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Frieda, Ani Noor, Prastyoningsih Aris, Wisnu P. Kanita Maria, et al. "Socialization Android-Based Electronic Screening Test Epidemiology (E-Steco 19) In Indonesia and Timor Leste." RA JOURNAL OF APPLIED RESEARCH 09, no. 02 (2023): 66–76. https://doi.org/10.5281/zenodo.7614525.
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<strong>Introduction: </strong>A statement of being in good health is one of the prerequisites as a lecturer or employee at Kusuma Husada University, in which an employee must be declared in good health to prove that an employee is healthy and ready to work, without any serious unhealthy obstacles. Health checks by computerized epidemiology screening test will speed up the process and assist the doctor The development of health applications using the epidemiological screening test method for lecturers and Studends, at the Kusuma Husada University Health Clinic Indonesia Health screening tests quick and efficient in accordance with existing data, and give the result of employee health data directly. The purpose of this study is to make it easier to detect the health status of Kusuma Husada University lecturers and Studends, which is used for one of the prerequisites for a work contract with a health application using an epidemiological screening test method. <strong>Methods: </strong>Data collection was done by qualitative method and carried out from February to August 2022. lecturers and Studends data retrieval is done using a digital questionnaire (online). Sample of this research is a studends, and employee at Kusuma Husada University Indonesia, the results is used to make applications (e-Steco). <strong>Results: </strong>E-Steco 19 is an Android-based application that is used to detect a person's health using the screening test method. Data will be stored using Google Sheet and Google App Script. Thus, the data that has been collected from users who fill out a list of questions in the application can be downloaded and processed according to the needs. Data can be downloaded via the link. The e-Steco result can be processed in such a way that the data can be used as material for decision making. E-Steco 19 consists of 14 questions for health screening. Here we show the results of data processing from e-Steco 19 Result in the form of a pie chart. <strong>Results: </strong>The e-Steco 19 application has been completed and has also been running a test, so that it can make it easier to detect the health of lecturers and Studends, at Kusuma Husada University Indonesia. This application is still possible to be developed and used in Timor Leste University and Health Clinic.
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LIN, ANDREW, LAM NGUYEN, LAURIE M. CLOTILDE, JULIE A. KASE, INSOOK SON, and CAROL R. LAUZON. "Isolation of Shiga Toxin–Producing Escherichia coli from Fresh Produce Using STEC Heart Infusion Washed Blood Agar with Mitomycin-C†." Journal of Food Protection 75, no. 11 (2012): 2028–30. http://dx.doi.org/10.4315/0362-028x.jfp-12-157.
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The ability to detect and isolate Shiga toxin–producing Escherichia coli (STEC) remains a major challenge for food microbiologists. Although methods based on nucleic acids and antibodies have improved detection of STECs in foods, isolation of these bacteria remains arduous. STEC isolation is necessary for matching food, environmental, and clinical isolates during outbreak investigations and for distinguishing between pathogenic and nonpathogenic organisms. STEC heart infusion washed blood agar with mitomycin-C (SHIBAM) is a modification of washed sheep blood agar prepared by adding mitomycin-C and optimizing both the washed blood and base agar to better isolate STECs. Most STEC isolates produce a zone of hemolysis on SHIBAM plates and are easily distinguishable from background microbiota. Here, we present data supporting the use of SHIBAM to isolate STECs from fresh produce. SHIBAM was tested for accuracy in identifying STECs (365 of 410 STEC strains were hemolytic, and 63 of 73 E. coli strains that did not produce Shiga toxin were not hemolytic) and for recovery from artificially inoculated fresh produce (11 of 24 romaine lettuce samples and 6 of 24 tomato samples). STEC recovery with SHIBAM agar was greatly improved when compared with recovery on Levine's eosin–methylene blue agar as a reference method.
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SMITH, JAMES L., and PINA M. FRATAMICO. "Effect of Stress on Non-O157 Shiga Toxin–Producing Escherichia coli†." Journal of Food Protection 75, no. 12 (2012): 2241–50. http://dx.doi.org/10.4315/0362-028x.jfp-12-255.
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Non-O157 Shiga toxin–producing Escherichia coli (non-O157 STEC) strains have emerged as important foodborne pathogens worldwide. Non-O157 STEC serogroups O26, O45, O103, O111, O121, and O145 have been declared as adulterants in beef by the U.S. Department of Agriculture Food Safety and Inspection Service. While documentation is limited, treatments including heat and acid that have been shown to inactivate E. coli O157:H7 will likely also destroy non-O157 STEC; however, non-O157 STEC strains show variability in their responses to stress. It has been shown that non-O157 STEC may survive in fermented sausages and cheeses, and treatments such as high pressure may be necessary to eliminate non-O157 STEC from these products. The mechanisms used by non-O157 STEC to resist acid environments are similar to those used by O157:H7 strains and include the acid tolerance response, the oxidative system, and the glutamate and arginine decarboxylase systems. However, one study demonstrated that some non-O157 STEC strains utilize a chaperone-based acid stress response (HdeA and HdeB) to combat acidic conditions, which is lacking in E. coli O157:H7. Genomic studies suggest that while non-O157 STEC can cause diseases similar to those caused by E. coli O157:H7, O157 and non-O157 STECs have different evolutionary histories. Non-O157 STECs are a heterogeneous group of organisms, and there is currently a limited amount of information on their virulence, fitness, and stress responses, rendering it difficult to draw firm conclusions on their behavior when exposed to stress in the environment, in food, and during processing.
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Wiriyaprom, Ratchakul, Ruttayaporn Ngasaman, Domechai Kaewnoi, and Sakaoporn Prachantasena. "Prevalence and Virulent Gene Profiles of Sorbitol Non-Fermenting Shiga Toxin-Producing Escherichia coli Isolated from Goats in Southern Thailand." Tropical Medicine and Infectious Disease 7, no. 11 (2022): 357. http://dx.doi.org/10.3390/tropicalmed7110357.
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Shiga toxin-producing Escherichia coli (STEC) is the pathogenic E. coli causing disease in humans via the consumption or handling of animal food products. The high prevalence of these organisms in ruminants has been widely reported. Among STECs, O157 is one of the most lethal serotypes causing serious disease in humans. The present study investigated the prevalence of sorbitol non-fermenting STECs in goats reared in the lower region of southern Thailand and described the virulent factors carried by those isolates. Sorbitol non-fermenting (SNF)-STECs were found in 57 out of 646 goats (8.82%; 95% CI 6.75% to 11.28%). Molecular identification revealed that 0.77% of SNF-STEC isolates were the O157 serotype. Shiga toxin genes (stx1 and stx2) and other virulent genes (i.e., eaeA, ehxA, and saa) were detected by molecular techniques. The presence of stx1 (75.44%) was significantly higher than that of stx2 (22.81%), whereas 1.75% of the total isolates carried both stx1 and stx2. Most of the isolates carried ehxA for 75.44%, followed by saa (42.11%) and eaeA (12.28%). In addition, 21.05% of STEC isolates did not carry any eaeA, ehxA, or saa. The first investigation on SNF-STECs in goat was conducted in the lower region of southern Thailand. The present study revealed that goats could be one of the potential carriers of SNF-STECs in the observing area.
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Ghaderi, P., E. Ahmadi, A. M. Farrokhi, et al. "Prevalence and molecular characterisation of Shiga toxin-producing Escherichia coli in sheep farms of Sanandaj, Iran." BULGARIAN JOURNAL OF VETERINARY MEDICINE 27, no. 2 (2024): 206–14. http://dx.doi.org/10.15547/bjvm.2022-0056.
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Shiga toxin-producing Escherichia coli (STEC) strains have emerged as important foodborne pathogens of global public health concern, causing life-threatening diseases. Animals and their products have been documented as important reservoirs for STECs, especially E. coli O157. The aim of this study was to investigate STECs from healthy and diarrhoeic sheep in Sanandaj, Iran. In the current study, a total of 81 sheep faecal samples were taken (22 from diarrhoeic sheep and 59 from healthy sheep). E. coli and subsequently STEC strains was detected according to standard protocol (cultural characterisation and PCR assays). Finally, the frequency of Shiga-toxin producing gene(s) (stx1, stx2), intimin (eaeA) and enterohaemolysin (hlyA) was detected among STEC isolates using duplex PCR. Totally, 42 E. coli were isolated from 81 faecal samples (51.85% contamination). Of these, 34 isolates (80.9%) were identified as STEC patotypes based on Sorbitol-MacConkey (SMAC) medium culturing and also the presence of stx1 and/or stx2. Of these, only 3 isolates (7.1%) were identified as serotype O157:H7 based on PCR assay. In addition, the results showed that STEC bacteria were significantly more prevalent in diarrhoeic samples than in healthy samples (50% vs. 22.1%). Overall, the PCR results showed that 33 (97%), 12 (35.3%) and 8 (23.5%) isolates carried stx1, stx2 and hlyA, respectively. The eaeA gene was not found in any isolate. The number of isolated STEC bacteria in spring (10 isolates) and winter (14 isolates) were significantly higher than those in summer (4 isolates) and autumn (6 isolates) (P=0.039). Also, the number of STEC in diarrhoea samples was significantly higher compared to non-diarrhoea samples (P=0.032). In conclusion, the present study revealed high prevalence rate of STEC including serotype O157:H7 and non-O157:H7 in sheep faeces which highlights the importance of sheep as a reservoir of STEC pathogen in Sanandaj region. Therefore, additional control and preventive measures must be undertaken to control the contamination by this pathogen.
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Nehoya, Kaarina N., Ndinomholo Hamatui, Renatus P. Shilangale, Harris Onywera, Jeya Kennedy, and Lamech M. Mwapagha. "Characterization of Shiga toxin-producing Escherichia coli in raw beef from informal and commercial abattoirs." PLOS ONE 15, no. 12 (2020): e0243828. http://dx.doi.org/10.1371/journal.pone.0243828.
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Shiga toxin-producing Escherichia coli are foodborne pathogens that are mostly associated with beef products and have been implicated in human illness. E.coli-associated illness range from asymptomatic conditions of mild diarrhoea to haemorrhagic colitis which can progress into life threatening haemolytic uremic syndrome (HUS). Beef from cattle are regarded as the main reservoir of Shiga toxin-producing E. coli (STEC) pathogen. The aim of this study was to assess the level and sources of contamination of raw beef with STEC, and determine the incidences of STEC strains in raw beef from informal and commercial abattoirs in Windhoek, Namibia. A total of 204 raw beef samples, 37 equipment and 29 hand swabs were collected and tested for STEC. The meat samples were first enriched with pre-warmed buffered peptone water, cultured on Tryptone Bile X-Glucuronide and CHROMagar STEC, and then sub-cultured on nutrient agar. The presence of E.coli in the samples was confirmed by using VITEK 2 E.coli identification cards and PCR. The overall prevalence of STEC in the meat samples from both the abattoirs was 41.66% raw beef samples; 5.40% equipment swabs; and none of the hand swabs was STEC positive. From the STEC positive meat samples 29.41% contained one of the major STEC strains. Moreover, 52% of the 25 samples that contained the major STECs were characterised by eae and stx1, 8% characterised by eae and stx2 while 40% were characterised by eae, stx1 and stx2 virulence genes. This study has revealed the necessity for proper training on meat safety (for meat handlers) as well as the development, implementation and maintenance of effective sanitary dressing procedures at abattoirs to eliminate beef contamination by STECs thereby ensuring the production of wholesome meat, and to prevent the occurrences of STEC infections.
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Brooks, Dane, Benjamin Bastin, Erin Crowley, et al. "Modification and Matrix Extension of the Bio-Rad iQ-Check E. coli O157:H7, STEC VirX, and STEC SerO Test Kits for the Detection of Shiga Toxin–Producing Escherichia coli (STEC) and Escherichia coli O157 From a Single Enrichment." Journal of AOAC INTERNATIONAL 103, no. 1 (2020): 161–75. http://dx.doi.org/10.5740/jaoacint.19-0254.
