• Journal of Microbiology and Biotechnology
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• v.27 no.6
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• pp.1163-1170
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• 2017

Clostridium difficile releases two exotoxins, toxin A and toxin B, which disrupt the epithelial cell barrier in the gut to increase mucosal permeability and trigger inflammation with severe diarrhea. Many studies have suggested that enteric nerves are also directly involved in the progression of this toxin-mediated inflammation and diarrhea. C. difficile toxin A is known to enhance neurotransmitter secretion, increase gut motility, and suppress sympathetic neurotransmission in the guinea pig colitis model. Although previous studies have examined the pathophysiological role of enteric nerves in gut inflammation, the direct effect of toxins on neuronal cells and the molecular mechanisms underlying toxin-induced neuronal stress remained to be unveiled. Here, we examined the toxicity of C. difficile toxin A against neuronal cells (SH-SY5Y). We found that toxin A treatment time- and dose-dependently decreased cell viability and triggered apoptosis accompanied by caspase-3 activation in this cell line. These effects were found to depend on the up-regulation of reactive oxygen species (ROS) and the subsequent activation of p38 MAPK and induction of $p21^{Cip1/Waf1}$. Moreover, the N-acetyl-$\text\tiny L$-cysteine (NAC)-induced down-regulation of ROS could recover the viability loss and apoptosis of toxin A-treated neuronal cells. These results collectively suggest that C. difficile toxin A is toxic for neuronal cells, and that this is associated with rapid ROS generation and subsequent p38 MAPK activation and $p21^{Cip1/Waf1}$ up-regulation. Moreover, our data suggest that NAC could inhibit the toxicity of C. difficile toxin A toward enteric neurons.

• Journal of Food Hygiene and Safety
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• v.32 no.6
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• pp.531-535
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• 2017

In this study, Leuconostoc mesenteroides CJNU0041 was isolated from Korean traditional food kimchi and antimicrobial activity of fermented broccoli with the isolate was tested against pathogenic Clostridium difficile. L. mesenteroides CJNU0041 showed higher glucosidase activity than other isolates. As the results of physiological properties such as pH and viable cell count during broccoli fermentation with L. mesenteroides CJNU0041, we confirmed that 48 hours was optimal fermentation time. As the results of metabolite analysis by HPLC, metabolites were changed during the fermentation. Especially, the growth of C. difficile was inhibited by the fermented broccoli. Therefore, L. mesenteroides CJNU0041 might be a candidate for improving the functionality of natural materials by lactic acid fermentation.

• Journal of Microbiology and Biotechnology
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• v.26 no.4
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• pp.693-699
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• 2016

Clostridium difficile toxin A is known to cause deacetylation of tubulin proteins, which blocks microtubule formation and triggers barrier dysfunction in the gut. Based on our previous finding that the Clostridium difficile toxin A-dependent activation of histone deacetylase 6 (HDAC-6) is responsible for tubulin deacetylation and subsequent microtubule disassembly, we herein examined the possible effect of potassium acetate (PA; whose acetyl group prevents the binding of tubulin to HDAC-6) as a competitive/false substrate. Our results revealed that PA inhibited toxin A-induced deacetylation of tubulin and recovered toxin A-induced microtubule disassembly. In addition, PA treatment significantly decreased the production of IL-6 (a marker of inflamed tissue) in the toxin A-induced mouse enteritis model. An in vitro HDAC assay revealed that PA directly inhibited HDAC-6-mediated tubulin deacetylation, indicating that PA acted as a false substrate for HDAC-6. These results collectively indicate that PA treatment inhibits HDAC-6, thereby reducing the cytotoxicity and inflammatory responses caused by C. difficile toxin A.

• Quality Improvement in Health Care
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• v.8 no.1
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• pp.10-21
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• 2001

Background : The Clostridium difficile is the most important identifiable cause of nosocomial infectious diarrhea and colitis, which lengthens hospital stay. Recently incidence of C. difficile has been increasing in an university hospital, and an intervention for prevention and control of C. difficile associated diarrhea (CDAD) was in prompt need. Methods : Subjects were the patients in the neurosurgical intensive care unit(NCU) where C. difficile was most frequently isolated. To increase participation of various departments, we used the CQI method, because management of CDAD requires a wholistic approach including control of antibiotics, barrier precaution and environmental cleaning and disinfection. Duration of the CQI activities was 9 months from April to December 1999. Results : The identified problems were misuse and overuse of antibiotics, lack of consciousness of medical personnels and the possibility of transmission from the contaminated environment and tube feeding. Education for proper use of antibiotics and management of C. difficile infection, use of precaution stickers, supplement of handwashing equipments, emphasis on environmental disinfection, and the change of the process of tube feeding were done. The CDAD rate in NCU was significantly decreased after the CQI program (8.6 case per 1,000 patient days from January to April 1999 vs 4.8 from May to December 1999). The distribution of neurosurgical wards including NCU among the total number of isolated C. difficile from the clinical specimens dropped from 49.4% in January to April to 33,7% in May to December. The average hospital stay of the neurosurgical department changed from 19.6 days to 15.2 days. Also, the effect of the CQI activities for C. difficile may have affected the incidence of vancomycin resistant enterococci (VRE). Duration and dosage of certain antibiotics used in the NS department were decreased. The distribution of neurosurgical department in the number of VRE isolated patients declined from 18.4% to 11.1%. Conclusion : Infection control of resistant organisms such as C. difficile is likely to be successful when management of environmental contamination an collaborative efforts of decreasing the patients' risk factors such as antibiotics management and decreasing the length of hospital stay come simultaneously. For this work, related departments need to actively participate in the entire process under a common target through discussions for identifying problems and bringing up solutions. In this respect, making use of a CQI team is an efficient method of infection control for gathering participation and cooperation of related departments.

