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Development of a multiplex-PCR for the rapid detection of Escherichia coli O157:H7 from raw beef (쇠고기중 Escherichia coli O157:H7 신속검출을 위한 multiplex - PCR 기법 개발)

  • Jung, Suk-chan;Jung, Byeong-yeal;Yoon, Jang-won;Cho, Yun-sang;Kim, Jong-yeom;Park, Yong-ho
    • Korean Journal of Veterinary Research
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    • v.38 no.1
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    • pp.173-181
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    • 1998
  • Esherichia coli O157 : H7의 slt I, slt II, uid A 및 eaeA 4종 유전자를 동시에 검출하기 위한 multiplex PCR 기법을 확립하고 쇠고기중 직접 E coli O157 : H7 검출시험을 실시하였다. 4 set의 primers를 이용한 multiplex PCR 기법으로 31종의 장내세균에 대한 특이성을 조사한 결과 E coli O157 : H7 에서 1,087bp (eae A), 584bp (slt II), 348bp (slt I) 또는 252bp (uid A)크기의 DNA를 동시에 특이적으로 검출할 수 있었다. E coli O157 : H7 15주는 모두 uid A 및 eae A 유전자가 동시에 검출되었고, 다른 장내세균에서는 검출되지 않았다. slt I 또는 slt II 유전자를 가지고 있는 E coli 표준균주 24종을 이용하여 multiplex PCR 기법과 Vero cell cytotoxicity assay을 비교검사한 결과 베로톡신 산생능과 PCR법의 결과는 일치하였다. mutiplex PCR 기법의 쇠고기중 검출한계는 modified EC(mEC)에서 증균없이는 E coli O157 : H7균 $10^4cells/g$ 이상에서 검출이 가능하였으나 mEC에 1차 증균후 modified TSB 증균하였을 경우에는 10cells/g이하까지도 검출이 가능하였다. 개발된 multiplex PCR 기법을 쇠고기 40종에 직접 적용한 결과 E coli O157 : H7은 검출되지 않았으나 slt I 및 slt II유전자를 가지고 있는 E coli 4종이 검출되었으며, 이들의 혈청형은 O6, O112, O115 및 O139 였다. 이 연구에서 개발된 multiplex PCR은 쇠고기중 E coli O157 : H7을 신속하고 특이적으로 검출하는데 사용할 수 있을 것으로 사료된다.

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Molecular Cloning of a Thermostable $\alpha$-Amylase Gene from Bacillus stearothermophilus and Its Expressions in E. coli (Bacillus stearothermophilus의 열안정성 $\alpha$-amylase 유전자의 E. coli내에서의 cloning과 발현)

  • Huh, Tae-Lin;Koh, Suk-Hoon;Lee, Se-Yong
    • Microbiology and Biotechnology Letters
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    • v.13 no.4
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    • pp.349-354
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    • 1985
  • A 4.7 kb Hind III fragment containing $\alpha$-amylase gene of Bacillus stearothermophilus IAM 11062 was cloned in Escherichia coil HB101, using plasmid pBR322 and runaway plasmid pSY343 as a vector. The cloned gene was stably maintained and expressed In E.coli. The constructed strain of E. coli have at least 3 times higher amylase activity than the donor strain, of B. stearothermophilus. About 75% of the $\alpha$-amylase produced by the constructed strain of E. coli was localized in the periplasm and it was found that the enzymes can be released by an osmotic shock using EDTA. The enzymatic properties of L-amylase produced in E. coli were very similar to those produced by B. stearothermophilus in terms of optimum temperature, heat stability and molecular weight.

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Virulence factors and antimicrobial resistance patterns of pathogenic Escherichia coli isolated from fecal samples of piglets (자돈 분변 유래 병원성 대장균의 병원성 인자 및 항생제 내성 양상)