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Abstract Background: The iQ-Check Real-Time PCR kits use PCR technology based on gene amplification and detection by a real-time PCR thermalcycler for the detection of target analytes in select food matrices. The iQ-Check E. coli O157:H7 [Performance Tested MethodSM (PTM) 020801] and STEC VirX and STEC SerO (combined PTM 121203) methods were previously validated for different matrices under different enrichment schemes. Objective: To modify the current iQ-Check E. coli O157:H7 Kit for the detection of Escherichia coli O157:H7 from 25 to 375 g for raw ground beef (17% fat), raw beef trim, and fresh spinach. In addition, a matrix extension was validated for iQ-Check E. coli O157:H7 for raw chicken breast without skin (25 g), raw chicken thigh with skin (25 g), mechanically separated chicken (25 g), and raw ground pork (25 g). The study also included the modification of the iQ-Check STEC VirX and SerO Kits for the detection of non-O157 Shiga toxin–producing E. coli (STEC) for raw ground beef (375 g), raw beef trim (375 g), and fresh spinach (375 g) from STEC Enrichment Broth to buffered peptone water (BPW). All tests were carried out at 8–22 h (10–22 h for fresh spinach). Methods: Ground beef, beef trim, and spinach were co-inoculated with E. coli O157:H7, non-O157 STECs, and Salmonella spp. and analyzed for E. coli O157:H7 and non-O157 STECs after an 8-22 h enrichment in BPW for the beef matrices and after a 10–22 h enrichment in BPW for spinach. The chicken matrices were inoculated with E. coli O157:H7 only and analyzed after an 8–22 h enrichment in BPW. The iQ-Check Free DNA Removal Solution workflow was utilized for all matrices. Confirmations at the 22 h time point and method comparisons were conducted with the appropriate reference method as outlined in the U.S. Food and Drug Administration Bacteriological Analytical Manual Chapter 4A or the U.S. Department of Agriculture Food Safety and Inspection Service Microbiology Laboratory Guidebook Chapters 5.09 and 5B.05. For the iQ-Check STEC VirX and STEC SerO Kits, inclusivity and exclusivity were also performed. Results: The two inclusivity and exclusivity evaluations indicated that the test methods can accurately detect the target analytes and correctly excluded nontarget organisms after 8 h of enrichment. In the method comparison study, the iQ-Check E. coli O157:H7 and STEC VirX and STEC SerO test kits demonstrated no statistically significant differences between candidate and reference method results or between presumptive and confirmed results for all food matrices analyzed and the two time points (8 or 10 and 22 h). Both time points produced the same results, with no discrepancies. Conclusions: The iQ-Check real-time PCR kits are effective methods for the detection of E. coli O157 and non-O157 STECs (both the virulence factors and the O groups) from raw ground beef, raw beef trim, and fresh spinach in 375 g samples enriched in BPW for 8–22 h (10–22 h for fresh spinach). In addition, the iQ-Check E. coli O157 Kit is effective in detecting E. coli O157 in 25 g samples of raw chicken breast without skin, raw chicken thigh with skin, mechanically separated chicken, and raw ground pork. The iQ-Check test kits allow the end user to pair enrichments for multiple target analytes, allowing the user to prepare a single enrichment and perform a single DNA extraction. The Free DNA Removal Solution removes free DNA from samples prior to PCR analysis, protecting DNA from intact and living cells. Highlights: The method modifications were granted based on the data collected.
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Babolhavaeji, Kiandokht, Leili Shokoohizadeh, Morteza Yavari, Abbas Moradi, and Mohammad Yousef Alikhani. "Prevalence of Shiga Toxin-Producing Escherichia coli O157 and Non-O157 Serogroups Isolated from Fresh Raw Beef Meat Samples in an Industrial Slaughterhouse." International Journal of Microbiology 2021 (December 15, 2021): 1–6. http://dx.doi.org/10.1155/2021/1978952.
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Background. The aims of the current study are the identification of O157 and non-O157 Shiga Toxin-Producing Escherichia coli (STEC) serogroups isolated from fresh raw beef meat samples in an industrial slaughterhouse, determination of antimicrobial resistance patterns, and genetic linkage of STEC isolates. Materials and Methods. A total of 110 beef samples were collected from the depth of the rump of cattle slaughtered at Hamadan industrial slaughterhouse. After detection of E. coli isolates, STEC strains were identified according to PCR for stx1, stx2, eaeA, and hlyA virulence genes, and STEC serogroups (O157 and non-O157) were identified by PCR. The genetic linkage of STEC isolates was analyzed by the ERIC- (Enterobacterial Repetitive Intergenic Consensus-) PCR method. The antimicrobial susceptibility of STEC isolates was detected by the disk diffusion method according to CLSI guidelines. Results. Among 110 collected beef samples, 77 (70%) were positive for E. coli. The prevalence of STEC in E. coli isolates was 8 (10.4%). The overall prevalence of O157 and non-O157 STEC isolates was 12.5% (one isolate) and 87.5% (7 isolates), respectively. The hemolysin gene was detected in 25% (2 isolates) of STEC strains. Evaluation of antibiotic resistance indicated that 100% of STEC isolates were resistant to ampicillin, ampicillin-sulbactam, amoxicillin-clavulanic acid, and cefazolin. Resistance to tetracycline and ciprofloxacin was detected in 62.5% and 12.5% of isolates, respectively. The analysis of the ERIC-PCR results showed five different ERIC types among the STEC isolates. Conclusion. The isolation of different clones STECs from beef and the presence of antibiotic-resistant isolates indicate that more attention should be paid to the hygiene of slaughterhouses.
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Yang, Xi, Yannong Wu, Qian Liu, et al. "Genomic Characteristics of Stx2e-Producing Escherichia coli Strains Derived from Humans, Animals, and Meats." Pathogens 10, no. 12 (2021): 1551. http://dx.doi.org/10.3390/pathogens10121551.
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Shiga toxin (Stx) can be classified into two types, Stx1 and Stx2, and different subtypes. Stx2e is a subtype commonly causing porcine edema disease and rarely reported in humans. The purpose of this study was to analyze the prevalence and genetic characteristics of Stx2e-producing Escherichia coli (Stx2e-STEC) strains from humans compared to strains from animals and meats in China. Stx2e-STEC strains were screened from our STEC collection, and whole-genome sequencing was performed to characterize their genetic features. Our study showed a wide distribution of Stx2e-STEC among diverse hosts and a higher proportion of Stx2e-STEC among human STEC strains in China. Three human Stx2e-STEC isolates belonged to O100:H30, Onovel26:H30, and O8:H9 serotypes and varied in genetic features. Human Stx2e-STECs phylogenetically clustered with animal- and food-derived strains. Stx2e-STEC strains from animals and meat showed multidrug resistance, while human strains were only resistant to azithromycin and tetracycline. Of note, a high proportion (55.9%) of Stx2e-STEC strains, including one human strain, carried the heat-stable and heat-labile enterotoxin-encoding genes st and lt, exhibiting a STEC/enterotoxigenic E. coli (ETEC) hybrid pathotype. Given that no distinct genetic feature was found in Stx2e-STEC strains from different sources, animal- and food-derived strains may pose the risk of causing human disease.
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Maguire, Meghan, Julie A. Kase, Dwayne Roberson, et al. "Precision long-read metagenomics sequencing for food safety by detection and assembly of Shiga toxin-producing Escherichia coli in irrigation water." PLOS ONE 16, no. 1 (2021): e0245172. http://dx.doi.org/10.1371/journal.pone.0245172.
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Shiga toxin-producing Escherichia coli (STEC) contamination of agricultural water might be an important factor to recent foodborne illness and outbreaks involving leafy greens. Closed bacterial genomes from whole genome sequencing play an important role in source tracking. We aimed to determine the limits of detection and classification of STECs by qPCR and nanopore sequencing using 24 hour enriched irrigation water artificially contaminated with E. coli O157:H7 (EDL933). We determined the limit of STEC detection by qPCR to be 30 CFU/reaction, which is equivalent to 105 CFU/ml in the enrichment. By using Oxford Nanopore’s EPI2ME WIMP workflow and de novo assembly with Flye followed by taxon classification with a k-mer analysis software (Kraken2), E. coli O157:H7 could be detected at 103 CFU/ml (68 reads) and a complete fragmented E. coli O157:H7 metagenome-assembled genome (MAG) was obtained at 105−108 CFU/ml. Using a custom script to extract the E. coli reads, a completely closed MAG was obtained at 107−108 CFU/ml and a complete, fragmented MAG was obtained at 105−106 CFU/ml. In silico virulence detection for E. coli MAGs for 105−108 CFU/ml showed that the virulotype was indistinguishable from the spiked E. coli O157:H7 strain. We further identified the bacterial species in the un-spiked enrichment, including antimicrobial resistance genes, which could have important implications to food safety. We propose this workflow provides proof of concept for faster detection and complete genomic characterization of STECs from a complex microbial sample compared to current reporting protocols and could be applied to determine the limit of detection and assembly of other foodborne bacterial pathogens.
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Cloke, Jonathan, Sharon Matheny, Michelle Swimley, et al. "Validation of the Applied Biosystems RapidFinder Shiga Toxin–Producing E. coli (STEC) Detection Workflow." Journal of AOAC INTERNATIONAL 99, no. 6 (2016): 1537–54. http://dx.doi.org/10.5740/jaoacint.16-0235.
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Abstract The Applied Biosystems™ RapidFinder™ STEC Detection Workflow (Thermo Fisher Scientific) is a complete protocol for the rapid qualitative detection of Escherichia coli (E. coli) O157:H7 and the “Big 6” non-O157 Shiga-like toxin-producing E. coli (STEC) serotypes (defined as serogroups: O26, O45, O103, O111, O121, and O145). The RapidFinder STEC Detection Workflow makes use of either the automated preparation of PCR-ready DNA using the Applied Biosystems PrepSEQ™ Nucleic Acid Extraction Kit in conjunction with the Applied Biosystems MagMAX™ Express 96-well magnetic particle processor or the Applied Biosystems PrepSEQ Rapid Spin kit for manual preparation of PCR-ready DNA. Two separate assays comprise the RapidFinder STEC Detection Workflow, the Applied Biosystems RapidFinder STEC Screening Assay and the Applied Biosystems RapidFinder STEC Confirmation Assay. The RapidFinder STEC Screening Assay includes primers and probes to detect the presence of stx1 (Shiga toxin 1), stx2 (Shiga toxin 2), eae (intimin), and E. coli O157 gene targets. The RapidFinder STEC Confirmation Assay includes primers and probes for the “Big 6” non-O157 STEC and E. coli O157:H7. The use of these two assays in tandem allows a user to detect accurately the presence of the “Big 6” STECs and E. coli O157:H7. The performance of the RapidFinder STEC Detection Workflow was evaluated in a method comparison study, in inclusivity and exclusivity studies, and in a robustness evaluation. The assays were compared to the U.S. Department of Agriculture (USDA), Food Safety and Inspection Service (FSIS) Microbiology Laboratory Guidebook (MLG) 5.09: Detection, Isolation and Identification of Escherichia coli O157:H7 from Meat Products and Carcass and Environmental Sponges for raw ground beef (73% lean) and USDA/FSIS-MLG 5B.05: Detection, Isolation and Identification of Escherichia coli non-O157:H7 from Meat Products and Carcass and Environmental Sponges for raw beef trim. No statistically significant differences were observed between the reference method and the individual or combined kits forming the candidate assay using either of the DNA preparation kits (manual or automated extraction). For the inclusivity and exclusivity evaluation, the RapidFinder STEC Detection Workflow, comprising both RapidFinder STEC screening and confirmation kits, correctly identified all 50 target organism isolates and correctly excluded all 30 nontarget strains for both of the assays evaluated. The results of these studies demonstrate the sensitivity and selectivity of the RapidFinder STEC Detection Workflow for the detection of E. coli O157:H7 and the “Big 6” STEC serotypes in both raw ground beef and beef trim. The robustness testing demonstrated that minor variations in the method parameters did not impact the accuracy of the assay and highlighted the importance of following the correct incubation temperatures
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TRIPLETT, ODBERT A., JIEKUN XUAN, STEVEN FOLEY, RAJESH NAYAK, and WILLIAM H. TOLLESON. "Immunomagnetic Capture of Big Six Shiga Toxin–Producing Escherichia coli Strains in Apple Juice with Detection by Multiplex Real-Time PCR Eliminates Interference from the Food Matrix." Journal of Food Protection 82, no. 9 (2019): 1512–23. http://dx.doi.org/10.4315/0362-028x.jfp-19-134.
