scholarly journals Performance of the ImmunoCard STAT! E. coli O157:H7 Test for Detection of Escherichia coli O157:H7 in Stools

2000 ◽  
Vol 38 (5) ◽  
pp. 1866-1868 ◽  
Author(s):  
Andrew Mackenzie ◽  
Elaine Orrbine ◽  
Lucie Hyde ◽  
Michelle Benoit ◽  
Frank Chan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
High Sensitivity ◽  
False Positives ◽  
Broth Culture ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Stool Specimens ◽  
Multicenter Prospective Study ◽  
Culture Positive ◽  
Culture Negative

ImmunoCard STAT! E. coli O157:H7 (Meridian Diagnostics, Inc., Cincinnati, Ohio) is a novel rapid (10-min) test for the presence of Escherichia coli O157:H7 in stools. The test may be performed either directly on stool specimens or on an overnight broth culture of stool. In a multicenter prospective study, 14 of 14 specimens positive by culture for E. coli O157:H7 were positive by the ImmunoCard STAT! O157:H7 test, and there were no false positives from 263 culture-negative specimens. In a retrospective study, the test was positive in 339 (81%) of 417 stored culture-positive specimens and the specificity was 95% (98 of 103 specimens). No false positives were associated with alternate stool pathogens. The ImmunoCard STAT! O157:H7 test has high sensitivity and specificity.

scholarly journals Escherichia coli O157:H7 in Environments of Culture-Positive Cattle

2005 ◽  
Vol 71 (11) ◽  
pp. 6816-6822 ◽  
Author(s):  
Margaret A. Davis ◽  
Karen A. Cloud-Hansen ◽  
John Carpenter ◽  
Carolyn J. Hovde
Keyword(s):  
Escherichia Coli ◽  
Room Temperature ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Different Temperatures ◽  
Farm Structures ◽  
On Farm ◽  
Culture Positive ◽  
Culture Negative ◽  
Over Time

ABSTRACT Outbreaks of Escherichia coli O157:H7 disease associated with animal exhibits have been reported with increasing frequency. Transmission can occur through contact with contaminated haircoats, bedding, farm structures, or water. We investigated the distribution and survival of E. coli O157:H7 in the immediate environments of individually housed, experimentally inoculated cattle by systematically culturing feed, bedding, water, haircoat, and feed bunk walls for E. coli O157:H7 for 3 months. Cedar chip bedding was the most frequently culture-positive environmental sample tested (27/96 or 28.15%). Among these, 12 (44.0%) of positive bedding samples were collected when the penned animal was fecal culture negative. Survival of E. coli O157:H7 in experimentally inoculated cedar chip bedding and in grass hay feed was determined at different temperatures. Survival was longest in feed at room temperature (60 days), but bacterial counts decreased over time. The possibility that urine plays a role in the environmental survival of E. coli O157:H7 was investigated. Cedar chip bedding moistened with sterile water or bovine urine was inoculated with E. coli O157:H7. Bedding moistened with urine supported growth of E. coli O157:H7, whereas inoculated bedding moistened with only water yielded decreasing numbers of bacteria over time. The findings that environmental samples were frequently positive for E. coli O157:H7 at times when animals were culture negative and that urine provided a substrate for E. coli O157:H7 growth have implications for understanding the on-farm ecology of this pathogen and for the safety of ruminant animal exhibits, particularly petting zoos and farms where children may enter animal pens.

scholarly journals PCR Detection of Escherichia coli O157:H7 Directly from Stools: Evaluation of Commercial Extraction Methods for Purifying Fecal DNA

2000 ◽  
Vol 38 (11) ◽  
pp. 4108-4113 ◽  
Author(s):  
J. L. Holland ◽  
L. Louie ◽  
A. E. Simor ◽  
M. Louie
Keyword(s):  
Escherichia Coli ◽  
False Negative ◽  
Extraction Methods ◽  
Rapid Identification ◽  
E Coli ◽  
Stool Specimens ◽  
Commercial Kits ◽  
Dna Template ◽  
Culture Positive ◽  
Culture Negative

Rapid identification of Escherichia coli O157:H7 is important for patient management and for prompt epidemiological investigations. We evaluated one in-house method and three commercially available kits for their ability to extract E. coli O157:H7 DNA directly from stool specimens for PCR. Of the 153 stool specimens tested, 107 were culture positive and 46 were culture negative. The sensitivities and specificities of the in-house enrichment method, IsoQuick kit, NucliSens kit, and QIAamp kit were comparable, as follows: 83 and 98%, 85 and 100%, 74 and 98%, and 86 and 100%, respectively. False-negative PCR results may be due to the presence of either inherent inhibitors or small numbers of organisms. The presence of large amounts of bacteria relative to the amount of the E. coli O157:H7 target may result in the lower sensitivities of the assays. All commercial kits were rapid and easy to use, although DNA extracted with the QIAamp kit did not require further dilution of the DNA template prior to PCR.

