Wei Si

Northeast Agricultural University, Charbin, Heilongjiang Sheng, China

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Publications (16)42.51 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: This study evaluated maternal immunity against Salmonella enterica serovar Enteritidis acquired through the egg yolk. Two-hundred 19-week-old specific pathogen free (SPF) broiler breeders which were randomly divided into two groups of equal size were injected with S. Enteritidis ghosts (5 x 109 colony forming units in 0.1 ml per hen) and phosphate-buffered saline (PBS, 0.01 mol⋅L-1, pH 7.4) twice, respectively, with an interval of 2 weeks. An indirect Enzyme-Linked Immunosorbent Assay (ELISA) was applied to detect specific antibodies against S. Enteritidis. S. Enteritidis-specific antibody levels in the vaccinated group increased over time and were significantly higher than those of the control group on day 28 (p < 0.001) and 35 (p < 0.001) post-vaccination. Ten 7-day-old chicks from hens that were vaccinated with a S. Enteritidis ghost vaccine were challenged at 14 days of age with 5 x 109 CFU of S. Enteritidis DH091 (homologous to the vaccine strain), 8/10 (80%) chicks from vaccinated hens survived, whereas 3/10 (30%) chicks from unvaccinated hens survived. The chicks acquired high levels of serum antibodies against S. Enteritidis. These results reveal that maternal antibodies in chicks acquired from vaccinated hens through eggs can confer a significant protection against S. Enteritidis infection.
    Research in Veterinary Science. 01/2014;
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    ABSTRACT: Newcastle disease virus (NDV) is a member of the Paramyxovirinae subfamily and can infect most species of birds. It has been a great threat for the poultry industry all around the world. In this report, we successfully produced infectious pseudotyped pNL4-3-Luc-R-E- (HIV-Luc) viruses with the HN and F envelope proteins of NDV. Further investigation revealed the cytoplasmic domains of HN and F, especially HN, plays a significant role in the infection efficiency of these pseudotyped HIV-Luc viruses. Replacement of, or direct fusion to the cytoplasmic domain of the HN protein by that of vesicular stomatitis virus G (VSV-G) could greatly enhance or destroy the infective potential of HN and F-pseudotyped (NDV-pseudotyped) HIV-Luc virus. We further established a novel neutralization assay to evaluate neutralizing antibodies against NDV with the NDV-pseudotyped HIV-Luc viruses. Comparative neutralization data indicate that the results determined by using the NDV-pseudotyped HIV-Luc viruses are as reliable as those by the conventional virus-neutralization assay (VN test) with native NDV. Moreover, the results show that the novel neutralization assay is more sensitive than the VN test.
    PLoS ONE 01/2014; 9(6):e99905. · 3.53 Impact Factor
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    ABSTRACT: The occurrence and counts of Listeria monocytogenes were investigated in a total of 526 retail raw food samples. All L. monocytogenes isolates were further analyzed by serotyping and antimicrobial susceptibility assays. The molecular basis of tetracycline resistance of each isolate and the genetic relatedness were determined. L. monocytogenes isolates were found in 12.4% (65/526) of the samples, with counts below 102 CFU/g. L. monocytogenes was most commonly isolated from pork (20%, 20/100), seafood (13.8%, 15/109), chicken (13.2%, 14/106), and beef (10.3%, 11/107). In addition, L. monocytogenes was also detected in 4.8% (5/104) of raw mutton samples. Four serogroups were identified among the 65 L. monocytogenes isolates, with serogroups 1/2a-3a (60%) and 4b-4d-4e (24.6%) being dominant. Most L. monocytogenes isolates were resistant to cefotaxime (54.6%), fosfomycin (51.5%), and clarithromycin (36.4%). Some isolates showed intermediate resistance to streptomycin (12.1%), norfloxacin (13.6%), ciprofloxacin (13.6%), and nitrofurantoin (9.1%). Multiple resistances were observed in 72.3% of isolates. Genetic relatedness analysis revealed that there were no prominent associations between specific food types, serotypes, antimicrobial susceptibility profiles and Pulsed-field gel electrophoresis (PFGE) patterns. In addition, these isolates were multiresistant and belonged to the epidemiologically important serotypes 1/2a and 4b, implying a potential public health risk.
