• 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.

• Food Science of Animal Resources
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• v.32 no.1
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• pp.62-67
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• 2012

This study calculated kinetic parameters of Escherichia coli O157:H7 and developed a probabilistic model to estimate growth probabilities of E. coli O157:H7 on polyethylene cutting boards as a function of temperature and time. The surfaces of polyethylene coupons ($3{\times}5$ cm) were inoculated with E. coli O157:H7 NCCP11142 at 4 Log $CFU/cm^2$. The coupons were stored at 13 to $35^{\circ}C$ for 12 h, and cell counts of E. coli O157:H7 were enumerated on McConkey II with sorbitol agar every 2 h. Kinetic parameters (maximum specific growth rate, Log $CFU/cm^2/h$; lag phase duration, h; lower asymptote, Log $CFU/cm^2$; upper asymptote, Log $CFU/cm^2$) were calculated with the modified Gompertz model. Of 56 combinations (temperature${\times}$time), the combinations that had ${\geq}$0.5 Log $CFU/cm^2$ of bacterial growth were designated with the value of 1, and the combinations that had increases of <0.5 Log $CFU/cm^2$ were given the value 0. These growth response data were fitted to the logistic regression to develop the model predicting probabilities of E. coli O157:H7 growth. Specific growth rate and growth data showed that E. coli O157:H7 cells were grown at $28-35^{\circ}C$, but there were no obvious growth of the pathogen below $25^{\circ}C$. Moreover, the developed probabilistic model showed acceptable performance to calculate growth probability of E. coli O157:H7. Therefore, the results should be useful in determining upper limits of working temperature and time, inhibiting E. coli O157:H7 growth on polyethylene cutting board.

• Journal of Food Hygiene and Safety
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• v.12 no.3
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• pp.175-180
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• 1997

Escherichia coli O157:H7 and Salmonella ser. typhimurium are pathogens involved in food poisoning in numerous countries. This study aimed to obtain knowleges on the survival of E. coli O157:H7 KSC 109 and S. ser. typhimurium ATCC 14028 in fermentedmilk products which were on sale in Suwon Yakult supplier. To the final concentration of 103~104 cfu/$m\ell$ of E. coli O157:H7 KSC 109 or S. wer. typhimurium ATCC 14028 in the fermented milks, Metchnikoff, Ace, Yakult, Mastoni and Super 100 were inoculated with these pathogens and then were stored at 4$^{\circ}C$ and viable cells of these pathogens were periodically counted. The results showed that the survival of two pathogens differed in the different types of fermented milks tested. Number of suriviving E. coli O157:H7 KSC 109 and S. ser. typimurium ATCC 14028 cells (initial inoculum, 103~104 cfu/$m\ell$) were decreased to 101, 102 cfu/$m\ell$ in Ace after 100 hours, and were decreased gradually to 101 cfu/$m\ell$ in Yakult after 250 hours. In the other fermented milks, viable cells of E. coli O157:H7 KSC 109 was not drastically decreased but those of S. ser. typhimurium ATCC 14028 was decreased gradually to 102 (Mastoni), and to 101 cfu/$m\ell$ (Super 100) after 250 hours. It appeared that S. ser. typhimurium ATCC 14028 was more susceptible than E. coli O157:H7 KSC 109 at low pH. Vibale cells of E. coli O157:H7 KSC 109 was not drastically decreased in most of fermented milks tested except Ace and Yakult, but in general, S. ser. typhimurium ATCC 14028 was drastically decreased in most of the fermented milks. The major ingibition factor against these pathogens in the fermented milks during storage at 4$^{\circ}C$ appeared to be the acidity and the metabolites produced by the starters bacteria used in fermented milk products.

• Microbiology and Biotechnology Letters
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• v.27 no.5
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• pp.419-425
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• 1999

Prophylactic effects of Bifidobacterium longum HY8001, Korean isolate, against Escherichia coli O157:H7 and Salmonella typhimurium DT104 enteric infection were examined at four groups of specific pathogen free(SPF)-ICR mouse for each pathogen. B. longum HY8001+B. typhimurium DT104+B. longum HY8001(BL+ST+BL) group and B. longum HY8001+E. coli O157:H7+B. longum HY8001(BL+E+BL) group were fed with B. longum HY8001 before and after E. coli O157:H7 or s. typhimurium DT104 challenge, while B. longum HY8001+S. typhimurium DT104(BL+ST) and B. longum HY8001+e. coli O157:H7(BL+E) groups were fed with B. longum HY8001 only before E. coli O157:H7 or S. typhimurium DT104 challenge. E. coli O157:H7(E) and S. typhimurium DT104(ST) groups were challenged with each pathogen without B. longum HY8001 administration and control groups were administered with phosphate buffered solution(PBS). After the oral administration with B. longum HY8001(109cfu), th emice were challenged with E. coli O157:H7(2$\times$1010cfu) or S. typhimurium DT104(108cfu) and the mortality rate and the fecal shedding of challenged pathogen were also examined define the reactivity of the B. longum HY8001. Production of toxin neutralizing substance(s) of B. longum HY8001 was determined by cell cytotoxicity assay using Vero cells. Fecal shedding of th eS. typhimurium DT104 was significantly decreased in BL+ST+BL group fed with B. longum HY8--1 before and after challenge(p<0.05), while the fecal shedding s of S. typhimurium DT104 in BL+ST and St groups remained more than 106cfu. the protective effect of the B. longum HY8001 against E. coli O157:H7 was significantly high only in BL+E+BL group fed with b. longum Hy8001 before and after E. coli O157:H7 challenge from the result of fecal E. coli O157:H7 isolation rate, mortality rate, and intestinal contents culture to detect E. coli O157:H7. the mortality rate of the BL+e and E groups. The cytopathic effect (CPE) of the Vero cytotoxin (Shiga like toxin I & II) in Vero cell was neutralized in B. longum HY8001 culture supernatant added wells which indicate the presence of soluble Vero cytotxin neutralizing substance(s) in B. longum HY8001 culture suprnatant.