  • Hyun Sook Shin;Keun-Ho Kim;Jin Sung Seo;Young Wook Kim;Suk-Kyung Lim;Byeong Yeal Jung
    • Korean Journal of Veterinary Service
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    • v.46 no.1
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    • pp.35-45
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    • 2023
  • Pathogenic Escherichia coli is the cause of a wide range of diseases in pigs, including diarrhea, edema disease, and septicemia. Diarrhea caused E. coli may result in significant economic losses, making pathogenic E. coli an important pathogen for the swine industry. This study investigated the prevalence of virulence factor genes, antimicrobial resistance phenotypes, and resistance genes in E. coli isolated from feces of piglets in Korea between 2017 and 2020. As a result, 119 pathogenic E. coli isolates were obtained from 601 fecal samples. The F4 adhesin gene and the STb enterotoxin gene were commonly present in E. coli isolated from diarrhea samples. The dominant virulotypes of isolates from diarrhea samples were STb, Stx2e, and F4:LT:STb. More than 80% of the screened isolates were resistant to ampicillin, sulfisoxazole, chloramphenicol, or tetracycline. To confirm the resistance mechanisms for β-lactam or quinolone, we investigated the genotypic factors of resistance. Each of the ceftiofur-resistant E. coli produced an extended-spectrum β-lactamase encoded by blaCTX-M-14, blaCTX-M-27, and blaCTX-M-55. And all ciprofloxacin-resistant E. coli harbored mutations in quinoloneresistance-determining-regions. In addition, some of the ciprofloxacin-resistant E. coli contained the plasmid-mediated-quinolone-resistance genes such as qepA, qnrB1, or qnrD. This study has confirmed that the F4 fimbria and the STb enterotoxin are the most predominant in pathogenic E. coli isolated from piglets with diarrhea in Korea and there is a great need for responsible and prudent use of antimicrobials to treat colibacillosis.

Survival and Cross-contamination of Escherichia coli O157:H7 on Various Agricultural Product-Contact Surfaces (농산물 접촉 표면 재질에 따른 Escherichia coli O157:H7의 생존 및 상추로의 교차오염도 조사)

  • Kim, Se-Ri;Choi, Song-Yi;Seo, Min-Kyoung;Kim, Won-Il;Chung, Duck-Hwa;Ryu, Kyoung Yul;Yun, Jong-Chul;Kim, Byung-Seok
    • Journal of Food Hygiene and Safety
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    • v.28 no.3
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    • pp.272-278
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    • 2013
  • To evaluate the effect of surface contaminated with Escherichia coli O157:H7 (E. coli O157:H7) on the microbiological safety of lettuce, this study was conducted to investigate the attachment, biofilm producing, survival, and cross-contamination of E. coli O157:H7 on stainless steel and polyvinyl chloride (PVC). The attachment rate of E. coli O157:H7 on PVC was 10 times higher than that on stainless steel after exposure 1 h in cell suspension. However, there was not a difference between two types of surface after exposure for 6 h and 24h. The biofilm producing of E. coli O157:H7 was TSB > 10% lettuce extracts > 1% lettuce extracts > phosphate buffer. When two kinds of materials were stored at various conditions ($20^{\circ}C$ and $30^{\circ}C$, relative humidity (RH) 43%, 69%, and 100%), the numbers of E. coli O157:H7 at $30^{\circ}C$, RH 43% or RH 69% were reduced by 5.0 log CFU/coupon within 12 h regardless of material type. Conversely, the survival of E. coli O157:H7 at RH 100% was lasted more than 5 days. In addition, the reduction rate of E. coli O157:H7 was decreased in the presence of organic matter. The transfer efficiency of E. coli O157:H7 from the contaminated surface to lettuce was dependent upon the water amount of the surface of lettuce. Especially, the transfer rate of E. coli O157:H7 was increased by 10 times in the presence of water on the lettuce surface. From this study, the retention of E. coli O157:H7 on produce contact surfaces increase the risk cross-contamination of this pathogen to produce. Thus, it is important that the surface in post harvest facility is properly washed and sanitized after working for prevention of cross-contamination from surface.

Isolation of Escherichia coli O157 in Children with Diarrhea (소아설사 환아에서의 Escherichia coli O157 분리)

  • Song, Wonkeun;Kim, Hyoun Tae;Lee, Kyu Man;Cha, Jae Kook;Lee, Kon Hee
    • Pediatric Infection and Vaccine
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    • v.4 no.1
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    • pp.73-78
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    • 1997
  • Purpose : Escherichia coli O157 can produce diarrhea as well as hemorrhagic colitis and hemolytic uremic syndrome. In many parts of North America, E. coli O157 often is the second or third most commonly isolated enteric bacterial pathogens. Recently, intakes of fast food, including hamburgers have increased in Korea. Therefore, E. coli O157 infection in Korea are likely to be increased. Methods : Stool samples from 317 pediatric diarrheal patients were analyzed by culture on sorbitol-MacConkey agar. Sorbitol-negative colonies were teated by E. coli O157 latex agglutination test. Results : Of the 317 specimens, one (0.3%) were E. coli O157:NM that not produced Shiga toxin. The 7 year old male patient who had complained of abdominal pain, vomiting and non-bloody diarrhea for 2 days. The patient was improved for 2 days after admission. Conclusions 1 Only one (0.3%) of all fecal samples were isolated E. coli O157 that not produced Shiga toxin. Therefore, routine stool culture for the isolation of E. coli O157 was not likely to be neccessary so far.