    Food Control 07/2013; 32(1):153–158. · 2.74 Impact Factor
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    ABSTRACT: The collagenase cathepsin K has been shown important in the pathogenesis of rheumatoid arthritis (RA). Icariin is the major pharmacologically active flavonol diglycoside of Herba Epimedii, an herb used in Chinese traditional medicine to treat arthritis. We investigated whether icariin can inhibit the protease activity of cathepsin K and its effects on a murine model of collagen-induced arthritis (CIA). Six-week old female BALB/C mice were immunized with type II collagen and treated with vehicle alone icariin (25mg/kg) for 21 days; a control remained untreated. Serum concentrations of type I collagen C-terminal telopeptide (CTX-I) and cartilage oligomeric matrix protein (COMP) and urinary concentrations of deoxypyridinoline (DPD) were measured, and disease severity was assessed. Compared with immunized, untreated mice, immunized icariin-treated mice had significantly lower urinary DPD (~25%, p<0.01) and serum COMP (~11.9%, p<0.01) concentrations, with serum CTX-1 (RatLaps) concentrations being significantly lower in immunized, icariin treated mice than in immunized, vehicle treated (p<0.01) and non-immunized (p<0.005) mice. Icariin also reduced the clinical signs of arthritis. Icariin inhibited cathpesin K activity in vitro and was effective in a mouse model of CIA similar to human RA, suggesting that this agent may have promise in the treatment of patients with RA.
    Phytomedicine: international journal of phytotherapy and phytopharmacology 06/2013; · 2.97 Impact Factor
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    ABSTRACT: Commercial bacterins for Glässers disease are widely used for prevention this disease caused by Haemophilus parasuis, however, the protective efficacy varied depending on the strains and serovars. Bacterial ghosts (BGs) are empty bacterial envelopes, unlike classical bacterins, suffer no denaturing steps during their production. These properties may lead to a superior protection. In this study, a BGs vaccine generated from the Haemophilus parasuis serovar 5 reference strain Nagasaki was prepared and used to inoculate piglets. The efficacy of the BGs vaccine was evaluated by clinical, bacteriological, serological, and post-mortem examinations. Inactivated bacterin (IB) and a placebo control (PC) were compared with the BGs vaccine in this study. The results showed that the piglets inoculated with the BGs vaccine developed a higher antibody activity and higher IFN-γ and IL-4 levels than those vaccinated with IB or the PC group after primary and secondary exposure to the antigens and challenge. The CD4(+) T lymphocytes were observed to increase following secondary immunization in the BGs-vaccinated group more than in the IB (p<0.05) and PC (p<0.05) groups. The CD8(+) T lymphocytes increased dramatically in all three groups after challenge, and the differences between each group all were significant (p<0.05). There were fewer tissue lesions and lower bacterial load in the tissue homogenates observed in the BGs group after challenge. The results suggested that higher CD4(+) T lymphocytes rate and both the CD4(+) MHC-II restricted Th1 type and Th2 type immune responses in the BGs group appear relevant to protection.
    Clinical and vaccine Immunology: CVI 03/2013; · 2.60 Impact Factor
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    ABSTRACT: Bacterial ghosts offer a new avenue for the study of inactivated vaccines. However, for many years the mechanism of genetic inactivation was controversial. To obtain mouse monoclonal antibodies (mAbs) against protein E will allow the observation of its location and dynamic expression to expand understanding of the lysis mechanism. In this study, a His-tagged ΔE fusion protein expressed in bacteria was used as the immunogen, and mAbs targeting protein E were produced by the hybridoma technique and selected by enzyme-linked immunosorbent assay using GST-E and GST-ΔE as coating proteins. Purified mAbs from mouse ascites were screened against a phage-displayed random dodecapeptide library (Ph.D.-12). After three rounds of biopanning, 30 phage clones were randomly selected and sequenced, and their amino acids were deduced. One epitope showed a good match with protein E at 57-63aa (KPLN--R) and the synthetic decapeptide Epep (VRLKPLNCSR) strongly inhibited combination of E-A5 with E fusion proteins. Immunofluorescence microscopy indicated specific immunoreactivity of E-A5 with Escherichia coli-expressed native protein E. The novel mAbs may be of great potential value in analysis of the function and lysis pathway of protein E and other relative phages and in evaluation of E as potential marker of bacterial ghosts.
