본 연구는 정향추출물의 E. coli O157:H7에 대한 항균 효과와 E. coli O157:H7 감염 마우스에 대한 치료효과를 평가하기 위해 수행되었다. 정향 메탄올 추출물(FSAE)을 이용하여 E. coli O157:H7 에 대한 항균효과 확인 시험을 수행한 결과, 배양 후 24 시간째에, FSAE를 첨가한 모든 군들에서 E. coli O157:H7 의 생존율이 무투여 대조군에 비해 통계적으로 유의성 있게 감소하는 결과를 보여(0.269 mg/ml, p < 0.05; 0.538과 1.075 mg/ml, p < 0.001), FSAE가 E. coli O157:H7의 증식 억제 효과가 뛰어난 것으로 확인되었다. 또한, E. coli O157:H7을 감염시킨 마우스에 FSAE를 경구로 투여한 결과, 투여 후 3일째에, FSAE를 1.075 (p < 0.05)와 2.15 mg/ ml (p < 0.01)로 투여한 군들에서 대조군과 비교하여 분변 내 E. coli O157:H7의 균수가 유의성 있게 감소하였으며, 투여 7일째에는, 모든 FSAE 투여군들에서 대조군과 비교 하여 E. coli O157:H7의 균수가 통계적으로 유의성 있게 감소하였다(0.538 mg/ml, p < 0.05; 1.075와 2.15 mg/ml, p < 0.001). 이상의 연구결과로부터, FSAE를 E. coli O157: H7에 감염된 마우스에 경구로 투여할 경우, 감염증상을 완화 시킬 수 있을 것으로 기대된다.

This study was conducted to investigate the effect of cold plasma combined with UV-C irradiation against Escherichia coli O157:H7, Salmonella enterica serovar Typhimurium, and Listeria monocytogenes on lettuce. E. coli O157:H7, S. Typhimurium, and L. monocytogenes, corresponding to approximately 5.82, 5.09, 5.65 log CFU/ g, were inoculated on lettuce, respectively. Then, the lettuce was treated with cold plasma, UV-C and combination (cold plasma + UV-C), respectively. The treated lettuce was stored for 9 days at 4oC for microbiological analysis and sensory evaluation. Cold plasma reduced the populations of E. coli O157:H7, S. Typhimurium, and L. monocytogenes by 0.26, 0.65, and 0.93 log CFU/g, respectively. Each microorganism were reduced by 0.87, 0.88, and 1.14 log CFU/ g after UV-C treatment. And, the combined treatment that was treated by cold plasma after UV-C treatment reduced the populations of inoculated microorganisms by 1.44, 2.70, 1.62 log CFU/g, respectively. The all treatment significantly (p < 0.05) reduced the populations of all inoculated bacteria compared to untreated lettuce. UV-C combined with cold plasma was the most effective for reducing the pathogenic bacteria on lettuce, by showing log-reductions of ≥ 2.0 log CFU/g. All treatment was not significantly different until 6 day storage compared to control group in terms of appearance, texture and overall acceptability. Therefore, the combined treatment will be an effective intervention method to control the bacteria on lettuce.

Various methods for the detection of E. coli O157:H7 in food have been developed in the past decades. However, current detection methods require specialized instruments and lengthy preparation time. In an effort to achieve a rapid and sensitive detection, we developed a radial chromatography (RC) biosensor integrated with gold nanoparticle (AuNP) conjugated with antibody for the detection of E. coli O157:H7. The immuno-AuNP binds to E. coli O157:H7 creating AuNP-E. coli O157:H7 complexes by specific antigen-antibody interaction. The AuNP-E. coli O157:H7 complexes can be identified clearly from free AuNP by RC based on their mobility on porous matrix. Thus, the AuNP complexed with target bacteria, E. coli O157:H7 could be discriminated from free AuNP by radial chromatography. The results showed that the developed RC biosensor is highly selective to E. coli O157:H7 over non-target bacteria, including Bacillus cereus, Staphylococcus aureus and Vibrio cholerae with a detection limit of 105 CFU/ml. When combined with a pre-concentration step using immunomagnetic beads, we could further enhance the detection limit down to 103 CFU/ml. In this study, we developed a novel method that is rapid, sensitive and applicable for qualitative and quantitative detection of E. coli O157:H7. The detection procedure is simple and the results can be easily determined by naked eyes, suggesting that this system is practical and can be applied to the field diagnosis in food industry for the detection of pathogenic bacteria.

