Stx2 Genes Research Articles

Pathogenic E. coli from Cattle as a Reservoir of Resistance Genes to Various Groups of Antibiotics

Antimicrobial resistance has become a worldwide concern in all public health domains and reducing the spread has become a global priority. Pathogenic E. coli is responsible for a number of illnesses in humans and outbreaks in the past have been correlated with the consumption of contaminated bovine products. This is why surveillance in all the steps of production is essential. This study focused on identifying the pathogenic strains of E. coli in two large bovine abattoirs from Romania and France, and on associating them with the antimicrobial resistance patterns. A total of 250 samples from intestinal content were aseptically collected during the evisceration step of the cattle slaughtering process, from which 242 E. coli strains were isolated. Seventeen percent of all samples tested positive to at least one E. coli isolate carrying eaeA, stx1 and stx2 genes. The most prevalent genetic profile found in the E. coli strains tested was Stx1-positive and Stx2/eaeA-negative. More than 68% of the pathogenic E. coli isolated in Romania showed multi-drug resistance (MDR) and in France, the percentage was significantly lower (38%). The MDR profiles showed a high gene diversity for antibiotic resistance, which represents a great risk for environmental spread and human health. Our results indicate that in Romania, bovines can represent a reservoir for MDR E. coli and, hence, a surveillance system for antimicrobials usage in farm animals is highly needed.

MOLECULAR DETECTION OF SHIGA TOXIN (stx1 and stx2) AND INTIMIN (eaeA) GENES IN ESCHERICHIA COLI ISOLATED FROM FECAL SAMPLES OF CATTLE, SHEEP, AND HUMAN IN BASRAH GOVERNORATE

The present study aims to isolate and identify Escherichia coli from fecal samples of farmanimals and human, also, it aims to molecular detection of shigatoxin and intimin genes inisolates. A total of (264) fecal samples and swabs were collected from different parts of Basrahin the period extending from September 2018 to January 2019. These samples were composed of(85) samples from cows, (94) samples from humanand (85) samples from sheep. Differenttechniques were used in this study to detect the presence of E. coli; these techniques includedconventional microbiological assays and molecular techniques (amplification of uidA gene byusing polymerase chain reaction).The results of these techniques indicated 50 (18.9%) were E. coli from the tested samples.These isolates were subjected to PCR to detect Shiga toxins and intimin genes (stx1, stx2, andeaeA). The results of PCR confirmed all (50) isolates were harbor at least one virulence gene.Out of 50 isolates 20 (40%) carried stx2 gene alone, the percentages of the carrier were (66.7 %,41.7% and 23.5%) from human, sheep and cattle samples, respectively. The genes (stx1 andstx2) were detected together in 9/50 (18%), represent (52.9%) of cattle isolates. The intimin gene(eaeA) alone was detected in 2/50 (4%), represent (11.8%) of cattle isolates. (28%) of isolatesharbor (stx2 and eaeA) genes, the isolates belong to human and sheep isolates (33.3%) and(45.8%), respectively. Presence of the genes (stx1, stx2, and eaeA) were discovered in (10%)of isolates, (11.8%) of cattle and (12.5%) of sheep. The isolates were resistant to ampicillin, tetracycline (92%, 74%), respectively. However, theisolates were susceptible to imipenem, gentamycin, chloramphenicol, ciprofloxacin, andcefotaxime with a ratio of 100%, 92%, 78%, 68%, and 58%, respectively.

Evaluation of the effect of Saccharomyces cerevisiae on the expression of enterotoxin genes in Escherichia coli O157: H7 (EHEC) and Escherichia coli H10407 (ETEC)

