Subunit vaccines are being developed as a potential therapy for preventing microbial pathogen infection. In this study, the immunogenicity of recombinant Brucella (B.) abortus Fe/Mn superoxide dismutase (rFe/Mn SOD) protein as a subunit vaccine against B. abortus was investigated in BALB/c mice model. Brucella Fe/Mn SOD gene was cloned into a pcold-TF DNA vector. The bacterial recombinant protein was expressed using the Escherichia coli DH5α strain with a size of 82.50 kDa. The western blotting assay showed that rFe/Mn SOD reacted with Brucella-positive serum, indicating the potential immunoreactivity of this recombinant protein. After the second and third vaccinations, the peripheral CD4+ T cell population was increased significantly in the rFe/Mn SOD-immunized mice group compared to the PBS control group. Moreover, immunization of this recombinant protein increased the CD4+ T cell population from the first vaccination to the third vaccination. Meanwhile, the CD8+ T cells were slightly enhanced after the second vaccination compared to the first vaccination and compared to control groups. Fourteen days after the bacterial infection, the splenomegaly and the number of bacteria in the spleen were evaluated. The result showed that both rFe/Mn SOD and positive control RB51 decreased the bacterial replication in the spleen and the splenomegaly compared to control groups. Altogether, these results suggested that rFe/Mn SOD could induce host immunity against B. abortus infection.

Chlorine dioxide (ClO2) has recently emerged as an ideal disinfectant and has shown a wide range of antimicrobial activities in various pathogenic microorganisms. In this study, the virucidal effect of ClO2 at low concentration (0.02 ppm) and higher concentration (0.06 – 0.09 ppm) against Adenovirus and Herpesvirus was evaluated based on the NF T 72-281 and ASTM 1053-11 standard methods at different exposure times. The virus suspension was dried onto the carrier and then exposed to gaseous ClO2 (gClO2) at 22 ± 2∘C. For Adenovirus, exposure at a low concentration of ClO2 at the middle height resulted in the average log10 reduction of 0.95, 2.65, and 5.30 after 1, 3, and 6 h post-exposure (pe), respectively. Moreover, more than 4-log10 reduction was achieved at 4 and 6 h pe with higher concentrations of ClO2. On the other hand, the antiviral activity of gClO2 at the middle height was also effective against Herpesvirus. In particular, at 1 h pe, a less than 4-log10 reduction was observed at all examined concentrations of ClO2, whereas exposure for 3 and 6 h (with low concentration) or 2 h (with higher concentration) inactivated completely viruses attached to the carrier. These results suggested that ClO2 fumigation is a potential alternative method for disinfecting healthcare facilities, high-containment laboratories, and households with a safe concentration for human health.

The structural safety of prototype transport and storage containers for very low-level radioactive liquid waste was experimentally estimated for its localization development. Transport containers for radioactive liquid waste have been researched and developed, however, there are no standardized commercial containers for very low-level radioactive waste in Korea. In this study, the structural safety of the designated IP-2 type container capable of transporting and temporarily storing large amounts of very low-level liquid waste, which is generated during the operation and decommissioning of nuclear power plants, was demonstrated. The stacking and drop tests, which were conducted to determine the structural integrity of the container, verified that there was no external leakage of the contents in spite of its structural deformation due to the drop impact. This study shows the effort required for the localization of the technology used in manufacturing transport and storage containers for very low-level radioactive liquid waste, and the additional structural reinforcement of the container in which the commercial intermediate bulk container (IBC) external frame was coupled.