    Journal of virological methods 03/2013; · 2.13 Impact Factor
  • Journal of Antimicrobial Chemotherapy 02/2013; · 5.34 Impact Factor
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    ABSTRACT: In this study, a hybridoma-based technique and phage display technology were used to obtain mouse monoclonal antibodies (mAb) against cold-shock DEAD-box protein A (CsdA) from Mycobacterium tuberculosis and to determine the location of the relevant epitope. One highly specific mAb, named A3G5, was developed against the recombinant CsdA protein (rCsdA) and could detect rCsdA protein in enzyme-linked immunosorbent assays (ELISA) and Western blot assays. By screening a phage displayed library of random 12-mers (Ph.D.-12), 10 positive phage clones were randomly selected after three rounds of bio-panning and identified by ELISA. Eight of these clones were sequenced, and their amino acid sequences were deduced. One B-cell epitope (-APDPPLSRR-) in the rCsdA protein was identified with mAb A3G5. A synthetic peptide (-MAPDPPLSRR-) (Cpep) matched well with the CsdA sequence at 443-451 aa and was confirmed by affinity ELISA, competitive inhibition assays and the development of an immune response in mice. These results may be of great potential value in the further analysis of the function and structure of the CsdA protein from M. tuberculosis.
    Research in Veterinary Science 12/2012; · 1.77 Impact Factor
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    ABSTRACT: In this study, we immunized mice with prokaryotically expressed recombinant surface layer protein, SapA, of Campylobacter fetus, generated hybridomas secreting mouse monoclonal antibodies (mAb) targeting SapA, and purified the mAb A2D5 from mouse ascites using saturated ammonium sulfate solution. The mAb A2D5, coated onto ELISA plates, was used to screen the phage random 12-peptide library through three rounds of panning. Following panning, 15 phage clones were randomly chosen and tested for reactivity with mAb A2D5 by indirect ELISA. Single-stranded DNA from positive clones was sequenced and compared with the sequence of SapA to predict the key epitope. ELISA and/or Western blot analyses further validated that synthetic peptides and recombinant peptide mimotopes all interact with mAb A2D5. Nine of ten positive phage clones identified by screening were sequenced successfully. Seven clones shared the same sequence HYDRHNYHWWHT; one had the sequence LSKNLPLTALGN; and the final one had the sequence SGMKEPELRSYS. These three sequences shared high homology with SapA J05577 in the region GNEKDFVTKIYSIALGNTSDVDGINYW, in which the underlined amino acids may serve as key residues in the epitope. ELISA and/or Western blot analyses showed that mAb A2D5 not only interacted with the four synthetic peptide mimotopes, but also with 14 prokaryotically expressed recombinant peptide mimotopes. The mimotopes identified in this study will aid future studies into the pathological processes and immune mechanisms of the SapA protein of C. fetus.
    Research in Veterinary Science 03/2012; 93(3):1274-80. · 1.77 Impact Factor
  • Journal of Antimicrobial Chemotherapy 02/2012; 67(5):1287-9. · 5.34 Impact Factor
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    ABSTRACT: We studied throat swabs and corresponding serum samples collected from 1067 protein purified derivative (PPD)-tuberculin skin test (TST) positive cattle from different regions of China. The 1067 throat swabs were inoculated onto modified Löwenstein-Jensen medium for the isolation and culture of Mycobacteria. Acid-fast bacilli were identified using traditional biochemical methods, polymerase chain reaction (PCR) amplification and multiplex PCR. They were distinguished as Mycobacterium tuberculosis complex (MTBC) and non-tuberculous mycobacteria (NTM) strains. An indirect Enzyme-Linked Immunosorbent Assay (ELISA) was applied to detect specific antibodies against bovine TB (bTB). Correlations among the ELISA, bacteriology and TST were analyzed and compared. Spoligotyping and variable number tandem repeats-mycobacterial interspersed repetitive unit (VNTR-MIRU) analysis were used to genotype the MTBC. In total, 111 strains of Mycobacteria were cultured from the 1067 throat swab samples, including 43 stains of MTBC (14 strains of Mycobacterium bovis and 29 of Mycobacterium tuberculosis) and 68 strains of NTM. Thirty-eight MTBC strains and four NTM strains were isolated from 72 throat swab samples that the ELISA determined were antibody positive; five MTBC strains and 64 NTM strains were isolated from 995 throat swab samples that were antibody negative on the ELISA. The positive isolation rates of MTBC and NTM were 38.7% (43/111) and 61.3% (68/111), respectively. The concordance rate of cultured MTBC with a positive result on the indirect ELISA for antibody was 52.8% (38/72), which was much higher than the positive rate for TST (4.0%; 43/1067). Genotyping of the 43 strains of MTBC isolated, using spoligotyping and VNTR-MIRU, showed that the 43 isolates had 26 genotypes; 16 strains had a unique genotype. Two groups of six strains and two strains, respectively, showed the same spoligotyping pattern, and belonged to the Beijing family and Beijing-like family, respectively. Combined application of spoligotyping and VNTR-MIRU typing would improve the molecular epidemiological investigation and monitoring of the etiology of bTB in China.
