2017 Published: July 18, 2017 ABSTRACT Avian-origin H5N1 and H7N9 influenza A viruses are capable of causing lethal infection in humans, with serious lung pathology and leading to acute respiratory distress syndrome. The contribution of host response associated with the poor prognosis of H5N1 and H7N9 infections remains unclear. The aim of this study was to identify the host factors involved in the high pathogenicity of H5N1 and H7N9 by a systematical meta-analysis. The RNA-seq datasets related to H5N1, H7N9, and H1N1 infections with time series were retrieved from GEO. After merging the data from different series, ComBat was used to adjust the known variances from different batches. The transcription factors binding the genes in each cluster were predicted by PASTAA. We figured out the genes that were differentially expressed at any time point in samples infected with H5N1, H7N9, or H1N1. The analysis of biological function showed that genes related with cytokine were up-regulated in all three viruses. However, genes associated with carbon metabolism were found exclusively down-regulated in H7N9 and the extracellular matrix pathway were only enriched in H5N1 and H7N9. To summary, our study suggested that the extracellular matrix might be associated with the high fatality of H5N1 and H7N9 viruses in humans.

Platanus acerifolia (P. acerifolia) is an important cause of pollinosis in cities. The use of allergen extracts on patients with allergic diseases is the most commonly applied method to attempt to treat pollinosis. Pla a 3, a non‑specific lipid transfer protein, is a major allergen present in P. acerifolia pollen extracts. In the present study, the Pla a 3 gene was sub‑cloned into a pSUMO‑Mut vector using Stu I and Xho I sites and transformed into the Arctic Express™ (DE3) RP E. coli host strain. The purified Pla a 3 allergen was analyzed by western blotting and the results revealed that the Pla a 3 allergen has the ability to bind IgE in the P. acerifolia pollen of allergic patients' sera. Moreover, the authors predicted the potential B cell epitopes of the Pla a 3 allergen using the DNAStar Protean system, the Bioinformatics Predicted Antigenic Peptides system and the BepiPred 1.0 server. In addition, the T cell epitopes were predicted by the SYFPEITHI database and the NetMHCII‑2.2 server. As a result, two B cell epitopes (35‑45 and 81‑86) and four potential T cell epitopes including 2‑15, 45‑50, 55‑61 and 67‑73 were predicted in the present study. The current results can be used to contribute to allergen immunotherapies and useful in peptide‑based vaccine designs of pollen allergy.

Platanus acerifolia pollen is considered an important source of airborne allergens in numerous cities. Pla a 1 is a major allergen from P. acerifolia pollen. The present study aimed to express and purify Pla a 1, and to prepare its monoclonal antibody. In the present study, the Pla a 1 gene was subcloned into a pET‑28a vector and transformed into the ArcticExpress™ (DE3) RP Escherichia coli host strain. The purified Pla a 1 was then used to immunize BALB/c mice. When serum detection was positive, spleen cells were isolated from the mice and fused with SP2/0 myeloma cells at a ratio of 10:1. Hybridoma cells were screened by indirect ELISA and limiting dilution. Positive cells were used to induce the formation of antibody‑containing ascites fluid, and the antibodies were purified using protein A‑agarose. The results of the present study demonstrated that recombinant Pla a 1 was successfully expressed and purified, and exhibited positive immunoglobulin E‑binding to serum from patients allergic to P. acerifolia. A total of 11 hybridomas that steadily secreted anti‑Pla a 1 antibody were obtained and an immunoblotting analysis indicated that all of these monoclonal antibodies specifically recognized the Pla a 1 protein. These results suggested that specific anti‑Pla a 1 antibodies may be obtained, which can be used for the rapid detection of Pla a 1 allergens and in the preparation of vaccines against P. acerifolia pollen.

PB2-627K is an important amino acid that determines the virulence of some influenza A viruses. However, it has not been experimentally investigated in the H3N2 swine influenza virus. To explore the potential role of PB2-K627E substitution in H3N2 swine influenza virus, the growth properties and pathogenicity between H3N2 swine influenza virus and its PB2-K627E mutant were compared. For the first time, our results showed that PB2-K627E mutation attenuates H3N2 swine influenza virus in mammalian cells and in mice, suggesting that PB2-627K is required for viral replication and pathogenicity of H3N2 swine influenza virus.