We investigated the effect of a synthetic complement peptide C3a on the outcome of Brucella abortus 544 infection in a murine macrophage cell line RAW264.7 cell. First, we determined the highest non-cytotoxic concentration of the peptide in the cell line. We also found that the peptide significantly increased the growth of the bacteria at 8 and 24 h. Although the number of bacterial CFU was also elevated at 48 and 72 h, the increases were not significant as compared to controls. We further investigated the effect of C3a peptide on the growth of Brucella by pre-incubating the peptide at various temperatures and found that the effect was reversed at 24 h post-incubation suggesting that incubation of peptide at high temperatures including 65°C or 95°C could inactivate its action. This also could indicate the beneficial effect of high temperature during infection. Although several studies reported the inhibitory effect of different antimicrobial peptides including C3a, the present study preliminarily revealed that it had no positive contribution on the control of B. abortus 544 infection in vitro and indirectly to its receptor, CD88, which belongs to GPCR. Moreover, the encouraged further exploration of the effect of other similar peptides would be performed for the purpose of finding Brucella-host cell interaction for the control of disease progression.

This study aims to investigate the effects of exogenous succinic acid (SCA) on Brucella (B.) abortus infection in macrophage RAW 264.7 cells and ICR mice. Firstly, the in vitro experiment was conducted by MTT cytotoxicity and bacterial internalization assay to evaluate the uptake of B. abortus into macrophage cells. Two non-cytotoxic concentrations of SCA demonstrated attenuated invasion of Brucella into macrophages at 30 and 45 min post- infection (pi). Secondly, ICR mice were treated with SCA and infected with B. abortus. On day-14 pi, spleen and blood serum were collected to evaluate the bacterial burden and total spleen weight as well as the production of cytokine/chemokine, respectively. The results showed that SCA treatment promoted bacterial growth and reduced the total spleen weight in mice. Furthermore, SCA treatment increased the level of IL-10 cytokine in the sera, while dampening the production of MCP-1 chemokine compared to the control. The results of bacterial load in spleen and spleen weight together with cytokine/chemokine production profile in the sera indicated that SCA induced the host anti-inflammatory response which is beneficial for the survival of Brucella. Therefore, these findings suggest that SCA contributed to host immunity against Brucella infection and the emerging potential topic-immunometabolism should be invested for further investigations.

We investigated the cytotoxic potential of three different commercially available absorbent feminine hygiene products and one transdermal patch using direct contact and extract exposure methods. Two different cell lines were used – mouse fibroblast L929 and normal human skin fibroblast CCD-986sk cell lines. The test samples were extracted using three different methods in accordance to International Organization for Standardization (ISO). Viability of cells was analyzed using MTT assay and morphology of the cells were also observed using phase contrast microscopy. Overall, the direct contact method using L929 cells showed that all the test samples had no toxic effect when exposed to extracts for 1 h. For the exposure method, no toxic effect was observed in both L929 and CCD986sk cells incubated with all the test samples regardless of the extraction methods used.

This study investigated the efficacy of four Brucella (B.) abortus recombinant proteins, namely adenylate kinase (Adk), nucleoside diphosphate kinase (Ndk), 50S ribosomal protein (L7/L12) and preprotein translocase subunit (SecB), as a combined subunit vaccine (CSV) against B. abortus infection in BALB/c mice. Immunoblotting assay showed that these four recombinant proteins as well as pcold-TF vector reacted individually with Brucella-positive serum, but not with Brucella-negative serum. The peripheral blood CD4+ T cell population was increased in CSV-immunized mice compared to PBS and pcold-TF vector groups. In addition, CSV and pcold-TF groups displayed induced IgG1 and IgG2a antibodies production compared to PBS and RB51 group, whereas IgG2a titer was higher than IgG1 titer in CSV group. The secretion profiles of IgG1 and IgG2a production together with an enhancement of CD4+ T cell population suggested that CSV did not only induce T helper 1 (Th1) T cell immunity but also humoral immunity. Therein, Th1 T cell immunity is more predominant in eliminating intracellular bacteria B. abortus. Furthermore, CSV immunization significantly reduced the bacterial burden in the spleen as well as the spleen weight in comparison to PBS and pcold-TF groups. Altogether, combination of these antigens could be potential to induce protective immunity against B. abortus infection in animals.