• Korean Journal of Clinical Pharmacy
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• v.21 no.3
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• pp.228-236
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• 2011

배경: 약물로 인한 Clostridium difficile-associated diarrhea (CDAD)는 널리 알려져 있으며 우리나라에서 항생제와 프로톤 펌프 억제제 소모량을 고려할 때 질환 치료과정에서의 CDAD 발생빈도 및 CDAD 유발 이전에 투여한 약물의 사용빈도와 CDAD의 치료방법을 조사할 필요성이 있다. 방법: 경상대학교 병원에서 2011년 1월부터 6월까지의 입원환자를 대상으로 대변 독소 검사에 의해 CDAD로 판명된 환자의 성별, 연령분포, 질환명, 입원병동, 재발률을 조사하였으며 CDAD 판명이전에 투여한 약제 및 CDAD 판명후 치료약제를 조사하였다. 결과: 연구기간 동안 CDAD 대변 독소 검사 의뢰된 환자수는 1,500명이었으며 CDAD 양성은 111명(9.3%)이었고, 재발은 29명(26.1%)이었다. CDAD를 주소로 입원한 환자는 17명 (15.3%)이었고, 나머지는 입원기간 중에 발생하였다. CDAD 양성인 환자의 연령대는 60대에서 32.4% (36/111명) 이었고, 내과병동에서 34.2%를 나타내었고, 재발률은 외과계 병동에서 41.4%로 가장 높게 나타났다. CDAD 환자의 17% (19/111명)은 항암제 투여 동안 발생하였으며 CDAD 발생 전 사용약물로는 세팔로스포린계 항생제가 162회로 가장 빈번하게 사용 되었으며, 히스타민2 수용체길항제 107회, 스테로이드 82회, 비 스테로이드 항염제 79회, 프로톤 펌프 억제제 77회, 하제 59회, 항암제가 33회 처방되었다. CDAD 치료약제로는 8종의 약제가 241회 처방 되었으며 metronidazole이 99회로 가장 빈번하게 사용되었고, vancomycin이 37회로 나타났다. 결론: 입원환자에 있어서 CDAD양성은 특히 고령의 암환자가 많아 항암제 투여 시에는 CDAD 발생에 주의해야 할것으로 보인다. CDAD의 치료약제로는 metronidazole이 vancomycin 보다 많이 사용되는 것으로 나타났다.

• Korean Journal of Clinical Pharmacy
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• v.27 no.2
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• pp.77-82
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• 2017

Background: Clostridium difficile associated diarrhea (CDAD) is a leading cause of hospital-associated gastrointestinal illness. Risk factors for CDAD include advanced age, long-term admission, antibiotics, proton-pump inhibitor or $H_2$ blocker use and immunosuppression. The practice guideline of American Journal of Gastroenterology (2013) suggests metronidazole for the first-line therapy of mild-moderate CDAD as well as vancomycin for severe CDAD. MICU inpatients receiving stress ulcer prophylaxis and antibiotics are susceptible to nosocomial CDAD. Therefore, this study aimed to evaluate occurrence and treatment of CDAD in MICU. Methods: Patients who were admitted to the MICU and had CDAD from August 2012 to August 2015 were analyzed retrospectively. Results: Of the 90 patients with CDAD, 20 patients (2.22%) had mild-moderate CDAD (16 received metronidazole and 4 received vancomycin therapy) and 70 patients (77.8%) had severe CDAD(54 received metronidazole and 16 received vancomycin therapy). Among the patients with mild- moderate CDAD, treatment with metronidazole or vancomycin resulted in same clinical cure in 50% of the patients (p=1.00). Among the patients with severe CDAD, treatment with metronidazole or vancomycin resulted in clinical cure in 40.7% and 50.0% of the patients, respectively (p=0.511). Clinical symptoms recurred in 7.4% of the severe CDAD patients treated with metronidazole and 6.3% of those treated with vancomycin(p=0.875). Conclusion: Our findings suggest that metronidazole and vancomycin are equally effective for the treatment of mild-moderate CDAD; however, vancomycin demonstrated higher clinical cure rate and lower recurrence rate for severe CDAD, although the difference was not statistically significant. For better clinical outcomes, appropriate medication use by disease severity is needed.