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ABSTRACT Having reliable methods for detecting Shiga toxin–producing Escherichia coli (STEC) in foods is an important food safety goal. The majority of STEC outbreaks have involved either the O157:H7 serotype or one of six non-O157 serogroups, O26, O45, O103, O111, O121, and O145, termed “The Big Six.” We have compared detection by PCR of the Shiga toxin genes stx1a and stx2a from STEC bacteria isolated from unclarified apple juice by simple centrifugation with the use of an immunocapture technique to minimize contaminants (such as pectin and polyphenols that may copurify with DNA) that may interfere with DNA amplification efficiencies and limit sensitivity. An internal control for successful immunocapture, DNA extraction, and PCR amplification was generated by introducing the pmRaspberry plasmid into an stx null strain, yielding an E. coli O45 pmRaspberry derivative that can be added to food samples directly. Using serial dilutions of a representative Big Six STEC in apple juice, our immunocapture method resulted in a 50% probability of detection value of 3.34, 2.25, and 4.25 CFU for detection by multiplex real-time PCR, growth on solid agar, and multiplex endpoint PCR, respectively. The time to result was 6.5 h, 9.5 h, and 1.5 days for immunocapture of Big Six STECs and detection by multiplex real-time PCR, endpoint PCR, and growth on solid agar, respectively. A set of 52 Big Six STEC isolates and 30 non–Big Six STEC strains was used to establish the inclusivity and exclusivity of the method. Finally, the ability to detect Big Six STEC contamination reliably was confirmed at 4.5 and 45 CFU/25-mL portions of refrigerated apple juice.
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MEDINA, MARJORIE B., WEILIN L. SHELVER, PINA M. FRATAMICO, et al. "Latex Agglutination Assays for Detection of Non-O157 Shiga Toxin–Producing Escherichia coli Serogroups O26, O45, O103, O111, O121, and O145†." Journal of Food Protection 75, no. 5 (2012): 819–26. http://dx.doi.org/10.4315/0362-028x.jfp-11-430.
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Latex agglutination assays utilizing polyclonal antibodies were developed for the top six non-O157 Shiga toxin–producing Escherichia coli (STEC) serogroups. Rabbit antisera were affinity purified through protein A/G columns, and the isolated immunoglobulins (IgGs) were covalently immobilized onto polystyrene latex particles. The resulting latex-IgG complex had a protein (IgG) load of 0.20 to 0.28 mg/ml in a 1% latex suspension. Optimum conditions for the agglutination assay consisted of utilizing 20 μl of latex-IgG reagent containing 2.0 to 2.8 μg IgG in a 0.5% latex suspension. Agglutination or flocculation was observed almost instantly after mixing the colonies with the latex-IgG, indicating STEC strains. More than 100 target and nontarget strains were tested in more than 3,000 test replicates. All target organisms produced positive results, but three antisera (anti-O26, anti-O103, and anti-O145) cross-reacted with some other STECs. The anti-O103 and anti-O145 latex reagents cross-reacted with O26 strains, and the anti-O26 cross-reacted with O103 strains. The latex-IgG reagents are stable for at least 1 year and are easy to prepare. These agglutination assays can be used for identification of presumptive non-O157 STEC colonies from agar media. The techniques used to prepare the latex reagents also can be utilized for testing other STEC serogroups, other E. coli serotypes, or other pathogens to ensure safe foods to consumers.
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Baiko, S. V. "Epidemiology and pathophysiology of hemolytic uremic syndrome associated with shiga toxin (literature review)." Nephrology (Saint-Petersburg) 25, no. 3 (2021): 36–42. http://dx.doi.org/10.36485/1561-6274-2021-25-3-36-42.
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Hemolytic uremic syndrome (HUS) associated with shiga toxin E. coli(STEC) is one of the most common causes of acute kidney injury in young children. The share of STEC-HUS among all HUS variants is up to 90%. Not all STECs are pathogenic to humans, and those that cause disease (hemorrhagic colitis, HUS) are referred to as enterohemorrhagic E. coli(EHEC). The main pathogens causing STEC-HUS include the serotype E. coliO157: H7, less often serotypes O26, O80, O103, O121, O145. EHEC exist as normal microbiota in cattle, but can also be found in goats, sheep, pigs, chickens, dogs, and rats. Infection can occur when using undercooked ground beef, unpasteurized milk, water, including tap water and from open ponds and pools, from an infected person and when visiting farms and zoos. The epidemiological history should be carefully assessed in each patient with HUS, taking into account the annual outbreaks of this disease in different regions of the world. In recent years actively discussed the issue of the transfer of shiga toxin (Stx) from the intestine to the blood and from the blood to target organs in the form of microvesicles, the wall of which is the outer shell of E.coliand blood cells. This allows Stx to escape the response of the human immune system. The article describes in detail the mechanisms of infection and expression of pathogenic genes of EHEC, the effect of Stx on endothelial cells, on expression of adhesion molecules and inflammatory chemokines, activation of the alternative complement pathway, which determine the development of HUS.
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Alkaby, SabahM Mohammed, and Mohammed Jebur. "Genetic Characterization of Escherichia coli O157 Isolated From Human Stool Specimens in Wassit Province of Iraq." Journal of Techniques 2, no. 3 (2020): 1–8. http://dx.doi.org/10.51173/jt.v2i3.206.
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The goal of this study was to assess the severity of diarrhea caused by Shiga toxins produced by Escherichia coli ( STEC) isolated from human infections in Wassit province of Iraq. Stool specimens from (161) sporadic cases of diarrheal Patients with the mean age of (20 years), and range from 1 month to 75 years, were collected in AL-Karamma Teaching Hospital between March to July 2019. Then they were processed by culture, microscopic tests and VITEK which were used for the identification as E. coli. PCR was performed for detecting shiga toxin genes in E. coli isolates, and rfb gene (encoding O-antigene ) of STEC- O157. DNA sequencing was done on some positive isolates. The results of PCR detected stx genes in 19 (12 %) culture isolates of E. coli isolated from human diarrheal specimens. While 9 cases were positive for stx genes and have rfb gene. DNA sequences that depend on the sequence of the vtx2 gene have shown a highly homologous sequencing identity with NCBI-Blast Escherichia coli strain( O157:H7) isolates from humans and animals. The phylogenetic study revealed a clear genetic relationship between human and animal E. coli isolates and then gene sequencing was deposited with accession number into NCBI-Genbank (0MN944014.1). In conclusion, prevalence of E. coli O157 in humans remains undiagnosed because there are no traditional O157 detection methods in all our hospitals. Our study showed that the use of molecular methods in the detection can be used to detect STECs which cannot be detected using routine methods.
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DENG, KAIPING, XUE WANG, LI-HAN YEN, HONGLIU DING, and MARY LOU TORTORELLO. "Behavior of Shiga Toxigenic Escherichia coli Relevant to Lettuce Washing Processes and Consideration of Factors for Evaluating Washing Process Surrogates." Journal of Food Protection 77, no. 11 (2014): 1860–67. http://dx.doi.org/10.4315/0362-028x.jfp-14-220.
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Postharvest processes for fresh produce commonly include washing in water containing antimicrobial chemicals, such as chlorine; however, if the antimicrobials are not present in sufficient levels, washing can promote the spread of contamination that might be present. To understand cross-contamination risk during washing, we tested a collection of Shiga toxigenic Escherichia coli (STEC), including O157:H7 and other non-O157 strains, for certain traits during washing of fresh-cut lettuce, i.e., sensitivity to sublethal chlorine levels and ability to cross-contaminate (detach from and attach to) lettuce in the presence of sublethal chlorine levels. Nonpathogenic E. coli Nissle 1917 (EcN) and Pediococcus pentosaceus lactic acid bacterial species (LAB) were included as potential washing process validation surrogates. As measured by extension of the lag phase of growth in media containing 0.15 ppm of chlorine, chlorine sensitivity varied among the STECs. Cross-contamination was assessed by evaluating transfer of bacteria from inoculated to uninoculated leaves during washing. Without chlorine, similar transfer to wash water and uninoculated leaves was shown. In 1 ppm of chlorine, cross-contamination was not detected with most strains, except for the substantial transfer by a STEC O111 strain and EcN in some replicates. Strain O111 and EcN showed less inactivation in 0.25 ppm of chlorine water compared with O157 (P &lt; 0.05). LAB showed similar transfer and similar chlorine inactivation to O157. Considering together the sublethal chlorine sensitivity and detachment/attachment traits, neither EcN nor LAB displayed optimal characteristics as washing process surrogates for the STEC strains, although further evaluation is needed. This work demonstrated a range of behaviors of STEC strains during lettuce washing and may be helpful in hazard characterization, identifying factors to consider for evaluating washing process efficacy, and identifying phenotypic traits to select surrogates to validate washing processes.
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da Silva, Caroline Rodrigues, Matheus Silva Sanches, Kawana Hiromori Macedo, et al. "Molecular and phenotypic characterization of diarrheagenic Escherichia coli isolated from groundwater in rural areas in southern Brazil." Journal of Water and Health 17, no. 4 (2019): 597–608. http://dx.doi.org/10.2166/wh.2019.142.
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Abstract Water-borne diseases like diarrheagenic Escherichia coli (DEC)-induced gastroenteritis are major public health problems in developing countries. In this study, the microbiological quality of water from mines and shallow wells was analyzed for human consumption. Genotypic and phenotypic characterization of DEC strains was performed. A total of 210 water samples was analyzed, of which 153 (72.9%) contained total coliforms and 96 (45.7%) E. coli. Of the E. coli isolates, 27 (28.1%) contained DEC genes. The DEC isolates included 48.1% Shiga toxin-producing E. coli (STEC), 29.6% enteroaggregative E. coli (EAEC), 14.9% enteropathogenic E. coli (EPEC), 3.7% enterotoxigenic E. coli (ETEC), and 3.7% enteroinvasive E. coli (EIEC). All the STECs had cytotoxic effects on Vero cells and 14.8% of the DEC isolates were resistant to at least one of the antibiotics tested. All DEC formed biofilms and 92.6% adhered to HEp-2 cells with a prevalence of aggregative adhesion (74%). We identified 25 different serotypes. One EPEC isolate was serotype O44037:H7, reported for the first time in Brazil. Phylogenetically, 63% of the strains belonged to group B1. The analyzed waters were potential reservoirs for DEC and could act as a source for infection of humans. Preventive measures are needed to avoid such contamination.
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Jaudou, Sandra, Carlus Deneke, Mai-Lan Tran, et al. "Exploring Long-Read Metagenomics for Full Characterization of Shiga Toxin-Producing Escherichia coli in Presence of Commensal E. coli." Microorganisms 11, no. 8 (2023): 2043. http://dx.doi.org/10.3390/microorganisms11082043.
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The characterization of Shiga toxin-producing Escherichia coli (STEC) is necessary to assess their pathogenic potential, but isolation of the strain from complex matrices such as milk remains challenging. In previous work, we have shown the potential of long-read metagenomics to characterize eae-positive STEC from artificially contaminated raw milk without isolating the strain. The presence of multiple E. coli strains in the sample was shown to potentially hinder the correct characterization of the STEC strain. Here, we aimed at determining the STEC:commensal ratio that would prevent the characterization of the STEC. We artificially contaminated pasteurized milk with different ratios of an eae-positive STEC and a commensal E. coli and applied the method previously developed. Results showed that the STEC strain growth was better than the commensal E. coli after enrichment in acriflavine-supplemented BPW. The STEC was successfully characterized in all samples with at least 10 times more STEC post-enrichment compared to the commensal E. coli. However, the presence of equivalent proportions of STEC and commensal E. coli prevented the full characterization of the STEC strain. This study confirms the potential of long-read metagenomics for STEC characterization in an isolation-free manner while refining its limit regarding the presence of background E. coli strains.
