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( Hannah Leah Tadeja Simborio ),( Alisha Wehdnesday Bernardo Reyes ),( Huynh Tan Hop ),( Lauren Togonon Arayan ),( Wongi Min ),( Hu Jang Lee ),( Jin Ju Lee ),( Hong Hee Chang ),( Suk Kim ) 한국미생물 · 생명공학회 2016 Journal of microbiology and biotechnology Vol.26 No.3
Brucellosis affects a wide range of host species, including humans and many livestock animals. Chronic infections of the disease make antibiotic treatment costly, and the current vaccine used in livestock has not been approved for human use. This study investigated the possible use of the Brucella abortus outer membrane protein A (OmpA) as a candidate subunit vaccine in an infected mouse model. The ompA gene was cloned and overexpressed, and the recombinant OmpA (rOmpA) protein fused to maltose binding protein (MBP) was purified in Escherichia coli. Immunogenicity was verified through western blotting, and mice were immunized and challenged to evaluate its protective effect. Mice treated with rOmpA exhibited induced humoral and host cell-mediated responses, with a significant increase in immunoglobulin G (IgG1 and IgG2a) and cytokine levels, especially TNF-α and IL-12, compared with the control groups treated with either MBP or PBS. In conclusion, rOmpA should be highly considered as a future subunit vaccine for brucellosis, and further studies regarding rOmpA and its protective ability are suggested.
Review Article : Strategies for the development of an effective vaccine against Brucellosis
( Hannah Leah Tadeja Simborio ),( Alisha Wehdnesday Bemardo Reyes ),( Huynh Tan Hop ),( Lauren Togonon Arayan ),( Won Gi Min ),( Hu Jang Lee ),( Hong Hee Chang ),( Suk Kim ) 한국예방수의학회 2014 예방수의학회지 Vol.38 No.2
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.
Development and Evaluation of a Rapid Dipstick Assay for Serodiagnosis of Bovine Brucellosis
Hannah Leah Tadeja Simborio,Alisha Wehdnesday Bernardo Reyes,Huynh Tan Hop,Lauren Togonon Arayan,Kwang Myun Cheong,김석 경상대학교 농업생명과학연구원 2016 농업생명과학연구 Vol.50 No.6
Fast, cheap and sufficient serodiagnostic tools needs to be developed for the early detectionof brucellosis. Currently the tools cannot differentiate an active infection from vaccinated, norcan it differentiate other bacterial infections with lipopolysaccharides, especially Yersiniainfections. In this study, we purified recombinant outer membrane protein 10 and 28(rOmp10,rOmp28), and a dipstick assay(indirect or sandwich) was constructed with single(rOmp10 orrOmp28) and combined rOmps(rOmp10 and rOmp28) from Brucella(B.) abortus 544 to evaluatebovine Brucella positive serum collected during the beginning of the Korean outbreak from2006 to 2015. In application with single rOmp, rOmp10(70%; indirect, 92.11%; sandwichdipstick) and rOmp28(72.5%; indirect, 86.84%; sandwich dipstick) had comparable results. Inaddition, results indicated that dipstick with combined rOmps(rOmps10 and rOmp28) weresuperior in detecting positive serum samples, at 85% indirect and 100% sandwich dipstick. Surprisingly, the results were the same in detecting negative results at 97.78% for both singleand combined indirect dipsticks. The dipstick tools with rOmp10 and rOmp28 would be usefulfor a rapid screen method for bovine brucellosis.
Antibacterial effects of Coptis chinensis Franch against Brucella abortus
김대근,Hannah Leah Tadeja Simborio,Alisha Wehdnesday Bernardo Reyes,민원기,이후장,이진주,장홍희,김동혁,김석 경상대학교 농업생명과학연구원 2014 농업생명과학연구 Vol.48 No.1
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.
REYESALISHA WEHDNESDAY,Hannah Leah Tadeja Simborio,Huynh Tan Hop,Lauren Togonon Arayan,민원기,이후장,이만휘,장홍희,김석 대한수의학회 2016 Journal of Veterinary Science Vol.17 No.3
Korean red ginseng (KRG) has long been used in traditional Korean and Oriental medicine. However, the anti-bacterial mechanism and therapeutic efficiency of KGR for intracellular Brucella infection are still unclear. In this study, the bactericidal activity of Korean red ginseng acidic polysaccharide (RGAP) on Brucella (B.) abortus and its cytotoxic effects on RAW 264.7 cells were evaluated. In addition, B. abortus internalization and intracellular replication in macrophages were investigated after RGAP treatment. RGAP-incubated cells displayed a marked reduction in the adherence, internalization and intracellular growth of B. abortus in macrophages. Furthermore, decreased F-actin fluorescence was observed relative to untreated B. abortus-infected cells. Western blot analysis of intracellular signaling proteins revealed reduced ERK, JNK and p38a phosphorylation levels in B. abortus-infected RGAP-treated cells compared to the control. Moreover, elevated co-localization of B. abortus-containing phagosomes with lysosome-associated membrane protein 1 (LAMP-1) were observed in RGAP-treated cells compared with the control. Overall, the results of this study suggest that RGAP can disrupt phagocytic activity of B. abortus via suppression of mitogen-activated protein kinases (MAPKs) signaling proteins ERK, JNK and p38 levels and inhibit intracellular replication of B. abortus by enhancing phagolysosome fusion, which may provide an alternative control of brucellosis.
