Usability and Performance of CHROMagar STEC Medium in Detection of Shiga Toxin-Producing Escherichia coli Strains

The performance and usability of CHROMagar STEC medium (CHROMagar Microbiology, Paris, France) for routine detection of Shiga toxin-producing Escherichia coli (STEC) strains were examined. The ability of the medium to selectively propagate STEC strains differing by their serotypes and virulence genes was studied with a collection of diarrheagenic E. coli isolates (n = 365) consisting of 49 different serotypes and with non-STEC and other bacterial isolates (n = 264). A total of 272 diarrheagenic E. coli (75.0%) isolates covering 24 different serotypes grew on CHROMagar STEC. The highest detection sensitivities were observed within the STEC serogroups O26 (90.0%), O111 (100.0%), O121 (100.0%), O145 (100.0%), and O157 (84.9%), and growth on CHROMagar STEC was highly associated with the presence of the tellurite resistance gene (terD). The specificity of the medium was 98.9%. In addition, CHROMagar STEC was used in parallel with a Shiga toxin-detecting immunoassay (Ridaquick Verotoxin/O157 Combi; R-biopharm, Darmstadt, Germany) to screen fecal specimens (n = 47) collected from patients suffering from hemorrhagic diarrhea. Positive growth on CHROMagar STEC was confirmed by the Premier EHEC enzyme immunoassay (Meridian Bioscience, Inc., Cincinnati, OH), and discrepant results between the two screening methods were confirmed by stx gene-detecting PCR. All 16 of the 47 stool samples that showed positive growth on CHROMagar STEC were also positive in the confirmatory tests. CHROMagar STEC proved to be an interesting option for STEC screening, allowing good detection sensitivity and specificity and permitting strain isolation for further outbreak investigations when required.     

Four-Year Experience with Simultaneous Culture and Shiga Toxin Testing for Detection of Shiga Toxin-Producing Escherichia coli in Stool Samples

We report our experience with universal Shiga toxin-producing Escherichia coli (STEC) screening using culture and Shiga toxin antigen testing over 4 years. Twelve cases were detected—8 detected by both culture and Shiga toxin immunochromatographic assay (IA), 3 by culture, and 1 by IA only. The addition of Shiga toxin testing is of questionable benefit over culture alone for detection of STEC in areas of low prevalence.     

Rapid and Sensitive Detection of Shiga Toxin-Producing Escherichia coli from Nonenriched Stool Specimens by Real-Time PCR in Comparison to Enzyme Immunoassay and Culture

