• Korean Journal of Microbiology
• /
• v.36 no.4
• /
• pp.254-258
• /
• 2000

Invasion process of bacterial cell into intestinal epithelium is important in Salmonella infection. The invasion is induced by the proteins secreted by type III secretion appratus of Salmonella. It has been known that the proteins do not have N-terminal signal peptide existing in general secreted proteins. Recent studies on Yersinia reported that secretion signal of type III appratus may lie on 5'end secondary structure of mRNA of secreted protein. In this study, we constructed translational fusion of ompR and sopE, encoding type III secretion protein of Salmonella, and observed secretion of the fusion protein for investigating the secretion signal of Salmonella type III appratus. The sopE DNA fragments of the translational fusion contain the region of promoter and from start code to tenth or to fifth code. These translational fusions indicate that type III secretion signal of Salmonella is located on 5'end of mRNA encoding secreted protein. We constructed prototype of secretion vector using this signal to produce useful foreign protein.

• Journal of Microbiology and Biotechnology
• /
• v.26 no.11
• /
• pp.1881-1890
• /
• 2016

Manganese superoxide dismutase (MnSOD) is a vital enzyme that protects cells from free radicals through eliminating superoxide radicals ($O^{2-}$). Hirudin, a kind of small active peptide molecule, is one of the strongest anticoagulants that can effectively cure thrombus diseases. In this study, we fused Hirudin to the C terminus of human MnSOD with the GGGGS linker to generate a novel dual-feature fusion protein, denoted as hMnSOD-Hirudin. The hMnSOD-Hirudin gene fragment was cloned into the pET15b (SmaI, CIAP) vector, forming a recombinant pET15b-hMnSOD-Hirudin plasmid, and then was transferred into Escherichia coli strain Rosetta-gami for expression. SDS-PAGE was used to detect the fusion protein, which was expected to be about 30 kDa upon IPTG induction. Furthermore, the hMnSOD-Hirudin protein was heavily detected as a soluble form in the supernatant. The purification rate observed after Ni NTA affinity chromatography was above 95%. The hMnSOD-Hirudin protein yield reached 67.25 mg per liter of bacterial culture. The identity of the purified protein was confirmed by western blotting. The hMnSOD-Hirudin protein activity assay evinced that the antioxidation activity of the hMnSOD-Hirudin protein obtained was $2,444.0{\pm}96.0U/mg$, and the anticoagulant activity of the hMnSOD-Hirudin protein was $599.0{\pm}35.0ATU/mg$. In addition, in vitro bioactivity assay showed that the hMnSOD-Hirudin protein had no or little cytotoxicity in H9c2, HK-2, and H9 (human $CD_4{^+}$, T cell) cell lines. Transwell migration assay and invasion assay showed that the hMnSOD-Hirudin protein could suppress human lung cancer 95-D cell metastasis and invasion in vitro.

• Korean Journal Plant Pathology
• /
• v.10 no.3
• /
• pp.163-168
• /
• 1994

Erwinia rhapontici causes soft-rot disease in a number of plants such as rhubarb, onion, hyacinth and garlic. Pectate lyase (Pel) depolymerizes pectin and other polygalacturonates, which is though to play a role in bacterial invasion of plants. Pel activity was not detected in E. rhapontici cultured in a minimal salts medium containing glycerol, polygalacturonate, or citrus pectin as a carbon source. However, when sterilized potato tuber and Chinese cabbage slices were added to minimal salts polygalacturonate (0.5%) medium, E. rhapontici produced pectate lyase enzyme. Also Pel activity was consistently detected from macerated potato tubers, Chinese cabbage leaves, lettuce leaves and celery petioles tissue. Pel in the extract of macerated Chinese cabbage caused by E. rhapontici strain 1, resulted in electrolyte loss, tissue maceration and cell death of potato tuber tissue. These results indicate that E. rhapontici produces pectate lyase only in the presence of non-diffusible plant components, and that this enzyme probably contributes to its pathogenicity.