Enterotoxigenic (ETEC) and enterohemorrhagic Escherichia coli (EHEC) are the most important intestinal pathogens. Probiotics play an effective role in reducing the expression of virulence factor genes in intestinal pathogenic bacteria. The aim of the present study is to investigate the effect of probiotic Saccharomyces cerevisiae S3 on the expression of enterotoxin genes in both ETEC and EHEC. Supernatant and lysate of S. cerevisiae S3 are prepared. Subminimal inhibitory concentrations (sub-MIC) of supernatant and lysate are individually exerted to O157: H7 and H10407. The genes' expression of enterotoxins (elt, est, stx1, and stx2) are then determined using real-time PCR technique. The results showed, the yeast supernatant could decrease the expression of the elt gene in ETEC and that of stx1 in EHEC. Of note, in other cases, stx1 and est genes’ expression increased. The lysate had no inhibitory effect on the expressions of elt, est, and stx2 genes, but it increased the expression of genes in both ETEC and EHEC. Lysate extract only decreased the expression of stx1 in O157: H7. Our study shows some interesting results regarding the effectiveness of the compounds produced by S. cerevisiae S3 in the expressions of toxin genes in both ETEC and EHEC. We recommend more similar studies be performed in this regard.

Validation of the 3M™ Molecular Detection Assay 2 - STEC Gene Screen (stx) for the Detection of Shiga Toxin Gene (stx1 and/or stx2) in Additional Matrixes: AOAC Performance Tested MethodSM 071903.

BackgroundThe 3M™ Molecular Detection Assay 2 - STEC Gene Screen (stx) method is based on gene amplification by the use of real-time loop-mediated isothermal amplification when used with the 3M Molecular Detection System for the rapid and specific detection of Shiga toxin gene (stx1 and/or stx2) from Shiga toxin-producing enterohemorrhagic Escherichia coli (STEC) in enriched foods.ObjectiveThe 3M Molecular Detection Assay 2 - STEC Gene Screen (stx) method was evaluated as a Level 2 method modification to add new matrixes to the certified claim: 25 g fresh raw ground beef (approximately 75% lean), 375 g raw beef trim (approximately 75% lean), 375 g fresh raw ground pork (approximately 70% lean), 375 g fresh raw poultry parts, and 25 g sprouts.MethodsMatrix studies were conducted to assess the method’s performance compared to the US Department of Agriculture Food Safety and Inspection Service Microbiology Laboratory Guidebook, 5C.00 for meat and poultry, and to the US Food and Drug Administration Bacteriological Analytical Manual, Ch. 4A for sprouts, using an unpaired study design.ResultsThe 3M Molecular Detection Assay 2 - STEC Gene Screen (stx) method demonstrated no significant differences between presumptive and confirmed results or between candidate and reference method results for any of the matrices tested.Conclusion and HighlightsThe data collected in these studies demonstrate that the 3M Molecular Detection Assay 2 - STEC Gene Screen (stx) is a reliable method for the rapid and specific detection of STEC in fresh raw ground beef (approximately 75% lean), fresh raw beef trim (approximately 75% lean), fresh raw ground pork (approximately 70% lean), fresh raw poultry parts, and sprouts.

Validation of the 3M™ Molecular Detection Assay 2 - STEC Gene Screen (stx and eae) for the Detection of Shiga Toxin Gene (stx1 and/or stx2) and Intimin Gene (eae): AOAC Performance Tested MethodSM 071902.

BackgroundThe 3M™ Molecular Detection Assay 2 - STEC Gene Screen (stx and eae) method is based on gene amplification by the use of real-time loop-mediated isothermal amplification when used with the 3M Molecular Detection System for the rapid and specific detection of Shiga toxin gene (stx1 and/or stx2) and intimin gene (eae) from Shiga toxin-producing enterohemorrhagic Escherichia coli (STEC) in enriched foods.ObjectiveThe 3M Molecular Detection Assay 2 - STEC Gene Screen (stx and eae) method was evaluated as a Level 2 method modification to add new matrixes to the certified claim: 25 g fresh raw ground beef (approximately 75% lean), 375 g fresh raw ground pork (approximately 70% lean), 375 g fresh raw poultry parts, and 25 g sprouts.MethodsMatrix studies were conducted to assess the method’s performance compared to the U. S. Department of Agriculture Food Safety and Inspection Service Microbiology Laboratory Guidebook, 5C.00 for meat and poultry, and to the U. S. Food and Drug Administration Bacteriological Analytical Manual, Ch. 4A for sprouts, using an unpaired study design.ResultsThe 3M Molecular Detection Assay 2 - STEC Gene Screen (stx and eae) method demonstrated no significant differences between presumptive and confirmed results or between candidate and reference method results for any of the matrixes tested.Conclusion and HighlightsThe data collected in these studies demonstrate that the 3M Molecular Detection Assay 2 - STEC Gene Screen (stx and eae) is a reliable method for the rapid and specific detection of STEC in fresh raw ground beef (approximately 75% lean), fresh raw ground pork (approximately 70% lean), fresh raw poultry parts, and sprouts.

