Child ; China ; Diarrhea ; diagnosis ; microbiology ; Escherichia coli Infections ; diagnosis ; Humans ; Shiga-Toxigenic Escherichia coli ; isolation & purification

Verotoxin-producing Escherichia coli O157 is a primary cause of severe and bloody diarrhea. Campylobacter spp. are one of the commonly reported bacterial cause of gastrointestinal infections throughout the world. Only a few cases involving both E. coli O157 and Campylobacter species have been reported. The authors simultaneously isolated verotoxin-producing E. coli O157 and Campylobacter species from the stool of a 3 year-old male with bloody diarrhea, fever and abdominal pain.
Abdominal Pain ; Campylobacter* ; Child, Preschool ; Diarrhea ; Escherichia coli O157* ; Escherichia coli* ; Escherichia* ; Fever ; Gastroenteritis* ; Humans ; Male ; Shiga Toxins ; Shiga-Toxigenic Escherichia coli

BACKGROUND: The Polymerase Chain Reaction (PCR) for the Shiga toxin has been widely used for diagnosis of Shiga toxin-producing Escherichia coli (STEC) infection including Escherichia coli O157 (O157) instead of using a culture. However, bacteriological isolation must be followed for final diagnosis. Our study was aimed to compare the detection limit between the culture after the acid treatment and the PCR after enrichment. METHODS: The standard strain of O157 was cultured, diluted and mixed with the stool of normal adult in order to make a final concentration of the 10(1)-10(5) colony forming unit (CFU)/g of stool. Each concentration of samples was enriched in a trypticase soy broth for 6 hours at 42degrees C and treated with acid to suppress normal flora. Then it was streaked on cefixime-tellurite-sorbitol MacConkey (CT-SMAC) agar evenly and cultured for 18 hours at 37degrees C. The same concentrations of bacterial suspension in the stool were enriched in a Luria-Bertani (LB) broth overnight at 37degrees C. The centrifuged pellets from 1 mL of each concentration of the samples were boiled and DNA was extracted using the resin method and PCR was performed to amplify stx2. RESULTS: The detection limit for the culture after acid treatment was 10(3) CFU/g of the stool and that for PCR after enrichment was 101 CFU/g of the stool. CONCLUSIONS: Culture after acid treatment for O157 would be an effective method for isolation of O157 from a patient's stool. However, this method is less sensitive than the PCR after enrichment as far as detection limit is concerned. A combination of both methods would be an effective method for detecting O157 from patient stools.
Adult ; Agar ; Diagnosis ; DNA ; Escherichia coli O157* ; Escherichia coli* ; Escherichia* ; Humans ; Limit of Detection ; Polymerase Chain Reaction* ; Shiga Toxin ; Shiga-Toxigenic Escherichia coli ; Stem Cells

We encountered a patient with hemolytic uremic syndrome (HUS) with persistent isolation of shiga toxin-producing Escherichia coli (STEC) for 3 weeks despite of having no clinical symptoms. STEC has been recognized as an important food-borne pathogen that causes severe diseases such as HUS. We characterized this STEC strain via a polymerase chain reaction, reverse-passive latex agglutination and the slide agglutination method. In this STEC strain, stx2 (shiga toxin), eaeA, tir, iha (adherence genes), espADB (type III secretion genes), and hlyA, ehxA, clyA (hemolysin genes) were present. The O antigen of the strain was non-typable.
Child, Preschool ; Female ; Hemolytic-Uremic Syndrome/diagnosis/*microbiology ; Humans ; Shiga-Toxigenic Escherichia coli/*isolation & purification/*pathogenicity

Salmonella (S.) enterica and Shiga toxin-producing Escherichia coli (STEC) are foodborne pathogens. Here, we report the prevalence of S. enterica and STEC in feces of 316 zoo animals belonging to 61 species from Chile. S. enterica and STEC strains were detected in 7.5% and 4.4% of animals, respectively. All Salmonella isolates corresponded to the serotype Enteritidis. To the best of our knowledge, this is the first report of S. Enteritidis in the culpeo fox (Lycalopex culpaeus), black-capped capuchin (Sapajus apella) and Peruvian pelican (Pelecanus thagus) and the first STEC report in Thomson's gazelle (Eudorcas thomsonii).
Animals ; Animals, Zoo* ; Chile* ; Feces ; Prevalence* ; Salmonella enterica* ; Salmonella* ; Serogroup ; Shiga-Toxigenic Escherichia coli*

