• Journal of Microbiology
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• 제42권3호
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• pp.216-222
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• 2004

In a previous paper, the ogdH gene that encodes 2-oxoglutarat dehydrogenase was isolated from Salmonella typhimurium. The catalytic N-terminal region in the enzyme was found to be very specific for the Salmonella species. Therefore, the aim of the present study was to detect S. typhimurium in food sources using primers designed for OGDH-l and OGDH-2 which were based on the salmonella-specific region of the ogdH gene. A simple polymerase chain reaction (PCR) detection method was developed to detect low numbers of S. typhimurium in a chicken meat microbial consortium. Using the ogdH-specific primers under stringent amplification conditions and for gene probe analysis, fewer than 100 colony-forming units (CFUs) were detectable when pure cultures were employed. When the PCR assay was run on S. typhimurium-contaminated meat contents, only the positive meat samples containing as few as 200 CFUs reacted to the assay. The method employed for sample processing is simple and it was determined to provide a sensitive means of detecting trace amounts of S. typhimurium-specific sequences in the presence of mixed meat microbial populations. When compared with six representative intestinal gram-negative bacterial strains in foods, including Vibrio parahaemolyticus, V. vulnificus, Enterobacter cloacae, E. coli O157:H7, Pseudomonas aeruginosa, and Proteus sp., S. typhimurium had a unique and distinct PCR product (796 bp). In conclusion, the two OGDH primers were found to be rapid and sensitive detectors of Salmonella spp for the PCR method.

• International Journal of Industrial Entomology and Biomaterials
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• 제37권1호
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• pp.21-28
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• 2018

CRISPR/Cas9 gene editing system is an efficient method to mutation in a sequence specific manner. Here we report the direct transfection of the Cas9 nuclease and gene specific guide RNA can be used in BM-N cell line derived from Bombyx mori ovarian tissue to enfeeble function of endogenous gene in vitro. We have used gene editing system to negative regulation components of major signaling cascade, the Toll pathway, which controls B. mori resistance to microbe infections, such as fungi and gram positive bacteria. We demonstrate that the $I{\kappa}B-like$ protein Cactus may controls the activation of transcription factors such as Rel A and Rel B. The direct transfection of Cas9 nuclease and Cactus-specific guide-RNA complex may be used in BM-N cells to disrupt the function of endogenous genes in vitro. A mutation frequency of 30-40% was observed in the transfected cells, and various mutations caused the target region. Moreover, RT-PCR analysis revealed that Cactus gene was down regulated after these mutations. More importantly, mutation of BmCactus stimulated expression of lysozyme, moricin, and lebocin genes. These results suggest that the CRISPR/Cas9 systems are expected to efficiently induce site-specific mutations and it was possible to produce antimicrobial peptide through the gene editing.

• Journal of Microbiology and Biotechnology
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• 제26권3호
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• pp.461-468
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• 2016

In recent years, various naturally occurring defence peptides such as plectasin have attracted considerable research interest because they could serve as alternatives to antibiotics. However, the production of plectasin from natural microorganisms is still not commercially feasible because of its low expression levels and weak stability. A tandemly arrayed plectasin gene (1,002 bp) from Pseudoplectania nigrella was generated using the isoschizomer construction method, and was inserted into the pPICZαA vector and expressed in Pichia pastoris. The selected P. pastoris strain yielded 143 μg/ml recombinant plectasin (Ple) under the control of the methanol-inducible alcohol oxidase 1 (AOX1) promoter. Ple was estimated by SDS-PAGE to be 41 kDa. In vitro studies have shown that Ple efficiently inhibited the growth of several gram-positive bacteria such as Streptococcus suis and Staphylococcus aureus. S. suis is the most sensitive bacterial species to Ple, with a minimum inhibitory concentration (MIC) of 4 μg/ml. Importantly, Ple exhibited resistance to pepsin but it was quite sensitive to trypsin and maintained antimicrobial activity over a wide pH range (pH 2.0 to 10.0). P. pastoris offers an attractive system for the cost-effective production of Ple. The antimicrobial activity of Ple suggested that it could be a potential alternative to antibiotics against S. suis and S. aureus infections.