We developed a Amylose magnetic beads (AMBs) based detection system for high efficient separation, concentration and detection of E. coli O157:H7 in real sample. AMBs were synthesized by amylosucrase from Deinococcus geothermalis (DgAS) with iron-oxide nanoparticle (NP). The design of amylose magnetic beads (AMBs) have studied by an enzymatic synthesis with optimized reaction condition such as substrate, sucrose, and iron-oxide NP. AMBs have specific feature. AMBs decorated with functional fusion protein, which consists in a maltose binding protein (MBP) and a streptococcal protein G (SPG). Amylose chains has maltose, thus MBP-SPG binds to the AMB. In addition, SPG specifically binds to the Fc part of antibody. That was used as a linker to immobilize antibody to the surface of AMBs. The resulting AMBs were efficiently separated and concentrated target bacteria, E. coli O157:H7. Concentrated sample is qualitatively analyzed by PCR. Our studies demonstrated that AMB-based PCR significantly reduced the limitation of detection as low as 10 1 CFU/mL, compared to that of conventional PCR. The principle of this system can be served as a high efficiency for detection method of any pathogenic bacteria. In addition, AMBs and MBP-SPG cross-linker protein developed in this study is expected to be applicable to the portable food based biological processing monitoring system.

Shiga toxins (Stxs), some of the most important virulence factors in enterohemorrhagic Escherichia coli (EHEC) O157:H7, are known to be induced and released by various environmental cues, such as DNA damage responses and stress-inducing chemicals. In order to investigate the possible effects of growth media on Stxs expression, we analyzed the growth kinetics and expression of Stxs (Stx1 and 2) in cells grown in Luria-Bertani (LB) and E. coli (EC) media, which are widely used for EHEC O157:H7. Through direct plating and quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR), it was revealed that, when EHEC O157:H7 was grown in EC medium, the total bacterial count was reduced significantly and the stx1 transcription was greatly increased during the stationary growth phase than that in LB. Here we report that bile salts and lactose, which are the two only components in EC medium that are absent in LB, function as negative and positive regulatory signals, respectively, for the transcription of both stx1 and stx2. Indeed, stx transcription was significantly increased (~5.7 and ~21.8 fold for stx1 and stx2, respectively; p < 0.05) in an EC medium lacking bile salts when compared to the normal EC. In contrast, EHEC O157:H7 grown in an EC medium lacking lactose did significantly decrease these transcriptions (~93.5 and ~4.3 fold for stx1 and stx2, respectively; p < 0.05). Consistently, stx transcription was drastically increased in an LB medium supplemented with lactose, implying that lactose might be an environmental trigger for the expression of Stxs.

본 연구는 마늘의 물추출물(AGE)의 E. coli O157:H7,S. typhimurium, 그리고 S. aureus에 대한 항균효과를 조사하였다. AGE의 E. coli O157:H7, S. typhimurium, and S.aureus에 대한 최소억제농도(MIC)는 각각 24, 48 그리고 24 mg/mL이었으며, 최소살균농도(MBC)는 모든 균에 대하여 96 mg/mL이었다. AGE 24 mg/mL을 E. coli O157:H7에 투여하고 배양 24시간 후에, 균의 증식이 무처리 대조군과 비교하여 유의성 있게 감소하였다(p < 0.01). 그러나 AGE 24 mg/mL을 S. aureus에 투여하고 배양 24시간 후에, 대조군과 비교하여 유의성 있는 균의 증식억제 효과가 나타나지 않았으나, AGE 96 mg/mL을 투여한 경우에서는 배양 24시간 후에 대조군과 비교하여 유의성 있는 균의 증식억제가 나타났다(p < 0.01). AGE 48 (p < 0.05)과 96 mg/mL (p < 0.001)을 S. typhimurim에 투여하고 24시간배양 후에, 대조군과 비교하여 통계적으로 유의성 있는 균의 증식억제 효과가 나타났다. 본 연구의 결과로부터, 마늘의 물추출물은 항생제와 화학적 식품보존제의 대체제로서 사용이 가능할 것으로 사료된다.