Background Breast cancer is the most common cancer among women worldwide, and approximately 70% of breast cancers are hormone receptor-positive and express estrogen receptor-α (ERα) or/and progesterone receptor. Therapies targeting ERα have been successfully used in patients with ERα+ breast cancer. However, intrinsic or acquired resistance to anti-estrogen therapy presents a major challenge. The Wnt/β-catenin signaling pathway regulates various processes that are important for cancer progression, and emerging evidences have shown a close interaction between Wnt/β-catenin and ERα signaling. miR-190 is also involved in ER signaling and our previous study indicated that miR-190 suppresses breast cancer metastasis. Methods The effect of miR-190 on breast cancer anti-estrogen sensitivity was investigated both in vitro and in vivo. The protein expression levels and localization were analyzed by western blotting and immunofluorescence, respectively. Chromatin immunoprecipitation and dual-luciferase reporter assays were used to validate the regulation of the zinc-finger E-box binding homeobox 1/ ERα-miR-190-SRY-related high mobility group box 9 (ZEB1/ERα-miR-190-SOX9) axis. Results miR-190 increased the anti-estrogen sensitivity of breast cancer cells both in vitro and in vivo. miR-190 inhibited Wnt/β-catenin signaling by targeting SOX9, and its expression inversely correlated with that of SOX9 in breast cancer samples. Furthermore, ERα and ZEB1 competitively regulated miR-190 expression. Conclusions Our data uncover the ZEB1/ERα-miR-190-SOX9 axis and suggest a mechanism by which the Wnt/β-catenin signaling pathway is involved in breast cancer anti-estrogen therapy. Electronic supplementary material The online version of this article (10.1186/s13046-019-1039-9) contains supplementary material, which is available to authorized users.

PA-X is a fusion protein encoded by a +1 frameshifted open reading frame (X-ORF) in PA gene. The X-ORF can be translated in full-length (61 amino acids, aa) or truncated (41 aa) form. However, the role of C-Terminal 20 aa of PA-X in virus function has not yet been fully elucidated. To this end, we constructed the contemporary influenza viruses with full and truncated PA-X by reverse genetics to compare their replication and pathogenicity. The full-length PA-X virus in MDCK and human A549 cells conferred 10- to 100-fold increase in viral replication, and more virulent and caused more severe inflammatory responses in mice relative to corresponding truncated PA-X virus, suggesting that the terminal 20 aa could play a role in enhancing viral replication and contribute to virulence.

Introduction: A real-time RT-PCR method for identification and quantification of porcine epidemic diarrhoea virus (PEDV) strains in China was developed. Material and Methods: Based on the conserved sequence of the PEDV nucleocapsid (N) gene, a primer pair and probe were designed to establish a TaqMan-MGB real-time RT-PCR assay for quantitative detection of the virus. The sequence was cloned into the pMD18-T vector and a series of diluted recombinant plasmids were used to generate a standard curve with an R2 value of 0.999. Results: The developed quantitative PCR assay detected viral titres as low as 0.1 TCID50 with high specificity and no cross-reaction with other porcine viruses (PoRV, TGEV, PRRSV, or CSFV). The intra-batch and inter-batch coefficients of variation were both less than 1%, which indicated good reproducibility. Thirty clinical diarrhoea samples obtained from pigs in Shanghai and Fujian were analysed using this quantitative PCR assay. Out of these samples, 93.3% were found to be PEDV positive. Conclusion: This approach is suitable for clinical sample identification and pathogenesis studies.