• Journal of Microbiology and Biotechnology
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• 제26권3호
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• pp.596-602
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• 2016

Staphylococcus aureus, like other gram-positive pathogens, has evolved a large repertoire of virulence factors as a powerful weapon to subvert the host immune system, among which alpha-hemolysin (Hla), a secreted pore-forming cytotoxin, plays a preeminent role. We observed a concentration-dependent reduction in Hla production by S. aureus in the presence of sub-inhibitory concentrations of isorhamnetin, a flavonoid from the fruits of Hippophae rhamnoides L., which has little antibacterial activity. We further evaluate the effect of isorhamnetin on the transcription of the Hla-encoding gene hla and RNAIII, an effector molecule in the agr system. Isorhamnetin significantly down-regulated RNAIII expression and subsequently inhibited hla transcription. In a co-culture of S. aureus and lung cells, topical isorhamnetin treatment protected against S. aureus-induced cell injury. Isorhamnetin may represent a leading compound for the development of anti-virulence drugs against S. aureus infections.

• International Journal of Oral Biology
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• 제35권2호
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• pp.43-49
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• 2010

Enterococcus faecalis, a gram-positive bacterium, has been implicated in endodontic infections, particularly in chronic apical periodontitis. Proinflammatory cytokines, including tumor necrosis factor-$\alpha$ (TNF-$\alpha$), are involved in the pathogenesis of these apical lesions. E. faecalis has been reported to stimulate macrophages to produce TNF-$\alpha$. The present study investigated the mechanisms involved in TNF-$\alpha$ production by a murine macrophage cell line, RAW 264.7 in response to exposure to E. faecalis. Both live and heat-killed E. faecalis induced high levels of gene expression and protein release of TNF-$\alpha$. Treatment of RAW 264.7 cells with cytochalasin D, an inhibitor of endocytosis, prevented the mRNA up-regulation of TNF-$\alpha$ by E. faecalis. In addition, antioxidant treatment reduced TNF-$\alpha$ production to baseline levels. Inhibition of extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein (MAP) kinase also significantly attenuated E. faecalis-induced TNF-$\alpha$ expression by RAW 264.7 cells. Furthermore, activation of NF-${\kappa}B$ and AP-1 in RAW 264.7 cells was also stimulated by E. faecalis. These results suggest that the phagocytic uptake of bacteria is necessary for the induction of TNF-$\alpha$ in E. faecalis-stimulated macrophages, and that the underlying intracellular signaling pathways involve reactive oxygen species, ERK, p38 MAP kinase, NF-${\kappa}B$, and AP-1.

• 대한의생명과학회지
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• 제26권3호
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• pp.244-248
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• 2020

Skin and soft tissue infections caused by methicillin-resistant Staphylococcus aureus (MRSA) can occur especially in community populations such as military training camps. We investigated antimicrobial resistance patterns and molecular epidemiological characteristics of MRSA isolated from nasal swabs in healthy army trainees. From January 2018 to March 2018, one MRSA strain was isolated from nasal swab and hand of healthy army trainees. mecA gene detection, SCCmec and mec complex typing were performed to analyze the antimicrobial resistance patterns and molecular epidemiological characteristics of MRSA isolates. As a result, SCCmec and mec complex type of MRSA isolate from military trainees was not-typeable (n=1). In conclusion, not-typeable subtype of MRSA isolate from military trainees need to be confirmed by continuous follow-up study to determine whether there is a different genotype or a new subtype of genotype present in the Republic of Korea.