• Journal of Microbiology and Biotechnology
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• v.16 no.8
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• pp.1169-1173
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• 2006

Imaging ellipsometry (IE) for detection of binding of Escherichia coli O157:H7 (E. coli O157:H7) to an immunosensor is reported. A protein G layer, chemically bound to a self-assembled layer of 11-mercaptoundecanoic acid (11-MUA), was adopted for immobilization of monoclonal antibody against E. coli O157:H7 (Mab). The immobilization of antibody was investigated using surface plasmon resonance. To fabricate antibody spots on a gold surface, protein G solution was spotted onto the gold surface modified with an 11-MUA layer, followed by immobilizing Mab on the protein G spot. Ellipsometric images of the protein G spot, the Mab spot, and Mab spots with binding of E. coli O157:H7 in various concentrations were acquired using the IE system. The change of mean optical intensity of the Mab spots in the ellipsometric images indicated that the lowest detection limit was $10^3$CFU/ml for E. coli O157:H7. Thus, IE can be applied to an immunosensor for detection of E. coli O157:H7 as a detection method with the advantages of allowing label-free detection, high sensitivity, and operational simplicity.

• Journal of Environmental Health Sciences
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• v.33 no.5
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• pp.403-407
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• 2007

The inhibitory effect of the food processing agent on growth of Escherichia coli O157:H7, Salmonella Enteritidis, and Listeria monocytogenes was performed with organic acid, and combination of citric acid, acetic acid, propionic acid and vanillic acid. The minimun inhibitory concentration(MIC) of propionic acid was 5,000 ppm in E. coli O157:H7, 2,500 ppm in Salmonella Enteritidis and Listeria monocytogenes. MIC of citric acid was 10,000 ppm in E. coli O157:H7 and Salmonella Enteritidis, 2,500 ppm in Listeria monocytogenes. MIC of acetic acid was 2,500ppm, while in vanillic acid was 5,000 ppm in Escherichia coli O157:H7, Salmonella Enteritidis, and Listeria monocytogenes. MIC of combined organc acid in E. coli O157:H7 were 2,500ppm in PC, 1,250 ppm in PA, PV, CA, CV and AV. MIC of combined organc acid in Salmonella Enteritidis were 2,500 ppm in PC, PA, PV, CA, and CV, 1,250 ppm in AV. MIC of combined organc acid in Listeria monocytogenes were 1,250 ppm in all treatment group. MIC of combined treatment of three organc acid in E. coli O157:H7, S. Enteritidis and L. monocytogenes were 1,250 ppm in PCA, PCV, PAV and CAV. The inhibitory effect of organc acid in E. coli O157:H7, S. Enteritidis and L. monocytogenes could be confirmed from the result of this experiment. Therefore, it was expected that the food process would increase or maintain by using organic acid.

• Korean Journal of Veterinary Research
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• v.39 no.4
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• pp.765-771
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• 1999

Unlike most Escherichia coli strains, E coli O157 : H7 didn't ferment sorbitol within 24h of incubation and showed a negative reaction for $\beta$-glucuronidase. We developed a new medium for the rapid isolation of E coli O157 : H7 using sorbitol MacConkey agar with cefixime, potassium tellurite and 4-methylumbelliferyl-${\beta}$-D-glucuronide (MUG) as a primary plating medium. The addition of $20{\mu}g/ml$ of vancomycin in enrichment broth for E coli O157 : H7 inhibited lots of Gram positive bacteria. Three strains (10.3%) of 29 non-O157 E coli strains and 3 strains (8.3%) of 36 Salmonella spp were inhibited at the $0.05{\mu}g/ml$ of cefixime and 23 strains (79.3%) of 29 non-O157 E coli strains and 12 strains (33.3%) of 36 Salmonella spp were inhibited at the $2.0{\mu}g/ml$ of potassium tellurite. But none of the E coli O157 : H7 was affected at these concentration. The addition of MUG at $100{\mu}g/ml$ level to sorbitol MacConkey agar with cefixime and potassium tellurite (CTM-SMAC) aided in the rapid isolation of E coli O157 : H7 from samples by checking sorbitol-negative and $\beta$-glucuronidase negative phenotypes simultaneously. In conclusion, inoculation of a positive in the O157 screening test from enrichment broth on CTM-SMAC appeared to be a rapid, cost-effective and sensitive method for the isolation of E coli O157 : H7.

