Enteropathogenic Escherichia coli (EPEC) have developed survival strategies to evade host defense systems. The intracellular level of guanosine tetraphosphate (ppGpp) controlled by RelA and SpoT can mediate immune evasion of EPEC. However, the impact of ppGpp-defective EPEC infection on phagocytes remains unknown. In this study, we report that disrupting relA and spoT of EPEC E2348/69 strain promotes its phagocytosis in porcine macrophages. Our experimental analysis showed that both uptake and killing of an E2348/69 ΔrelAΔspoT mutant by macrophages were increased compared to those of wildtype strain. These results suggest that ppGpp plays an essential role in evading phagocytosis during EPEC pathogenesis.

본 연구는 집단급식소에서 제공되는 빈도수가 높은 비 가열 및 가열조리 엽경채류에 사용되는 차아염소산나트륨 수에 대하여 미생물적 안전성을 평가하고자 수행되었다. 비병원성 대장균과 장출혈성 대장균의 칵테일(E. coli O157:H7)을 엽경채류(초기 균수 7-8 log CFU/g)에 인위적 으로 오염시킨 후 차아염소산나트륨을 5분간 침지 후 흐 르는 물에 3번 씻어서 생균수를 측정하였다. 실험 결과 초 기 오염물질에 비해 살균효과가 1-2 log CFU/g 저감화하 여 대조군에 대해 유의적인 차이가 있었다(P<0.05). 잎채 소의 특성에 따라 약간의 차이가 있었는데 표면적이 클수 록, 덜 거칠고 잎이 부드러울수록 살균효과가 높았다. 200 mg/kg으로 처리하였을 때 100 mg/kg에 비해 0.1-0.3 log CFU/g만큼 효과가 더 감소하였으나 농도 증가에 따 른 유의적 차이는 없었다(P>0.05). 그러므로 학교급식위생 관리지침에서 제시한 기준 이상으로 차아염소산나트륨 농 도를 높이는 것은 불필요하다고 판단된다. 그러나 잎채소 는 일반적으로 미생물의 초기 오염도가 높기 때문에 차아 염소산나트륨 처리만으로는 안전한 수준의 저감을 달성하 기 어려워 생물학적 위험이 잔존한다. 따라서 여름철에 가 열하지 않은 잎채소의 대체 조리방법을 개발하는 것이 안 전성에 보다 효과적인 것으로 판단된다.

Enteropathogenic Escherichia coli (EPEC) is one of the etiological agents that causes diarrhea in weaning pigs. In this study, we report that mutating both relA and spoT genes in EPEC E2348/69 can promote bacterial clearance in porcine gastrointestinal tract (GIT). Our experimental analyses showed that an E2348/69 ΔrelAΔspoT mutant strain was not detected in porcine feces after 1 day post-infection (dpi), whereas its parental strain was continuously detected in porcine feces within 10 dpi. Histologic assessment revealed that the mutant strain was unable to induce moderate pathologic lesions in porcine GIT when compared to those with the wildtype strain. Taken together, our data suggest that the relA and spoT genes in EPEC play an important role in bacterial survival and pathogenesis in porcine GIT.

Water disinfection using UV-LED(Light emitting diode) has many advantages, such as smaller footprint and power consumption as well as relatively longer lifespan than those of conventional mercury-UV lamps. Moreover, UV-LED disinfection is considered an environmentally benign process due to its mercury-free nature. In this study, disinfection using an LED module emitting 275nm UV was carried out. 384 UV-LEDs were put into a cylinder tube with a capacity of 1.7 liters. The UV intensity of the UV-LED module was controlled from 1.7 to 8.4 mW/cm2. The disinfection efficiency for the model microorganism solutions(E. coli ) was monitored. As the UV intensity(I) and contact time(t) varied, inactivation of the microorganisms from 2 to 4-log-removals(i.e., 99 to 99.99% of disinfection efficiency) was achieved. Disinfection using UV-LED was followed to 1st order reaction and the reaction rate constant, k was determined. In addition, the relationship between UV intensity(I) and contact time(t) in order to obtain 99.99% of disinfection efficiency was modeled: I1.2∙𝑡= 460, which indicates that the product of UV intensity and contact time requiring 4-log-removals is always constant.