Swine influenza viruses (SwIVs) cause considerable morbidity and mortality in domestic pigs, resulting in a significant economic burden. Moreover, pigs have been considered to be a possible mixing vessel in which novel strains loom. Here, we developed and evaluated a novel M2e-multiple antigenic peptide (M2e-MAP) as a supplemental antigen for inactivated H3N2 vaccine to provide cross-protection against two main subtypes of SwIVs, H1N1 and H3N2. The novel tetra-branched MAP was constructed by fusing four copies of M2e to one copy of foreign T helper cell epitopes. A high-yield reassortant H3N2 virus was generated by plasmid based reverse genetics. The efficacy of the novel H3N2 inactivated vaccines with or without M2e-MAP supplementation was evaluated in a mouse model. M2e-MAP conjugated vaccine induced strong antibody responses in mice. Complete protection against the heterologous swine H1N1 virus was observed in mice vaccinated with M2e-MAP combined vaccine. Moreover, this novel peptide confers protection against lethal challenge of A/Puerto Rico/8/34 (H1N1). Taken together, our results suggest the combined immunization of reassortant inactivated H3N2 vaccine and the novel M2e-MAP provided cross-protection against swine and human viruses and may serve as a promising approach for influenza vaccine development.

The classical swine (CS) H1N1 swine influenza virus (SIVs) emerged in humans as a reassortant virus that caused the H1N1 influenza virus pandemic in 2009, and the European avian-like (EA) H1N1 SIVs has caused several human infections in European and Asian countries. Development of the influenza vaccines that could provide effective protective efficacy against SIVs remains a challenge. In this study, the bivalent reassortant inactivated vaccine comprised of SH1/PR8 and G11/PR8 arboring the hemagglutinin (HA) and neuraminidase (NA) genes from prevalent CS and EA H1N1 SIVs and six internal genes from the A/Puerto Rico/8/34(PR8) virus was developed. The protective efficacy of this bivalent vaccine was evaluated in mice challenged with the lethal doses of CS and EA H1N1 SIVs. The result showed that univalent inactivated vaccine elicited high-level antibody against homologous H1N1 viruses while cross-reactive antibody responses to heterologous H1N1 viruses were not fully effective. In a mouse model, the bivalent inactivated vaccine conferred complete protection against lethal challenge doses of EA SH1 virus or CS G11 virus, whereas the univalent inactivated vaccine only produced insufficient protection against heterologous SIVs. In conclusion, our data demonstrated that the reassortant bivalent inactivated vaccine comprised of SH1/PR8 and G11/PR8 could provide effective protection against the prevalent EA and CS H1N1 subtype SIVs in mice.

Both classical swine fever (CSF) and pseudorabies are highly contagious, economically significant diseases of swine in China. Although vaccination with the C-strain against classical swine fever virus (CSFV) is widely carried out and severe outbreaks of CSF seldom occur in China, CSF is sporadic in many pig herds and novel sub-subgenotypes of CSFV endlessly emerge. Thus, new measures are needed to eradicate CSFV from Chinese farms. The emergence of a pseudorabies virus (PRV) variant also posed a new challenge for the control of swine pseudorabies. Here, the recombinant PRV strain JS-2012-ΔgE/gI-E2 expressing E2 protein of CSFV was developed by inserting the E2 expression cassette into the intergenic region between the gG and gD genes of the gE/gI-deletion PRV variant strain JS-2012-ΔgE/gI. The recombinant virus was stable when passaged in vitro. A single vaccination of JS-2012-ΔgE/gI-E2 via intramuscular injection fully protected against lethal challenges of PRV and CSFV. Vaccination of piglets with the recombinant JS-2012-ΔgE/gI-E2 in the presence of high levels of maternally derived antibodies (Abs) to PRV can provide partial protection against lethal challenge of CSFV. Vaccination of the recombinant PRV JS-2012-ΔgE/gI-E2 strain did not induce the production of Abs to the gE protein of PRV or to the CSFV proteins other than E2. Thus, JS-2012-ΔgE/gI-E2 appears to be a promising recombinant marker vaccine candidate against PRV and CSFV for the control and eradication of the PRV variant and CSFV.