• Proceedings of the Korean Society of Applied Pharmacology
• /
• 1998.11a
• /
• pp.95-98
• /
• 1998

Ginseng (the root of Panax ginseng C. A. MEYER, Araliaceae) has been used for traditional medicine in China, Korea, Japan and other Asian countries for the treatment of various diseases including psychiatric and neurologic diseases as well as diabetes mellitus. So far, ginseng saponins (ginsenosides) have been regarded as the principal components responsible for the pharmacological activities of ginseng. Ginsenosides are glycosides containing an aglycone (protopanaxadiol or protopanaxatriol) with a dammarane skeleton and have been shown to possess various biological activities including the enhancement of cholesterol biosynthesis, stimulation of serum protein synthesis, immuno- modulatory effects and anti-inflammatory activity. Several studies using ginsenosides have also reported anti-tumor effects, particularly the inhibition of tumor-induced angiogenesis, tumor invasion and metastasis, and the control of phenotypic expression and differentiation of tumor cells.

• The Journal of Korean Obstetrics and Gynecology
• /
• v.27 no.1
• /
• pp.65-80
• /
• 2014

Objectives: The object of this study was to observe the in vitro antibacterial effects of Gagam-seopyoungjeon aqueous extracts (GGSYJ) against Gardnerella vaginalis and the possible synergic combination effects with clindamycin. Methods: Antibacterial activities against Gardnerella vaginalis of GGSYJ were detected using minimal inhibition concentration (MIC), and the effects on the bacterial growth curve were also monitored at MIC and MIC${\times}$2 levels. The combination effects of GGSYJ with clindamycin were observed by checkboard microtiter assay, and the effects of bacterial growth curve treated with GGSYJ MIC+clindamycin MIC, 1/2 MIC and 1/4 MIC, respectively. The effects on the bacterial invasion and intracellular killing of GGSYJ were also observed using human vaginal epithelial (VK2) and murine macrophage (Raw264.7) cells with combination effects with clindamycin after treatment of GGSYJ MIC+clindamycin 1/2 MIC, 1/4 MIC and 1/6 MIC, respectively. Results: The MIC of clindamycin and GGSYJ against Gardnerella vaginalis were detected as $0.012{\pm}0.006$ (0.004~0.016)${\mu}g/ml$ and $1.016{\pm}0.524$ (0.391~1.563) mg/ml, respectively. Clindamycin and GGSYJ were also showed marked dosage-dependent inhibition of bacterial growth, and significant decreases of viable cells were detected in clindamycin MIC+GGSYJ MIC and clindamycin 1/2 MIC+GGSYJ MIC treatment as compared with each of single clindamycin MIC and GGSYJ MIC treatments. And significant decreases of intraepithelial and intra-macrophage viable bacteria numbers were detected in clindamycin 1/2 MIC+GGSYJ 1/2 MIC and clindamycin 1/4 MIC+GGSYJ 1/2 MIC treatment as compared with each of single clindamycin GGSYJ 1/2 MIC treatments, respectively. Conclusions: GGSYJ showed slight antibacterial effects against Gardnerella vaginalis, but they showed dosage-dependent inhibitory effects on the bacterial growth and VK2 epithelial invasions of bacteria with favorable accelerating effects of intracellular killing activities of macrophages. In addition, combination of GGSYJ also increased the inhibitory effects of clindamycin on the epithelial invasions of Gardnerella vaginalis and intracellular killing activities of macrophages against Gardnerella vaginalis as 2-fold higher as compared with clindamycin single treatment, respectively. Therefore, we expected that the clinical dosages of clindamycin can be reduced as 1/2 levels as combination with GGSYJ.

• The Journal of Korean Obstetrics and Gynecology
• /
• v.26 no.1
• /
• pp.1-24
• /
• 2013