Shiga toxin (Stx)-producing Escherichia coli (STEC) bacteria are a frequent cause of food-borne gastroenteritis, hemorrhagic colitis, and hemolytic uremic syndrome. Because antimicrobial agents are generally contraindicated in patients infected with STEC, a sensitive and specific diagnostic test with rapid turnaround is essential. Current culture methods may fail to detect non-O157 STEC. We evaluated a Stx gene real-time PCR assay using hybridization probes and the LightCycler instrument with 204 prospectively collected stool specimens, which were also tested for Stx by enzyme immunoassay (EIA) (ProSpecT STEC; Remel, Lenexa, KS) and by culturing on chromogenic agar (Chromagar O157; BD BBL, Sparks, MD). In addition, 85 archived stool specimens previously tested for Stx (by EIA) and/or E. coli O157:H7 (by culture) were tested by PCR. Sample preparation for PCR included mixing the stool in sterile water and extraction of nucleic acid using the MagNA Pure LC instrument (Roche Diagnostics). The PCR assay had 100% sensitivity and specificity compared to EIA and culture for specimens collected prospectively (4 of 204 specimens were positive) and compared to culture and/or EIA for archival specimens (42 of 85 specimens were positive). Both the EIA and PCR produced positive results from a specimen containing an O103 serotype STEC in the prospective specimens, and the PCR test detected three positive specimens that contained nonviable STEC in the archived specimens. The PCR assay demonstrated 100% sensitivity and specificity compared to EIA and/or culture and more rapid turnaround than either EIA or culture.Shiga toxin (Stx)-producing Escherichia coli (STEC) is a frequent cause of food-borne outbreaks of diarrhea (15). Disease caused by STEC is characterized by abdominal pain and bloody diarrhea, and 5 to 15% of those individuals infected with serotype O157:H7 develop hemolytic uremic syndrome (HUS), a potentially life-threatening condition consisting of hemolytic anemia, thrombocytopenia, and kidney failure caused primarily by Stx (8). STEC may carry genes for one or both types of Stx, Stx1 and Stx2 (17).Although STEC strains are a diverse group of pathogens, up to the present, the most common serotype in the United States has been O157:H7. A common association is that of E. coli O157:H7 contaminating ground beef (3, 7), but recent large outbreaks have involved a variety of other foods, including leafy greens (6, 29). The diversity of potentially contaminated food means that patients may acquire STEC infection from many foodstuffs, far beyond the stereotypical risk of undercooked ground beef. The common denominator of tainted food products seems to be direct or indirect contamination from bovine feces. To best detect infected patients and potential outbreaks, clinical laboratories must have tools to quickly and accurately detect STEC in stool specimens. Culture on sorbitol MacConkey agar is an inexpensive, effective, and widely used method based on lack of sorbitol fermentation by E. coli O157:H7. Several drawbacks limit the utility of culture, including slow turnaround, false-negative results in antibiotic-treated patients, and false-negative results due to emerging serotypes of non-O157 STEC that ferment sorbitol (1, 14, 16, 29). Alternatively, a method that is increasingly utilized is detection of Stx antigen from stool, either directly or after broth enrichment. Our experience concurs with enzyme immunoassay (EIA) product insert data that optimal sensitivity and specificity are achieved only when a broth enrichment step is employed; this results in slow turnaround.Here, we describe a real-time PCR assay that can detect STEC using nucleic acid extracts of stool specimens. We evaluated the performance of this assay using both archived stool specimens and prospectively collected specimens and compared the results to those of culture and Stx antigen detection.(This study was presented in part at the 48th Annual Interscience Conference on Antimicrobial Agents and Chemotherapy, Washington, DC, 25 to 28 October 2008.)     

Virulence Properties of Shiga Toxin-Producing Escherichia coli (STEC) Strains of Serogroup O118, a Major Group of STEC Pathogens in Calves

Shiga toxin-producing Escherichia coli (STEC) strains of serogroup O118 are the most prevalent group among STEC strains in diarrheic calves in Germany (L. H. Wieler, Ph.D. thesis, University of Giessen, 1997). To define their virulence properties, 42 O118 (O118:H16 [n = 38] and O118:H− [n = 4]) strains were characterized. The strains displayed three different Stx combinations (Stx1 [36 of 42], Stx1 and Stx2 [2 of 42], and Stx2 [4 of 42]). A total of 41 strains (97.6%) harbored a large virulence-associated plasmid containing hly_EHEC (hly from enterohemorrhagic E. coli). The strains’ adhesive properties varied in relation to the eukaryotic cells tested. Only 28 of 42 strains (66.7%) showed localized adhesion (LA) in the human HEp-2 cell line. In contrast, in bovine fetal calf lung (FCL) cells, the number of LA-positive strains was much higher (37 of 42 [88.1%]). The locus of enterocyte effacement (LEE) was detected in 41 strains (97.6%). However, not all LEE-positive strains reacted positively in the fluorescence actin-staining (FAS) test, which indicated the attaching and effacing (AE) lesion. In HEp-2 cells, only 22 strains (52.4%) were FAS positive, while in FCL cells, the number of FAS-positive strains was significantly higher (38 of 42 [90.5%; P < 0.001]). In conclusion, the vast majority of the O118 STEC strains from calves (41 of 42 [97.6%]) have a high virulence potential (stx, hly_EHEC, and LEE). This virulence potential and the high prevalence of STEC O118 strains in calves suggest that these strains could be a major health threat for humans in the future. In addition, the poor association between results of the geno- and phenotypical tests to screen for the AE ability of STEC strains calls the diagnostic value of the FAS test into question.     