• Journal of Microbiology and Biotechnology
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• v.27 no.4
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• pp.694-700
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• 2017

Clostridium difficile, which causes pseudomembranous colitis, releases toxin A and toxin B. These toxins are considered to be the main causative agents for the disease pathogenesis, and their expression is associated with a marked increase of apoptosis in mucosal epithelial cells. Colonic epithelial cells are believed to form a physical barrier between the lumen and the submucosa, and abnormally increased mucosal epithelial cell apoptosis is considered to be an initial step in gut inflammation responses. Therefore, one approach to treating pseudomembranous colitis would be to develop agents that block the mucosal epithelial cell apoptosis caused by toxin A, thus restoring barrier function and curing inflammatory responses in the gut. We recently isolated an antimicrobial peptide, Periplanetasin-2 (Peri-2, YPCKLNLKLGKVPFH) from the American cockroach, whose extracts have shown great potential for clinical use. Here, we assessed whether Peri-2 could inhibit the cell toxicity and inflammation caused by C. difficile toxin A. Indeed, in human colonocyte HT29 cells, Peri-2 inhibited the toxin A-induced decrease in cell proliferation and ameliorated the cell apoptosis induced by this toxin. Moreover, in the toxin A-induced mouse enteritis model, Peri-2 blocked the mucosal disruption and inflammatory response caused by toxin A. These results suggest that the American cockroach peptide Peri-2 could be a possible drug candidate for addressing the pseudomembranous colitis caused by C. difficile toxin A.

• YAKHAK HOEJI
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• v.40 no.3
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• pp.343-346
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• 1996

The influence of LB20304a, a new fluoroquinolone antibiotic agent, on microflora of caecum in mice was compared with those of ciprofloxacin and piperacillin after administration of drugs for 5 days. Selective medium (CCFMA) was used for the isolation of Clostridium difficile from the specimens of mouse caecum. The emergence of C. difficile in mouse caecum induced by LB20304a was lower than that by ciprofloxacin or piperacillin at day 1 and day 7 after completing administration of drugs.

• Journal of Microbiology and Biotechnology
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• v.29 no.10
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• pp.1675-1681
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• 2019

Clostridium difficile toxin A is known to cause colonic epithelial cell apoptosis, which is considered the main causative event that triggers inflammatory responses in the colon, reflecting the concept that the essential role of epithelial cells in the colon is to form a physical barrier in the gut. We previously showed that toxin A-induced colonocyte apoptosis and subsequent inflammation were dependent on prostaglandin E2 ($PGE_2$) produced in response to toxin A stimulation. However, the molecular mechanism by which $PGE_2$ mediates cell apoptosis in toxin A-exposed colonocytes has remained unclear. Here, we sought to identify the signaling pathway involved in toxin A-induced, $PGE_2$-mediated colonocyte apoptosis. In non-transformed NCM460 human colonocytes, toxin A exposure strongly upregulated expression of Bak, which is known to form mitochondrial outer membrane pores, resulting in apoptosis. RT-PCR analyses revealed that this increase in Bak expression was attributable to toxin A-induced transcriptional upregulation. We also found that toxin A upregulation of Bak expression was dependent on $PGE_2$ production, and further showed that this effect was recapitulated by an Prostaglandin E2(PGE2) receptor-1 receptor agonist, but not by agonists of other EP receptors. Collectively, these results suggest that toxin A-induced cell apoptosis involves $PGE_2$-upregulation of Bak through the EP1 receptor.

• Korean Journal of Clinical Laboratory Science
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• v.46 no.4
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• pp.140-142
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• 2014

To evaluate the recovery rates to increase toxigenic C. difficile, the selective enrichment broth culture methods were compared with commonly used cytotoxin assays and toxigenic culture. First, the enrichment culture, using the selective medium broth for 2 to 5 days, was performed and then, toxigenic C. difficile was confirmed by C. difficile toxin gene-specific PCR after being cultured on C. difficile selective agar. The sensitivity of C. difficile from the enrichment culture (100%) was higher than that of C. difficile selective agar culture (93.8%), while positive predictive values (PPV) were low; 72.7% (16/22) and 88.2% (15/17). PPV of the enrichment culture are not high. Recently, combinations of C. difficile selective agar culture, C. difficile A & B assays, glutamate dehydrogenase, and nucleic acid amplification method are widely used. The enrichment culture was disadvantageous in PPV, turn-around time, and cost. So, what we performed is not considered as a common method of diagnosis of C. difficile-associated diarrhea.