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Dean-Nystrom, Evelyn A., William C. Stoffregen, Brad T. Bosworth, Harley W. Moon, and Joachim F. Pohlenz. "Early Attachment Sites for Shiga-Toxigenic Escherichia coli O157:H7 in Experimentally Inoculated Weaned Calves." Applied and Environmental Microbiology 74, no. 20 (2008): 6378–84. http://dx.doi.org/10.1128/aem.00636-08.
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ABSTRACT Weaned 3- to 4-month-old calves were fasted for 48 h, inoculated with 1010 CFU of Shiga toxin-positive Escherichia coli (STEC) O157:H7 strain 86-24 (STEC O157) or STEC O91:H21 strain B2F1 (STEC O91), Shiga toxin-negative E. coli O157:H7 strain 87-23 (Stx− O157), or a nonpathogenic control E. coli strain, necropsied 4 days postinoculation, and examined bacteriologically and histologically. Some calves were treated with dexamethasone (DEX) for 5 days (3 days before, on the day of, and 1 day after inoculation). STEC O157 bacteria were recovered from feces, intestines, or gall bladders of 74% (40/55) of calves 4 days after they were inoculated with STEC O157. Colon and cecum were sites from which inoculum-type bacteria were most often recovered. Histologic lesions of attaching-and-effacing (A/E) O157+ bacteria were observed in 69% (38/55) of the STEC O157-inoculated calves. Rectum, ileocecal valve, and distal colon were sites most likely to contain A/E O157+ bacteria. Fecal and intestinal levels of STEC O157 bacteria were significantly higher and A/E O157+ bacteria were more common in DEX-treated calves than in nontreated calves inoculated with STEC O157. Fecal STEC O157 levels were significantly higher than Stx− O157, STEC O91, or control E. coli; only STEC O157 cells were recovered from tissues. Identifying the rectum, ileocecal valve, and distal colon as early STEC O157 colonization sites and finding that DEX treatment enhances the susceptibility of weaned calves to STEC O157 colonization will facilitate the identification and evaluation of interventions aimed at reducing STEC O157 infection in cattle.
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Zhang, Yujie, Yen-Te Liao, Xiaohong Sun, and Vivian C. H. Wu. "Is Shiga Toxin-Producing Escherichia coli O45 No Longer a Food Safety Threat? The Danger is Still Out There." Microorganisms 8, no. 5 (2020): 782. http://dx.doi.org/10.3390/microorganisms8050782.
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Many Shiga toxin-producing Escherichia coli (STEC) strains, including the serogroups of O157 and most of the top six non-O157 serotypes, are frequently associated with foodborne outbreaks. Therefore, they have been extensively studied using next-generation sequencing technology. However, related information regarding STEC O45 strains is scarce. In this study, three environmental E. coli O45:H16 strains (RM11911, RM13745, and RM13752) and one clinical E. coli O45:H2 strain (SJ7) were sequenced and used to characterize virulence factors using two reference E. coli O45:H2 strains of clinical origin. Subsequently, whole-genome-based phylogenetic analysis was conducted for the six STEC O45 strains and nine other reference STEC genomes, in order to evaluate their evolutionary relationship. The results show that one locus of enterocyte effacement pathogenicity island was found in all three STEC O45:H2 strains, but not in the STEC O45:H16 strains. Additionally, E. coli O45:H2 strains were evolutionarily close to E. coli O103:H2 strains, sharing high homology in terms of virulence factors, such as Stx prophages, but were distinct from E. coli O45:H16 strains. The findings show that E. coli O45:H2 may be as virulent as E. coli O103:H2, which is frequently associated with severe illness and can provide genomic evidence to facilitate STEC surveillance.
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Hosking, Edan, Brooke Roman, Susan Alles, et al. "NeoSeekTM STEC: A Multiplex Molecular Method for Detection and Identification of Select Shiga Toxin–Producing Escherichia coli in Beef." Journal of AOAC INTERNATIONAL 103, no. 2 (2020): 523–32. http://dx.doi.org/10.5740/jaoacint.19-0300.
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Abstract Background: NeoSeekTM STEC is a single-source, service-based method for detection and identification of select Shiga toxin–producing Escherichia coli (STEC), including E. coli O157:H7 and STEC of somatic groups O26, O45, O103, O111, O121, and O145. The method is a multiplex molecular method utilizing more than 80 genetic targets to identify STEC in complex matrices such as food enrichment cultures. Objective: A study was conducted to validate the NeoSeek method for detection of select STEC in raw beef trim. Methods: Performance of the NeoSeek STEC method was compared with that of the U.S. Department of Agriculture, Food Safety and Inspection Service reference methods for E. coli O157:H7 and non-O157 STEC for detection of E. coli O157:H7 and E. coli O26:H11 in raw beef trim. Additionally, inclusivity/exclusivity testing and method robustness testing were performed. Results: Results of raw beef trim testing showed no statistically significant differences in performance between the NeoSeek and reference methods in the ability to detect either E. coli O157:H7 or E.coli O26:H11, as determined by probability of detection analysis. Results of inclusivity and exclusivity testing showed 100% expected results with target and nontarget bacteria, with the exception of a single strain of E. coli O157:H7, which was subsequently verified to be stx-negative by PCR. Conclusions and Highlights: NeoSeek STEC is an accurate, reliable method for rapid detection and identification of select STEC in complex populations such as beef trim enrichment cultures.
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Barlow, Robert S., and Glen E. Mellor. "Foodborne pathogenic E. coli (focus on STEC)." Microbiology Australia 34, no. 2 (2013): 80. http://dx.doi.org/10.1071/ma13028.
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Lima, Paulo Gomes de, Thalita Martins da Silva, Luciana Maria Ramires Esper, Alice Gonçalves Martins Gonzalez, and Robson Maia Franco. "Viabilidade de Escherichia coli O153:H25, O113:H21 e O111:H8 (STEC não-O157) produtoras de toxina Shiga em queijo minas frescal." Ciência Rural 45, no. 1 (2015): 52–57. http://dx.doi.org/10.1590/0103-8478cr20131678.
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A existência de um reservatório animal é de grande importância na transmissão de Escherichia coli, produtora de toxina shiga (STEC) aos humanos. Epidemiologicamente, o sorotipo O157:H7 tem sido o mais envolvido em surtos de doença humana causada por STEC, porém surtos envolvendo STEC não pertencentes ao sorogrupo O157 (STEC não-O157) têm sido descritos. Inúmeros trabalhos constatam uma elevada ocorrência destes microrganismos em fezes de bovinos no Brasil, entretanto, pouco se sabe sobre a transmissão destes aos produtos de origem animal em nosso país. Neste trabalho, foi avaliada a viabilidade de E.coli O153:H25; O113:H21 e O111:H8 em Queijo Minas Frescal (QMF), produzido com inóculos de STEC não O157: H7 e armazenados a 8ºC. Realizaram-se contagens de E. coli e psicrotróficos totais após o processamento do queijo e com intervalos de sete e quinze dias. Foi observado aumento nas contagens de E. coli STEC não O157: H7 e psicrotróficos totais logo após o processamento do QMF, bem como durante o armazenamento a 8ºC, temperatura máxima recomendada pela legislação brasileira. Demonstra-se que, caso haja contaminação da matéria-prima com STEC não O157: H7 (deste estudo), o processamento do QMF não elimina os microrganismos e a temperatura máxima recomendada pela legislação não inibe a multiplicação bacteriana, mantendo-se o risco à população. Reforça-se, portanto, a atenção à qualidade da matéria-prima, das ferramentas de qualidade no campo e na indústria de alimentos para garantir a inocuidade do produto final
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THRAN, B. H., H. S. HUSSEIN, M. R. HALL, and S. F. KHAIBOULLINA. "Shiga Toxin–Producing Escherichia coli in Beef Heifers Grazing an Irrigated Pasture." Journal of Food Protection 64, no. 10 (2001): 1613–16. http://dx.doi.org/10.4315/0362-028x-64.10.1613.
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Shiga toxin–producing Escherichia coli (STEC) produce toxins that have been associated with several human illnesses. E. coli O157:H7 is the most well-studied STEC and was first associated with consumption of improperly cooked ground beef in 1982. E. coli O157:H7 is not the only foodborne STEC because other STEC serotypes are also associated with human illnesses. The objective of this study was to assess prevalence of STEC in 23 yearling beef (Angus) heifers grazing an irrigated grass pasture in spring (April), summer (July), fall (October), and winter (December) of 1999. A total of 86 fecal samples were rectally collected and were subjected to microbiological testing for the presence of STEC. Nine E. coli isolates from five heifers (one in spring and fall and three in winter) were toxic to Vero cells. Of these isolates, four were E. coli O157:H7, two belonged to the serogroup O6, one O39:NM, one O113:H−, and the final isolate was untypable. The STEC prevalence rate in our herd ranged from 4% (spring) to 15% (winter). Based on detecting both O157:H7 and non-O157:H7 STEC in our heifers, it is clear that screening fecal samples should not be limited to E. coli O157:H7. Identification of STEC-positive cattle prior to slaughter should help in reducing the risk of beef contamination with such foodborne pathogens if pre- and/or postharvest control measures are applied to such animals.
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Paquette, Sarah-Jo, Rahat Zaheer, Kim Stanford, James Thomas, and Tim Reuter. "Competition among Escherichia coli Strains for Space and Resources." Veterinary Sciences 5, no. 4 (2018): 93. http://dx.doi.org/10.3390/vetsci5040093.
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Shiga toxin-producing Escherichia coli (STEC) are a subgroup of E. coli causing human diseases. Methods to control STEC in livestock and humans are limited. These and other emerging pathogens are a global concern and novel mitigation strategies are required. Habitats populated by bacteria are subjected to competition pressures due to limited space and resources but they use various strategies to compete in natural environments. Our objective was to evaluate non-pathogenic E. coli strains isolated from cattle feces for their ability to out-compete STEC. Competitive fitness of non-pathogenic E. coli against STEC were assessed in competitions using liquid, agar, and nutrient limiting assays. Winners were determined by enumeration using O-serogroup specific quantitative PCR or a semi-quantitative grading. Initial liquid competitions identified two strong non-pathogenic competitors (O103F and O26E) capable of eliminating various STEC including O157 and O111. The strain O103F was dominant across permeable physical barriers for all tested E. coli and STEC strains indicating the diffusion of antimicrobial molecules. In direct contact and even with temporal disadvantages, O103F out-competed STEC O157E. The results suggest that O103F or the diffusible molecule(s) it produces have a potential to be used as an alternative STEC mitigation strategy, either in medicine or the food industry.
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Hughes, Anna C., Yuzhu Zhang, Xiangning Bai, et al. "Structural and Functional Characterization of Stx2k, a New Subtype of Shiga Toxin 2." Microorganisms 8, no. 1 (2019): 4. http://dx.doi.org/10.3390/microorganisms8010004.
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Shiga toxin (Stx) is the major virulence factor of Shiga toxin-producing Escherichia coli (STEC). Stx evolves rapidly and, as such, new subtypes continue to emerge that challenge the efficacy of existing disease management and surveillance strategies. A new subtype, Stx2k, was recently identified in E. coli isolated from a wide range of sources including diarrheal patients, animals, and raw meats, and was poorly detected by existing immunoassays. In this study, the structure of Stx2kE167Q was determined at 2.29 Å resolution and the conservation of structure with Stx2a was revealed. A novel polyclonal antibody capable of neutralizing Stx2k and an immunoassay, with a 10-fold increase in sensitivity compared to assays using extant antibodies, were developed. Stx2k is less toxic than Stx2a in Vero cell assays but is similar to Stx2a in receptor-binding preference, thermostability, and acid tolerance. Although Stx2k does not appear to be as potent as Stx2a to Vero cells, the wide distribution and blended virulence profiles of the Stx2k-producing strains suggest that horizontal gene transfer through Stx2k-converting phages could result in the emergence of new and highly virulent pathogens. This study provides useful information and tools for early detection and control of Stx2k-producing E. coli, which could reduce public risk of infection by less-known STECs.