The aim of study was to investigate the virulence profile of Escherichia coli O157:H7 bacteriophages isolated from sewage and livestock stools. Among 23 E. coli O157:H7 bacteriophages, 14 strains were isolated from sewage and 9 were from animal stools collected from 10 livestock farms in Korea. For each bacteriophage DNA sample, the presence of stx1, stx2, eae, aafII, ial, elt, estI, estII, astA, afa, and cnf was examined by polymerase chain reaction. The detection rate of eae, stx2, estI, astA, and ial was 100%, 69.6%, 13.0%, 13.0%, 8.7%, respectively. While all E. coli O157:H7 bacteriophages isolated from stools carried eae+stx2, stx2+eae, eae+astA, eae, stx2+eae+estI, eae+estI, stx2+eae+ial, and eae+ial were observed in bacteriophages isolated from sewage. As several plasmid-carrying virulence factors (estI, astA, and ial) were found in E. coli O157:H7 bacteriophages obtained from sewage and stools, the microbial safety of bacteriophages should be investigated in further study.

The objective of this study was to investigate the antimicrobial effects of carvacrol (CV) against Staphylococcus aureus (S. aureus) and Escherichia coli O157:H7 (E. coli O157:H7) strains in milk. The minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) of CV against S. aureus and E. coli O157:H7 were determined. In addition, bactericidal kinetics and antimicrobial activity of CV against the aforementioned pathogens in milk over a period of 2 weeks were investigated. CV exhibited antibacterial activity against both foodborne pathogens tested. The MIC and MBC of CV against S. aureus were 15.0 and 20 mg/mL, respectively, whereas those against E. coli O157:H7 were 16.0 and 32 mg/mL, respectively. In time-kill assays, CV at MBC reduced the number of S. aureus and E. coli O157:H7 in milk to undetectable levels within 24 hr. The antibacterial effects of CV persisted for 14 days without any loss of activity. Results of this study suggest that CV has a potential antibacterial activity against foodborne pathogens such as S. aureus and E. coli O157:H7 in milk.

Previously, we demonstrated the presence of a second copy of LPS myristoyl transferase in enterohemorrhagic Escherichia coli O157:H7, an important zoonotic diarrheagenic food-borne pathogen; the pO157-encoded ecf (an eae-conserved fragment) and the chromosomally-encoded lpxM (also referred to as msbM) genes. Although both genes share the same function as an LPS myristoyl transferase, the pO157-encoded ecf is thermoregulated via an intrinsically curved DNA while the chromosomal lpxM is regulated by the PhoP/Q two component regulatory system. However, it is unclear why E. coli O157: H7 carries two copies of LPS myristoyl transferase that are differentially regulated. In this study, a whole genome-scale transcriptome specific to E. coli O157:H7 was carried out for identification of the genes differentially expressed in the amyristoylated E. coli O157:H7. The results identified a total of 110 EHEC genes that were up- or down-regulated in the amyristoylated E. coli O157:H7 strain, including genes associated with virulence (26.36%), metabolism (20.91%), transport (10.91%), signal transduction (4.55%), genetic information processing (3.64%), stress response (2.73%), regulatory function (2.73%), motility/adherence (3.64%), cell envelope (2.73%), cell division (1.82%) and ORFs of unknown function (17.27%). Of particular interest, the expression of LEE pathogenicity island genes was significantly influenced by LPS structural defects.

본 연구는 polyvinyl chroride (PVC)와 stainless steel 표면에서 E. coli O157:H7의 부착, 바이오필름형성, 온·습도에 따른 생존율을 조사하고, E. coli O157:H7이 오염된 작업대 표면에서 상추로의 교차오염도 조사를 통하여 E. coli O157:H7에 오염된 작업대가 상추의 미생물학적 안전성에 미치는 영향을 평가하고자 수행하였다. 작업대 표면 재질인 PVC와 stainless steel에 E. coli O157:H7 배양액을 1시간 노출시켰을 때, PVC의 경우, stainless steel 보다 10배 정도 부착력이 높았으나, 6시간째는 두 재질간의 차이가 없었다. 또한, E. coli O157:H7의 바이오필름 형성은 TSB > 10% 상추추출액 > 1% 상추추출액 > phosphate buffer 순이었으며, 재질별로는 PVC에서 바이오필름형성능이 높은 것으로 나타났다. 온도 20oC, 30oC, 습도 43%, 69%, 100% 에서 각각 노출 시켰을 때, E. coli O157:H7은 온도 30oC, 습 도 43%, 69% 조건에서 12시간 내에 약 5.0 log CFU/coupon이 감소한데 반해, 상대습도 100%에서는 농도의 큰 변화 없이 5 일 이상 생존이 지속되었다. 또한, 유기물이 작업대 표면에 존재 시, E. coli O157:H7의 감소 속도가 느렸다. E. coli O157:H7 에 오염된 작업대에 상추를 접촉시키고 상추의 오염도를 조사한 결과, 상추 표면에 수분이 존재할 때 오염된 작업대 표면에서 E. coli O157:H7의 이동수준이 상추에 수분이 없을 때 보다 10 배 이상 증가하는 것으로 나타났다. 따라서 E. coli O157:H7에 오염된 작업대는 상추의 안전성에 직접적인 영향을 미칠 수 있어 작업 후에 세척, 소독을 통하여 위생적으로 관리하는 것이 필요하다.