The widespread emergence of antimicrobial-resistant E. coli is now being seen in humans and animals, and there is an increasing prevalence of multidrug-resistant (MDR) E. coli on a global. In this study, 31 MDR E. coli isolates recovered from pork meat at retail markets were analyzed to determine the phenotypic and genetic characteristics. The resistance to ampicillin (30 isolates, 96.8%) and tetracycline (28 isolates, 90.3%) were the most frequent among the MDR E. coli isolates. Thirty (96.8%) of MDE E. coli isolates harbored the ß-lactamase–encoding gene blaTEM. Twenty-six (83.9%) isolates harbored a class 1 integron, and 30 (96.8%) isolates carried from 2 to 6 resistance genes. Frep (45.2%) and FIB (22.6%) replicons were most common on the replicon typing. This study suggests that monitoring of the antimicrobial resistance of the pathogens found in pork meat should be continued in the future due to concerns about food safety issues.

Many β-lactam antimicrobials, including cephalosporins, have been used in both veterinary and human medicine in the treatment of zoonotic and infectious diseases. Especially, third-generation cephalosporins such as ceftiofur have been approved for systemic use in food-producing animals, which has resulted in the emergence of β-lactamase genes. This study aimed to investigate the occurrence of β-lactamase-producing E. coli isolated from commercial layers and characterized their antimicrobial resistance and virulence genes. Among the 85 cefotaxime (CTX)-resistant E. coli, all isolates showed resistance to at least one antimicrobial, and the rates of resistance to nalidixic acid, cephalothin, ampicillin, and cefazolin were more than 50.0%. In particular, 28 isolates were identified as containing b-lactamase genes. The extended-spectrum β-lactamase (ESBL) and plasmid-mediated AmpC genes blaCTX-M-1, blaCTX-M-14, blaCTX-M-15, and blaCMY-2 were detected in 1, 6, 5, and 4 isolates, respectively. The non-ESBL/pAmpC gene blaTEM-1 was detected in 12 isolates. The distribution of antimicrobial resistance genes in 28 β-lactamase-producing E. coli was as follows: aac(3)-II (64.3%), sul2 (32.1%), tetA (28.6%), sul1 (25.0%), cmlA gene (25.0%), and tetB (14.3%). In total, 6 virulence genes (astA, eaeA, escV, fimH, iucC, and papC) were also identified and the rates in virulence gene were as below: fimH (92.9%), iucC (25.0%), astA (21.4%), papC (10.7%), eaeA (7.1%) and escV (7.1%). Our findings suggest that antimicrobials used in commercial layer must be regulated in Korea, and comprehensive surveillance is necessary to prevent the dissemination of resistant isolates.

Chlorine dioxide gas is a relatively new sanitizer in the food industry that has more accessibility than its aqueous form. Depending on the method by which ClO2 gas is generated, there can be byproducts like chlorite and chlorate ions, which can decrease its disinfectant effect and purity. Recently, new technology that generates chlorine dioxide without using chlorine gas has been developed to remove those defects. This new electrochemical method generates gaseous chlorine dioxide from aqueous sodium chlorite (NaClO2). The present study was conducted to evaluate the effects of ClO2 gas generated by an electrochemical method against foodborne microorganisms. To accomplish this, ClO2 gas at different concentrations (1, 5, 10 and 20 ppm) was applied to E. coli and S. Typhimurium for different exposure times (1, 5, 10, 15 and 20 min) under room temperature conditions at <40% relative humidity. The results revealed ClO2 gas was highly effective for the inactivation of E. coli and S. Typhimurium and showed a reduction in populations of over 5 log CFU/ml under ambient conditions with low relative humidity (30–40%). In conclusion, ClO2 gas treatment is highly applicable to control of foodborne pathogens.

Rapid, simple, and sensitive detection of pathogen bacteria is a highly topical research area due to increasingly concerning of food safety and public health. Surface-enhanced Raman spectroscopy (SERS) is a promising and attractive technique offering fast, sensitive, comparatively low-cost, and in-suit detection of pathogenic bacteria. However, this technique requires the preparation step for reducing the noise derived from heterogeneous matrixes of food sample. Immunomagnetic separation (IMS) is widely used technique enabling separation and concentration of the target analyte. It can be used not only laboratory scale but also field diagnosis easily. Here, we synthesized gold-shelled starch magnetic microparticles (GS@SMMPs) for effective separation and concentration of Escherichia coli O157:H7, which were subsequently subjected to SERS integrated with gold-coated 3D-well substrate for bacterial detection in aqueous solution. GS@SMMPs were labelled by Anti-E. coli O157 monoclonal antibody through gold binding protein and staphylococcal protein G (GBP-SPG) fusion protein. In IMS experiment, the immuno-GS@SMMPs showed high capture efficiency over 90% to E. coli O157:H7, which resulted in 10 times decrease in detection limit in PCR assay. Through SERS assay, E. coli O157:H7 concentrated by immuno-GS@SMMPs were successfully detected even at an extremely low concentration of 101 CFU/ml the subjected to SERS. Moreover, by using sandwich method using SERS reporter consisting of GBP-SPG, we found that E. coli O157:H7 were able to be detected by SERS quantitatively through measuring the SERS intensity of GBP-SPG. This novel strategy combining SERS and IMS could be meaningful for extending the application in SERS for in-suit sensitive detection of pathogenic bacteria.