Objective Investigating the expression of the lnc RNAs screened above between normal and insulin resistant 3T3-L1 adipocytes. Addressing the mechanism underlying the regulation of inflammation response by lnc TINCR. Methods 3T3-L1 preadipocytes were induced to differentiate into mature adipocytes. Oil red O staining was used to find the fat droplets in mature adipocytes. Mature adipocytes were randomized to normal control group and Tri-DAP (NOD1 ligand) group. After the establishment of insulin resistance model, we used deep RNA sequencing(RNA-Seq) to identify lncRNAs that are regulated during NODI activation in mouse adipocytes. Real-time PCR was used to analyze the expression of lnc TINCR, proinflammatory IL-6, TNF-α, Cxcl1 and RIPK2 in the presence or absence of Tri-DAP(10 μg/ml). We employed siRNA against lnc TINCR to confirm its effects in inflammatory response. Results Deep RNA sequencing identified 81 lncRNAs and 167 coding genes that were significantly up-related while 78 lncRNAs and 82 coding genes that were significantly down-related greater than twofold during NOD1 activation in adipocytes. We discovered that lnc TINCR, termed lnc TINCR(Tri-DAP-inducible non-protein coding RNA) is greatly upregulated in Tri-DAP activated adipocytes. Moreover knockdown of lnc TINCR dampens the proinflammatory response (P Conclusions lnc TINCR is a positive regulator of inflammation-induced insulin resistance presumably via activation of NOD1 signaling pathways.

Dogs are a major source of indoor allergens. However, the prevalence of dog allergies in China remains unclear, especially in children. In the present study, Can f 7, a canine allergen belonging to the Niemann pick type C2 protein family, was selected to study its sensitization rate in Chinese children with dog allergies. The Can f 7 gene was subcloned into a pET‑28a vector and expressed in Escherichia coli BL21 (DE3) cells. Recombinant Can f 7 was purified by nickel affinity chromatography, identified by SDS‑PAGE electrophoresis, and had its allergenicity assessed by western blot, ELISA and basophil activation tests. Through a series of bioinformatical approaches, B‑cell epitopes, secondary structures, and 3 dimensional (3D) homology modeling of Can f 7 were predicted. The activity of the B cell epitopes was verified by ELISA. The recombinant Can f 7 showed a distinct band with a molecular weight of 14 kDa. Six of 20 sera from dog‑allergic children reacted positively to the Can f 7. Can f 7 induced an ~4.0‑fold increase in cluster of differentiation 63 and C‑C motif chemokine receptor R3 expression in basophils sensitized with the serum of dog‑allergic children compared with those of non‑allergic controls. The secondary structure analysis showed that Can f 7 contains 6 β‑sheets. Five B cell epitopes of Can f 7 were predicted, and two of these were confirmed by ELISA. These results indicate that Can f 7 is an important canine allergen in Chinese children and provide novel data for further research concerning the use of Can f 7 in the diagnosis and treatment of Chinese children with canine allergy symptoms.

The purified whole-virus proteins derived from A/swine/Shanghai/1/2014 (H1N1) (SH1) were chosen to immunize BALB/c mice to prepare the monoclonal antibody (MAb) against hemagglutinin (HA) protein of an European avian-like (EA) H1N1 swine influenza virus (SIV). After cloning three times by limiting dilution, one strain of hybridoma cells named 3C7 secreting anti-HA protein MAb was obtained by hybridoma technique. The results of indirect immunofluorescence assay and western blot analyses showed that the MAb 3C7 specifically reacted with the HA protein of EA H1N1 SIV. This work indicated that the MAb 3C7 would be a valuable tool as a specific diagnostic reagent for SIV epidemiological surveys and identification of HA protein epitopes of the EA H1N1 SIVs in the future.

Swine influenza A viruses (SIVs) causing outbreaks of acute, highly contagious respiratory disease in pigs also pose a potential threat to public health. European avian-like H1N1 (EA H1N1) SIVs are the predominant circulating viruses in pigs in China and also occasionally cause human infection. In this study, a high-growth reassortant virus (SH1/PR8), with HA and NA genes from a representative EA H1N1 isolate A/Swine/Shanghai/1/2014 (SH1) in China and six internal genes from the high-growth A/Puerto Rico/8/34 (PR8) virus, was generated by plasmid-based reverse genetics and tested as a candidate seed virus for the preparation of inactivated vaccine. The protective efficacy of inactivated SH1/PR8 was evaluated in mice and pigs challenged with wild-type SH1 virus. After primer and boost vaccination, the SH1/PR8 vaccine induced high-level hemagglutination inhibiting (HI) antibodies, IgG antibodies, and neutralization antibodies in mice and pigs. Mice and pigs in the vaccinated group showed less clinical phenomena and pathological changes than those in the unvaccinated group. In conclusion, the inactivated high-growth reassortant vaccine SH1/PR8 could induce high antibody levels and complete protection is expected against SH1 wild type SIV, and protection against heterologous EA H1N1 SIV needs further evaluation.