Evaluation and Implementation of a Chromogenic Agar Medium for Salmonella Detection in Stool in Routine Laboratory Diagnostics

We evaluated which chromogenic agar medium for Salmonella detection in stool would be most sensitive and specific in our culture protocol. The use of BBL CHROMagar Salmonella chromogenic medium combined with xylose-lysine-deoxycholate agar yielded a sensitivity of 100% and also reduced workload and costs.     

Clinical Evaluation of a Real-Time PCR Assay for Identification of Salmonella,Shigella, Campylobacter (Campylobacter jejuni and C. coli), and Shiga Toxin-Producing Escherichia coli Isolates in Stool Specimens

Enteric illness affects millions of individuals annually in the United States and results in >50,000 hospitalizations. The rapid and accurate identification of bacterial pathogens associated with gastroenteritis can aid acute patient management decisions, including the use of antibiotic therapy and infection control. This study compared the ProGastro SSCS multiplex real-time PCR assay (Gen-Probe Prodesse, San Diego, CA) to culture for the identification of Campylobacter spp. (Campylobacter jejuni and Campylobacter coli), Salmonella spp., and Shigella spp. and to broth enrichment followed by an FDA-cleared enzyme immunoassay (EIA) for the identification of Shiga toxin-producing Escherichia coli (STEC) isolates in stool specimens. Stool samples submitted in preservatives for routine culture and EIA were prospectively enrolled and tested at four clinical centers. Discrepancies between the ProGastro SSCS assay and culture or EIA were resolved using bidirectional sequencing. The overall prevalence of the pathogens as detected by culture was 5.6% (1.8% Campylobacter, 1.8% Salmonella, 1.3% Shigella, and 0.8% STEC). When results based on the ProGastro SSCS assay and bidirectional sequencing were applied, the overall prevalence increased to 8.3% (2.3% Campylobacter, 2.6% Salmonella, 1.8% Shigella, and 1.6% STEC). Following resolution of the discrepant results, the sensitivity of the ProGastro SSCS assay was 100% for all pathogens, and the specificities ranged from 99.4% to 100%. The sensitivity of culture compared to sequence-confirmed ProGastro SSCS results ranged from 52.9% to 76.9%, with the specificities ranging from 99.9% to 100%. Overall, these results suggest that the ProGastro SSCS assay is highly sensitive and specific in a clinical setting.     

Pathogenesis and Diagnosis of Shiga Toxin-Producing Escherichia coli Infections

Since their initial recognition 20 years ago, Shiga toxin-producing Escherichia coli (STEC) strains have emerged as an important cause of serious human gastrointestinal disease, which may result in life-threatening complications such as hemolytic-uremic syndrome. Food-borne outbreaks of STEC disease appear to be increasing and, when mass-produced and mass-distributed foods are concerned, can involve large numbers of people. Development of therapeutic and preventative strategies to combat STEC disease requires a thorough understanding of the mechanisms by which STEC organisms colonize the human intestinal tract and cause local and systemic pathological changes. While our knowledge remains incomplete, recent studies have improved our understanding of these processes, particularly the complex interaction between Shiga toxins and host cells, which is central to the pathogenesis of STEC disease. In addition, several putative accessory virulence factors have been identified and partly characterized. The capacity to limit the scale and severity of STEC disease is also dependent upon rapid and sensitive diagnostic procedures for analysis of human samples and suspect vehicles. The increased application of advanced molecular technologies in clinical laboratories has significantly improved our capacity to diagnose STEC infection early in the course of disease and to detect low levels of environmental contamination. This, in turn, has created a potential window of opportunity for future therapeutic intervention.     