Comparison of toxicities between Vibrio fluvialis and Vibrio furnissii strains isolated from environmental samples

Gastrointestinal episodes associated with Vibrio species have been rising worldwide in the last few years. In this study, toxicities of two very well characterized V. fluvialis and V. furnissii strains, isolated from environmental samples, were used to investigate the comparative pathogenicity of these strains. The results showed that there is a positive relationship between toxin production as well as a high interspecies correlation. In the analysis of toxicity, both species showed positive results with close similarity in cell death on BHK-21 cell line and destruction of RBC, which were indicative of cytotoxic and hemolytic activity, respectively. Besides, molecular analysis showed the absence of ctx, tdh, trh, stx1 and stx2 genes. On the other hand, reduced fluid accumulation ratio in rabbit ileal loop assay and paralysis of hind legs of Swiss Albino mice were observed for V. furnissi strain, which indicates the differences in pathogenicity between the two species.
 Bangladesh J Microbiol, Volume 38, Number 2, December 2021, pp 73-77

Prevalence and Genetic Diversity of Escherichia coli Isolated from Sewage Samples of Hospitals in Different Regions of Bangladesh

Escherichia coli are a large and diverse group of bacteria found in the environment, foods and intestines of humans and animals. This study was carried out to observe the genotypic variations in E. coli found at different regions in Bangladesh using Amplified Ribosomal DNA Restriction Analysis (ARDRA) of the 16S rDNA and RAPD (Random Amplified Polymorphic DNA). Sewage samples were collected from different medical college hospitals and a total of 118 samples were observed in this study of which 26 were identified as E. coli based on their cultural, biochemical and molecular characteristics. Antibiotic susceptibility test was carried out where most isolates were resistant to Amoxicillin, Rifampicin and Erythromycin; and some were resistant to kanamycin, ceftriaxone, Chloramphenicol and Azithromycin; besides 73% (N=19 out of 26) of the test isolates were multi-drug resistant. Results of Polymerase chain reaction (PCR) showed that none of the isolates carried Stx1, Stx2 and LT gene. Two genotypic groups were found from Amplified Ribosomal DNA Restriction Analysis (ARDRA) by Alu1 restriction enzyme and seven different Random Amplification of Polymorphic DNA (RAPD) patterns were found. After analyzing, it was found that 24 out of 26 isolates showed similarity with Escherichia coli strain NBRC 102203 and the remaining two isolates showed similarity with Escherichia coli strain Mui T5 based on the restriction patterns of ARDRA and sequences of 16S rDNA. From the study, it was concluded that Escherichia coli strain NBRC 102203 was more prevalent than Escherichia coli strain Mui T5 isolated from sewage sample of hospitals from different regions of Bangladesh.
 Bangladesh J Microbiol, Volume 38, Number 2, December 2021, pp 39-44

Occurrence of the Seven Most Common Serotypes of Shiga Toxin–Producing Escherichia coli in Beef Cuts Produced in Meat Processing Plants in the State of São Paulo, Brazil

Healthy cattle are considered the main reservoir of Shiga toxin-producing Escherichia coli (STEC) strains, so in some places in the world, products derived from beef are the most common source for disease outbreaks caused by these bacteria. Therefore, to guarantee that the beef produced by our slaughterhouses is safe, there is a need for continuous monitoring of these bacteria. In this study, 215 beef cuts were evaluated, including chilled vacuum-packed striploins (151 samples), rib eyes (30 samples), and knuckles (34 samples), from March to June 2018. These meat samples were collected from the slaughter of unconfined cattle, being arbitrarily collected from eight meat processing companies in São Paulo state, Brazil. Each sample was examined for the presence of STEC toxin type (stx1 and/or stx2 genes) and also the attaching and effacing E. coli (eae) gene, determined by a multiplex PCR assay. We show that the major seven STEC strains (O serogroups O26, O45, O103, O111, O121, O145, and O157) are not detected in any of the analyzed beef cut samples; however, three of them presented the virulence eae gene. Therefore, the absence of STEC strains in the beef samples may be an indication of the low prevalence of this pathogen in the cattle herd on the farm, associated with good hygiene and handling practices adopted by the meat industry.