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Xia, Xiaodong, Jianghong Meng, Patrick F. McDermott, et al. "Presence and Characterization of Shiga Toxin-Producing Escherichia coli and Other Potentially Diarrheagenic E. coli Strains in Retail Meats." Applied and Environmental Microbiology 76, no. 6 (2010): 1709–17. http://dx.doi.org/10.1128/aem.01968-09.
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ABSTRACT To determine the presence of Shiga toxin-producing Escherichia coli (STEC) and other potentially diarrheagenic E. coli strains in retail meats, 7,258 E. coli isolates collected by the U.S. National Antimicrobial Resistance Monitoring System (NARMS) retail meat program from 2002 to 2007 were screened for Shiga toxin genes. In addition, 1,275 of the E. coli isolates recovered in 2006 were examined for virulence genes specific for other diarrheagenic E. coli strains. Seventeen isolates (16 from ground beef and 1 from a pork chop) were positive for stx genes, including 5 positive for both stx 1 and stx 2, 2 positive for stx 1, and 10 positive for stx 2. The 17 STEC strains belonged to 10 serotypes: O83:H8, O8:H16, O15:H16, O15:H17, O88:H38, ONT:H51, ONT:H2, ONT:H10, ONT:H7, and ONT:H46. None of the STEC isolates contained eae, whereas seven carried enterohemorrhagic E. coli (EHEC) hlyA. All except one STEC isolate exhibited toxic effects on Vero cells. DNA sequence analysis showed that the stx 2 genes from five STEC isolates encoded mucus-activatable Stx2d. Subtyping of the 17 STEC isolates by pulsed-field gel electrophoresis (PFGE) yielded 14 distinct restriction patterns. Among the 1,275 isolates from 2006, 11 atypical enteropathogenic E. coli (EPEC) isolates were identified in addition to 3 STEC isolates. This study demonstrated that retail meats, mainly ground beef, were contaminated with diverse STEC strains. The presence of atypical EPEC strains in retail meat is also of concern due to their potential to cause human infections.
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ACHESON, DAVID W. K. "How Does Escherichia coli O157:H7 Testing in Meat Compare with What We Are Seeing Clinically?" Journal of Food Protection 63, no. 6 (2000): 819–21. http://dx.doi.org/10.4315/0362-028x-63.6.819.
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Escherichia coli O157:H7 is but one of a group of Shiga toxin-producing E. coli (STEC) that cause both intestinal disease such as bloody and nonbloody diarrhea and serious complications like hemolytic uremic syndrome (HUS). While E. coli O157: H7 is the most renowned STEC, over 200 different types of STEC have been documented in meat and animals, at least 60 of which have been linked with human disease. A number of studies have suggested that non-O157 STEC are associated with clinical disease, and non-O157 STEC are present in the food supply. Non-O157 STEC, such as O111 have caused large outbreaks and HUS in the United States and other countries. The current policy in the United States is to examine ground beef for O157:H7 only, but restricting the focus to O157 will miss other important human STEC pathogens.
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Cornick, N. A., S. L. Booher, T. A. Casey, and H. W. Moon. "Persistent Colonization of Sheep byEscherichia coli O157:H7 and Other E. coliPathotypes." Applied and Environmental Microbiology 66, no. 11 (2000): 4926–34. http://dx.doi.org/10.1128/aem.66.11.4926-4934.2000.
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ABSTRACT Shiga toxin-producing Escherichia coli (STEC) is an important cause of food-borne illness in humans. Ruminants appear to be more frequently colonized by STEC than are other animals, but the reason(s) for this is unknown. We compared the frequency, magnitude, duration, and transmissibility of colonization of sheep by E. coli O157:H7 to that by other pathotypes of E. coli. Young adult sheep were simultaneously inoculated with a cocktail consisting of two strains of E. coli O157:H7, two strains of enterotoxigenic E. coli (ETEC), and one strain of enteropathogenic E. coli. Both STEC strains and ETEC 2041 were given at either 107 or 1010CFU/strain/animal. The other strains were given only at 1010 CFU/strain. We found no consistent differences among pathotypes in the frequency, magnitude, and transmissibility of colonization. However, the STEC strains tended to persist to 2 weeks and 2 months postinoculation more frequently than did the other pathotypes. The tendency for persistence of the STEC strains was apparent following an inoculation dose of either 107 or 1010 CFU. One of the ETEC strains also persisted when inoculated at 1010 CFU. However, in contrast to the STEC strains, it did not persist when inoculated at 107 CFU. These results support the hypothesis that STEC is better adapted to persist in the alimentary tracts of sheep than are other pathotypes ofE. coli.
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Banerjee, Aparna, and Surajit Acharyya. "Molecular characterization of STEC isolated from Ducks and its relation to ESBL production." Ukrainian Journal of Veterinary and Agricultural Sciences 3, no. 2 (2020): 24–29. http://dx.doi.org/10.32718/ujvas3-2.04.
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The ESBL producing genes are responsible for bacterial resistance to number of antibiotics whereas Shiga toxin producing genes are responsible for bacterial virulence. The association between ESBL producing genes and Shiga toxin producing E. coli (STEC) may pose bigger threat in the battle of antibiotic resistance. This study was conducted to determine the prevalence of Shiga-toxin-producing Escherichia coli (STEC) in ducks reared in organized and unorganized sectors from different agro climatic zones of West Bengal, India and to study their relationship with extended spectrum beta-lactamase (ESBL) production. Total 202 cloacal swab samples were collected from both indigenous ducks reared in backyards sector (110 samples) and Khaki Campbell Ducks reared in organized farm (92 samples). Initially the samples were screened for detection of E. coli on the basis of their cultural, morphological and biochemical properties followed by PCR confirmation for E. coli 16S rRNA. E. coli isolates were subjected to multiplex PCR to detect the presence of shiga toxin producing genes such as stx1, stx2, eaeA and ehxA. STEC isolates were screened phenotypically for production of ESBL and ACBL by double disk diffusion method and subsequently PCR detection for blaCTX-M, blaTEM, blaSHV and blaAmpC genes were performed. Serotyping of all the STEC was also done. Out of 202 samples total 109 were confirmed to be E. coli positive. Out of them total 27 (24.77 %) E. coli isolates were detected to be positive for STEC. Higher prevalence of STEC was observed in unorganized sector compared to the organized sector. Positive association (P < 0.05) was observed between STEC and ESBL production. This study indicates that the duck may play an important role in transmission of Siga-toxin-producing E. coli (STEC) as well as antibiotic resistance genes to human beings, other birds, animals and environment also.
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TERAO, YOSHITAKA, KANA TAKESHITA, YASUTAKA NISHIYAMA, NAOKI MORISHITA, TAKASHI MATSUMOTO, and FUMIKI MORIMATSU. "Promising Nucleic Acid Lateral Flow Assay Plus PCR for Shiga Toxin–Producing Escherichia coli." Journal of Food Protection 78, no. 8 (2015): 1560–68. http://dx.doi.org/10.4315/0362-028x.jfp-14-495.
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Shiga toxin (Stx)–producing Escherichia coli (STEC) is a frequent cause of foodborne infections, and methods for rapid and reliable detection of STEC are needed. A nucleic acid lateral flow assay (NALFA) plus PCR was evaluated for detecting STEC after enrichment. When cell suspensions of 45 STEC strains, 14 non-STEC strains, and 13 non–E. coli strains were tested with the NALFA plus PCR, all of the STEC strains yielded positive results, and all of the non-STEC and non–E. coli strains yielded negative results. The lower detection limit for the STEC strains ranged from 0.1 to 1 pg of genomic DNA (about 20 to 200 CFU) per test, and the NALFA plus PCR was able to detect Stx1- and Stx2-producing E. coli strains with similar sensitivities. The ability of the NALFA plus PCR to detect STEC in enrichment cultures of radish sprouts, tomato, raw ground beef, and beef liver inoculated with 10-fold serially diluted STEC cultures was comparable to that of a real-time PCR assay (at a level of 100 to 100,000 CFU/ml in enrichment culture). The bacterial inoculation test in raw ground beef revealed that the lower detection limit of the NALFA plus PCR was also comparable to that obtained with a real-time PCR assay that followed the U.S. Department of Agriculture guidelines. Although further evaluation is required, these results suggest that the NALFA plus PCR is a specific and sensitive method for detecting STEC in a food manufacturing plant.
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Trung, Nguyen Vinh, Hoang Ngoc Nhung, Juan J. Carrique-Mas, et al. "Colonization of Enteroaggregative Escherichia coli and Shiga toxin-producing Escherichia coli in chickens and humans in southern Vietnam." BMC Microbiology 16, no. 1 (2016): 208. https://doi.org/10.1186/s12866-016-0827-z.
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<strong>Background: </strong>Enteroaggregative (EAEC) and Shiga-toxin producing <i>Escherichia coli</i> (STEC) are a major cause of diarrhea worldwide. <i>E. coli</i> carrying both virulence factors characteristic for EAEC and STEC and producing extended-spectrum beta-lactamase caused severe and protracted disease during an outbreak of <i>E. coli</i> O104:H4 in Europe in 2011. We assessed the opportunities for <i>E. coli</i> carrying the <i>aggR</i> and <i>stx</i> genes to emerge in ‘backyard’ farms in south-east Asia.<strong>Results: </strong>Faecal samples collected from 204 chicken farms; 204 farmers and 306 age- and gender-matched individuals not exposed to poultry farming were plated on MacConkey agar plates with and without antimicrobials being supplemented. Sweep samples obtained from MacConkey agar plates without supplemented antimicrobials were screened by multiplex PCR for the detection of the <i>stx1</i>, <i>stx2</i> and <i>aggR</i> genes. One chicken farm sample each (0.5 %) contained the <i>stx1</i> and the <i>aggR</i> gene. Eleven (2.4 %) human faecal samples contained the <i>stx1</i> gene, 2 samples (0.4 %) contained <i>stx2</i> gene, and 31 (6.8 %) contained the <i>aggR</i> gene. From 46 PCR-positive samples, 205 <i>E. coli</i> isolates were tested for the presence of <i>stx1</i>, <i>stx2</i>, <i>aggR</i>, <i>wzx</i> <sub>O104</sub> and <i>fliC</i> <sub>H4</sub> genes. None of the isolates simultaneously contained the four genetic markers associated with <i>E. coli</i> O104:H4 epidemic strain (<i>aggR</i>, <i>stx2</i>, <i>wzx</i> <sub> <i>O104</i> </sub> and <i>fliC</i> <sub> <i>H4</i> </sub>). Of 34 EAEC, 64.7 % were resistant to 3<sup>rd</sup>-generation cephalosporins.<strong>Conclusion: </strong>These results indicate that in southern Vietnam, the human population is a more likely reservoir of <i>aggR</i> and <i>stx</i> gene carrying <i>E. coli</i> than the chicken population. However, conditions for transmission of isolates and/or genes between human and animal reservoirs resulting in the emergence of highly virulent <i>E. coli</i> strains are still favorable, given the nature of‘backyard’ farms in Vietnam.
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CERNICCHIARO, N., D. G. RENTER, C. A. CULL, Z. D. PADDOCK, X. SHI, and T. G. NAGARAJA. "Fecal Shedding of Non-O157 Serogroups of Shiga Toxin–Producing Escherichia coli in Feedlot Cattle Vaccinated with an Escherichia coli O157:H7 SRP Vaccine or Fed a Lactobacillus-Based Direct-Fed Microbial†." Journal of Food Protection 77, no. 5 (2014): 732–37. http://dx.doi.org/10.4315/0362-028x.jfp-13-358.