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Fluctuation of Escherichia coli on the storage of Kimchi treated with Leuconostoc mesenteroides IFO 12060 and Nisin (저장김치의 Leuconostoc mesenteroides IFO 12060 및 Nisin첨가에 의한 Escherichia coli의 소장)

  • 최신양;이한웅;정건섭
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.21 no.4
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    • pp.414-417
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    • 1992
  • In order to study the addition effect of Leuconostoc mesenteroides and nisin on Escherichia coli and lactic acid bacteria during fermentation and storage of Kimchi, Kimchi was stored at 4$^{\circ}C$ for 7 days and then in-creased the temperature to $25^{\circ}C$. Lactic acid content on Kimchi fermentation at 4$^{\circ}C$ was maintained initial content which was increased upto 0.9% and lactic bacteria was also increased after switching to $25^{\circ}C$. E. coli, on the other hand, was a little decreased from the initial level, but a significant decrease was found for the those Kimchi of Leuconostoc muenteroides added and nisin treated group when the fermentation temperature was switched to $25^{\circ}C$.

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Effects of essential oil (blended and single essential oils) on anti-biofilm formation of Salmonella and Escherichia coli

  • Oh, S.Y.;Yun, W.;Lee, J.H.;Lee, C.H.;Kwak, W.K.;Cho, J.H.
    • Journal of Animal Science and Technology
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    • v.59 no.2
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    • pp.4.1-4.5
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    • 2017
  • Background: Biofilms were the third-dimensional structure in the solid surface of bacteria. Bacterial biofilms were difficult to control by host defenses and antibiotic therapies. Escherichia coli (E. coli) and Salmonella were popular pathogenic bacteria that live in human and animal intestines. Essential oils are aromatic oily liquids from plant materials and well known for their antibacterial activities. Method: This study was conducted to determine effect of essential oil on anti-biological biofilm formation of E. coli and Salmonella strains in in vitro experiment. Two kinds of bacterial strains were separated from 0.2 g pig feces. Bacterial strains were distributed in 24 plates per treatment and each plates as a replication. The sample was coated with a Bacterial biofilm formation was. Result: Photographic result, Escherichia coli (E. coli) and Salmonella bacteria colony surface were thick smooth surface in control. However, colony surface in blended and single essential oil treatment has shown crack surface layer compared with colony surfaces in control. Conclusion: In conclusion, this study could confirm that essential oils have some interesting effect on anti-biofilm formation of E. coli and Salmonella strains from pig feces.

A Study on Bioserotype and Drug Resistance of Salmonella and Escherichia coli Isolated from Feces in Zoological Animals (동물원의 야생동물 분변에서 분리한 살모넬라균과 대장균의 생물형, 혈청형 및 약제내성에 관한 연구)

  • 윤은선;박석기;문현칠;최원식
    • Korean Journal of Veterinary Service
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    • v.16 no.1
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    • pp.41-50
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    • 1993
  • This study was undertaken the bioserotype and drug resistance of Salmonella and Escherichia coli isolated from feces for the prevention and treatment of salmonellosis and colibacillosis in zoological animals. The results obtained from the research were as follows 1. Salmonella were isolated 19, or 4.7% from 408 samples and E. coli were isolated 12, or 40.0% from 30 diarrheal samples. 2. The biotypes in 19 Salmonella were Subspecies 1. 3. The serogroups of Salmonella isolated were 47.4% in B group, 31.6% in C, 5.3% in D and 15.8% in other, and serotype of E. coli was 100% in 0127a. 4. The antibiotic resistance of Salmonella and E. coli isolated were 13, or 68.4% and 7, or 58.3% strains, respectively 5. The multiple resistant patterns of antibiotics in Salmonella were 2drugs- and 3 drugs-resistance 30.8%, respectively, and those in E. coli were mono drug-, 2 drugs- and 7 drugs-resistance 28.6%, respectively. 6. The transferred rate of resistance to recipients (E. coli ML 1410 NA$^{r}$ ) in Salmonella was 38.5%, but that in E. coli was 71.4%.