Assessment of the Microbiological Quality and Safety of Unpasteurized Milk Cheese for Sale in England between 2019 and 2020

Cheese made with unpasteurized milk has been associated with outbreaks of illness. However, there are limited data on the prevalence of Shiga toxin-producing Escherichia coli (STEC) in these products and a lack of clarity over the significance of E. coli as a general indicator of hygiene in raw milk cheeses. The aim of this study was to provide further data to address both of these issues, as well as assessing the overall microbiological quality of raw milk cheeses available to consumers in England. A total of 629 samples of cheese were collected from retailers, catering premises, and manufacturers throughout England. The majority (80%) were made using cow's milk, with 14% made from sheep's milk and 5% from goat's milk. Samples were from 18 different countries of origin, with the majority originating from either the United Kingdom (40%) or France (35%). When interpreted against European Union microbiological criteria and United Kingdom guidance, 82% were considered to be of satisfactory microbiological quality, 5% were borderline, and 12% were unsatisfactory. Four samples (0.6%) were potentially injurious to health due to the isolation of STEC from one, >104 CFU/g of coagulase-positive staphylococci in two, and >100 CFU/g of Listeria monocytogenes in the fourth sample. Indicator E. coli and Listeria species were detected more frequently in soft compared with hard cheese. Higher levels of indicator E. coli were significantly associated with a greater likelihood of detecting Shiga toxin genes (stx1 and/or stx2).

Improved Genomic Identification, Clustering, and Serotyping of Shiga Toxin-Producing Escherichia coli Using Cluster/Serotype-Specific Gene Markers.

Shiga toxin-producing Escherichia coli (STEC) have more than 470 serotypes. The well-known STEC O157:H7 serotype is a leading cause of STEC infections in humans. However, the incidence of non-O157:H7 STEC serotypes associated with foodborne outbreaks and human infections has increased in recent years. Current detection and serotyping assays are focusing on O157 and top six (“Big six”) non-O157 STEC serogroups. In this study, we performed phylogenetic analysis of nearly 41,000 publicly available STEC genomes representing 460 different STEC serotypes and identified 19 major and 229 minor STEC clusters. STEC cluster-specific gene markers were then identified through comparative genomic analysis. We further identified serotype-specific gene markers for the top 10 most frequent non-O157:H7 STEC serotypes. The cluster or serotype specific gene markers had 99.54% accuracy and more than 97.25% specificity when tested using 38,534 STEC and 14,216 non-STEC E. coli genomes, respectively. In addition, we developed a freely available in silico serotyping pipeline named STECFinder that combined these robust gene markers with established E. coli serotype specific O and H antigen genes and stx genes for accurate identification, cluster determination and serotyping of STEC. STECFinder can assign 99.85% and 99.83% of 38,534 STEC isolates to STEC clusters using assembled genomes and Illumina reads respectively and can simultaneously predict stx subtypes and STEC serotypes. Using shotgun metagenomic sequencing reads of STEC spiked food samples from a published study, we demonstrated that STECFinder can detect the spiked STEC serotypes, accurately. The cluster/serotype-specific gene markers could also be adapted for culture independent typing, facilitating rapid STEC typing. STECFinder is available as an installable package (https://github.com/LanLab/STECFinder) and will be useful for in silico STEC cluster identification and serotyping using genome data.

Isolation and Characterization of Shiga-toxigenic <i>Escherichia coli</i> Isolated from Various Food Samples

Shiga toxigenic Escherichia coli (STEC) is a food-borne pathogen associated with community health concern. Contamination of food by STEC and toxin causes infectious diseases. Escherichia coli isolated from food samples such as yogurt, apples, tomatoes and milk by applying violet-red bile glucose agar, Triphenyl tetrazolium chloride agar (TTC) and eosin-methylene blue agar (EMB) test after pre-enrichment of food samples in buffered peptone water. Escherichia coli isolates were screened for the existence of shiga toxin genes Stx1 and Stx2 through Polymerase Chain Reaction (PCR). Apples (17%), tomatoes (26%) and milk (1%) of the samples examined showed positive results for Stx genes. Yoghurt samples examined in this study had no Stx1 or Stx2 gene. The presence of Shiga toxin that is protein in nature was confirmed by SDS-PAGE in milk sample. The present study has shown that foods from densely populated areas i.e. Harbanspura, Lahore could be sources of STEC.