It has been known that the ability of Shiga toxinproducing Escherichia coli (STEC) to produce Stx2e in culture media plays a role in the diagnosis of edema disease and determination of subunit vaccine candidates in STEC isolates. To examine the efficiency of Stx2e production in several commercial media, a Stx2e-producing strain (KEFS1302) was grown in four different media: ISO-Sensitest broth (ISB), E. coli broth (ECB), trypticase soy broth (TSB), and Mueller Hinton broth (MHB), with or without mitomycin C at 37°C (250 rpm) for 6 h. Toxin production was measured by enzyme-linked immunosorbent assay. In the presence of mitomycin C, ECB was found to be the most suitable medium, reaching a production peak (OD600 = 1.2) at 1 h; Stx2e was mostly produced during the logarithmic phase (within 3 h). On the other hand, toxin production in ISB reached a peak at 3 h after incubation in the absence of mitomycin C. Stx2e was purified by fast protein liquid chromatography (FPLC) using anion-exchange chromatography. The 43 kDa band of Stx2e was confirmed by western blot using the ECB supernatant. Our results showed that ECB and ISB media would be a suitable medium for mass production of Stx2e even if the toxin production is dependent on time.

The emergence of antimicrobial resistant Escherichia (E.) coli is a major problem in pig farms. To tackle this issue, in July 2011, the Korean government banned the use of antimicrobials for growth promotion of animals in farms. Moreover, E. coli encoding the Stx2e gene cause edema disease which results in high mortality and morbidity in pig farms. Therefore, the aim of this study was to investigate the prevalence of antimicrobial resistance among E. coli encoding the Stx2e gene isolated from weaned piglets with diarrhea before and after the ban on antibiotic growth promoters (AGPs) in Korea from 2007 to 2016. In this period, 479 E. coli isolates were obtained from weaned piglets with diarrhea, and of them, 144 E. coli isolates encoding the Stx2e gene were detected by polymerase chain reaction. The susceptibility of the E. coli isolates to antibiotics were tested using the standard Kirby-Bauer disk diffusion method. The most frequently observed resistances in isolates obtained from weaned piglets in the last 10 years were to tetracycline (92.4%) and chloramphenicol (88.9%). The prevalence of resistance to colistin (3.1% to 16.5%) and tetracycline (86.2% to 97.5%) was also observed to have increased over this period. Additionally, multi-drug resistance was also found to have increased (87.7% to 97.5%) after the ban on AGPs. These findings provide useful data for designing prevention and treatment strategies for postweaning diarrhea and edema disease, and can be used in future studies on antimicrobial resistance in Korea.

본 연구는 다양한 농업 환경에서 채집된 파리의 분비물에서 E. coli을 분리하고 분리된 E. coli의 병원성유전자 및 항생제내성을 조사하기 위하여 수행되였다. 파리는 과일농장(n = 19), 장류생산농장(n = 9), 생활쓰레기 야적장(n = 46), 축사(n = 66), 도축장(n = 38), 퇴비장(n = 10)에서 총 188 마리를 채집하여 토사물과 배설물로부터 E. coli을 분리 및 동정하였다. 그 결과, 채집된 파리의 63%(119/188)에서 E. coli이 검출되었으며 특히 도축장에서 채집된 파리에서 E. coli의 검출률이 89%(34/38)로 가장 높았다. 또한 분리된 E. coli을 대상으로 병원성 유전자 8종(ST, LT, VT1, VT2, aggR, bfpA, eaeA, ipaH)을 조사한 결과, 도축장에서 채집 된 파리에서 분리된 E. coli 중 91%(31/34)가 장독소를 생산할 수 있는 ST유전자를 보유하고 있었다. 분리된 E. coli 의 16%(31/188)가 1종 이상의 항생제에 내성을 보였다. 특 히, 항생제사용빈도가 높은 축사에서 채집된 파리의 E. coli 경우에는 59%(23/39)가 항생제 내성을 나타내었다. 분 리된 항생제 내성 E. coli 균주 중 10%(12/119)는 2종 이 상의 항생제에 내성을 보였고, 모두 축사 채집 파리에서 분리된 균주였으며, 이 중 2개 균주는 다재내성의 지표인 ESBL (Extended-spectrum beta-lactamase)에 양성을 나타내었다. ESBL 양성균주 중 1 균주는 7종의 항생제에 내성 을 보였이는 것으로 조사되었다. 본 연구의 결과, 축산환 경 서식 파리에서 분리된 E. coli은 병원성 유전자를 보유 하고 있을 가능성이 높을 뿐만 아니라 항생제에 내성을 나타낼 가능성도 높기 때문에 농식품을 생산하는 농장이나 식품공장은 가급적 축산환경으로부터 일정한 거리를 두거나 방충망 등의 차단조치가 필요할 것으로 판단된다.