Real-Time PCR Assay for Detection and Differentiation of Shiga Toxin-Producing Escherichia coli from Clinical Samples

Timely accurate diagnosis of Shiga toxin-producing Escherichia coli (STEC) infections is important. We evaluated a laboratory-developed real-time PCR (LD-PCR) assay targeting stx₁, stx₂, and rfbE_O157 with 2,386 qualifying stool samples submitted to the microbiology laboratory of a tertiary care pediatric center between July 2011 and December 2013. Broth cultures of PCR-positive samples were tested for Shiga toxins by enzyme immunoassay (EIA) (ImmunoCard STAT! enterohemorrhagic E. coli [EHEC]; Meridian Bioscience) and cultured in attempts to recover both O157 and non-O157 STEC. E. coli O157 and non-O157 STEC were detected in 35 and 18 cases, respectively. Hemolytic uremic syndrome (HUS) occurred in 12 patients (10 infected with STEC O157, one infected with STEC O125ac, and one with PCR evidence of STEC but no resulting isolate). Among the 59 PCR-positive STEC specimens from 53 patients, only 29 (54.7%) of the associated specimens were toxin positive by EIA. LD-PCR differentiated STEC O157 from non-O157 using rfbE_O157, and LD-PCR results prompted successful recovery of E. coli O157 (n = 25) and non-O157 STEC (n = 8) isolates, although the primary cultures and toxin assays were frequently negative. A rapid “mega”-multiplex PCR (FilmArray gastrointestinal panel; BioFire Diagnostics) was used retrospectively, and results correlated with LD-PCR findings in 25 (89%) of the 28 sorbitol-MacConkey agar culture-negative STEC cases. These findings demonstrate that PCR is more sensitive than EIA and/or culture and distinguishes between O157 and non-O157 STEC in clinical samples and that E. coli O157:H7 remains the predominant cause of HUS in our institution. PCR is highly recommended for rapid diagnosis of pediatric STEC infections.     

Characterization of Urinary Tract Infection-Associated Shiga Toxin-Producing Escherichia coli

Enterohemorrhagic Escherichia coli (EHEC), a subgroup of Shiga toxin (Stx)-producing E. coli (STEC), is a leading cause of diarrhea and hemolytic-uremic syndrome (HUS) in humans. However, urinary tract infections (UTIs) caused by this microorganism but not associated with diarrhea have occasionally been reported. We geno- and phenotypically characterized three EHEC isolates obtained from the urine of hospitalized patients suffering from UTIs. These isolates carried typical EHEC virulence markers and belonged to HUS-associated E. coli (HUSEC) clones, but they lacked virulence markers typical of uropathogenic E. coli. One isolate exhibited a localized adherence (LA)-like pattern on T24 urinary bladder epithelial cells. Since the glycosphingolipids (GSLs) globotriaosylceramide (Gb3Cer) and globotetraosylceramide (Gb4Cer) are well-known receptors for Stx but also for P fimbriae, a major virulence factor of extraintestinal pathogenic E. coli (ExPEC), the expression of Gb3Cer and Gb4Cer by T24 cells and in murine urinary bladder tissue was examined by thin-layer chromatography and mass spectrometry. We provide data indicating that Stxs released by the EHEC isolates bind to Gb3Cer and Gb4Cer isolated from T24 cells, which were susceptible to Stx. All three EHEC isolates expressed stx genes upon growth in urine. Two strains were able to cause UTI in a murine infection model and could not be outcompeted in urine in vitro by typical uropathogenic E. coli isolates. Our results indicate that despite the lack of ExPEC virulence markers, EHEC variants may exhibit in certain suitable hosts, e.g., in hospital patients, a uropathogenic potential. The contribution of EHEC virulence factors to uropathogenesis remains to be further investigated.     