Molecular Detection of Certain Virulence Genes of Avian Pathogenic Escherichia coli (APEC) in Chicken

In the present study, 86 avian pathogenic E. coli isolates from chicken were subjected to multiplex PCR to detect the presence of virulence genes stx1, stx2, eaeA, and hlyA. Forty-seven isolates harbored one or more virulence genes, stx1, stx2, and eaeA while gene hlyA was not detected in any of the samples. Of these, 15 (17.44%) isolates harbored stx1gene, 11 (12.79%) isolates carried stx2 gene, 18 (20.93%) isolates were positive for both stx1 and stx2 genes and 3 (3.49%) isolates carried eaeA gene.

Detection of SHIGA-TOXIN producing E. coli in some retail markets in Egypt using qPCR assay with special reference to serotyping

Background: Shiga toxin-producing Escherichia coli are dangerous foodborne pathogens that represent a severe public health issue worldwide. Raw foods are considered an important source of STEC infection in humans. Objective: In the current study, STEC contamination was investigated in 80 raw foods (chicken, beef, and milk) and water collected from different localities of Greater Cairo using Real-Time PCR. Moreover, the virulence genes of isolates were characterized. Results: STEC was detected in eight samples, (7 beef and 1 chicken), which represent about 10% of the tested samples. Positive beef and chicken samples show the presence of 13 and 2 STEC genes, respectively. Five samples were positive for eaeA (intemin), two samples were positive for stx1 gene and eight samples were positive for stx2 gene. Beef samples reveal the highest incidence of virulence gene stx2 (35%), followed by eaeA (20%), then stx1 (10%). The incidence of STEC was lower in chicken samples and the prevalence of virulence gene was 5% for stx2 and eaeA, respectively. Stx2 gene was the most prevalent subtype identified in beef samples. Serotyping of isolated STEC strains (14) revealed isolation of seven STEC strains belong to O157 serogroup, two strains belong to O111and five strains belong to serogroup O26. Conclusion: The current study concluded that recovery of STEC from raw chicken and beef samples is of important concern.

Prevalence of Verotoxigenic E.Coli in Locally Manufactured Dairy Products

Verotoxigenic E.coli (VTEC) is one of the emerging pathogens worldwide. The aim of this study was to investigate the prevalence of VTEC among One hundred from different dairy products samples (Kareish cheese and plain yogurt) were collected randomly from street-vendors, supermarkets and Small dairies at Alexandria City, Egypt. (50 of each). The Prevalence of Verotocytotoxigenic Escherichia coli in examined locally manufactured dairy products samples using Sorbitol MacConky agar give rise for 24% (6/25), 0% (0/25) and 12% (3/25), 12% (3/25) positive samples from street-vendor and supermarket Kareish cheese, ,and Supermarket Plain yogurt and Small dairies Plain yogurt, respectively while, its prevalence based on molecular identification revealed that only 8% (2/25) positive samples from street-vendor Kareish cheese. E.coli isolated from street-vendor Kareish cheese could be serologically identified into E.coli O26, O55 and O127, while that isolated from plain yoghurt samples into O55 and O86a. All screened E.coli strains had a determinant gene (phoA) while, stx1 and stx2 genes could be detected only in of selected street-vended Kareish cheese isolates., meanwhile both virulence genes could not be detected in all selected small dairies plain yoghurt isolates. On contrary, virulence gene eaeA could be detected in all test E.Coli isolates from street-vendor Kareish cheese and small dairies plain yoghurt. Prevalence of Verotoxigenic E.coli (VTEC) poses public health threats to consumers. Food safety concept should be implemented for all steps through manufacture in order to produce safe and good quality dairy products.