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The objectives of this study were to determine whether fecal shedding of non-O157 Shiga toxin–producing Escherichia coli (STEC) in feedlot cattle was affected by the use of an E. coli O157:H7 vaccine or a direct-fed microbial (DFM) and whether the shedding of a particular non-O157 STEC serogroup within feces was associated with shedding of O157 or other non-O157 STEC serogroups. A total of 17,148 cattle in 40 pens were randomized to receive one, both, or neither (control) of the two interventions: a vaccine based on the siderophore receptor and porin proteins (E. coli SRP vaccine, two doses) and a DFM product (low-dose Bovamine). Fresh fecal samples (30 samples per pen) were collected weekly from pen floors for four consecutive weeks beginning approximately 56 days after study allocation. DNA extracted from enriched samples was tested for STEC O157 and non-O157 serogroups O26, O45, O103, O111, O121, and O145 and for four major virulence genes (stx1, stx2, eae, and ehxA) using an 11-gene multiplex PCR assay. Generalized linear mixed models were used to analyze the effects of treatments and make within-sample comparisons of the presence of O-serogroup–specific genes. Results of cumulative prevalence measures indicated that O157 (14.6%), O26 (10.5%), and O103 (10.3%) were the most prevalent STEC O serogroups. However, the vaccine, DFM, or both had no significant effect (P &gt; 0.05) on fecal prevalence of the six non-O157 STEC serogroups in feedlot cattle. Within-sample comparisons of the presence of STEC serogroup–specific genes indicated that fecal shedding of E. coli O157 in cattle was associated with an increased probability (P &lt; 0.05) of fecal shedding of STEC O26, O45, O103, and O121. Our study revealed that neither the E. coli O157:H7 vaccine, which reduced STEC O157 fecal shedding, nor the DFM significantly affected fecal shedding of non-O157 STEC serogroups, despite the fact that the most prevalent non-O157 STEC serogroups tended to occur concurrently with O157 STEC strains within fecal samples.
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PAOLI, GEORGE C., CHANDI WIJEY, and GAYLEN A. UHLICH. "Genetically Marked Strains of Shiga Toxin–Producing O157:H7 and Non-O157 Escherichia coli: Tools for Detection and Modeling†." Journal of Food Protection 78, no. 5 (2015): 888–901. http://dx.doi.org/10.4315/0362-028x.jfp-14-472.
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Shiga toxin–producing E. coli (STEC) is an important group of foodborne pathogens in the United States and worldwide. Nearly half of STEC-induced diarrheal disease in the United States is caused by serotype O157:H7, while non-O157 STEC account for the remaining illnesses. Thus, the U.S. Department of Agriculture (USDA) Food Safety and Inspection Service has instituted regulatory testing of beef products and has a zero-tolerance policy for regulatory samples that test positive for STEC O157:H7 and six other non-O157 STEC (serogroups O26, O45, O103, O111, O121, and O145). In this study, positive control (PC) strains for the detection of STEC O157:H7 and the six USDA-regulated non-O157 STEC were constructed. To ensure that the food testing samples are not cross-contaminated by the PC sample, it is important that the STEC-PC strains are distinguishable from STEC isolated from test samples. The PC strains were constructed by integrating a unique DNA target sequence and a gene for spectinomycin (Sp) resistance into the chromosomes of the seven STEC strains. End-point and real-time PCR assays were developed for the specific detection of the PC strains and were tested using 93 strains of E. coli (38 STEC O157:H7, at least 6 strains of each of the USDA-regulated non-O157 STEC, and 2 commensal E. coli) and 51 strains of other bacteria (30 species from 20 genera). The PCR assays demonstrated high specificity for the unique target sequence. The target sequence was detectable by PCR after 10 culture passages (~100 generations), demonstrating the stability of the integrated target sequence. In addition, the strains were tested for their potential use in modeling the growth of STEC. Plating the PC strains mixed with ground beef flora on modified rainbow agar containing Sp eliminated the growth of the background flora that grew on modified rainbow agar without Sp. Thus, these strains could be used to enumerate and model the growth of STEC in the presence of foodborne background flora.
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LACHER, DAVID W., JAYANTHI GANGIREDLA, ISHA PATEL, CHRISTOPHER A. ELKINS, and PETER C. H. FENG. "Use of the Escherichia coli Identification Microarray for Characterizing the Health Risks of Shiga Toxin–Producing Escherichia coli Isolated from Foods." Journal of Food Protection 79, no. 10 (2016): 1656–62. http://dx.doi.org/10.4315/0362-028x.jfp-16-176.
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ABSTRACT More than 470 serotypes of Shiga toxin–producing Escherichia coli (STEC) have been identified, but not all cause severe illness in humans. Most STEC that cause severe diseases can adhere to epithelial cells, produce specific stx subtypes, and belong to certain serotypes; therefore, these traits appear to be critical STEC risk factors. However, testing for these traits is labor intensive, and serotyping is inadequate because of extensive variations among E. coli O and H antigen types. In the present study, the E. coli identification microarray, which tests for over 40,000 E. coli gene targets, was examined for its potential to quickly characterize STEC strains. Analysis of 47 E. coli isolates, including 31 STEC isolates, recovered from 39 foods revealed that the microarray effectively determined the presence or absence of adherence genes and identified the specific eae allele in 3 isolates. The array identified most of the stx subtypes carried by all the isolates but had some difficulties in discerning between stx2a, stx2c, and stx2d because of the genetic similarities of these subtypes. The array determined the O and H types of 68 and 96% of the isolates, respectively, and although most serotypes were unremarkable, a few known pathogenic serotypes were also found. These selected STEC traits provided a scientific basis for assessing the potential health risks of STEC strains and also showed the importance of H typing in determining health risks. However, the diversity of the STEC group, the complexity of virulence mechanisms, and the variation in pathotypes among strains continue to pose challenges to assessing the potential of STEC strains to cause severe illness.
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Imamovic, Lejla, Rosangela Tozzoli, Valeria Michelacci, et al. "OI-57, a Genomic Island of Escherichia coli O157, Is Present in Other Seropathotypes of Shiga Toxin-Producing E. coli Associated with Severe Human Disease." Infection and Immunity 78, no. 11 (2010): 4697–704. http://dx.doi.org/10.1128/iai.00512-10.
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ABSTRACT Strains of Shiga toxin-producing Escherichia coli (STEC) are a heterogeneous E. coli group that may cause severe disease in humans. STEC have been categorized into seropathotypes (SPTs) based on their phenotypic and molecular characteristics and the clinical features of the associated diseases. SPTs range from A to E, according to a decreasing rank of pathogenicity. To define the virulence gene asset (“virulome”) characterizing the highly pathogenic SPTs, we used microarray hybridization to compare the whole genomes of STEC belonging to SPTs B, C, and D with that of STEC O157 (SPT A). The presence of the open reading frames (ORFs) associated with SPTs A and B was subsequently investigated by PCR in a larger panel of STEC and in other E. coli strains. A genomic island termed OI-57 was present in SPTs A and B but not in the other SPTs. OI-57 harbors the putative virulence gene adfO, encoding a factor enhancing the adhesivity of STEC O157, and ckf, encoding a putative killing factor for the bacterial cell. PCR analyses showed that OI-57 was present in its entirety in the majority of the STEC genomes examined, indicating that it represents a stable acquisition of the positive clonal lineages. OI-57 was also present in a high proportion of the human enteropathogenic E. coli genomes assayed, suggesting that it could be involved in the attaching-and-effacing colonization of the intestinal mucosa. In conclusion, OI-57 appears to be part of the virulome of pathogenic STEC and further studies are needed to elucidate its role in the pathogenesis of STEC infections.
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Stromberg, Zachary R., Rick E. Masonbrink, and Melha Mellata. "Transcriptomic Analysis of Shiga Toxin-Producing Escherichia coli during Initial Contact with Cattle Colonic Explants." Microorganisms 8, no. 11 (2020): 1662. http://dx.doi.org/10.3390/microorganisms8111662.
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Foodborne pathogens are a public health threat globally. Shiga toxin-producing Escherichia coli (STEC), particularly O26, O111, and O157 STEC, are often associated with foodborne illness in humans. To create effective preharvest interventions, it is critical to understand which factors STEC strains use to colonize the gastrointestinal tract of cattle, which serves as the reservoir for these pathogens. Several colonization factors are known, but little is understood about initial STEC colonization factors. Our objective was to identify these factors via contrasting gene expression between nonpathogenic E. coli and STEC. Colonic explants were inoculated with nonpathogenic E. coli strain MG1655 or STEC strains (O26, O111, or O157), bacterial colonization levels were determined, and RNA was isolated and sequenced. STEC strains adhered to colonic explants at numerically but not significantly higher levels compared to MG1655. After incubation with colonic explants, flagellin (fliC) was upregulated (log2 fold-change = 4.0, p < 0.0001) in O157 STEC, and collectively, Lon protease (lon) was upregulated (log2 fold-change = 3.6, p = 0.0009) in STEC strains compared to MG1655. These results demonstrate that H7 flagellum and Lon protease may play roles in early colonization and could be potential targets to reduce colonization in cattle.
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Семейко, Г. В., Е. О. Самойлович, С. В. Байко, and М. Д. Чередниченко. "Etiological Diagnostics of Postdiarheal Hemolytic Uremic Syndrome in Children Using TaqMan Array Cards." Педиатрия. Восточная Европа, no. 4 (January 25, 2023): 510–19. http://dx.doi.org/10.34883/pi.2022.10.4.006.
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Введение. Применение микропроточных TaqMan Array Cards (TAC карт) стало одним из инновационных направлений микробиологии последних двух десятилетий, позволяя одномоментно определять целый спектр энтеропатогенов (бактерий, вирусов, простейших, гельминтов). Цель. Идентификация возбудителей кишечных инфекций у детей с постдиарейным гемолитико-уремическим синдромом (ГУС) с использованием метода ПЦР в реальном времени на основе ТАС карт. Материалы и методы. В исследование включено 54 ребенка в возрасте от 9 месяцев до 10 лет с диагнозом ГУС, госпитализированных в УЗ «2-я городская детская клиническая больница» г. Минска в 2021 г. Результаты. Гены энтеропатогенов, способных вызывать диарейный синдром, были выявлены у 39 (72,2%) из 54 обследованных детей: гены бактериальных патогенов (диареегенная E. сoli, Campylobacter, Clostridium difficile, Bacteroides fragilis) – у 23 (42,6%), вирусных патогенов (рота-, норо-, адено-, саповирусов) – у 11 (20,4%), у 2 детей (3,7%) выявлены гены простейшего (Cryptosporidium), у 3 детей (5,6%) – гены и вирусных и бактериальных патогенов. Среди 25 детей с выявленной диареегенной E. coli у 20 (80,0%) присутствовала шигатоксин-продуцирующая E. сoli (STEC). Во всех 20 случаях наличие STEC сопровождалось выявлением шигатоксина 2-го типа, в 8 их них одновременно присутствовал шигатоксин 1-го и 2-го типов. В 6 случаях выявлялись только STEC, в 8 – обнаружена энтерогеморрагическая E. coli (STEC/EHEC), у 6 – гибридные варианты (STEC/EAEC и STEC/EHEC/EAEC). 92,6% детей получали антибактериальные или кишечные противомикробные препараты на момент поступления в нефрологический центр. STEC выявлялись через 7 (5; 9) дней, максимально – 13 дней от начала диареи до забора фекалий. Выводы. Полученные с использованием ТАС карт данные подтверждают роль STEC как основного этиологического агента ГУС. Роль других E. сoli (не шигатоксин-продуцирующих штаммов), а также других патогенов в развитии ГУС требует дальнейшего изучения. Целесообразно расширение алгоритма этиологической диагностики ГУС и генетического анализа обнаруженных гибридных вариантов диареегенных E. coli. Introduction. Applying of microfluidic TaqMan Array Cards (TAC cards) has become one of the innovative directions in microbiology of the last two decades, allowing simultaneous detection of a wide range of enteropathogens (bacteria, viruses, protozoa, helminths). Purpose. Identification of the aetiology of diarrhoea in children with postdiarrheal hemolytic uremic syndrome (HUS) using real-time PCR based on TAC cards. Materials and methods. A total of 54 patients aged 9 months to 10 years with HUS hospitalized at the 2nd City Children’s Clinical Hospital in Minsk in 2021 were included in the study. Results. Genes of enteropathogens responsible for diarrheal syndrome were identified in 39 (72.2%) of 54 examined children: genes of bacterial pathogens (diarrheal E. coli, Campylobacter, Clostridium difficile, Bacteroides fragilis) – in 23 (42.6%), viral pathogens (Rota-, Noro-, Adeno-, Sapoviruses) – in 11 (20.4%); in 2 children (3.7%) genes of the protozoan (Cryptosporidium) were detected, and in 3 children (5.6%) – genes of both viral and bacterial pathogens. Among 25 children diagnosed with diarrheagenic E. coli, 20 (80.0%) had shigatoxin-producing E. coli (STEC). In all 20 cases shigatoxin type 2 (stx2) was detected, in 8 of them both shigatoxin types 1 and 2 were present. In 6 cases, only STEC was detected, whereas enterohemorrhagic E. coli (STEC/EHEC) in 8 cases and hybrid variants in 6 cases (STEC/EAEC and STEC/EHEC/EAEC). 92.6% of children received antibacterial or intestinal antimicrobials at the time of admission to the nephrology center. STEC were detected after 7 (5; 9) days, with a maximum of 13 days from the onset of diarrhea to fecal collection. Conclusions. The data obtained using TAC cards confirm the role of STEC as the main etiological agent of HUS. The role of other E. coli (non-shiga toxin-producing strains), as well as other pathogens, in the development of HUS requires further study. It is advisable to expand the algorithm for HUS etiological diagnosis of and genetic analysis of detected hybrid variants of diarrheal E. coli.