The Japanese encephalitis virus (JEV) envelope (E) protein, as one of mediators of virus entry into host cells, plays a critical role in determining virulence. The Glu-to-Lys mutation of residue 138 in E protein (E138) plays an important role in attenuating JEV vaccine strain SA14-14-2. However, it is not clear how E138 attenuates JEV. Here, we demonstrate that the Glu-to-Arg mutation of E138 also determines the attenuation of JEV strain 10S3. Likewise, for its parent strain (HEN0701), a virulence strain, the mutations of E138 are responsible for virulence alteration. Furthermore, we demonstrated that mutations of alkaline residues in E138 contributed to the attenuation of neurovirulence; in contrast, mutations of acidic residues enhanced the neurovirulence of the strains. Moreover, acidity in residue E47 had a similar effect on neurovirulence. Furthermore, the alkaline E138 residue enhanced susceptibility to heparin inhibition in vitro and limited JEV diffusion in mouse brain. These results suggest that the acidity/alkalinity of the E138 residue plays an important role in neurovirulence determination. IMPORTANCE The E protein is the only glycoprotein in mature JEV, and it plays an important role in viral neurovirulence. E protein mutations attenuate JEV neurovirulence through unclear mechanisms. Here, we discovered that E138 is a predominant determinant of JEV neurovirulence. We demonstrated that the alkalinity/acidity of E138 determines JEV neurovirulence. These data contribute to the characterization of the E protein and the rational development of novel JEV vaccines.

MicroRNAs (miRNAs) contribute to gene regulation at the post-transcriptional level and are capable of mRNA silencing by binding to target sites exhibiting high degrees of complementarity. Therefore, cloning host miRNA-recognition sequences into the genome of RNA viruses represents a rational strategy for manipulating viral replication. Here, we performed deep sequencing to obtain small-RNA (sRNA)-expression profiles from in vitro-cultured MARC-145 cells post infection with porcine reproductive and respiratory syndrome virus (PRRSV) and chose six candidate miRNAs of different abundance (miR-21, miR-140-3p, miR-185, miR-26a, miR-505, and miR-199a) for further study. Based on the full-length cDNA clone p7USC, we constructed a number of PRRSV mutants that provided complementary base-pairing target sites for the miRNAs in 3′ untranslated regions. Our results showed that all low- and moderate- abundant miRNA-target mutants showed similar growth properties, whereas the highest-abundant miRNA-target mutant blocked both viral transcription and replication. Discontinuous mutations in high-abundant miRNA-target sites subsequently recovered viral viability and propagation. These results demonstrated the copy number of endogenous miRNAs and the extent of sRNA complementarity were key factors to silence potential mRNA expression/translation, thereby determining PRRSV viability. Interestingly, the mutant containing miR-140-target sites (v140-t) showed strong suppression of viral replication from P1 to P3 in vitro, as shown by virus titer, plaque morphology, and qRT-PCR assays. To assess genetic stability, sequencing of v140-t (P1, P3, P5 and P10) revealed spontaneous mutations preferentially located among several nucleotides near the 3′ end of the insertion region and corresponding to the “seed region” of miR-140-3p, explaining the induced viral repression and the direction of virus evolution. This approach provided a general silencing strategy for limiting PRRSV replication by endogenous miRNAs in MARC-145 cells.