Molecular Characterization of Virulence and Antimicrobial Resistance Genes of E. coli Isolated from Different Animal Sources

The aim of the current study was to characterize E. coli isolates belonging to several serotypes in samples from different animal sources. The genetic diversity among the isolates was evaluated by multiplex PCR. Out of 30 suspected isolated E. coli which were identified microscopically, culturally and biochemically examined by VITEK2 and serologically identified there were 8 isolates showing low discrimination for E. coli and were non-typeable while 22 isolates positive, showing excellent level of identification for E. coli and were identified serologically. Thirteen different serogroups were identified, O26:H11, O91:H21, O78, O111:H2, O125:H21, O146:H21, O1:H7, O44:H18, O55:H7, O119:H6, O121:H7, O128:H2 and O159. Findings of multiplex PCR of virulence genes showed that the eaeA gene was successfully amplified in O26:H11; stx1 gene was successfully amplified in O26:H11, O55:H7, O91:H21, O111:H2, O119:H6, O128:H2 and O146:H21 serogroups but stx2 gene was successfully amplified in O1:H7, O26:H11, O78, O91:H21, O111:H2, O119:H6 and O125:H21. O91:H21, O111:H2 and O119:H6 serogroups carried both (stx1 and stx 2) genes, O26:H11 serogroup carried (stx 1, stx 2 and eae) genes and O44:H18, O121:H7 and O159 serogroups were negative for these genes. Findings of multiplex PCR of β-lactamase antimicrobial resistance genes showed that blaOXA gene was successfully amplified in O26:H11 only; blaCTX-M1 gene was successfully amplified in O78 and O121:H7 serogroups but blaTEM gene was successfully amplified in O26:H11, O91:H21, O111:H2 and O125: H21. O26:H11 serogroup carried both (blaOXA and blaTEM) genes and O1: H7, O44:H18, O55:H7, O119:H6, O128:H2, O146:H21 and O159 serogroups were negative for (blaOXA, blaCTX-M and blaTEM) genes. In conclusion, animals and animal products could be a significant reservoir of diverse E. coli virulence and antimicrobial resistance genes which is potentially hazardous to public health.