본 연구는 신선편이 식품에서 오염 가능성이 있는 병원성 식중독 균 E. coli O157:H7에 대해 파프리카에서 성장 예측 모델을 적용하고, 본 연구에서 개발된 성장 예측 모델을 내부 검증하였다. 이를 비교하여 신선편이 식품을 안 전하게 관리하기 위한 적절한 모델을 제시하고자 하였다. 파프리카에 E. coli O157:H7접종하여 온도에 따라 12, 24, 30, 36℃에 보관하여 성장을 측정하였다. Gompertz 식을 이용하여 온도에 따른 성장곡선을 그리고 LT와 SGR을 산출하였다. 산출된 LT와 SGR은 각각 Davey model 와 squareroot model를 이용하여 2차 모델을 개발하였다. 개발된 2차 모델에 대하여 LT 와 SGR model의 R2값은 각각 0.999, 0.994로 1에 근접하는 높은 적합성을 보였다. 또한 내부 검정 결과 LT 와 SGR model 의 Bf 값은 각각 1.01, 0.89, LT model은 안전하게 SGR model 위험하게 예측되었다. 파프리카의 LT와 SGR의 상대적인 오차 값은 모두 허용 가능한 오차 범위에 포함 되었다. 따라서 개발된 모델을 이용하여 온도에 따른 E. coli O157:H7성장을 추정할 수 있으며, 이를 위해평가 자료로 활용할 수 있을 것으로 보인다.

The present study was evaluated the antibacterial effect of the combination of Coptidis rhizoma, Glycyrrhiza uralensis Fischet, Schizandra chinensis and Corni Fructus(1:1:1) extracts(CGSC10). Furthermore, the effectiveness of CGSC10, sodium chlorate, and the combination of CGSC10 and sodium chlorate(CGSCS10) against E. coli O157:H7 infection was studied using ICR female mice. During the incubation period, the dose of 5, 10, and 20% CGSC10 was inhibited the growth of E. coli O157:H7 by 34.7, 60.2, and 76.4%, respectively. For 7 days after single challenge with E. coli O157:H7, forty female ICR mice were divided into four experimental groups which were administered in drinking water with saline, 10% CGSC10, 15 mM sodium chlorate, and CGSCS10, respectively. On the 3rd day, the number of E. coli O157:H7 in mouse feces was significantly decreased by administration of CGSC10, 15 mM sodium chlorate, and CGSCS10 (p < 0.001). On the 7th day-after administration, CGSC10, sodium chlorate, and CGSCS10 were decreased the number of E. coli O157:H7 by 27.1, 67.7, and 83.3%, respectively. According to the results of the present study, administration of CGSCS10 to mice can reduce the severity of E. coli O157:H7 infection. In addition, it is suggested that CGSCS10 represents a good candidate for the treatment of enteric infections in domestic animals.