    Research in Veterinary Science 06/2011; 90(3):385-91. · 1.77 Impact Factor
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    ABSTRACT: Bacterial ghosts that are generated using the regulated PhiX174 lysis gene E offer a new avenue for the study of inactivated vaccines. Here, we constructed a library of mutant gene E using a gene-shuffling technique. After screening and recombination with the prokaryotic non-fusion expression vector pBV220, two lysis plasmids were selected. Among which, a novel mutant E gene (named mE), consisting of a 74-bp non-encoding sequence at 5'-end and a 201-bp gene ΔE, significantly increased the lysis effect on prokaryotic Escherichia coli and Salmonella enteritidis. Moreover, lysis efficiency, as measured by the OD600 value, reached 1.0 (10⁹ CFU), avoiding the bottleneck problem observed with other bacterial lysis procedures, which results in a low concentration of bacteria in suspension, and consequent low production of bacterial ghosts. Our results may provide a promising avenue for the development of bacterial ghost vaccines.
    Virology Journal 01/2011; 8:206. · 2.09 Impact Factor
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    ABSTRACT: Bacterial ghosts (BGs) are empty bacterial envelopes generated by expulsion of the bacterial genome and cytoplasmic contents from bacterial cells, and the process is mediated by lysis protein E encoded on bacteriophage PhiX174. BGs represent a new approach in vaccine development and have been applied to a variety of gram-negative bacterial vaccine candidates. In this study, a BG vaccine generated from Salmonella enteritidis (S. enteritidis) strain DH091 was prepared using the highly efficient plasmid, pBV-mE. The efficacy of the BG vaccine was tested using 75 chicks (Gallus gallus) kept under specific pathogen-free (SPF) conditions. A comprehensive evaluation of the immune response, including humoral and cellular immune responses, interferon-γ (IFN-γ) and interleukin-4 (IL-4) production, and histopathology of various tissues, was performed in BG-vaccinated animals subsequently challenged with S. enteritidis. The results were compared with animals that were immunized with the inactivated vaccine. S. enteritidis ghosts not only promoted the generation of high titer antibodies and IFN-γ and IL-4 production but also stimulated a significant increase in CD8(+) and CD4(+) T lymphocytes. In particular, the dramatic increase in CD8(+) T cells indicated that the vaccine was able to induce clearance of intracellular Salmonella. The protective effects of BG vaccination in SPF chicks against 5×10(9) colony forming units of S. enteritidis were a result of the induction of a more effective immune response than that observed with the inactivated vaccine. These findings demonstrate the potential of S. enteritidis ghosts to be used as effective vaccines.