Comparing Real-Time and Conventional PCR to Culture-Based Methods for Detecting and Quantifying Escherichia coli O157 in Cattle Feces

2014 ◽  
Vol 77 (2) ◽  
pp. 314-319 ◽  
Author(s):  
M. E. JACOB ◽  
J. BAI ◽  
D. G. RENTER ◽  
A. T. ROGERS ◽  
X. SHI ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Real Time ◽  
Positive Culture ◽  
Feedlot Cattle ◽  
Escherichia Coli O157 ◽  
Conventional Pcr ◽  
Culture Methods ◽  
E Coli ◽  
Cattle Feces ◽  
Culture Negative

Detection of Escherichia coli O157 in cattle feces has traditionally used culture-based methods; PCR-based methods have been suggested as an alternative. We aimed to determine if multiplex real-time (mq) or conventional PCR methods could reliably detect cattle naturally shedding high (≥104 CFU/g of feces) and low (~102 CFU/g of feces) concentrations of E. coli O157. Feces were collected from pens of feedlot cattle and evaluated for E. coli O157 by culture methods. Samples were categorized as (i) high shedders, (ii) immunomagnetic separation (IMS) positive after enrichment, or (iii) culture negative. DNA was extracted pre- and postenrichment from 100 fecal samples from each category (high shedder, IMS positive, culture negative) and subjected to mqPCR and conventional PCR assays based on detecting three genes, rfbE, stx1, and stx2. In feces from cattle determined to be E. coli O157 high shedders by culture, 37% were positive by mqPCR prior to enrichment; 85% of samples were positive after enrichment. In IMS-positive samples, 4% were positive by mqPCR prior to enrichment, while 43% were positive after enrichment. In culture-negative feces, 7% were positive by mqPCR prior to enrichment, and 40% were positive after enrichment. The proportion of high shedder–positive and culture-positive (high shedder and IMS) samples were significantly different from mqPCR-positive samples before and after enrichment (P < 0.01). Similar results were observed for conventional PCR. Our data suggest that mqPCR and conventional PCR are most useful in identifying high shedder animals and may not be an appropriate substitute to culture-based methods for detection of E. coli O157 in cattle feces.

scholarly journals Use of In Vivo-Induced Antigen Technology for Identification of Escherichia coli O157:H7 Proteins Expressed during Human Infection

2005 ◽  
Vol 73 (5) ◽  
pp. 2665-2679 ◽  
Author(s):  
Manohar John ◽  
Indira T. Kudva ◽  
Robert W. Griffin ◽  
Allen W. Dodson ◽  
Bethany McManus ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Vaccine Development ◽  
Human Infection ◽  
Escherichia Coli O157 ◽  
Standard Laboratory ◽  
E Coli ◽  
Phase Culture ◽  
Stool Specimens

ABSTRACT Using in vivo-induced antigen technology (IVIAT), a modified immunoscreening technique that circumvents the need for animal models, we directly identified immunogenic Escherichia coli O157:H7 (O157) proteins expressed either specifically during human infection but not during growth under standard laboratory conditions or at significantly higher levels in vivo than in vitro. IVIAT identified 223 O157 proteins expressed during human infection, several of which were unique to this study. These in vivo-induced (ivi) proteins, encoded by ivi genes, mapped to the backbone, O islands (OIs), and pO157. Lack of in vitro expression of O157-specific ivi proteins was confirmed by proteomic analysis of a mid-exponential-phase culture of E. coli O157 grown in LB broth. Because ivi proteins are expressed in response to specific cues during infection and might help pathogens adapt to and counter hostile in vivo environments, those identified in this study are potential targets for drug and vaccine development. Also, such proteins may be exploited as markers of O157 infection in stool specimens.

scholarly journals Effect of Forage or Grain Diets with or without Monensin on Ruminal Persistence and Fecal Escherichia coli O157:H7 in Cattle

2004 ◽  
Vol 70 (9) ◽  
pp. 5336-5342 ◽  
Author(s):  
M. J. Van Baale ◽  
J. M. Sargeant ◽  
D. P. Gnad ◽  
B. M. DeBey ◽  
K. F. Lechtenberg ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Nalidixic Acid ◽  
Escherichia Coli O157 ◽  
Fecal Samples ◽  
Fecal Shedding ◽  
E Coli ◽  
Culture Positive ◽  
Forage Diets

