• Parasites, Hosts and Diseases
• /
• 제51권2호
• /
• pp.155-163
• /
• 2013

This study aimed to evaluate the efficacy of fructose-1,6-bis phosphate aldolase (SMALDO) DNA vaccination against Schistosoma mansoni infection using different routes of injection. The SMALDO has been cloned into the eukaryotic expression vector pcDNA3.1/V5-His TOPO-TA and was used in injecting Swiss albino mice intramuscularly (IM), subcutaneously (SC), or intraperitoneally (IP) ($50{\mu}g/mouse$). Mice vaccinated with non-recombinant pcDNA3.1 served as controls. Each group was immunized 4 times at weeks 0, 2, 4, and 6. Two weeks after the last booster dose, all mice groups were infected with 80 S. mansoni cercariae via tail immersion. At week 8 post-infection, animals were sacrificed for assessment of parasitological and histopathological parameters. High anti-SMALDO IgG antibody titers were detected in sera of all vaccinated groups (P<0.01) compared to the control group. Both the IP and SC vaccination routes resulted in a significant reduction in worm burden (46.2% and 28.9%, respectively, P<0.01). This was accompanied by a significant reduction in hepatic and intestinal egg counts (41.7% and 40.2%, respectively, P<0.01) in the IP group only. The number of dead eggs was significantly increased in both IP and IM groups (P<0.01). IP vaccination recorded the highest significant reduction in granuloma number and diameter (54.7% and 29.2%, respectively, P<0.01) and significant increase in dead miracidia (P<0.01). In conclusion, changing the injection route of SMALDO DNA vaccination significantly influenced the efficacy of vaccination. SMALDO DNA vaccination via IP route could be a promising protective and antipathology vaccine candidate against S. mansoni infection.

• 대한수의학회지
• /
• 제46권2호
• /
• pp.127-133
• /
• 2006

Pasteurella multocida is a terrible veterinary pathogen that causes widespread infections in husbandry. To induce homologous and/or heterologous immunity against the infections, outer membrane protein Hs (OmpH) in the envelope of different strains of P. multocida are thought to be attractive vaccine candidates. Previously we cloned and characterized a gene for OmpH from pathogenic P. multocida (A : 3) (In Press, Korean J. Microbiol. Biotechnol. 2005, 33, December). The gene is composed of 1,047 nucleotides (nt) coding 348 amino acids (aa) with signal peptide of 20 aa. The truncated ompH, a gene without nt coding for the signal peptide, was generated using pRSET A to name "pRSET A/OmpH-F2". This truncated ompH was well expressed in Escherichia coli BL21 (DE3). Truncated OmpH was purified for induction of immunity against live pathogen of fowl cholera (P. multocida A : 3) in mice. Some $50{\mu}g$ of the purified polypeptide was intraperitoneally injected into mice two times with 10 day interval. Lethal dose ($25{\mu}l$) of live P. multocida A : 3 was determined by directly injecting the pathogen into wild mice (n = 25). To demonstrate the vaccine candidate of the truncated OmpH, the live pathogen ($25{\mu}l$) was challenged with the OmpH-immunized mouse group as well as positive & negative controls (n = 80). The results show that the truncated OmpH can be used for an effective vaccine production to prevent fowl cholera caused by pathogenic P. multocida (A : 3).

• Fisheries and Aquatic Sciences
• /
• 제25권11호
• /
• pp.559-571
• /
• 2022

Galectins, a family of ß-galactoside-binding lectins, have emerged as soluble mediators in infected cells and pattern recognition receptors (PRRs) responsible for evoking and regulating innate immunity. The present study aimed to evaluate the role of galectin-1 in the host immune response of redlip mullet (Liza haematocheilia). We established a cDNA database for redlip mullet, and the cDNA sequence of galectin-1 (LhGal-1) was characterized. In silico analysis was performed, and the spatial and temporal expression patterns in gills and blood in response to lipopolysaccharide polyinosinic:polycytidylic acid, and Lactococcus garvieae were estimated via quantitative real-time PCR. Functional assays were conducted using recombinant protein to investigate carbohydrate binding, bacterial binding, and bacterial agglutination activity. LhGal-1 was composed of 135 amino acids. Conserved motifs (H-NPR, -N- and -W-E-R) within the carbohydrate recognition domain were found in LhGal-1. The tissue distribution revealed that the healthy stomach expressed high levels of LhGal-1. The temporal monitoring of LhGal-1 mRNA expression in the gill and blood showed its significant upregulation in response to immune challenges with different stimulants. rLhGal-1 exhibited binding activity in response to carbohydrates and bacteria. Moreover, the agglutination of rLhGal-1 against Escherichia coli was observed. Collectively, our findings suggest that LhGal-1 may function as a PRR in redlip mullet. Furthermore, LhGal-1 can be considered a significant gene to play a protective role in redlip mullet immune system.