Objectives: The object of this study was to observe the protective effect of Gamijipaesan aqueous extracts(GJS), which has been traditionally used in Korean medicine in obstetrics & gynecological fields as anti-infectious and anti-inflammatory agents, against mastitis induced by Staphylococcus aureus infection in a rat model through antibacterial, antiinflammatory, immunomodulatory, and anti-oxidant effects. Methods: Antibacterial activities of GJS against S. aureus were detected using standard agar microdilution methods, with the effects on the bacterial invasion and intracellular killing of individual test materials in human mammary gland carcinoma cell(MCF-7) and murine macrophages(Raw 264.7) at MIC1/2, MIC and MIC2 concentration levels. In addition, the effects on the cell viability, nitric oxide(NO), tumor necrosis factor(TNF)-${\alpha}$ and interleukin (IL)-6 productions of LPS activated Raw 264.7 cells. The changes on the mammary tissue viable bacterial numbers, myeloperoxidae(MPO), inducible nitric oxide synthetase(iNOS), TNF-${\alpha}$ and IL-6 contents were observed in the S. aureus in vivo intramammary infectious rat model. The anti-bacterial and anti-inflammatory effects were compared with ciprofloxacin and piroxicam, respectively in the present study. Results: MIC of GJS and ciprofloxacin against S. aureus were detected as $0.860{\pm}0.428$ (0.391-1.563) mg/ml and $0.371{\pm}0.262$(0.098-0.782) ${\mu}g/ml$, respectively. In addition, GJS and ciprofloxacin were also showed marked dosage-dependent inhibition of the both bacterial invasion and intracellular killing assays using MCF-7 and Raw 264.7 cells at MIC1/2, MIC and $MIC{\times}2$ concentrations, respectively. $ED_{50}$ against LPS-induced cell viabilities and NO, TNF-${\alpha}$ and IL-6 releases of GJS were detected as 0.72, 0.04, 0.08 and 0.11 mg/ml, and as 19.04, 4.18, 5.37 and 4.27 ${\mu}g/ml$ in piroxicam, respectively. 250 and 500 mg/kg of GJS also inhibit the intramammary bacterial growth, MPO, iNOS, TNF-${\alpha}$ and IL-6 contents in S. aureus in vivo intramammary infected rats, respectively. GJS 500 mg/kg showed quite similar antibacterial and anti-infectious effects as compared with ciprofloxacin 40 mg/kg and also showed similar anti-inflammatory effects as piroxicam 10 mg/kg, in S. aureus in vivo intramammary infectious models. Conclusions: The results obtained in this study suggest that over 250 mg/kg of GJS showed favorable anti-infectious effects against S. aureus infection in a rat model through their antibacterial, anti-inflammatory, immunomodulatory and anti-oxidant effects and therefore expected that GJS can be used as alternative therapies, having both anti-inflammatory and anti-infectious activities. However, more detail mechanism studies should be conducted in future with the efficacy tests of individual herbal composition of GJS and the screening of the biological active compounds in individual herbs. In the present study, GJS 500 mg/kg showed quite similar anti-infectious effects were detected as compared with ciprofloxacin 40 mg/kg treated rats, and also GJS shows quite similar anti-inflammatory effects as compared with piroxicam 10 mg/kg in S. aureus in vivo intramammary infectious rats, but ciprofloxacin did not showed any anti-inflammatory effects, and piroxicam did not showed anti-infectious effects in this study.

• Journal of Veterinary Science
• /
• v.24 no.6
• /
• pp.82.1-82.12
• /
• 2023

Background: The current conventional serotyping based on antigen-antisera agglutination could not provide a better understanding of the potential pathogenicity of Salmonella enterica subsp. enterica serovar Brancaster. Surveillance data from Malaysian poultry farms indicated an increase in its presence over the years. Objective: This study aims to investigate the virulence determinants and antimicrobial resistance in S. Brancaster isolated from chickens in Malaysia. Methods: One hundred strains of archived S. Brancaster isolated from chicken cloacal swabs and raw chicken meat from 2017 to 2022 were studied. Two sets of multiplex polymerase chain reaction (PCR) were conducted to identify eight virulence genes associated with pathogenicity in Salmonella (invasion protein gene [invA], Salmonella invasion protein gene [sipB], Salmonella-induced filament gene [sifA], cytolethal-distending toxin B gene [cdtB], Salmonella iron transporter gene [sitC], Salmonella pathogenicity islands gene [spiA], Salmonella plasmid virulence gene [spvB], and inositol phosphate phosphatase gene [sopB]). Antimicrobial susceptibility assessment was conducted by disc diffusion method on nine selected antibiotics for the S. Brancaster isolates. S. Brancaster, with the phenotypic ACSSuT-resistance pattern (ampicillin, chloramphenicol, streptomycin, sulphonamides, and tetracycline), was subjected to PCR to detect the corresponding resistance gene(s). Results: Virulence genes detected in S. Brancaster in this study were invA, sitC, spiA, sipB, sopB, sifA, cdtB, and spvB. A total of 36 antibiogram patterns of S. Brancaster with a high level of multidrug resistance were observed, with ampicillin exhibiting the highest resistance. Over a third of the isolates displayed ACSSuT-resistance, and seven resistance genes (β-lactamase temoneira [bla_TEM], florfenicol/chloramphenicol resistance gene [floR], streptomycin resistance gene [strA], aminoglycoside nucleotidyltransferase gene [ant(3")-Ia], sulfonamides resistance gene [sul-1, sul-2], and tetracycline resistance gene [tetA]) were detected. Conclusion: Multidrug-resistant S. Brancaster from chickens harbored an array of virulence-associated genes similar to other clinically significant and invasive non-typhoidal Salmonella serovars, placing it as another significant foodborne zoonosis.