ABSTRACT Twelve ruminally cannulated cattle, adapted to forage or grain diet with or without monensin, were used to investigate the effects of diet and monensin on concentration and duration of ruminal persistence and fecal shedding of E. coli O157:H7. Cattle were ruminally inoculated with a strain of E. coli O157:H7 (1010 CFU/animal) made resistant to nalidixic acid (Nalr). Ruminal and fecal samples were collected for 11 weeks, and then cattle were euthanized and necropsied and digesta from different gut locations were collected. Samples were cultured for detection and enumeration of Nalr E. coli O157:H7. Cattle fed forage diets were culture positive for E. coli O157:H7 in the feces for longer duration (P < 0.05) than cattle fed a grain diet. In forage-fed cattle, the duration they remained culture positive for E. coli O157:H7 was shorter (P < 0.05) when the diet included monensin. Generally, ruminal persistence of Nalr E. coli O157:H7 was not affected by diet or monensin. At necropsy, E. coli O157:H7 was detected in cecal and colonic digesta but not from the rumen. Our study showed that cattle fed a forage diet were culture positive longer and with higher numbers than cattle on a grain diet. Monensin supplementation decreased the duration of shedding with forage diet, and the cecum and colon were culture positive for E. coli O157:H7 more often than the rumen of cattle.

scholarly journals Comparison of Escherichia coli O157:H7 Antigen Detection in Stool and Broth Cultures to That in Sorbitol-MacConkey Agar Stool Cultures

2000 ◽  
Vol 38 (9) ◽  
pp. 3404-3406 ◽  
Author(s):  
Jennifer R. Stapp ◽  
Srdjan Jelacic ◽  
Yoo-Lee Yea ◽  
Eileen J. Klein ◽  
Marc Fischer ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antigen Detection ◽  
Detection Methods ◽  
Broth Culture ◽  
Agar Culture ◽  
E Coli ◽  
Direct Testing ◽  
Stool Specimens ◽  
Stool Cultures ◽  
Test Result

We evaluated the Meridian IC-STAT direct fecal and broth culture antigen detection methods with samples from children infected withEscherichia coli O157:H7 and correlated the antigen detection results with the culture results. Stools of 16 children who had recently had stool cultures positive for this pathogen (population A) and 102 children with diarrhea of unknown cause (population B) were tested with the IC-STAT device (direct testing). Fecal broth cultures were also tested with this device (broth testing). The results were correlated to a standard of the combined yield from direct culture of stools on sorbitol-MacConkey (SMAC) agar and culture of broth on SMAC agar. Eleven (69%) of the population A stool specimens yieldedE. coli O157:H7 when plated directly on SMAC agar. Two more specimens yielded this pathogen when the broth culture was similarly plated. Of these 13 stool specimens, 8 and 13 were positive by direct and broth testing (respective sensitivities, 62 and 100%). Compared to the sensitivity of a simultaneously performed SMAC agar culture, the sensitivity of direct testing was 73%. Three (3%) of the population B stool specimens contained E. coli O157:H7 on SMAC agar culture; one and three of these stool specimens were positive by direct and broth testing, respectively. The direct and broth IC-STAT tests were 100% specific with samples from children from population B. Direct IC-STAT testing of stools is rapid, easily performed, and specific but is insufficiently sensitive to exclude the possibility of infection with E. coli O157:H7. Performing the IC-STAT test with a broth culture increases its sensitivity. However, attempts to recover E. coli O157:H7 by culture should not be abandoned but, rather, should be increased when the IC-STAT test result is positive.

Escherichia coli O157:H7 Prevalence in Fecal Samples of Cattle from a Southeastern Beef Cow-Calf Herd

2003 ◽  
Vol 66 (10) ◽  
pp. 1778-1782 ◽  
Author(s):  
D. G. RILEY ◽  
J. T. GRAY ◽  
G. H. LONERAGAN ◽  
K. S. BARLING ◽  
C. C. CHASE
Keyword(s):  
Escherichia Coli ◽  
Control Point ◽  
Foodborne Pathogen ◽  
Beef Cows ◽  
Escherichia Coli O157 ◽  
Fecal Samples ◽  
E Coli ◽  
Critical Control Point ◽  
Cow Calf ◽  
Culture Positive

The proportion of fecal samples culture-positive for Escherichia coli O157:H7 was determined for samples collected from 296 beef cows on pasture in a single Florida herd in October, November, and December 2001. The overall proportion of samples that cultured positive was 0.03. The proportion of cows that were culture-positive on at least one occasion was 0.091. No effect of pregnancy status or nutritional regimen on the proportion of culture-positive samples for E. coli O157:H7 was detected. We detected a breed effect on the shedding of E. coli O157, with Romosinuano cows having a lower (P &lt; 0.01) proportion of samples culture-positive than Angus or Brahman cows. This difference might have resulted from the presence of confounding variables; however, it also might represent evidence of breed-to-breed genetic variation in E. coli O157 shedding. Further research is warranted to evaluate breed as a possible risk factor for shedding of this important foodborne pathogen. Further substantiated findings could indicate that breed is a cow-calf–level critical control point of E. coli O157:H7.