• Journal of Microbiology and Biotechnology
• /
• 제16권8호
• /
• pp.1180-1184
• /
• 2006

The capsule that surrounds Yersinia pestis cells is composed of a protein-polysacchride complex; the purified protein component is fraction I (F1) antigen. We report the cloning of the cafl gene and its expression in Escherichia coli using the vector pETl02/D-TOPO and the F1-specific monoclonal antibody. The recombinant F1 (rF1) antigen had a molecular size of 17.5 kDa, which was identical to that of the F1 antigen produced by Y. pestis. Recombinant F1 protein was found to react to polyclonal antiserum to Y. pestis Fl. Recombinant F1 was purified by ProBond purification system and induced a protective immune response in BALB/c mice challenged with up to 10$^5$ virulent Y. pestis. Purified rF1 protein was used in an ELISA to evaluate the ability of a method to detect antibodies to Y. pestis in animal sera. These results strongly indicated that the rF1 protein is a suitable species-specific immunodiagnostic antigen and vaccine candidate.

• Journal of Microbiology and Biotechnology
• /
• 제11권3호
• /
• pp.482-490
• /
• 2001

The gene for glycoprotein B (gB2) of HSV-2-strain G was subcloned, sequenced, recombinated into the lacZ-HcNPV, expressed in insect cells, and compared with the homologous gene of other HSV-2 strains. The ORF of the gB2 gene was 2,715 bp. The overall nucleotide sequence homology of te gB2 gene compared ith that of the two previously reported HSV-2 strains appeared to be over 98%. A recombinant virus named Baculo-gB2 protein in insect cells. The recombination was confirmed by a PCR and the expression was demonstrated by radio immunoprecipitation. Insect cells infected with the Baculo-gB2 virus synthesized and processed gB2 with approximately 120 kDa in the cells, and then secreted it into the culture media, where it reacted with a nomoclonal antibody to gB2. The gB2 polypeptide contained two main hydrophobic regions (a signal sequence from 1 to 23 amino acid residues, and a membrane anchor sequence from aa 745 to 798), eight N-glycosylation sites evenly distributed, and was rich in alanine (11.2%). Antibodies to this recombinant protein that were raised in mice recognized the viral gB2 and neutralized the infectivity of the HSV-2 in vitro. There results show that the gB2 protein was successfully porduced in insect cells and could be used to raise a protective neutralizing antibody. Accordingly, this particular recombinant protein may be useful in the development of a subunit vaccine.

• Journal of Preventive Veterinary Medicine
• /
• 제42권4호
• /
• pp.157-162
• /
• 2018

This study evaluated the protective effects of a combination of eight B. abortus recombinant proteins that were cloned and expressed into a pMal vector system and $DH5{\alpha}$: nucleoside diphosphate kinase (rNdk), 50S ribosomal protein (rL7/L12), malate dehydrogenase (rMDH), DNA starvation/stationary phase protection protein (rDps), elongation factor (rTsf), arginase (rRocF), superoxide dismutase (rSodC), and riboflavin synthase subunit beta (rRibH). The proteins were induced, purified, and administered intraperitoneally into BALB/c mice. The mice were immunized three times at weeks 0, 2, and 5 and then infected intraperitoneally (IP) with $5{\times}10^4CFU$ of virulent B. abortus 544 one week after the last immunization. The spleens were collected and the bacterial burden was evaluated at four weeks post-infection. The results showed that this combination produced a significant reduction of the bacterial burden in the spleen with a log reduction of 1.01 compared to the PBS group. Cytokine analysis revealed induction of the cell-mediated immune response in that TNF (tumor necrosis factor) and proinflammatory cytokines IL-6 (Interleukin 6) and MCP-1 (macrophage chemoattractant protein-1) were elevated significantly. In summary, vaccination with a combination of eight different proteins induced a significant protective effect indicative of a cell mediated immune response.