BALB/c mice were vaccinated with Brucella (B.) abortus recombinant protein L27 (50S ribosomal protein L27) cloned into a pMal vector system. L27 was induced, purified and injected intraperitoneally (IP). Mice were vaccinated on 0-, 15- and 35-day. Serum cytokines were evaluated on 36- and 49-day from first vaccination. Mice were intraperitoneally infected with 5×104 CFU of virulent B. abortus 544 on day-50 and sacrificed after two weeks from infection. Bacterial burden from the spleen was quantified and showed a 0.7- and 0.9-log reduction in vaccinated mice in comparison to PBS and MBP (maltose binding protein) groups respectively. Cytokines in the serum demonstrated increased interferon-gamma (IFN-γ) and other pro-inflammatory cytokines such as macrophage chemoattractant protein-1 (MCP-1) and interleukin 6 (IL-6). On the other hand, interleukin 10 (IL-10) was attenuated in the sera of vaccinated mice. This cytokine profile is indicative of a cell-mediated type of immune response which is favorable for the eradication of intracellular infections. The current study showed the potential of another B. abortus ribosomal protein in inducing protective immunity against B. abortus infection.

In this study, we examined the protective immunity of a combination of seven Brucella abortus recombinant proteins; superoxide dismutase (rSodC), riboflavin synthase subunit beta (rRibH), 50S ribosomal protein (50s rL7/L12), nucleoside diphosphate kinase (rNdk), malate dehydrogenase (rMDH), arginase (rRocF), and elongation factor (rTsf) cloned in a pMal vector system and expressed in DH5α. Mice groups were immunized thrice with a combined subunit vaccine (CSV-7) at 0, 2, and 5 weeks and subsequently challenged with B. abortus at 5 × 104 CFU at 6 weeks. At four weeks post-infection, the mice were sacrificed and the bacterial burden in their spleens was quantified. Results revealed bacterial log reductions of 0.63 and 0.34 in comparison to PBS and maltose-binding protein (MBP), respectively. Cytokine profiling revealed a marked increase in IFN-γ (interferon-gamma), MCP-1 (macrophage chemoattractant protein-1) and IL-6 (interleukin 6) cytokines at 5-weeks post-immunization. On the other hand, only TNF was heightened at 7-weeks post-immunization. In general, this cytokine profile is consistently reflective of a Th1 immune response, which is beneficial for host immunoresistance.

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 DH5α: 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 5×104 CFU 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.

To date, most serodiagnostic methods for brucellosis screening are based on antibodies against lipopolysaccharides of Brucella spp. However, this approach has the drawback of yielding false-positive results due to cross-reactivity with lipopolysaccharides of other related pathogens, especially Yersinia enterocolitica O:9. In this study, Brucella abortus AspC was cloned and expressed by PCR amplification into a pCold TF expression system to obtain recombinant AspC (rAspC). The immunogenicity of rAspC was confirmed by western blotting of Brucella-positive bovine serum. rAspC-based ELISA was performed to determine whether rAspC could be used in the serodiagnosis of bovine brucellosis. rAspC reacted strongly with anti-Brucella antibodies in positive sera in the tube agglutination test (TAT), but did not show strong reaction with most negative samples. In particular, the average OD492 value at the highest TAT titer showed a 1.4-fold increase with respect to the cutoff value. The accuracy, specificity, and sensitivity of rAspC were 71.88%, 78.33%, and 68%, respectively. These findings suggest that rAspC might be valuable for the serological diagnosis of bovine brucellosis.