Recent Progress of Shiga Toxin Neutralizer for Treatment of Infections by Shiga Toxin-Producing Escherichia coli

Infection with Shiga toxin (Stx)-producing Escherichia coli (STEC), including O157:H7, causes bloody diarrhea and hemorrhagic colitis in humans, occasionally resulting in fatal systemic complications, such as neurological damage and hemolytic-uremic syndrome. Because Stx is a major virulence factor of the infectious disease, a series of Shiga toxin neutralizers with various structural characteristics has been developed as promising therapeutic agents. Most of these agents function to bind to the toxin directly and inhibit the binding to its receptor present on the target cells. Other neutralizers do not inhibit receptor binding but induce aberrant intracellular transport of the toxin, resulting in effective detoxification. Such a novel type of Stx neutralizer provides a new therapeutic strategy against STEC infections. Here, recent progress of the development of Stx neutralizers is reviewed.     

Characterization of Third-Generation-Cephalosporin-Resistant Shiga Toxin-Producing Strains of Escherichia coli O157:H7 in Japan

We isolated Shiga toxin-producing Escherichia coli O157:H7 strains resistant to third-generation cephalosporins. The resistant strains harbored bla_CMY-2, a plasmid-mediated AmpC β-lactamase. Genotyping of isolates revealed the possible spread of this problematic bacterium. Results suggested the importance of the investigation and surveillance of enterobacteria with plasmids harboring bla_CMY-2.     

Evaluation of Enzyme Immunoassays and Real-Time PCR for Detecting Shiga Toxin-Producing Escherichia coli in Southern Alberta,Canada

Two immunoassays (Shiga Toxin Chek and Shiga Toxin Quik Chek) and real-time PCR were used to detect Shiga toxin-producing Escherichia coli. For enriched culture, the sensitivity and specificity of the three methods ranged from 80.0% to 98.2% and 98.0% to 100.0%, respectively. STEC isolates were identified in 2.6% of the 784 samples.     

Evaluation of a New Selective Medium,BD BBL CHROMagar MRSA II,for Detection of Methicillin-Resistant Staphylococcus aureus in Stool Specimens

We compared the recovery of methicillin-resistant Staphylococcus aureus (MRSA) on a new selective chromogenic agar, BD BBL CHROMagar MRSA II (CMRSAII), to that on traditional culture media with 293 stool specimens. The recovery of MRSA was greater on the CMRSAII agar. Screening of stool samples can identify patients who were previously unknown carriers of MRSA.Active surveillance cultures to identify patients colonized with methicillin-resistant Staphylococcus aureus (MRSA) are recommended as part of an intensive program to control the spread of the organism in health care settings (10, 14). Although the anterior nares are currently the most common site from which to obtain culture specimens to test for MRSA colonization (4, 7, 13, 15), other sites, including the intestinal tract, can be potential sources of unrecognized colonization by this organism (1, 2, 6). Patients with S. aureus (including MRSA) colonization of the intestinal tract are associated with greater contamination of health care workers'' hands and the environment (3). Patients with intestinal colonization by MRSA and concurrent diarrhea contaminate their environment to a significantly greater extent than patients colonized at other body sites (5). Screening stool samples for MRSA can detect patients who are previously unknown carriers and who may not otherwise be cared for using contact precautions, which could potentially lead to increased health care-associated transmission and subsequent MRSA infections (1, 2, 6). It has also been suggested that undetected intestinal colonization by MRSA could be the cause of persistent colonization or recolonization in patients after successful decontamination (8, 11).The use of chromogenic agars has been shown to increase sensitivity and decrease the time to detection of MRSA compared with traditional culture methods (9). However, to date, all currently available chromogenic media used for screening patients for MRSA are recommended only for use with specimens from the anterior nares. We evaluated a new selective and differential chromogenic medium, BD BBL CHROMagar MRSA II (CMRSAII) (BD Diagnostics, Sparks, MD), for its ability to detect MRSA in specimens from multiple body sites, including stool specimens, as part of a large multicenter trial (14a). We report on the evaluation of this medium for detection of MRSA from stool samples collected at our institution.The study was conducted during the period from December 2007 to February 2008 at a 500-bed university-affiliated hospital. A total of 293 stool specimens collected from all wards in the hospital and submitted to the laboratory for Clostridium difficile toxin assay (CDT) were included in the study. Stools were not rejected on the basis of specimen consistency. The specimens were inoculated onto colistin-nalidixic acid (CNA) agar and immediately afterward onto CMRSAII. The plates were incubated in ambient air at 36°C and examined at 24 h (range, 18 to 28 h), and if negative, they were reincubated and examined at 48 h (range, 36 to 52 h). CMRSAII was incubated in the dark to avoid exposure to light. Colonies morphologically consistent with S. aureus recovered on CNA plates were confirmed as S. aureus with a positive coagulase test (Staphaureux, Remel, Lenexa, KS) and confirmed as MRSA by cefoxitin disk diffusion. Mauve-colored colonies present on CMRSAII at 24 and 48 h were presumed to be MRSA and were confirmed as S. aureus with a positive coagulase test. A cefoxitin disk diffusion test was performed if MRSA was not recovered from the CNA plate. Differences in proportions were analyzed using McNemar''s test.MRSA was recovered from 62 (21.2%) of 293 stool specimens. The recovery rate was 60/62 specimens (96.8%) for CMRSAII and 47/62 (75.8%) specimens for CNA media (P = 0.004) (Table ​(Table1).1). Of the 60 MRSA isolates recovered on CMRSAII, 44/60 (73.3%) of MRSA isolates were identified at 24 h and 14/60 (26.7%) at 48 h, whereas the traditional culture method took between 48 to 96 h to confirm an isolate as MRSA. When CMRSAII was compared with cefoxitin disk diffusion, the overall agreement was 97.3% (285/293) at 24 h and 99.3% (291/293) at 48 h. Compared to cefoxitin disk diffusion, CMRSAII had a sensitivity of 84.6% (44/52) at 24 h and 96.8% (60/62) at 48 h and a specificity of 100% at both 24 and 48 h. Traditional culture had a sensitivity of 90.4% (47/52) at 24 h and 75.8% (47/62) at 48 h and a specificity of 100% at both 24 and 48 h compared to the cefoxitin disk diffusion method (Table ​(Table2).2). The sensitivity of the traditional culture method at 48 h was lower than that at 24 h because additional stools were positive on CMRSAII at 48 h and were confirmed by cefoxitin disk diffusion, but none of these were positive at 48 h on traditional media. No false-positive results were identified on CMRSAII after 48 h of incubation.