• Journal of Microbiology
• /
• 제44권5호
• /
• pp.548-555
• /
• 2006

FaeG is the key factor in the infection process of K88ad enterotoxigenic Escherichia coli (ETEC) fimbrial adhesin. In an attempt to determine the possibility of expressing recombinant FaeG with immunogenicity for a new safe and high-production vaccine in E. coli, we constructed the recombinant strain, BL21 (DE3+K88), which harbors an expression vector with a DNA fragment of faeG, without a signal peptide. Results of 15% SDS-polyacrylamide slab gel analysis showed that FaeG can be stably over-expressed in BL21 (DE3+K88) as inclusion bodies without FaeE. Immunoglobulin G (IgG) and M (IgM) responses in pregnant pigs, with boost injections of the purified recombinant FaeG, were detected 4 weeks later in the sera and colostrum. An in vitro villius-adhesion assay verified that the elicited antibodies in the sera of vaccinated pigs were capable of preventing the adhesion of K88ad ETEC to porcine intestinal receptors. The protective effect on the mortality rates of suckling piglets born to vaccinated mothers was also observed one week after oral challenge with the virulent ETEC strain, $C_{83907}$ (K88ad, $CT^+,\;ST^+$). The results of this study proved that the adhesin of proteinaceous bacterial fimbriae or pili could be overexpressed in engineered E. coli strains, with protective immune responses to the pathogen.

• 대한바이러스학회지
• /
• 제28권1호
• /
• pp.71-78
• /
• 1998

Vaccinia virus is the prototype orthopoxvirus that has been used as a vaccine strain for small pox. This virus has been used to express a variety of cellular and viral genes in mammalian cells at high levels. Interleukin-4 (IL-4) has been found to stimulate the proliferation of T cells and enhance the cytolytic activity of cytotoxic T lymphocytes. To test the immunotherapeutic potential of IL-4 delivered in vivo by poxvirus, a recombinant vaccinia virus expressing the murine IL-4 gene (RVVmIL-4) was constructed. A high level of IL-4 production was confirmed by infecting HeLa cells and measuring IL-4 in cell culture supernatant by ELISA. As a tumor model, two cell lines were used; the murine T leukemic line P388 and the murine breast cancer line TS/A. CDF1 mice were intraperitoneally inoculated with $1\;{\times}\;10^5$ cells of P388. Mice were injected at the same site with $5\;{\times}\;10^5\;PFU$ of recombinant vaccinia virus; first, 3 days after the injection of tumor cells and thereafter once every week for 3 weeks. Intraperitoneal injections of RVVmIL-4 significantly prolonged the survival time of mice inoculated with tumor cells. All mice injected with RVVmIL-4 remained alive for 30 days after the postinoculation of tumor cells, while 100% and 70% of the animals injected with saline or wild type vaccinia virus died, respectively. In another tumor model using TS/A, tumor was established by subcutaneously inoculating $2{\times}10^5$ tumor cells to BALB/c mice. After tumor formation was confirmed on day 4 in all mice, $5\;{\times}\;10^6\;PFU$ of RVVmIL-4 was inoculated subcutaneously three times, once every week for 3 weeks. The TS/A tumor was eradicated in two of the nine mice. Seven of the nine mice treated with RVVmIL-4 developed a tumor, but tumor growth was significantly delayed compared to those treated with saline or wild type vaccinia virus. These results indicate that recombinant vaccinia viruses may be used as a convenient tool for delivering immunomodulator genes to a variety of tumors.

• 대한의생명과학회지
• /
• 제18권3호
• /
• pp.210-217
• /
• 2012

Oncolytic viral vectors have shown good candidates for cancer treatment but have many limitations. To improve the therapeutic potential of oncolytic vaccinia virus, we developed a recombinant vaccinia virus expressing the 4-1BBL co-stimulatory molecule or CCL21. 4-1BBL and CCL21 expression was identified by FACS analysis and immunoblotting. rV-4-1BBL vaccination shows significant tumor regression compared to rV-LacZ, but rV-CCL21 shows rapid tumor growth compared to rV-LacZ in the poorly immunogenic B16 murine melanoma model. 4-1BBL expression resulted in the increase of the number of CD8+ T cells and especially the increase of effector (CD62L-CD44+) CD8+ T cells. These data suggest 4-1BBL may be the potential target for enhancement of tumor immunotherapy.

• Journal of Animal Science and Technology
• /
• 제57권4호
• /
• pp.10.1-10.7
• /
• 2015

Control and prevention of foot and mouth disease (FMD) by vaccination remains unsatisfactory in endemic countries. Indeed, consistent and new FMD epidemics in previously disease-free countries have precipitated the need for a worldwide control strategy. Outbreaks in vaccinated animals require that a new and safe vaccine be developed against foot and mouth virus (FMDV). FMDV can be eradicated worldwide based on previous scientific information about its spread using existing and modern control strategies.