Brucellosis is an important and re-emerging zoonotic disease worldwide. The prevention of human infection is achieved predominantly through the control of brucellosis in agricultural animals, which in turn depends on accurate diagnosis and vaccination. However, conventional serological diagnosis of brucellosis has several limitations, and currently available vaccines for animals have several drawbacks, including the ability to cause infection in humans. Phosphoglycerate kinase (Pgk) is one of the specific proteins reactive with mouse sera in the early stage of Brucella infection, and deletion of the pgk gene in B. abortus strain 2308 resulted in extreme attenuation of this strain in vitro and in vivo. Furthermore, the B. abortus pgk mutant has been used as a live vaccine, and in challenge experiments, it induced protection that was superior to that conferred by commercial strains. In this study, the pgk gene from Brucella abortus 544 was successfully amplified and cloned into a maltose binding protein fusion protein expression vector (pMAL). The recombinant protein was expressed in Escherichia coli DH5α and purified. The immunogenicity of purified recombinant B. abortus 544 Pgk (rPgk) was evaluated by western blot analysis using Brucella-positive mouse sera. rPgk could be used as an antigenic component for future serological tests and potential vaccine development.

Brucellosis is a notorious zoonotic disease with global implications. Efforts to control the spread of the disease have been restricted to the agricultural livestock. Increasing incidences of accidental human infection have motivated researches to start working on alternative vaccines. At present, live attenuated vaccines are the only accepted type of vaccines used in developed countries for the prevention of brucellosis. Although serodiagnosis is occasionally unreliable, some countries have already claimed to have eradicated the disease, based on this testing. Live attenuated vaccines are not suitable for use in pregnant and immune-depressed animals. Moreover, these vaccines are not tolerated in humans. Therefore, many researches have been striving to discover alternative methods of vaccination. Most research has focused on the generation of subcellular, subunit, and DNA vaccines that are as efficient as the live attenuated vaccines. At present, none of the available vaccines has been able to replace the live attenuated vaccines. Therefore, additional research is necessary in order to discover a new brucellosis vaccine that is suitable for human use.

Brucellosis is an important bacterial zoonotic infection worldwide responsible for economic losses in livestock industries which represents a considerable and increasing public health burden. The disease is caused by Brucella which possess the ability to invade and replicate within phagocytes and may result to chronic infection, leading to difficulty in medical therapy of the disease. The treatment for brucellosis employs conventional principles that have been applied for a long time and a combination of antibiotics is currently used due to low efficacy of monotherapies. However, vital effects with respect to health and safety are neglected. Nevertheless, the preventive methods and treatments for brucellosis using traditional medicine have not yet been thoroughly studied, hence, alternative therapies such as the use of natural plant extracts as traditional medicine that are safe, efficient and economical should be explored in order to identify candidates that eliminate complications due to brucellosis. Coptis chinensis Franch (Huanglian) is a traditional Chinese medicinal herb and its extract has been known to possess strong antibacterial activity. In this study, the antibacterial effects of C. chinensis Franch extracts, and the major components of the herb namely berberine and palmatine, were investigated on B. abortus. The C. chinensis Franch ethanol extracts (CCFE) showed bactericidal effects at 1,000 μg/ml concentration and berberine at 100 μg/ml concentration. However, C. chinensis Franch and its components did not affect invasion and intracellular growth of B. abortus in RAW 264.7 cells. In conclusion, C. chinensis Franch ethanol extracts, water extract and its major components such as berberine and palmatine would be a beneficial antimicrobial agent without affecting phagocytic pathway within macrophages, and further study for the precise mechanisms of antibacterial effect would be necessary.

To profile the proteome in porcine plasma, blood samples were collected from adult male barrows and those plasma were retrieved. For the depletion or pre-fractionation of high-abundance proteins, plasma samples were treated with commercial kits. Then, protein profiling was initiated using one and two-dimensional electrophoresis. Proteins were spotted and then identified by MALDI-TOF-TOF and LC-MS-MS. In the results, more than forty six proteins were identified and the reference map was constructed. The pre-treatment for the removal of high-abundance proteins caused the changes in 2-DE images and some of the proteins were newly uncovered after the most of high abundant proteins were removed. However, it is expected for further steps necessary to identify more low-abundance proteins that may contain potential bio-markers.