• Food Science of Animal Resources
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• v.31 no.2
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• pp.158-165
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• 2011

A sensitive reverse transcriptase-PCR (RT-PCR) method to detect viable Escherichia coli O157:H7 in milk was established. The primer sets were designed based on the nucleotide sequences of the rfbE (per) and wbdN genes in the O157 antigen gene cluster of E. coli O157:H7. RT-PCR using five different primer sets yielded DNA with sizes of 655, 518, 450, and 149-bp, respectively. All five of the E. coli O157:H7 strains were detected by RT-PCR, but 11 other bacterial species were not. The sensitivity of RT-PCR was improved by adding yeast tRNA as a carrier to the crude RNA extract. The RT-PCR amplifying the 149-bp DNA fragment was the most sensitive for detecting E. coli O157:H7 and the most refractory to the bactericidal treatments. Heat treatment at $65^{\circ}C$ for 30 min was the least inhibitory of all bactericidal treatments. Treatment with RNase A strongly inhibited the RT-PCR of heated milk but not unheated milk. This study described RT-PCR methods that are specific and sensitive with a detection limit of 10 E. coli O157:H7 cells, and showed that pre-treating milk samples with RNase A improved the specificity to detect viable bacteria by RT-PCR.

• Journal of environmental and Sanitary engineering
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• v.19 no.4 s.54
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• pp.69-75
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• 2004

The inhibitory effect of the food processing agent on growth of Escherichia coli O157:H7, Salmonella Enteritidis, and Listeria monocytogenes was performed with hydrogen peroxide and lactic acid, and combination of hydrogen peroxide and lactic acid. The minimun inhibitory concentration (MIC) of hydrogen peroxide in E coli O157:H7 was 100 ppm at pH 5.0, 6.0, 6.5 and 7.0, while in Listeria monocytogenes 25 ppm at PH 5.5, 6.0 and 50 ppm at PH 6.5, 75ppm at pH 7.0. MIC of lactic acid in E coli O157:H7 was 2500 ppm at pH 5.0, 6.0, 6.5 and 7.0. MIC of lactic acid in S. Enteritidis was 1250 ppm at pH 5.0, 2500 ppm at pH 5.5, 6.0, 5.5 and 7.0, while in L monocytogenes 625 ppm at pH 5.5 and 125 ppm at pH 6.0, 6.5 and 7.0. MIC of combined hydrogen Peroxide and lactic acid in E. coli O157:H7, S. Enteritidis, and L. monocytogenes was 75 ppm of hydrogen peroxide with 2500 ppm of lactic acid at pH 6.5. The correlations between MICs of hydrogen peroxide and lactic acid in E. coli O157:H7, S. Enteritidis and L. monocytogene were obtained through the coefficient of $determination(R^2)$. $R^2$ value were 0.9994, 0.9935 and 0.9283, respectively. The inhibitory effect of hydrogen peroxide and lactic acid in E. coli O157:H7, S. Enteritidis and L. monocytogenes could be confirmed from the result of this experiment. Therefore, it was expected that the food process would increase or maintain by using lactic acid together with hydrogen peroxide.

• Journal of Microbiology and Biotechnology
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• v.13 no.4
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• pp.509-516
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• 2003

A model system for the immnunochemical detection of Escherichia coli O157:H7 using a combined immunomagnetic separation (IMS) and test-strip liposome immunoassay (LIA) procedure was developed. Immunomagnetic beads coated with anti-E. coli O157 IgG antibodies were used to separate the E. coli O157 (including the H7 serotype) from culture. Immunoliposomes, whose surface was conjugated to goat anti-E. coli O157:H7 IgG and which encapsulated the marker dye, sulforhodamine B, were used as a detection label. The test strip, onto which antibodies to goat IgG were immobilized, was the immunosensor capturing immunoliposomes that did not bind to E. coli O157:H7 on the immunomagnetic bead-E. coli O157:H7 complexes. In experiments, pure cell culture suspensions of $10^5 E.$ coli O157:H7 organisms per ml produced a measurable signal inhibition, whereas a weak yet detectable signal inhibition occurred with $10^3CFU/ml$. The inhibition signals increased, when the incubation time for IMS was extended to 90 min and higher IgG-tag density (0.4mol%) was used on the liposomes. With 0.2 and 0.4mol% IgG-tagged liposomes, the IMS-LIA procedure showed more improved signal inhibitions than those of a direct (no IMS) LIA. The combined assay, which measures the instantaneous signal from immunoliposomes, can be completed within 90 min, making it significantly faster than conventional plating methods and enzyme-linked immunosorbent assay (ELISA). Accordingly, it is quite feasible to use the combined immunoassay format of IMS and dye-loaded immunoliposomes for the detection of E. coli O157:H7.