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Clinical Significance of Extended-Spectrum ${\beta}$-Lactamase Producing $Escherichia$ $coli$ in Pediatric Patients with Febrile Urinary Tract Infection (발열성 소아 요로감염에서 Extended-Spectrum ${\beta}$-Lactamase 생성 $Escherichia$ $coli$의 임상적 의의)

  • Park, Cheol;Kim, Min-Sang;Kim, Mi-Kyung;Yim, Hyung-Eun;Yoo, Kee-Hwan;Hong, Young-Sook;Lee, Joo-Won
    • Childhood Kidney Diseases
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    • v.16 no.1
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    • pp.38-45
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    • 2012
  • Purpose: The incidence of community-acquired urinary tract infection (UTI) due to extended-spectrum ${\beta}$-lactamase producing $Escherichia$ $coli$ (ESBL(+) $E.$ $coli$) has increased worldwide. ESBL causes resistance to various types of the newer ${\beta}$-lactam antibiotics, including the expanded spectrum cephalosporins and monobactams. We aimed to investigate the severity of UTI and associated genitourinary malformations in children with febrile UTI caused by ESBL(+) $E.$ $coli$. Methods: We retrospectively reviewed the medical records of 290 patients diagnosed as febrile UTI caused by $E.$ $coli$ between January 2008 and October 2010 at Korea University Medical center. We classified the patients into two groups with ESBL(+) and ESBL(-) $E.$ $coli$ group according to the sensitivity of urine culture. Fever duration, admission period, white blood cell (WBC) counts and C-reactive protein (CRP) in peripheral blood, the presence of hydronephrosis, cortical defects, vesicoureteral reflux (VUR) and renal scar were compared between the two groups. Results: Patients with ESBL(+) $E.$ $coli$ were 32, and those with ESBL(-) $E.$ $coli$ were 258. If we excluded those tested with a sterile urine bag, patients with ESBL(+) $E.$ $coli$ were 22, and those with ESBL(-) $E.$ $coli$ were 212. Whether the results of sterile urine bag tests were included or not, there was no significant difference in all parameters between the two groups statistically. Conclusion: Our data shows that ESBL(+) $E.$ $coli$ may not be related to the severity of UTI and associated genitourinary malformations.

Comparison of Biochemical Identification to Detect Pathogenic Escherichia coli in Fresh Vegetables (신선편이 엽채류의 병원성 E. coli 검출을 위한 생화학적동정법 비교 분석)

  • Choi, Yukyung;Lee, Heeyoung;Lee, Soomin;Kim, Sejeong;Ha, Jimyeng;Lee, Jeeyeon;Oh, Hyemin;Yoon, Yohan
    • Journal of Food Hygiene and Safety
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    • v.31 no.6
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    • pp.393-398
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    • 2016
  • The objective of this study was to isolate pathogenic Escherichia coli from fresh vegetables with selective media and Petrifilm, and identify a suspicious colony using biochemical identification. Twenty gram of lettuce, twenty gram of cabbage and ten gram of sprout were prepared, and a 5-strain mixture of pathogenic E. coli (Enterohemorrhagic E. coli NCCP11142, Enterotoxigenic E. coli NCCP14037, Enteropathogenic E. coli NCCP14038, Enteroaggregative E. coli NCCP14039, Enteropathogenic E. coli NCCP15661) was inoculated to obtain 1, 2 and 3 log CFU/g. Eighty to ninety milliliter of buffered peptone water (BPW) was placed and pummeled for 60 s. As a results, the Petrifilm method was all positive, but enrichment method of qualitative analysis was negative except for 3-log CFU/g inoculated lettuce. Regarding biochemical identification of pathogenic E. coli, the identification rates were dependent on type of methods and vegetables; lettuce: API 20E 100% (44/44), Microgen GNA 100% (44/44) and Food System 66.7% (10/15), cabbage: API 20E 64.7% (22/34), Microgen GNA 50% (16/32) and Food System 60% (9/15), sprout: API 20E 65.1% (28/43), Microgen GNA 62.3% (27/43) and Food System 53.3% (8/15). These results could be useful in determining an appropriate method to detect pathogenic E. coli in fresh vegetables.