REYESALISHA WEHDNESDAY,Hannah Leah Tadeja Simborio,Huynh Tan Hop,Lauren Togonon Arayan,김석 대한수의학회 2016 Journal of Veterinary Science Vol.17 No.1
The Brucella mdh gene was successfully cloned and expressed in E. coli. The purified recombinant malate dehydrogenase protein (rMDH) was reactive to Brucella-positive bovine serum in the early stage, but not reactive in the middle or late stage, and was reactive to Brucella-positive mouse serum in the late stage, but not in the early or middle stage of infection. In addition, rMDH did not react with Brucella-negative bovine or mouse sera. These results suggest that rMDH has the potential for use as a specific antigen in serological diagnosis for early detection of bovine brucellosis.
( Alisha Wehdnesday Bernardo Reyes ),( Hannah Leah Tadeja Simborio ),( Huynhtanhop ),( Lauren Togonon Arayan ),( Won Gi Min ),( Hu Jang Lee ),( Hong Hee Chang ),( Suk Kim ) 한국예방수의학회 2014 예방수의학회지 Vol.38 No.4
Brucellosis is an important and re-emerging zoonotic disease worldwide. The prevention of human infection isachieved predominantly through the control of brucellosis in agricultural animals, which in turn depends on accurate diagnosisand vaccination. However, conventional serological diagnosis of brucellosis has several limitations, and currently availablevaccines 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 thepgk 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 superiorto that conferred by commercial strains. In this study, the pgk gene from Brucella abortus 544 was successfully amplified andcloned into a maltose binding protein fusion protein expression vector (pMAL). The recombinant protein was expressed inEscherichia coli DH5α and purified. The immunogenicity of purified recombinant B. abortus 544 Pgk (rPgk) was evaluatedby western blot analysis using Brucella-positive mouse sera. rPgk could be used as an antigenic component for futureserological tests and potential vaccine development.
Review Article : The key roles of toll-like receptor (TLR) for intracellular survival of Brucella
( Dae Geun Kim ),( Hannah Leah Tadeja Simborio ),( Alisha Wehdnesday Bernardo Reyes ),( Won Gi Min ),( Hu Jang Lee ),( Jin Ju Lee ),( Suk Kim ) 한국예방수의학회 2013 예방수의학회지 Vol.37 No.4
Brucella spp. are facultative intracellular pathogens that have the ability to survive and multiply in professional and nonprofessional phagocytes and cause abortion in domestic animals and undulant fever in humans. Brucella species can survive in a variety of cells, including macrophages and their virulence and chronic infections are thought to be due to their ability to avoid the killing mechanisms within macrophages. Inhibition of phagosome-lysosome fusion has been proposed as a mechanism for intracellular survival of Brucella in professional and nonprofessional phagocytes. Toll-like receptors (TLRs) are part of a skillful system for detection of invasion by microbial pathogens. Recognition of microbial components by TLRs triggers signaling pathways that promote expression of genes and regulate innate immune responses. Recent studies for the interaction between TLRs-Brucella have indicated the importance of control of Brucella infection. Here, we review selected aspects of TLRs-Brucella interaction, which may be helpful to understanding the mechanism of Brucella pathogenesis.
( Alisha Wehdnesday Bernardo Reyes ),( Hannah Leah Tadeja Simborio ),( Huynh Tan Hop ),( Lauren Togonon Arayan ),( Tran Xuan Ngoc Huy ),( Wongi Min ),( Suk Kim ) 한국예방수의학회(구 한국수의공중보건학회) 2015 예방수의학회지 Vol.39 No.4
The bacterial lipopolysaccharide (LPS) mainly contributes to the structural integrity, survival and protection barrier against harsh environments. Therefore, the early stages in LPS or lipid A biosynthesis are attractive targets in the identification and development of inhibitors which would be effective against infections caused by Gram-negative bacteria. The bacterial outer membrane proteins (OMPs) meanwhile function as maintenance for structure, adhesion to other cells and substances, as well as development of resistance to antimicrobials. The LPS and LPS-related molecules, and OMPs are important immunogenic components of several important pathogens including Brucella, which have been extensively used in immunological studies and in the diagnosis of diseases. Here we review the importance, structure, functions and immunogenic aspects of LPS and OMPs particularly of Brucella which can be targeted for the prevention and diagnosis of brucellosis.