Here, we present an approach of blood plasma proteome profiling and their comparisons between the young and the adult pigs as prerequisite for the identification of bio-markers related to the health conditions, growth performance and meat quality. To profile the proteome in porcine plasma, blood samples were collected from 19 young piglets and 20 adult male barrows and the plasma was retrieved. Then, protein profiling was initiated using one and two-di-mensional electrophoresis. Proteins were spotted and then identified by MALDI-TOF-TOF and LC-MS-MS. In the re-sults, more than thirty-six and twenty eight protein spots were selected in young piglets and adult pigs, respectively and twenty three proteins were identified. The proteome profile images were compared between those ones using Image Master Version 7.0. The image of expressed proteome showed that most of proteins from plasma of young pig-let separated clearly and concentrated in 2DE display compared to ones from adult. Image analysis in detail was car-ried out to look for the specific proteins related to age progression. It demonstrated that the characteristics of proteome expression could be distinct to their age stages. Further investigations needed to proceed to understand the age de-pendent change of protein conformation and biological meaning of those differences in proteome expression between young and mature adult pigs.

Muscle satellite cell (SC) is responsible for postnatal muscle growth, repair, and regeneration. Satellite cell is an im-portant source of multi-potent stem cell process and differentiation into adipogenic, myogenic, and osteoblastogenic. The objective of this study was to identify alter of transcriptome during differentiation in porcine satellite cell and to elevated transcriptome at different stages of postnatal development to gain insight into the differences in differ-entiated PSC. We used RNA-seq technique to investigate the transcriptomes during differentiation in pig muscle. Sequence reads were obtained from Illumina HiSeq2000. Differentially expressed genes (DEG) were detected by EdgeR. Gene ontology (GO) terms are powerful tool for unification among representation genes or products. In study of GO biological terms, functional annotation clustering involved in cell cycle, apoptosis, extracellular matrix, phosphoryla- tion, proteolysis, and cell signaling in differences stage. Taken together, these results would be contributed to a better understanding of muscle biology and processes underlying differentiation. Our results suggest that the source of DEGs could be better understanding of the mechanism of muscle differentiation and transdifferentiation.

Satellite cells were derived from muscular tissue in postnatal pig. Satellite cell is an important to growth and development in animal tissues or organs. However, the progress underlying induced differentiation is not clear. The aim of this study was to evaluate the morphologic and the transcriptome changes in porcine satellite cell (PSC) treated with insulin, rosiglitazone, or dexamethasone respectively. PSC was obtained from postnatal muscle tissue. In study 1, for study the effect of insulin and FBS on the differentiated satellite cells, cells were cultured at absence or presence of insulin treated with FBS. Total RNA was extracted for determining the expression levels of myo-genic PAX3, PAX7, Myf5, MyoD, and myogenin genes by real-time PCR. Myogenic genes decreased expression levels of mRNA in treated with insulin. In study 2, in order to clarify the relationship between rosiglitazone and lipid in differentiated satellite cells, we further examined the effect of FBS on lipid accumulation in the presence or absence of the rosiglitazone and lipid. Significant differences were observed between rosiglitazone and lipid by FBS. The mRNA of FABP4 and PPARγ increased in rosiglitazone treatment. In study 3, we examined the effect of dexame-thasone on osteogenic differentiation in PSC. The mRNA was increased osteoblasotgenic ALP and ON genes treated with dexamethasone in 2% FBS. Dexamethasone induces osteoblastogenesis in differentiated PSC. Taken together, in differentiated PSCs, FABP4 and PPARγ increased to rosiglitazone. Whereas, no differences to FBS and lipid. These results were not comparable with previous reports. Our results suggest that adipogenic, myogenic, and osteoblasto-genic could be isolated from porcine skeletal muscle, and identify culture conditions which optimize proliferation and differentiation formation of PSC.