This study was to evaluate the growth potential of E. coli O157:H7 in lettuce leaf extracts and on lettuce leaf surface at various temperatures. The pathogen can survive and multiply in the extracts and leaf surface of lettuce. The population of E. coli O157:H7 in the lettuce extracts reached to 4.79 log CFU/mL at 37℃. The multiplication of pathogen in lettuce extracts initiated within 10 hours of inoculation over 15℃ conditions. And it can survive in the lettuce leaf extracts at 4℃ for 100 hours at least. And this pathogen can multiply on lettuce leaf surface and the population of pathogen on the lettuce leaf surface increased to 1.82 log CFU/g at 25℃. At 37℃, the pathogen density increased to 1.53 CFU/g within 3 days after inoculation. At all temperature, irrespective of the inoculation level, similar trends in growth of E. coli O157:H7 were observed. These results emphasize the growth potential of E. coli O157:H7 in lettuce leaf extract and on lettuce leaf surface. To reduce the risk of outbreak, it is important to maintain the cold chain system during storage before the consumption.

In this study, two duplex real-time PCR approach with melting curve analysis is presented for the detection of Escherichia coli O157:H7, Listeria monocytogenes, Salmonella spp. and Staphylococcus aureus, which are important food-borne bacterial pathogens usually present in fresh and/or minimally processed vegetables. Reaction conditions were adjusted for the simultaneous amplification and detection of specific fragments in the β-glucuronidase (uidA, E. coli), thermonuclease (nuc, S. aureus), hemolycin (hly, L. monocytogenes) and tetrathionate reductase (ttr, Salmonella spp.) genes. Melting curve analysis using a SYBR Green I real-time PCR approach showed characteristic Tm values demonstrating the specific and efficient amplification of the four pathogens; 80.6 ± 0.9 ℃,86.9 ± 0.5 ℃, 80.4 ± 0.6 ℃ and 88.1 ± 0.11 ℃ for S. aureus, E. coli O157:H7, L. monocytogenes and Salmonella spp.,respectively. For all the pathogens, the two duplex, real-time PCR was equally sensitive to uniplex real-time PCR,using same amounts of purified DNA, and allowed detection of 10 genome equivalents. When our established duplex real-time PCR assay was applied to artificially inoculated fresh lettuce, the detection limit was 10³ CFU/g for each of these pathogens without enrichment. The results from this study showed that the developed duplex real-time PCR with melting curve analysis is promising as a rapid and cost-effective test method for improving food safety.

The present study was evaluated the antibacterial effect of the combination of Coptidis rhizoma,Lonicerae Flos, and Paeonia japonica (1:1:1) extracts (CLP1000). Also, the effectiveness of CLP1000, dioctahedral smectite (DHS), and the combination of CLP1000 and DHS (CLPS1000) against E. coli O157:H7 infection was studied using ICR female mice. During the incubation period, the dose of 10% and 20% CLP1000 were inhibited the growth of E. coli O157:H7 by 30% and 47%, respectively. For 7 days after single challenge with E. coli O157:H7, forty female ICR mice were divided into four experimental groups which were orally administered with saline, 10% CLP1000, 10% DHS, and 10% CLPS1000, respectively. On the 3rd day, the number of E. coli O157:H7 in mouse feces was significantly decreased by administration of CLP1000 (p < 0.05), DHS (p < 0.05) and CLPS1000 (p < 0.001). On the 7th day, CLP1000 (p < 0.05) and CLPS1000 (p < 0.001) administration significantly decreased the number of E. coli O157:H7. According to the results of the present study, administration of CLPS1000 to mice can reduce the severity of E. coli O157:H7 infection. Also, it is suggested that CLPS100 represents a good candidate for the treatment of enteric infections in domestic animals.

This study was conducted to develop a model for describing the effect of storage temperature (4, 10, 15, 20, 25, 30 and 35℃) on the growth of Escherichia coli O157 : H7 in ready-to-eat (RTE) lettuce treated with or without (control) alkaline electrolyzed water (AIEW). The growth curves were well fitted with the Gompertz equation, which was used to determine the specific growth rate (SGR) and lag time (LT) of E. coli O157 : H7 (R2 = 0.994). Results showed that the obtained SGR and LT were dependent on the storage temperature. The growth rate increased with increasing temperature from 4 to 35℃. The square root models were used to evaluate the effect of storage temperature on the growth of E. coli O157 : H7 in lettuce samples treated without or with AIEW. The coefficient of determination (R2), adjusted determination coefficient (R2 Adj), and mean square error (MSE) were employed to validate the established models. It showed that R2 and R2 Adj were close to 1 (> 0.93), and MSE calculated from models of untreated and treated lettuce were 0.031 and 0.025, respectively. The results demonstrated that the overall predictions of the growth of E. coli O157 : H7 agreed with the observed data.