본 연구에서는 선택배지와 건조필름을 이용하여 신선편이 엽채류 3종(양상추, 양배추, 어린잎채소)에서 병원성 E. coli를 분리하였고, 의심집락 동정을 위해 생화학적 분석법을 사용하여 동정한 후 결과를 비교 분석하였다. 양상추 20 g, 양배추 20 g, 어린잎채소 10 g에 병원성 E. coli 혼합 균액(Enterohemorrhagic E. coli NCCP11142, Enterotoxigenic E. coli NCCP14037, Enteropathogenic E. coli NCCP14038, Enteroaggregative E. coli NCCP14039, Enteropathogenic E. coli NCCP15661)을 최종농도가 1, 2, 3 log CFU/g이 되도록 접종하였고, BPW (80~90 ml)을 넣은 후60초 동안 균질화하여 분석하였다. 연구결과, 모든 시료에서 건조필름 시험 양성, 양상추 시료 일부(최종농도 3 log CFU/g 접종시료)를 제외한 모든 시료에서 증균배양법을 이용한 정성시험결과가 음성으로 나타났다. 증균배양과 건조필름 시험을 통해 분리한 병원성 E. coli를 이용하여 생화학적 분석을 실시한 결과, 양상추에서 분리한 병원성 E. coli의 경우, API 20E 100% (44/44), Microgen GNA 100% (44/44), Food System 66.7% (10/15)의 동정률이 나타났다. 양배추의 경우, API 20E 64.7% (22/34), Microgen GNA 50% (16/32), Food System 60% (9/15), 어린잎채소의 경우, API 20E 65.1% (28/43), Microgen GNA 62.3% (27/43), Food System 53.3% (8/15)가 병원성 E. coli로 동정되었다. 본 연구의 결과는 신선편이 엽채류에 대한 병원성 E. coli 검출법 선택에 유용하게 이용될 것으로 판단된다.

Disinfection of microorganisms using UV light is widely used in the field of water supply and wastewater treatment plant, In spite of high germicidal effect and relatively clean by-product, UV disinfection has fundamental defeat that is accumulation of fouling materials at the interface of water and lamp sleeve. Non-contact type of UV photoreactor which can avoid this fouling generation was developed and the experimental performance evaluation of the system was carried out in this study. Log inactivation rate of E. coli was selected as a disinfection index. The concentration of E. coli of second clarifier effluent was 8.2×101 - 8.2×103 colony per mL and was well inactivated by the non-contact type of UV photoreactor. Under the UV intensity condition of 2.1 - 2.5mW/cm2, E. coli removal rate was observed in the range of 54 - 95% when the HRT was increased from 10 to 52 seconds. Experimental results showed that log inactivation of E. coli was proportional to UV dosage and 200mJ/cm2 of UV dose is expected for the 2.0 log inactivation of E. coli from the second clarifier effluent. Between the two parameters of UV intensity and contact time which are consist of UV dose, UV intensity was 4 times more effective than contact time.s

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.

Antibodies are used for preventing several infectious diseases in food animals. Though, antibiotics can deal with infection however, their widespread usage may cause developments of resistance and can also get transferred to humans through animal products. Therefore, production of antibodies against infectious agents in the egg yolk could be an interesting alternative. The present study was conducted with a focus on producing the antibody against Escherichia coli E68 and H28 in egg yolk of laying hens, which are believed to be a cause for diarrhea and beriberi in piglets. The result received from the experiment were promising and have shown efficiency of antibody production in the range of 2-7.3% in the groups which are infected with E68 only and the one which was infected with both strains. The outcome of present research has revealed the potentiality of egg yolk in production of antibody for laying hens, could open a new approach for production of antibodies to manage diarrhea and beriberi