Herein, we report the pathogenic and phylogenetic characteristics of seven Shiga toxin (Stx)-producing Escherichia coli (STEC) isolates from 434 retail meats collected in Korea during 2006 to 2012. The experimental analyses revealed that all isolates (i) were identified as non-O157 STEC, including O91:H14 (3 isolates), O121:H10 (2 isolates), O91:H21 (1 isolate), and O18:H20 (1 isolate), (ii) carried diverse Stx subtype genes (stx₁, stx(2c), stx(2e), or stx₁ + stx(2b)) whose expression levels varied strain by strain, and (iii) lacked the locus of enterocyte effacement (LEE) pathogenicity island, a major virulence factor of STEC, but they possessed one or more alternative virulence genes encoding cytotoxins (Cdt and SubAB) and/or adhesins (Saa, Iha, and EcpA). Notably, a significant heterogeneity in glutamate-induced acid resistance was observed among the STEC isolates (p < 0.05). In addition, phylogenetic analyses demonstrated that all three STEC O91:H14 isolates were categorized into sequence type (ST) 33, of which two beef isolates were identical in their pulsotypes. Similar results were observed with two O121:H10 pork isolates (ST641; 88.2% similarity). Interestingly, 96.0% of the 100 human STEC isolates collected in Korea during 2003 to 2014 were serotyped as O91:H14, and the ST33 lineage was confirmed in approximately 72.2% (13/18 isolates) of human STEC O91:H14 isolates from diarrheal patients.
Cytotoxins ; Enterocytes ; Escherichia coli ; Genomic Islands ; Humans ; Korea ; Meat ; Population Characteristics ; Red Meat ; Shiga Toxin ; Shiga-Toxigenic Escherichia coli ; Virulence ; Virulence Factors

Sequence type (ST) 33 of Shiga toxin-producing Escherichia coli (STEC) strain O91:H14 has been proposed as a potential domestic clone of STEC in Korea because of its high prevalence among human patients with mild diarrhea or asymptomatic carriers. Herein, the clonal diversity of 17 STEC O91:H14 isolates of ST33 during 2003 to 2014 was analyzed by pulsed-field gel electrophoresis, including 14 isolates from human patients and 3 from retail meats. Their virulence characteristics, acid resistance, and antimicrobial susceptibility were also determined. Our results showed that all isolates were clustered mainly into three different pulsotypes and were likely low pathogenic without antimicrobial resistance.
Clone Cells ; Diarrhea ; Electrophoresis, Gel, Pulsed-Field ; Escherichia coli ; Humans ; Korea ; Meat ; Molecular Epidemiology ; Prevalence ; Shiga Toxin ; Shiga-Toxigenic Escherichia coli ; Virulence

In Gwangju, Korea, over the last 4 years, human gastrointestinal infection caused by shiga toxin-producing E. coli (STEC) increased. The aim of this study was to ascertain the genetic relatedness of STEC strains by pulsed-field gel electrophoresis (PFGE), as no data on the molecular epidemiology of STEC in Gwangju has yet been published. The PFGE banding patterns were defined for 62 of the 67 STEC strains isolated from cattle and human. There were 11 clonal types in the 11 STEC strains of cattle origin. Among the 11 STEC strains from asymptomatic person, four O91 strains were 100% similarity in band profiles. In the STEC strains isolated from diarrhea patients, same serogroups were grouped to the same cluster; O111 stains were 89.5% similarity, O157 strains 80%, O26 strains 81.5%, and O103 strains 91% similarity, respectively. In conclusion, this is the first report that a large collection of STEC strains from Korea has been analyzed, and a high degree of diversity was observed among the strains analyzed by this technique. PFGE analysis revealed that the strains isolated from human and cattle were closely related within serotypes, and it was useful for epidemiological analysis of STEC. The importance and usefulness of active laboratory surveillance of STEC such as PFGE should be recommended.
Animals ; Cattle ; Coloring Agents ; Diarrhea ; Electrophoresis ; Electrophoresis, Gel, Pulsed-Field ; Humans ; Korea ; Molecular Epidemiology ; Shiga-Toxigenic Escherichia coli