• Parasites, Hosts and Diseases
• /
• v.54 no.4
• /
• pp.385-391
• /
• 2016

The discovery and understanding of antigenic proteins are essential for development of a vaccine against malaria. In Plasmodium falciparum, Pf92 have been characterized as a merozoite surface protein, and this protein is expressed at the late schizont stage, but no study of Pv92, the orthologue of Pf92 in P. vivax, has been reported. Thus, the protein structure of Pv92 was analyzed, and the gene sequence was aligned with that of other Plasmodium spp. using bioinformatics tools. The recombinant Pv92 protein was expressed and purified using bacterial expression system and used for immunization of mice to gain the polyclonal antibody and for evaluation of antigenicity by protein array. Also, the antibody against Pv92 was used for subcellular analysis by immunofluorescence assay. The Pv92 protein has a signal peptide and a sexual stage s48/45 domain, and the cysteine residues at the N-terminal of Pv92 were completely conserved. The N-terminal of Pv92 was successfully expressed as soluble form using a bacterial expression system. The antibody raised against Pv92 recognized the parasites and completely merged with PvMSP1-19, indicating that Pv92 was localized on the merozoite surface. Evaluation of the human humoral immune response to Pv92 indicated moderate antigenicity, with 65% sensitivity and 95% specificity by protein array. Taken together, the merozoite surface localization and antigenicity of Pv92 implicate that it might be involved in attachment and invasion of a merozoite to a new host cell or immune evasion during invasion process.

• Journal of Dental Rehabilitation and Applied Science
• /
• v.40 no.1
• /
• pp.31-38
• /
• 2024

Dens in dente is a developmental anomaly resulting from infolding of the enamel organ into dental papilla prior to calcification of dental tissue. The pulpal tissue of the tooth can be vulnerable for bacterial invasion through direct exposure to the oral cavity or through defective enamel and dentin of the infolding part, thereby increasing the possibility of pulpal necrosis and subsequent apical periodontitis. Treatment planning of teeth with dens invaginatus may be difficult due to the complex root canal morphology. Therefore, thorough knowledge of anatomical variations of dens invaginatus is of great importance for proper treatment planning. The focus of this case report is on Oehler's type II and III dens invaginatus. The infolding of type III dens invaginatus extends beyond the crown and CEJ. Bacterial invasion through the infolding can easily cause inflammation of the pulpal and periradicular tissue. This case report presents endodontic treatment of type II and III dens invaginatus with the aid of CBCT.

• Molecules and Cells
• /
• v.38 no.3
• /
• pp.273-278
• /
• 2015

The induction of angiogenesis is a crucial step in tumor progression, and therefore, efficient inhibition of angiogenesis is considered a powerful strategy for the treatment of cancer. In the present study, we report that the lipophilic antimicrobial peptides from EML-CAP3, a new endophytic bacterial strain isolated from red pepper leaf (Capsicum annuum L.), exhibit potent antiangiogenic activity both in vitro and in vivo. The newly obtained antimicrobial peptides effectively inhibited the proliferation of human umbilical vein endothelial cells at subtoxic doses. Furthermore, the peptides suppressed the in vitro characteristics of angiogenesis such as endothelial cell invasion and tube formation stimulated by vascular endothelial growth factor, as well as neovascularization of the chorioallantoic membrane of growing chick embryos in vivo without showing cytotoxicity. Notably, the angiostatic peptides blocked tumor cell-induced angiogenesis by suppressing the expression levels of hypoxia-inducible $factor-1{\alpha}$ and its target gene, vascular endothelial growth factor (VEGF). To our knowledge, our findings demonstrate for the first time that the antimicrobial peptides from EML-CAP3 possess antiangiogenic potential and may thus be used for the treatment of hypervascularized tumors.