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Tolen, Tamra, Yicheng Xie, Thomas Hairgrove, Jason Gill, and T. Taylor. "Evaluation of Commercial Prototype Bacteriophage Intervention Designed for Reducing O157 and Non-O157 Shiga-Toxigenic Escherichia coli (STEC) on Beef Cattle Hide." Foods 7, no. 7 (2018): 114. http://dx.doi.org/10.3390/foods7070114.
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Microbiological safety of beef products can be protected by application of antimicrobial interventions throughout the beef chain. This study evaluated a commercial prototype antimicrobial intervention comprised of lytic bacteriophages formulated to reduce O157 and non-O157 Shiga-toxigenic Escherichia coli (STEC) on beef cattle hide pieces, simulating commercial pre-harvest hide decontamination. STEC reduction in vitro by individual and cocktailed phages was determined by efficiency of plating (EOP). Following STEC inoculation onto hide pieces, the phage intervention was applied and hide pieces were analyzed to quantify reductions in STEC counts. Phage intervention treatment resulted in 0.4 to 0.7 log10 CFU/cm2 (p < 0.01) E. coli O157, O121, and O103 reduction. Conversely, E. coli O111 and O45 did not show any significant reduction after application of bacteriophage intervention (p > 0.05). Multiplicity of infection (MOI) evaluation indicated E. coli O157 and O121 isolates required the fewest numbers of phages per host cell to produce host lysis. STEC-attacking phages may be applied to assist in preventing STEC transmission to beef products.
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Bower, John R. "Foodborne Diseases: Shiga Toxin Producing E. coli (STEC)." Pediatric Infectious Disease Journal 18, no. 10 (1999): 909–10. http://dx.doi.org/10.1097/00006454-199910000-00014.
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ATALLA, HEBA NASHED, ROGER JOHNSON, SCOTT MCEWEN, R. W. USBORNE, and C. L. GYLES. "Use of a Shiga Toxin (Stx)-Enzyme-Linked Immunosorbent Assay and Immunoblot for Detection and Isolation of Stx-Producing Escherichia coli from Naturally Contaminated Beef." Journal of Food Protection 63, no. 9 (2000): 1167–72. http://dx.doi.org/10.4315/0362-028x-63.9.1167.
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The purpose of this study was to evaluate an enzyme-linked immunosorbent assay (ELISA) and an immunoblot procedure for detection and isolation of Shiga toxin-producing Escherichia coli (STEC) from beef, and to correlate the presence of STEC in beef with E. coli and total coliform counts. A total of 120 samples of boneless beef supplied to a meat processor in southern Ontario were tested for the presence of STEC, E. coli, and total coliforms. Following enrichment in modified tryptic soy broth, samples were screened for Shiga toxin (Stx) by a Stx-ELISA and a Vero cell assay (VCA). Samples that were positive in the Stx-ELISA were subjected to the Stx-immunoblot for STEC isolation. Overall, 33.3% of samples were positive in the VCA, and 34.2% were positive in the Stx-ELISA. There was almost complete agreement between the Stx-ELISA and the VCA results (kappa = 0.98). The sensitivity and specificity of the Stx-ELISA with respect to the VCA were 100% and 98.75%, respectively. STEC were isolated by the Stx-immunoblot from 87.8% of the samples that were positive in the Stx-ELISA. The STEC isolates belonged to 19 serotypes, with serotype O113:H21 accounting for 10 of 41 isolates. No STEC of serotype O157:H7 were isolated. There was a significant correlation between E. coli counts and total coliform counts (Spearman correlation coefficient = 0.68, P &lt; 0.01). The E. coli count was positively correlated with detection of STEC by both the Stx-ELISA and the VCA (P &lt; 0.01).
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HARRIES, M., J. DREESMAN, S. RETTENBACHER-RIEFLER та E. MERTENS. "Faecal carriage of extended-spectrum β-lactamase-producing Enterobacteriaceae and Shiga toxin-producing Escherichia coli in asymptomatic nursery children in Lower Saxony (Germany), 2014". Epidemiology and Infection 144, № 16 (2016): 3540–48. http://dx.doi.org/10.1017/s0950268816001837.
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SUMMARYChildren may be at higher risk for carriage of antimicrobial-resistant bacteria because of higher usage of antimicrobials. They also have higher rates of Shiga toxin-producing Escherichia coli (STEC) infections than other population groups. Some infections, particularly in children, are asymptomatic, but still lead to the excretion of large numbers of bacteria and viruses that may cause clinical disease in other individuals. That is one reason why, in Lower Saxony as in other German federal states – asymptomatic carriers of STEC are excluded from nurseries and schools until three consecutive stool samples test negative in order to prevent secondary cases. The prevalence of children who are asymptomatic STEC carriers is unknown. But if it is high, this measure would have substantial socioeconomic effects on families. Infections with extended-spectrum β-lactamase-producing Enterobacteriaceae (ESBL-E) are an increasing problem for public health, especially for hospitals. However, there are no reliable estimates of the prevalence of asymptomatic ESBL-E carriers in Lower Saxony, as there is no mandatory requirement to report these carriers. In order to discuss the exclusion policies for children attending nurseries and ascertain a baseline of ESBL-E carriers, we conducted a cross-sectional study. The aim was to determine the prevalence of ESBL-E and STEC and identify risk factors for carriage in nursery children without diarrhoea (asymptomatic) aged 0–6 years in four selected districts in Northern Germany. During April–September 2014, we collected stool specimens with the support of voluntarily participating nurseries. We tested for STEC by PCR and for ESBL-E on chromogenic agar. Questionnaires answered by parents contained data on eating and drinking habits, outdoor activities, prior antibiotic treatment and animal contact for each participating child. We compared the epidemiological characteristics of ESBL-E carriers vs. non-carriers by using univariable analysis (P value, odds ratio and 95% confidence interval). We could not perform a statistical analysis for STEC carriers due to the low numbers of positive STEC specimens. Of 224 asymptomatic nursery children, we found a prevalence of 2·3% for ESBL-E carriage and 0·5% for STEC carriage. Asymptomatic ESBL-E carriers were more likely to have consumed raw milk, have had contact with pet rodents, or to have taken antibiotics during the preceding 6 months. We also found a high proportion of raw milk consumption (11%). We suggest that the low STEC prevalence in asymptomatic children supports the current practice of excluding STEC carriers from nurseries. The association between ESBL-E carriage and raw milk consumption and contact with pet rodents needs further investigation.
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Lorenz, Sandra C., Insook Son, Anna Maounounen-Laasri, Andrew Lin, Markus Fischer, and Julie A. Kase. "Prevalence of Hemolysin Genes and Comparison ofehxASubtype Patterns in Shiga Toxin-Producing Escherichia coli (STEC) and Non-STEC Strains from Clinical, Food, and Animal Sources." Applied and Environmental Microbiology 79, no. 20 (2013): 6301–11. http://dx.doi.org/10.1128/aem.02200-13.
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ABSTRACTShiga toxin-producingEscherichia coli(STEC) belonging to certain serogroups (e.g., O157 and O26) can cause serious conditions like hemolytic-uremic syndrome (HUS), but other strains might be equally pathogenic. While virulence factors, likestxandeae, have been well studied, little is known about the prevalence of theE. colihemolysin genes (hlyA,ehxA,e-hlyA, andsheA) in association with these factors. Hemolysins are potential virulence factors, andehxAandhlyAhave been associated with human illness, but the significance ofsheAis unknown. Hence, 435E. colistrains belonging to 62 different O serogroups were characterized to investigate gene presence and phenotypic expression of hemolysis. We further investigatedehxAsubtype patterns inE. coliisolates from clinical, animal, and food sources. WhilesheAandehxAwere widely distributed,e-hlyAandhlyAwere rarely found. Most strains (86.7%) were hemolytic, and significantly more hemolytic (95%) than nonhemolytic strains (49%) carriedstxand/oreae(P< 0.0001).ehxAsubtyping, as performed by using PCR in combination with restriction fragment length polymorphism analysis, resulted in six closely related subtypes (>94.2%), with subtypes A/D beingeae-negative STECs and subtypes B, C, E, and Feaepositive. Unexpectedly,ehxAsubtype patterns differed significantly between isolates collected from different sources (P< 0.0001), suggesting that simple linear models of exposure and transmission need modification; animal isolates carried mostly subtypes A/C (39.3%/42.9%), food isolates carried mainly subtype A (81.9%), and clinical isolates carried mainly subtype C (66.4%). Certain O serogroups correlated with particularehxAsubtypes: subtype A with O104, O113, and O8; B exclusively with O157; C with O26, O111, and O121.
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Kudva, Indira T., Erika N. Biernbaum, and Julian M. Trachsel. "Growth dynamics and protein-expression of Escherichia coli serotypes O26:H11, O111:H8 and O145:NM in the bovine rumen." PLOS One 20, no. 6 (2025): e0313978. https://doi.org/10.1371/journal.pone.0313978.
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To adapt to the ruminal environment, Shiga toxin-producing Escherichia coli (STEC) O157:H7 (O157) expresses proteins involved in survival rather than virulence. Additionally, STEC O157 strains exhibit distinct in vitro but shared in vivo survival patterns in rumen fluids that sets them apart from non-STEC, commensal E. coli. To determine if similar responses would be observed with other STEC, we evaluated three non-O157 serotypes, O26:H11, O111:H8 and O145:NM, along with a non-STEC E. coli, under the growth conditions used for STEC O157: (i) anaerobically, in vitro, in rumen fluid from cattle on a lactation (low fiber, high protein) or maintenance (high fiber, low protein) diet, at 39oC for 48 hr and (ii) in vivo for 48 hr within the rumen of cattle on the same diets using a non-terminal, rumen-fistulated animal model that allows for introduction of bacteria without ruminal contamination. On the lactation diet, the ruminal pH was acidic ranging from 5.2–6.0 and the total volatile fatty acids (VFA) concentration, 141–230 μM/ml. On the maintenance diet, the ruminal fluid pH was close to neutral ranging from 6.0–7.0, with a total VFA of 87–190 μM/ml. Unlike STEC O157, the three non-O157 serotypes demonstrated survival patterns similar to each other and the control non-STEC E. coli. A greater reduction in viable STEC counts was observed in vitro in rumen fluid from cattle fed the lactation diet than in vivo, corroborating previous reports that in vitro conditions cannot mimic those observed in vivo. Like STEC O157, the non-O157 serotypes mainly expressed proteins supporting ruminal adaptation although not all proteins matched those expressed by STEC O157, and included the virulence protein, intimin. Explorative studies such as this could provide insights into common conditions/ targets that may have application in broader STEC control in cattle. Importance of this study This study demonstrates that non-O157 serotypes O26:H11, O111:H8, O145:NM have similar survival and protein expression patterns in rumen fluid with variations being influenced primarily by rumen fluid composition associated with diet. Unlike STEC O157, under in vivo conditions, the growth dynamics of the non-O157 serotypes were comparable to that of non-STEC, commensal E. coli. Hence, exploring bacterial protein expression within the host is critical in discerning therapeutic targets, unique to/shared between STEC, for broad control strategies. In addition, this study further validates the value of using a non-terminal animal model for rumen studies that reduces number of animals used for an experiment.