• Pediatric Infection and Vaccine
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• 제27권2호
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• pp.127-133
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• 2020

Parvimonas micra는 아포 비형성 혐기성 그람 양성 알균으로, 피부, 잇몸, 질, 위장관 등에 정상 상재균으로 존재하며 주로 치과 치료 등의 침습적 치료 후 기회 감염을 일으킬 수 있다. 저자들은 최근 1년 이내 치과 질환을 포함한 특이 과거력이 없는 11세 소아에서 P. micra에 의해 발생한 뇌농양을 경험하여 보고하고자 한다. 환자는 내원 7일 전부터 두통, 구토가 발생하였으며, 내원일 복시가 발생하였다. 뇌 자기공명영상에서 좌측 전두정엽 뇌농양으로 진단한 후 수술적 배농을 시행하였으며, 수술 시 흡인한 검체의 혐기성 세균 배양 검사에서 P. micra가 동정되었다. 수술 후 6주간 ceftriaxone과 metronidazole 항생제 병합요법으로 치료한 후 재발 없이 안정된 상태를 보이고 있다.

• 생명과학회지
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• 제17권4호
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• pp.581-586
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• 2007

본 연구에서는 피부상재균이며 기회병원균이기도 한 Staphylococcus epidermidis ATCC12228로부터 urease효소를 4단계 크로마토그라피 방법을 사용하여 1,127배 정제하고 그 생화학적인 특성을 규명하였다. 정제된 urease 효소는 SDS-PACE 전기영동분석 및 gel-filtration 크로마토그라피를 이용한 천연분자량 분석결과, 67, 16.1 및 12.7 kDa의 3개 subunit가 3량체로 회합되어 존재하는 것으로 나타났으며 catalytic unit 당 2.2개의 니켈 원소를 함유하는 것으로 측정되었다. 정제된 효소의 비활성은 993.8 U/mg, $K_m$값은 8.5mM로 각각 산출되었다.

• 대한방사선기술학회지:방사선기술과학
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• 제38권4호
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• pp.395-401
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• 2015

영상의학과의 흉부엑스선 검사에서 감염관리에 대한 기기의 오염 정도를 세균학적으로 확인하고 균을 동정하여 병원감염의 예방과 감염관리방안을 마련하는데 목적이 있다. 영상의학과 흉부 엑스선 검사에서 환자의 기기 접촉부위인 어깨, 손, 턱, 흉부 측면촬영의 손잡이 부위를 70% 이소프로필 알코올로 소독하기 전과 후를 면봉으로 검체를 채취하여 세균을 동정하여 비교하여 분석하였다. 흉부 엑스선 기기에서 소독전의 측면촬영의 손잡이에서 그람 양성의 Staphylococcus가 분리되었다. 최종동정을 위해 노보바이오신 (nobobiocin) 항생제 시험을 실시하여 민감성으로 Staphylococcus epidermidis로 확정하였다. 흉부촬영 기기에서 측면촬영의 손잡이 부위를 소독 전에 Staphylococcus epidermidis 균이 검출되었다. 흉부엑스선 검사에서 기기를 주기적인 소독 및 멸균을 실시하여 병원감염을 예방하는 관리방안을 마련하여야 한다.

• Bulletin of the Korean Chemical Society
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• 제28권8호
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• pp.1335-1340
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• 2007

β-Ketoacyl acyl carrier protein synthase (KAS) III is a particularly attractive target in the type II fatty acid synthetic pathway, since it is central to the initiation of fatty acid synthesis. Enterococcus faecalis, a Grampositive bacterium, is one of the major causes of hospital acquired infections. The rise of multidrug-resistant of most bacteria requires the development of new antibiotics, such as inhibition of the KAS III. In order to block the fatty acid synthesis by inhibition of KAS III, at first, three dimensional structure of Enterococcus faecalis KAS III (efKAS III) was determined by comparative homology modeling using MODELLER based on x-ray structure of Staphylococcus aureus KAS III (saKAS III) which is a gram-positive bacteria and is 36.1% identical in amino acid sequences with efKAS III. Since His-Asn-Cys catalytic triad is conserved in efKAS III and saKAS III, substrate specificity of efKAS III and saKAS III and the size of primer binding pocket of these two proteins are expected to be similar. Ligand docking study of efKAS III with naringenin and apigenin showed that naringenin docked more strongly with efKAS III than apigenin, resulting in the intensive hydrogen bond network between naringenin and efKAS III. Also, only naringenin showed antibacterial activity against E. faecalis at 256 μg/mL. This study may give practical implications of flavonoids for antimicrobial effects against E. faecalis.