    Immunobiology 10/2010; 216(5):558-65. · 2.81 Impact Factor
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    ABSTRACT: Campylobacteriosis is a zoonosis that occurs worldwide. Infection with Campylobacter fetus (C. fetus) causes infertility and abortion in sheep and cattle. The current study focuses on the SapA gene of C. fetus that encodes surface array proteins and plays an important role in the virulence of C. fetus. The SapA-N (1398bp) and SapA-C (1422bp) fragments were amplified from the C. fetusSapA gene using polymerase chain reaction (PCR), and the corresponding recombinant proteins rSapA-N and rSapA-C were expressed in Escherichia. coli BL21 cells. Results of Western blotting and enzyme-linked immunosorbent assay (ELISA) showed that the immunological activity of rSapA-N was higher than that of rSapA-C (P<0.05). Therefore, rSapA-N was selected to establish an indirect ELISA for detecting antibodies against C. fetus. The diagnostic criteria were as follows: S/P0.45: positive; S/P<0.4: negative; 0.45>S/P0.4: suspected. The specificity and sensitivity of our method were 94.3% and 88.6%, respectively. Moreover, no cross-reactions were observed between rSapA-N and serum samples that were positive for other bovine bacterial pathogens diseases such as Mycobacterium avium subspecies paratuberculosis. One hundred and two serum samples from cows that had experienced abortion were tested. Four and 2 C. fetus-positive serum samples were found among the 70 bovine brucellosis-positive samples and the 32 infectious bovine rhinotracheitis (IBR)-positive samples, respectively. The findings suggest that the rSapA-N-based ELISA method has immense potential in future applications.
    Research in Veterinary Science 06/2010; 88(3):446-51. · 1.77 Impact Factor
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    ABSTRACT: A multiplex reverse transcription-nested polymerase chain reaction (RT-nPCR) method was developed for the detection and differentiation of wild-type and vaccine strains of canine distemper virus (CDV). A pair of primers (P1 and P4) specific for CDV corresponding to the highly conserved region of the CDV genome were used as a common primer pair in the first-round PCR of the nested PCR. Primers P2 specific for CDV wild-type strains, were used as the forward primer together with the common reverse primer P4 in the second round of nested PCR. Primers P3, P5 specific for CDV wild-type strain or vaccine strain, were used as the forward primer together with the common reverse primer P4+P6 in the second round of nested PCR. A fragment of 177 bp was amplified from vaccine strain genomic RNA, and a fragment of 247 bp from wild-type strain genomic RNA in the RT-nPCR, and two fragments of 247 bp and 177 bp were amplified from the mixed samples of vaccine and wild-type strains. No amplification was achieved for uninfected cells, or cells infected with Newcastle disease virus (NDV), canine parvovirus (CPV), canine coronavirus (CCV), rabies virus (RV), or canine adenovirus (CAV). The RT-nPCR method was used to detect 30 field samples suspected of canine distemper from Heilongjiang and Jilin Provinces, and 51 samples in Shandong province. As a result of 30 samples, were found to be wild-type-like, and 5 to be vaccine-strain-like. The RT-nPCR method can be used to effectively detect and differentiate wild-type CDV-infected dogs from dogs vaccinated with CDV vaccine, and thus can be used in clinical detection and epidemiological surveillance.
    Virology Journal 01/2010; 7:86. · 2.09 Impact Factor
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    ABSTRACT: The role of in vivo-induced ApxIV toxin of Actinobacillus pleuropneumoniae in protective immunity was evaluated in pigs by administering it alone or added to a multicomponent recombinant subunit vaccine composed of recombinant ApxI, ApxII, ApxIII toxin, and 42-kDa outer membrane protein (OMP). The pigs were immunized with vaccine I (rApxIVN), vaccine II (rApxI+rApxII+rApxIII+rApxIVN+rOMP), vaccine III (rApxI+rApxII+rApxIII+rOMP), or placebo (phosphate-buffered saline+adjuvant). A. pleuropneumoniae serovar 1 field isolate JMS 06 and serovar 2 field strain FX 01 were used as the challenge strains. Pigs that were immunized with vaccine I or vaccine II all developed high antibody titers against rApxIVN. The antibody titers against rApxI, rApxII, rApxIII, and rOMP in pigs immunized with vaccine II were higher than those in pigs vaccinated with vaccine III. Following the challenge, the pigs immunized with rApxIVN alone showed similar results to the pigs in the control group, such as severe respiratory symptoms and severe lung lesions. Pigs that had been immunized with vaccine II or vaccine III were protected against challenge with A. pleuropneumoniae serovar 1 and serovar 2. The pigs immunized with vaccine II had slighter lung lesions and fewer bacterial recovery than those of pigs immunized with vaccine III. These results indicate that rApxIVN contributes to the production of high level of antibodies directed against the vaccination antigens, and thus confers strong protection against challenges with different serovars of A. pleuropneumoniae.
    Vaccine 09/2009; 27(42):5816-21. · 3.77 Impact Factor