Bloody Diarrhea and Shiga Toxin Screening

Source: Ardissino G, Vignati C, Masia C, et al. Bloody diarrhea and Shiga toxin-producing Escherichia coli hemolytic uremic syndrome in children: data from the ItalKid-HUS network. J Pediatr. 2021; 237:34-40. e1; doi: 10.1016/j.jpeds.2021.06.048Investigators from multiple institutions conducted an analysis of data from a screening program for Shiga toxin (Stx) in children with bloody diarrhea. The screening program was developed for identification of children with Shiga toxin-Escherichia coli (STEC) infection for early diagnosis of STEC hemolytic uremic syndrome (STEC-HUS) by the ItalKid-HUS Network. The network includes 63 pediatric units in Northern Italy. Stool samples in children with acute bloody diarrhea were tested for Stx genes (Stx1 and Stx2). Patients who were Stx gene positive were monitored for hemoglobinuria as an early sign of STEC-HUS. Main study outcomes included rate of Stx positivity among children with bloody diarrhea and development of STEC-HUS. Criteria for a diagnosis of STEC-HUS were platelet count <150,000 mm3 or 50% acute reduction in platelet count, non-immune-mediated hemolysis, and evidence of renal damage (elevated serum creatinine [Cr], proteinuria, or hematuria). The utility of hemoglobinuria as a sign of STEC-HUS in children with Stx gene-positive bloody diarrhea was determined. Finally, the analysis also included children diagnosed with ongoing STEC-HUS who were not screened for Stx. Laboratory findings and long-term adverse outcomes (chronic kidney disease [CKD], extrarenal sequelae, or death) in children diagnosed through the screening program were compared to those diagnosed with ongoing STEC-HUS with chi square tests.A total of 4,767 children with bloody diarrhea were screened for the Stx gene from 2010 through 2019. The median age of these children was 3.4 years. Among the 4,767 patients screened, 214 (4.5%) were positive for Stx, including 62 with Stx1, 97 with Stx2, and 55 with both genes. In the 214 children who were Stx positive, 34 developed STEC-HUS (15.9% of Stx positive patients and 0.71% of screened children with bloody diarrhea). All of those who developed STEC-HUS were positive for Stx2. Overall, the risk of STEC-HUS was 0% for those with Stx1, 23.7% for those with Stx2, and 12.7% for those with both. In 63 patients with Stx2 who were screened, the sensitivity of hemoglobinuria as a marker for STEC-HUS was 100% (95% CI, 95%, 100%). specificity was 85% (95% CI, 77%, 91%), positive predictive value was 68% (95% CI, 55%, 79%), and negative predictive value was 100% (95% CI, 93%, 100%). There were an additional 95 children diagnosed with ongoing STEC-HUS. These children had a significantly higher Cr at HUS diagnosis than those identified through the screening program; the rate of long-term adverse outcomes was 15.8% in these patients vs 2.9% in those screened (P = 0.051).The authors conclude that 0.71% of children with bloody diarrhea who were screened for Stx developed STEC-HUS.Dr Rosenthal has disclosed no financial relationship relevant to this commentary. This commentary does not contain a discussion of an unapproved/investigative use of a commercial product/device.Bloody diarrhea dictates an evaluation of its cause. Among the most common causes in Western countries, STEC has been associated with HUS.1 The current researchers screened 4,767 children with bloody diarrhea for Stx; 214 were positive for Stx genes; 62 with Stx1, 97 with Stx2, and 55 with both genes. Notably, STEC-HUS developed only in children with Stx2 alone or in combination. In 63 patients with Stx2 who were screened, the sensitivity of hemoglobinuria as a marker for STEC-HUS was 100%.HUS is associated with a nonimmune hemolytic anemia, low platelet count, renal impairment, and multiorgan injury.1 The anemia is severe and microangiopathic, with fragmented red blood cells in the peripheral smear, circulating free hemoglobin, and elevated reticulocytes. Platelet count is <60,000/mm3 in most cases. Acute renal failure occurs in 55-70% of cases.2In 70% of cases in North America, STEC-HUS is secondary to infection with the E. coli serotype O157:H7.3 This serotype does not ferment sorbitol, making it distinguishable from most other fecal E. coli.4Unfortunately, there is no treatment of confirmed value for STEC-HUS, and care during the acute phase of the illness remains supportive. The use of antibiotics is controversial.5What have we learned from the current report? Among children with bloody diarrhea, STEC-HUS occurred only in those with Stx2 alone or in combination with Stx 1. Monitoring STEC patients with Stx2 with urine dipstick for hemoglobinuria can aid in recognizing renal complications early to optimize supportive care.Screen patients with bloody diarrhea for Stx, and if Stx2 positive, monitor for hemoglobinuria.STEC-HUS remains the leading cause of acute kidney injury in young children.6 Of import, early administration of intravenous fluids after timely diagnosis of STEC infection (sought by the current investigators) appears to decrease disease severity. (See AAP Grand Rounds. 2016;[5]:50.)6

Virulence Genes and Antimicrobial Resistance Profile of Escherichia Coli Isolates from Diarrhoeic Neonatal Dromedary Camels

In the present study, Escherichia coli were isolated from rectal swabs of total 48 (20.68%) diarrhoeic neonatal dromedary camels during the five years study period in an organised farm. The PCR for amplification of virulence genes revealed that 31 (64.58%) isolates harboured at least one virulence gene. The detection rates of stx1, stx2, eae, F41, K99 and sta virulence genes were 4.16%, 2.08%, 35.41%, 14.58%, 18.75% and 16.66%, respectively. Based on occurrence of these virulence genes the isolates were pathotyped into shigatoxigenic E. coli (STEC) (6.25%), enteropathogenic E. coli (EPEC) (20.83%) and enterotoxigenic E. coli (ETEC) (29.16%). Atypical combinations of EPEC+ETEC (8.33%) were also detected. The E. coli isolates from all three neonatal camels having acute haemorrhagic enteritis and mortality were found to be of STEC type. In antibiotic sensitivity test, most prevalent resistance was observed against amoxicillin, cloxacillin, erythromycin and lincomycin whereas lowest resistance was observed against gentamicin and amikacin. Findings of this study indicate that neonatal camels are the probable reservoir of multidrug resistant and zoonotic STEC. Young age (below 7 days), housing system with loose sandy ground and winter season were identified as important risk factors for high incidence of neonatal camel calf diarrhoea in the present study.