TABLE 1.

Performance of BD BBL CHROMagar MRSA II compared to colistin-nalidixic acid agar

Comparative virulence characterization of the Shiga toxin phage-cured Escherichia coli O104:H4 and enteroaggregative Escherichia coli

Escherichia coli O104:H4 (E. coli O104:H4), which caused in 2011 a massive foodborne outbreak in Germany, is characterized by an unusual combination of virulence traits. E. coli O104:H4 contains a prophage-encoded Shiga toxin (Stx) gene, which is the cardinal virulence factor of enterohemorrhagic E. coli (EHEC). However, the outbreak strain shares highest DNA sequence similarity with enteroaggregative E. coli (EAEC) and displays the EAEC-characteristic tight adherence to epithelial cells. The virulence potential of the underlying EAEC background has not been investigated and it is therefore not clear whether E. coli O104:H4 displays distinct virulence characteristics in comparison to prototypical EAEC. In this study, we performed a detailed comparative phenotypic characterization of the Stx phage-cured E. coli O104:H4 strain C227-11φcu, the closely related EAEC strain 55989 and two other well-characterized EAEC strains 042 and 17-2 with focus on virulence traits. C227-11φcu displayed superior aggregative adherence phenotype to cultured HCT-8 epithelial cells, adhering with 3–6 times more bacteria per epithelial cells than the tested EAEC strains. Otherwise, C227-11φcu showed similar virulence characteristics to its closest relative 55989, i.e. strong acid resistance, good biofilm formation and cytotoxic culture supernatants. Furthermore, C227-11φcu was characterized by significantly weaker motility and pro-inflammatory properties than 55989 and 042, nevertheless stronger than 17-2. Taken together, C227-11φcu displayed mostly robust, but not outstanding virulence characteristics in comparison to the tested EAEC. Therefore, it appears likely that the combination of Stx production and EAEC characteristics in general, rather than an exceptionally potent EAEC background resulted in the unusual virulence of the E. coli O104:H4. Thus, the emergence of such hypervirulent strains in the future might be more likely than previously anticipated.     