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Hornitzky, Michael A., Kim Mercieca, Karl A. Bettelheim, and Steven P. Djordjevic. "Bovine Feces from Animals with Gastrointestinal Infections Are a Source of Serologically Diverse Atypical Enteropathogenic Escherichia coli and Shiga Toxin-Producing E. coli Strains That Commonly Possess Intimin." Applied and Environmental Microbiology 71, no. 7 (2005): 3405–12. http://dx.doi.org/10.1128/aem.71.7.3405-3412.2005.
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ABSTRACT Shiga toxin-producing Escherichia coli (STEC) and enteropathogenic E. coli (EPEC) cells were isolated from 191 fecal samples from cattle with gastrointestinal infections (diagnostic samples) collected in New South Wales, Australia. By using a multiplex PCR, E. coli cells possessing combinations of stx 1, stx 2, eae, and ehxA were detected by a combination of direct culture and enrichment in E. coli (EC) (modified) broth followed by plating on vancomycin-cefixime-cefsulodin blood (BVCC) agar for the presence of enterohemolytic colonies and on sorbitol MacConkey agar for the presence of non-sorbitol-fermenting colonies. The high prevalence of the intimin gene eae was a feature of the STEC (35 [29.2%] of 120 isolates) and contrasted with the low prevalence (9 [0.5%] of 1,692 fecal samples possessed STEC with eae) of this gene among STEC recovered during extensive sampling of feces from healthy slaughter-age cattle in Australia (M. Hornitzky, B. A. Vanselow, K. Walker, K. A. Bettelheim, B. Corney, P. Gill, G. Bailey, and S. P. Djordjevic, Appl. Environ. Microbiol. 68:6439-6445, 2002). Forty-seven STEC serotypes were identified, including O5:H−, O8:H19, O26:H−, O26:H11, O113:H21, O157:H7, O157:H− and Ont:H− which are known to cause severe disease in humans and 23 previously unreported STEC serotypes. Serotypes Ont:H− and O113:H21 represented the two most frequently isolated STEC isolates and were cultured from nine (4.7%) and seven (3.7%) animals, respectively. Fifteen eae-positive E. coli serotypes, considered to represent atypical EPEC, were identified, with O111:H− representing the most prevalent. Using both techniques, STEC cells were cultured from 69 (36.1%) samples and EPEC cells were cultured from 30 (15.7%) samples, including 9 (4.7%) samples which yielded both STEC and EPEC. Culture on BVCC agar following enrichment in EC (modified) broth was the most successful method for the isolation of STEC (24.1% of samples), and direct culture on BVCC agar was the most successful method for the isolation of EPEC (14.1% samples). These studies show that diarrheagenic calves and cattle represent important reservoirs of eae-positive E. coli.
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Stevens, Mark P., Olivier Marchès, June Campbell, et al. "Intimin, Tir, and Shiga Toxin 1 Do Not Influence Enteropathogenic Responses to Shiga Toxin-Producing Escherichia coli in Bovine Ligated Intestinal Loops." Infection and Immunity 70, no. 2 (2002): 945–52. http://dx.doi.org/10.1128/iai.70.2.945-952.2002.
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ABSTRACT Shiga toxin-producing Escherchia coli (STEC) comprises a group of attaching and effacing (A/E) enteric pathogens of animals and humans. Natural and experimental infection of calves with STEC may result in acute enteritis or subclinical infection, depending on serotype- and host-specific factors. To quantify intestinal secretory and inflammatory responses to STEC in the bovine intestine, serotypes that are associated with human disease (O103:H2 and O157:H7) were introduced into ligated mid-ileal loops in gnotobiotic and conventional calves, and fluid accumulation and recruitment of radiolabeled neutrophils were measured after 12 h. STEC serotype O103:H2, but not serotype O157:H7, elicited strong enteropathogenic responses. To determine if the inflammatory response to STEC O103:H2 in calves requires Shiga toxin 1 or intimate bacterial attachment to the intestinal epithelium, defined mutations were made in the stx1, eae, and tir genes. Our data indicate that some STEC induce intestinal inflammatory responses in calves by a mechanism that is independent of A/E-lesion formation, intimin, or Shiga toxin 1. This may have implications for strategies to reduce STEC carriage in cattle.
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Szczerba-Turek, Anna, and Bernard Kordas. "Fallow Deer (Dama dama) as a Reservoir of Shiga Toxin-Producing Escherichia coli (STEC)." Animals 10, no. 5 (2020): 881. http://dx.doi.org/10.3390/ani10050881.
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Shiga toxin-producing Escherichia (E.) coli (STEC) are responsible for the outbreaks of serious diseases in humans. Only a few reports on fallow deer as a reservoir of foodborne pathogens have been published to date. The purpose of this study was to determine the occurrence of STEC strains in the fallow deer population in Poland. In all, 94 fallow deer swabs were tested. Polymerase chain reaction (PCR) was performed to detect the virulence profile of stx1, stx2 and eae or aggR genes, to identify the subtypes of stx1 and stx2 genes and to perform O and H serotyping. STEC and attaching and effacing (AE)-STEC were identified in 13 isolates (13.83%). The most hazardous virulence profile was detected in three strains, namely stx2d serotype O103:HNM, eae/stx1a serotype O26:HNM and eae/stx1a serotype O157:H7. The predominant stx gene was stx2, which was identified in 76.92% of isolates. E. coli O157 was detected in 4/94 (4.26%). Other E. coli serogroups, O26, O103, O111 and O145, were identified in 14/94 fallow deer (14.89%). The present findings suggest that fallow deer are carriers of STEC/AE-STEC that are potentially pathogenic to humans.
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Sang, Willie Kipkemboi, Hamadi Iddi Boga, Peter Gichuhi Waiyaki, David Schnabel, Njeri C. Wamae, and Sam M. Kariuki. "Prevalence and genetic characteristics of Shigatoxigenic Escherichia coli from patients with diarrhoea in Maasailand, Kenya." Journal of Infection in Developing Countries 6, no. 02 (2011): 102–8. http://dx.doi.org/10.3855/jidc.1750.
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Introduction: Shigatoxigenic Escherichia coli strains are food-borne bacterial pathogens that may cause haemorrhagic colitis (HC) in humans which can lead to life-threatening systemic complication, including haemolytic uremic syndrome (HUS). This study aimed to characterize and analyze virulence properties of pathogenic E. coli isolates among patients with diarrhoea from a Maasai community in Kenya. Methodology: Stool samples from 380 patients of all ages from the Kajiado and Narok districts of Kenya were investigated for the presence of enteric bacterial pathogens by conventional and molecular methods. Results: Bacterial diarrhoea was diagnosed in 141/380 (37.1%) cases, of which enterotoxigenic E. coli (ETEC) compromised 29.8%, shigatoxigenic E. coli (STEC) 24.1%, enteroaggregative E. coli (EAEC) 14.2%, enteroinvasive E. coli (EIEC) 12.8% and enteropathogenic E. coli (EPEC) 3.5%. Gene analysis for STEC virulence factors showed that 52.9% isolates carried stx1, 29.4% possessed stx2, 14.7% carried both stx1 and stx2, and 2.9% had stx2e. 23.5% isolates carried enterohaemolysin and 20.5% isolates possessed the Intimin gene. From 9 strains that exhibited adherence, 7 contained both Intimin and Haemolysin genes. Infections with Intimin-positive STEC strains (46%) were more frequent in patients with bloody diarrhoea, especially in children under 5 years of age, whereas Intimin-negative STEC infections dominated in adults. Conclusion: Although STEC infection as a cause of bloody diarrhoea has not attracted much attention as a medical problem in Kenya, our findings indicate that this is a problem that must be investigated. The 24.1% isolation rate of STEC among the Maasai is one of the highest reported rates worldwide.
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Girardeau, Jean Pierre, Yolande Bertin, and Christine Martin. "Genomic analysis of the PAI ICL3 locus in pathogenic LEE-negative Shiga toxin-producing Escherichia coli and Citrobacter rodentium." Microbiology 155, no. 4 (2009): 1016–27. http://dx.doi.org/10.1099/mic.0.026807-0.
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Shiga toxin-producing Escherichia coli (STEC) causes a spectrum of human illnesses such as haemorrhagic colitis and haemolytic–uraemic syndrome. Although the locus of enterocyte effacement (LEE) seems to confer enhanced virulence, LEE-negative STEC strains are also associated with severe human disease, suggesting that other unknown factors enhance the virulence potential of STEC strains. A novel hybrid pathogenicity island, termed PAI ICL3, has been previously characterized in the LEE-negative O113 : H21 STEC strain CL3. Screening for the presence of PAI ICL3 elements in 469 strains of E. coli, including attaching and effacing (A/E) pathogens [enteropathogenic E. coli (EPEC) and enterohaemorrhagic E. coli (EHEC)], non-A/E pathogens [LEE-negative STEC, extra-intestinal pathogenic E. coli (ExPEC), enterotoxigenic E. coli (ETEC) and enteroaggregative E. coli (EAEC)] and commensal E. coli isolates, showed that PAI ICL3 is unique to LEE-negative STEC strains linked to disease, providing a new marker for these strains. We also showed that a PAI ICL3-equivalent gene cluster is present in the genome of Citrobacter rodentium, on a 53 kb genomic island inserted into the pheV tRNA locus. While the C. rodentium PAI ICL3 shows high similarities at the nucleotide level and in organization with the E. coli PAI ICL3, the genetic context of the integration differs completely. In addition, blast searches revealed that other E. coli pathotypes (O157 : H7 EHEC, ExPEC, EPEC and EAEC) possess incomplete PAI ICL3 elements that contain only the genes located at the extremities of the island. Six of the 16 sequenced E. coli genomes showed deleted PAI ICL3 gene clusters which are carried on mobile genetic elements inserted into pheV, selC or serW tRNA loci, which is compatible with the idea that the PAI ICL3 gene cluster entered E. coli and C. rodentium at multiple times through independent events. The phylogenetic distribution of the PAI ICL3 variants suggests that a B1 genetic background is necessary for the maintenance of the full complement of PAI ICL3 genes in E. coli.
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KANKI, MASASHI, KAZUKO SETO, and YUKO KUMEDA. "Simultaneous Immunomagnetic Separation Method for the Detection of Escherichia coli O26, O111, and O157 from Food Samples." Journal of Food Protection 77, no. 1 (2014): 15–22. http://dx.doi.org/10.4315/0362-028x.jfp-13-225.
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We performed a simultaneous immunomagnetic separation (IMS) assay on Shiga toxin–producing Escherichia coli (STEC) serogroups O26, O111, and O157 with immunobeads coated with O26, O111, or O157 antibodies that were simultaneously added to an aliquot of food culture. We also compared the usefulness of CHROMagar STEC medium against various selective isolation agars designed to test for the three serogroups. Samples of sliced beef, ground beef, and radish sprout were artificially contaminated with STEC O26, O111, and O157 strains after incubation in enrichment broth and were subjected to conventional and simultaneous IMS assays. Simultaneous IMS did not affect the sensitivity of target cell detection. For STEC O26, O111, and O157 inoculated into the enriched samples of sliced beef and radish sprout, the detection ability of CHROMagar STEC was similar to or exceeded that of other isolation agars. However for STEC O157 inoculated into ground beef cultures, cefixime tellurite sorbitol MacConkey agar (CT-SMAC) was the superior detection medium. The performance of simultaneous IMS combined with CT-SMAC and CHROMagar STEC detection is similar to that of the Japanese standard method for isolating E. coli O26, O111, and O157. However, the proposed approach involves the same time, materials, and labor costs as the standard E. coli O157 reference procedure but allows detection of three E. coli serotypes rather than a single strain.