Porcine reproductive and respiratory syndrome (PRRS) and classical swine fever (CSF) are economically significant diseases that affect the swine industry worldwide. However, the current vaccination strategy, which uses two single live attenuated vaccines, can result in interference for each other. In addition, the universally used CSFV vaccine C-strain does not allow for differentiation of infected and vaccinated animals. In this study, rPRRSV-E2, PRRS virus (PRRSV) expressing CSF virus (CSFV) E2, was constructed by reverse genetics. The E2 gene of CSFV was inserted between ORF1b and ORF2 in the genome of the PRRS vaccine virus, HuN4-F112. A copy of transcriptional regulatory sequence 6 was inserted at the 3' terminal of the exogenous gene to produce CSFV E2 as a unique subgenomic mRNA transcript. The rPRRSV-E2 was stable for at least 25 serial cell passages. Single-shot intramuscular immunization of rPRRSV-E2 into pigs induced PRRSV-specific and CSFV-specific antibodies and fully protected pigs from lethal challenge with highly-pathogenic PRRSV and CSFV. These results demonstrate that a novel strategy for recombinant PRRSV production is effective, and suggest that rPRRSV-E2 is a promising live, virus-vectored vaccine against PRRS and a marker vaccine against CSF.

Recently pseudorabies outbreaks have occurred in many vaccinated farms in China. To identify genetic characteristics of pseudorabies virus (PRV) strains, we obtained the genomic sequences of PRV strains HeN1 and JS, which were compared to 4 PRV genomes and 729 partial gene sequences. PRV strains isolated in China showed marked sequence divergence compared to European and American strains. Phylogenetic analysis revealed that for the first time PRV can be divided into 2 distinct clusters, with Chinese strains being genotype II and PRVs isolated from other countries being genotype I. Restriction fragment length polymorphism analysis confirmed differences between HeN1 and Bartha strains, as did the presence of unique insertion/deletion polymorphisms and microsatellites. This divergence between the two genotypes may have been generated from long-term, independent evolution, which could also explain the low efficacy of the Bartha vaccine in protecting pigs infected with genotype II PRV.

Tannin grafted on mesoporous silica nanoparticles (tannin-MSNs) was synthesized by the amidation reaction of carboxyl benzyl borate with amino group modified MSN. Tannin-MSNs were an MCM-41 type of mesoporous material with a uniform diameter of about 150 nm. The XRD and N2 adsorption/desorption isotherm results demonstrated that the rhodamine was effectively loaded and the pores of MSN were blocked by tannin. Rhodamine as a model cargo was used to evaluate the loading and releasing behaviors of tannin-MSNs. The drug release rate of tannin-MSNs was pH dependent. An in vitro cellular cytotoxicity test indicated that tannin-MSNs were highly biocompatible and suitable to be used as drug carriers. The cellular uptake was investigated by confocal laser scan microscopy. The results showed that tannin-MSNs are promising platforms to construct a pH-responsive controlled drug delivery system for cancer and stomach therapy.

Since the highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) variant emerged in 2006, it has caused death in more than 20 million pigs in China and other Southeast Asian countries, making it the most destructive swine pathogen currently in existence. To characterize the cellular responses to HP-PRRSV infection, the gene expression profile of porcine alveolar macrophage (PAM) cells, the primary target cells of PRRSV, was analyzed in HP-PRRSV-infected and uninfected PAMs by suppression subtractive hybridization. After confirmation by Southern blot, genes that were differentially expressed in the HP-PRRSV-infected and uninfected PAMs were sequenced and annotated. Genes that were upregulated mainly in HP-PRRSV-infected PAM cells were related to immunity and cell signaling. Among the differentially expressed genes, Mx1 and HSP70 protein expression was confirmed by western blotting, and IL-8 expression was confirmed by ELISA. In PAM cells isolated from HP-PRRSV-infected piglets, the differential expression of 21 genes, including IL-16, TGF-beta type 1 receptor, epidermal growth factor, MHC-I SLA, Toll-like receptor, hepatoma-derived growth factor, FTH1, and MHC-II SLA-DRB1, was confirmed by real-time PCR. To our knowledge, this is the first study to demonstrate differential gene expression between HP-PRRSV-infected and uninfected PAMs in vivo. The results indicate that HP-PRRSV infection excessively stimulates genes involved in the innate immune response, including proinflammatory cytokines and chemokines.