Molecular detection and phylogenetic analysis of a Shiga toxin-producing strain Escherichia coli (partial rfbE and fliCh7 gene), serotype O157:H7 isolated from a living chicken of a traditional market in Indonesia

The objective of this study was to identify a Shiga toxin-producing strain Escherichia coli partial rfbE and fliCh7 gene of O157:H7 isolated from a faeces sample collected from a live chicken in a traditional market in Bogor, Indonesia. The isolate was investigated using multiplex polymerase chain reaction (PCR) to detect stx1, stx2, rfbE, and fliCh7 gene of STEC O157:H7. Then, sequencing was applied to identify the antigen markers, which are the rfbE and fliCh7 genes. In the present study, the isolate which was obtained from a live chicken was successfully identified as STEC O157:H7 strain. This conclusion was based on multiplex PCR and a nucleotide sequence analysis. This pathogen was not only found in the live chicken, but it was further suggested that the rfbE and fliCh7 genes can be used as alternative targets for molecular identification of this pathogen.

Molecular characteristics of Escherichia coli from bulk tank milk in Korea.

BackgroundEscherichia coli, which causes subclinical or clinical mastitis in cattle, is responsible for transmitting antimicrobial resistance via human consumption of raw milk or raw milk products.ObjectivesThe objective of this study was to investigate the molecular characteristics of 183 E. coli from bulk tank milk of five different dairy factories in Korea.MethodsThe molecular characteristics of E. coli such as serogroup, virulence, antimicrobial resistance, and integron genes were detected using polymerase chain reaction and antimicrobial susceptibility were tested using the disk diffusion test.ResultsIn the distribution of phylogenetic groups, group D was the most prevalent (59.6%) and followed by group B1 (25.1%). The most predominant serogroup was O173 (15.3%), and a total of 46 different serotypes were detected. The virulence gene found most often was fimH (73.2%), and stx1, fimH, incC, fyuA, and iutA genes were significantly higher in isolates of phylogenetic group B1 compared to phylogenetic groups A, B2, and D (p < 0.05). Among 64 E. coli isolates that showed resistance to at least one antimicrobial, the highest resistance rate was observed for tetracyclines (37.5%). All 18 integron-positive E. coli carried the integron class I (int1) gene, and three different gene cassette arrangements, dfrA12+aadA2 (2 isolates), aac(6′)-Ib3+aac(6′)-Ib-cr+aadA4 (2 isolates), and dfrA17+aadA5 (1 isolate) were detected.ConclusionsThese data suggest that the E. coli from bulk tank milk can be an indicator for dissemination of antimicrobial resistance and virulence factors via cross-contamination.

Prevalence of Shiga toxin-producing Escherichia coli in Yezo sika deer (Cervus nippon yesoensis) in the Tokachi sub-prefecture of Hokkaido, Japan.

In food hygiene, the surveillance of foodborne pathogens in wild animals is indispensable because we cannot control hygienic status of them. Yezo sika deer (Cervus nippon yesoensis), which are found only on the island of Hokkaido, Japan, are the most common game animal in the country. In this study, we analyzed the incidence of Shiga toxin-producing Escherichia coli (STEC) in Yezo sika deer hunted in the Tokachi sub-prefecture, which is one of the densest zones for the sub-species. Real-time polymerase chain reaction testing detected STEC in 18.3% of fecal samples (59/323) collected from deer hunted between 2016 and 2017, whereas no Shigella and Salmonella markers were detected. No correlation was found between STEC detection from fecal samples and characteristics of carcasses, such as hunting area, age, and fascioliasis. From 59 STEC-positive fecal samples, we isolated 37 STEC strains, including 34 O- and H-genotyped strains, in which 16 different serogroups were detected. Genetic analysis revealed that our isolates included various stx gene types (stx1+/stx2−, stx1+/stx2+, and stx1−/stx2+) and carried eae. This study demonstrated that STEC strains with various features colonized the Yezo sika deer, similar to other subspecies of sika deer. We conclude that continuous surveillance activity is important to monitor the suitability of game animals as a food source and to assess the validity of the food safety management system for game meat production.