BACKGROUND: Causative bacterial agents of infectious diarrheal disease were traditionally diagnosed by stool cultures. Stool culture, however, has a problem because of relatively low sensitivity and long turnaround time. In this study, we evaluated multiplex PCR applied on stool specimens directly to diagnose enteropathogenic bacteria. METHODS: From June to September 2009, 173 diarrheal stools submitted for stool cultures were tested by Seeplex(R) Diarrhea ACE Detection kit (Seegene, Korea) to detect 10 enteropathogenic bacteria. Specimens were cultured for Salmonella, Shigella, Vibrio, and Yersinia. Late 50 specimens were also cultured for Campylobacter. The specimens positive for verotoxin-producing Escherichia coli (VTEC) were further subcultured for detecting enterohaemorrhagic Escherichia coli O157:H7. Electronic medical records were reviewed for clinical and laboratory findings. RESULTS: Of 173 specimens, multiplex PCR and cultures identified enteropathogens in 36 (20.8%) and 8 specimens (4.6%), respectively. While multiplex PCR detected 5 Salmonella, 15 Campylobacter, 1 Vibrio, 4 Clostridium difficiles toxin B, 5 Clostridium perfringens, 1 Yersinia enterocolitica, 5 Aeromonas, and 2 VTEC, cultures detected 5 Salmonella, 1 Vibrio, 1 Y. enterocolitica, 1 Aeromonas, and 2 E. coli O157:H7. CONCLUSION: Multiplex PCR would be useful to detect Campylobacter, VTEC and C. perfringens, as well as have equivalent sensitivity to conventional culture for ordinary enteropathogens such as Salmonella, Shigella, Vibrio, Y. enterocolitica. Direct application of multiplex PCR combined with conventional cultures on stool warrants remarkable improvement of sensitivity to diagnose enteropathogenic bacteria.
Aeromonas ; Bacteria ; Campylobacter ; Clostridium ; Clostridium perfringens ; Diarrhea ; Dysentery ; Electronic Health Records ; Escherichia coli ; Multiplex Polymerase Chain Reaction ; Salmonella ; Shiga-Toxigenic Escherichia coli ; Shigella ; Vibrio ; Yersinia ; Yersinia enterocolitica

Enterohemorrhagic Escherichia coli (EHEC) is an important cause of bloody diarrhea in children, but is considered to be rare in infants. Herein, a case of infant hemorrhagic colitis of verotoxin-producing E. coli O157:H7 diagnosed by multiplex PCR is reported. A nine-month-old boy was admitted to our hospital with bloody diarrhea for the previous two days. Multiplex PCR using Seeplex(R) Diarrhea ACE Detection Kit (Seegene, Seoul, Korea) was directly applied to the stool specimens. Amplified bands specific for verotoxin, O157, and H7 indicated the presence of O157:H7 EHEC. The stool specimens were inoculated on sorbitol-MacConkey agar (SMA) and tryptic soy broth containing mitomycin C (TSB-M). Colorless colonies on sorbitol-MacConkey agar were O157-positive. TSB-M enrichment cultures of the stool specimen and the isolates were positive for verotoxin according to an enzyme immunoassay (EIA). The prepared ingredients of baby foods for the patient including ground meat, chopped carrot, chopped cabbage, and white rice porridge showed no EHEC on TSB-M and SMA. The patient's parents and three-year-old sister did not recently have any gastrointestinal symptoms. Cefdinir was administered for one day and was ceased after diagnosis of EHEC colitis. The stool culture and verotoxin assay were negative on the second day of hospitalization. Application of multiplex PCR and verotoxin EIA directly to diarrheal stool warrants the rapid diagnosis and appropriate treatment of EHEC colitis.
Agar ; Brassica ; Caseins ; Cephalosporins ; Child ; Colitis ; Daucus carota ; Diarrhea ; Enterohemorrhagic Escherichia coli ; European Continental Ancestry Group ; Hospitalization ; Humans ; Immunoenzyme Techniques ; Infant ; Meat ; Mitomycin ; Multiplex Polymerase Chain Reaction ; Parents ; Protein Hydrolysates ; Shiga Toxins ; Shiga-Toxigenic Escherichia coli ; Siblings