The sterile alpha motif and HD domain 1 (SAMHD1) protein is a novel innate immunity restriction factor that inhibits HIV-1 infection in myeloid cells. Here, we cloned the full-length SAMHD1 complementary DNA (cDNA) from porcine peripheral blood lymphocytes. The porcine SAMHD1 cDNA was of 3951 bp with an open reading frame of 1884 bp, encoding a polypeptide of 627 amino acids. Porcine SAMHD1 mRNA was detected in all swine tissues examined, with the higher expression in the tonsil, lung, liver, and lymph node tissues. The SAMHD1 protein was localized to the nucleus. Overexpression of SAMHD1 blocked the proliferation of HuN4, a highly pathogenic strain of porcine reproductive and respiratory syndrome virus (HP-PRRSV), in MARC-145 cells, by inhibiting the synthesis of the HuN4 complement RNA. The antiviral effects of the simian SAMHD1 protein were nearly equivalent to those of porcine SAMHD1 in the HuN4-infected MARC-145 cells. Phosphorylation analysis of SAMHD1 showed that overexpressed SAMHD1 protein was in primarily an unphosphorylated state. SAMHD1 overexpression increased the transcript abundance of IFN-stimulated genes ISG15 and ISG56. The mRNA levels of SAMHD1 and ISGs were significantly increased in porcine alveolar macrophages infected with HP-PRRSV. SAMHD1 protein level was also elevated, and the protein was not phosphorylated during infection. Collectively, our data indicate that SAMHDI inhibits HP-PRRSV proliferation through inhibiting the replication of HP-PRRSV. SAMHD1 might be the protein participating in the IFN signaling and is thus an important immunoregulatory protein in innate immunity.

Platanus acerifolia is one of the major sources of outdoor allergens to humans, and can induce allergic asthma, rhinitis, dermatitis and other allergic diseases. Pla a 2 is a polygalacturonase and represents the major allergen identified in P. acerifolia pollen. The aim of the present study was to express and purify Pla a 2, and to predict B and T cell epitopes of Pla a 2. The gene encoding Pla a 2 was cloned into the pET28a vector and subsequently transfected into ArcticExpress™ (DE3) Escherichia coli cells; purified Pla a 2 was analyzed by western blot analysis. The results of the present study revealed that the Pla a 2 allergen has the ability to bind immunoglobulin E within the sera of patients allergic to P. acerifolia pollen. In addition, the B cell epitopes of Pla a 2 were predicted using the DNAStar Protean system, Bioinformatics Predicted Antigenic Peptides and BepiPred 1.0 software; T cell epitopes were predicted using NetMHCIIpan ‑3.0 and ‑2.2. In total, eight B cell epitopes (15‑24, 60‑66, 78‑86, 109‑124, 232‑240, 260‑269, 298‑306 and 315‑322) and five T cell epitopes (62‑67, 86‑91, 125‑132, 217‑222 and 343‑350) were predicted in the present study. These findings may be used to improve allergen immunotherapies and reduce the frequency of pollen‑associated allergic reactions.

Swine influenza viruses have been circulating in pigs throughout world and might be potential threats to human health. PA-X protein is a newly discovered protein produced from the PA gene by ribosomal frameshifting and the effects of PA-X on the 1918 H1N1, the pandemic 2009 H1N1, the highly pathogenic avian H5N1 and the avian H9N2 influenza viruses have been reported. However, the role of PA-X in the pathogenesis of swine influenza virus is still unknown. In this study, we rescued the H1N1 wild-type (WT) classical swine influenza virus (A/Swine/Guangdong/1/2011 (H1N1)) and H1N1 PA-X deficient virus containing mutations at the frameshift motif, and compared their replication properties and pathogenicity of swine influenza virus in vitro and in vivo. Our results show that the expression of PA-X inhibits virus replication and polymerase activity in cultured cells and decreases virulence in mouse models. Therefore, our study demonstrates that PA-X protein acts as a negative virulence regulator for classical H1N1 swine influenza virus and decreases virulence by inhibiting viral replication and polymerase activity, deepening our understanding of the pathogenesis of swine influenza virus.