Associations between Virulence Factors of Shiga Toxin-Producing Escherichia coli and Disease in Humans

Associations between known or putative virulence factors of Shiga toxin-producing Escherichia coli and disease in humans were investigated. Univariate analysis and multivariate logistic regression analysis of a set of 237 isolates from 118 serotypes showed significant associations between the presence of genes for intimin (eae) and Shiga toxin 2 (stx₂) and isolates from serotypes reported in humans. Similar associations were found with isolates from serotypes reported in hemorrhagic colitis and hemolytic-uremic syndrome. The enterohemorrhagic E. coli (EHEC) hemolysin gene was significantly associated with isolates from serotypes found in severe diseases in univariate analysis but not in multivariate logistic regression models. A strong association between the intimin and EHEC-hemolysin genes may explain the lack of statistical significance of EHEC hemolysin in these multivariate models, but a true lack of biological significance of the hemolysin in humans or in disease cannot be excluded. This result warrants further investigations of this topic. Multivariate analysis revealed an interaction between the eae and stx₂ genes, thus supporting the hypothesis of the synergism between the adhesin intimin and Shiga toxin 2. A strong statistical association was observed between the stx₂ gene and severity of disease for a set of 112 human isolates from eight major serotypes. A comparison of 77 isolates of bovine origin and 91 human isolates belonging to six major serotypes showed significant associations of the genes for Shiga toxin 1 and EspP protease with bovine isolates and an increased adherence on HEp-2 cell cultures for human isolates, particularly from diarrheic patients and healthy persons.     

Hemolytic Uremic Syndrome following Infection with O111 Shiga Toxin-Producing Escherichia coli Revealed through Molecular Diagnostics

We report a case of hemolytic uremic syndrome in a 69-year-old woman due to Shiga toxin-producing Escherichia coli, possibly serotype O111, to illustrate the potentially deleterious implications of a Campylobacter enzyme immunoassay (EIA) result and the increasing importance of molecular testing when conventional methods are limited.     

PCR-Based Detection and Molecular Characterization of Shiga Toxin-Producing Escherichia coli Strains in a Routine Microbiology Laboratory over 16 years

Shiga toxin-producing Escherichia coli (STEC) is a heterogeneous group of bacteria causing disease ranging from asymptomatic carriage and mild infection to hemolytic uremic syndrome (HUS). Here, we describe patients with STEC infection and characterize the STEC strains detected in our laboratory by use of PCR for stx₁, stx₂, and eae from 1996 through 2011. Patient information was collected from referral forms and from the Norwegian Surveillance System for Communicable Diseases. STEC isolates were characterized with respect to serogroup or serotype, selected potential virulence genes, and multilocus variable-number tandem-repeat analysis (MLVA) genotype. STEC strains were isolated from 138 (1.09%) of 12,651 patients tested. STEC strains of serogroups O26, O103, O121, O145, and O157 were the most frequent. These serogroups, except non-sorbitol-fermenting O157, were also the most frequent among the 11 patients (all ≤5 years old) who developed HUS. Twenty-four STEC strains were classified as being HUS associated based on an epidemiological link to a HUS case, including an MLVA genotype identical to that of the STEC strain. The age of the patient (≤5 years) and the genes eae and stx_2a were significantly associated with HUS-associated STEC (P < 0.05 for each parameter), while stx₁ was associated with non-HUS-associated STEC (P < 0.05). All of the potential virulence genes analyzed, except ehxA, were significantly more frequent among HUS-associated than non-HUS-associated strains (P < 0.05 for each gene). However, these genes were also present in some non-HUS-associated STEC strains and could therefore not reliably differentiate between HUS-associated and non-HUS-associated STEC strains.