Effects of Farm Manure-Handling Practices on Escherichia coli O157 Prevalence in Cattle

1997 ◽  
Vol 60 (4) ◽  
pp. 363-366 ◽  
Author(s):  
DALE D. HANCOCK ◽  
DANIEL H. RICE ◽  
DONALD E. HERRIOTT ◽  
THOMAS E. BESSER ◽  
ERIC D. EBEL ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Dairy Herds ◽  
Escherichia Coli O157 ◽  
Forage Crops ◽  
Fecal Samples ◽  
Herd Prevalence ◽  
E Coli ◽  
High Prevalence ◽  
Handling Practices ◽  
Culture Positive

Thirty-six dairy herds in Idaho, Oregon, and Washington were selected on the basis of cattle housing and manure-handling practices. Approximately 60 fecal samples from heifers were collected monthly in each herd for 6 months and cultured for Escherichia coli O157. One hundred seventy-nine of 12,664 (1.41%) individual fecal samples from 27 of the 36 herds (75%) were culture positive for E. coli O157. Within-herd prevalence ranged from 0% to 5.5% with a strong clustering toward the lower end of this range. A tendency was observed for herds to maintain either a relatively low or high prevalence of E. coli O157. Prevalence of E. coli O157 was similar in herds which housed heifers in dry lots and on pasture with and without application of manure. Also, application of manure to cattle forage crops was not associated with the prevalence of E. coli O157 in dairy herds.

Detection of Escherichia coli O157:H7 in 10- and 25-Gram Ground Beef Samples with an Evanescent-Wave Biosensor with Silica and Polystyrene Waveguides

2002 ◽  
Vol 65 (4) ◽  
pp. 596-602 ◽  
Author(s):  
DANIEL R. DeMARCO ◽  
DANIEL V. LIM
Keyword(s):  
Escherichia Coli ◽  
Optical Fiber ◽  
Evanescent Wave ◽  
Ground Beef ◽  
False Positives ◽  
Evanescent Field ◽  
Escherichia Coli O157 ◽  
Fiber Probe ◽  
Fluorescent Signal ◽  
E Coli

A portable evanescent-wave fiber-optic biosensor was used to detect Escherichia coli O157:H7 in seeded 10- and 25-g ground beef samples. The biosensor works by launching light from a 635-nm laser diode into specially designed optical fiber probes, generating an evanescent field that extends approximately 1,000 nm from the fiber surface. Fluorescent molecules within the evanescent field are excited, and a portion of their emission recouples into the fiber probe. The return path emission is transported by an optical fiber to a photodiode within the biosensor that detects and quantifies the fluorescent signal. A sandwich immunoassay was performed on the fiber probes with cyanine 5 dye–labeled polyclonal anti–E. coli O157:H7 antibodies for generation of the specific fluorescent signal. Biotin-streptavidin interactions were used to attach polyclonal antiE. coli O157:H7 antibodies to the surface of the fiber probe. A centrifugation method was developed to obtain samples suitable for biosensor analysis from 10- and 25-g ground beef samples. The assay was shown to be sensitive and repeatable. One hundred percent correct identification of positive samples was demonstrated at 9.0 × 103 CFU/g for 25-g ground beef samples with silica waveguides and at 5.2 × 102 CFU/g for 10-g ground beef samples with polystyrene waveguides. The reaction was highly specific. No false positives were observed for 10-g ground beef samples not spiked with the pathogen. In addition, when samples were spiked with high concentrations of a variety of non–E. coli O157:H7 organisms, no false positives were observed. The method was rapid, with results being obtained within 25 min of sample processing.

scholarly journals Application of Bacteriophages To Control Intestinal Escherichia coli O157:H7 Levels in Ruminants

2006 ◽  
Vol 72 (8) ◽  
pp. 5359-5366 ◽  
Author(s):  
Haiqing Sheng ◽  
Hannah J. Knecht ◽  
Indira T. Kudva ◽  
Carolyn J. Hovde
Keyword(s):  
Escherichia Coli ◽  
Phage Therapy ◽  
Oral Treatment ◽  
Treatment Protocol ◽  
Escherichia Coli O157 ◽  
Lytic Bacteriophage ◽  
E Coli ◽  
Culture Positive ◽  
Phage Treatment

ABSTRACT A previously characterized O157-specific lytic bacteriophage KH1 and a newly isolated phage designated SH1 were tested, alone or in combination, for reducing intestinal Escherichia coli O157:H7 in animals. Oral treatment with phage KH1 did not reduce the intestinal E. coli O157:H7 in sheep. Phage SH1 formed clear and relatively larger plaques on lawns of all 12 E. coli O157:H7 isolates tested and had a broader host range than phage KH1, lysing O55:H6 and 18 of 120 non-O157 E. coli isolates tested. In vitro, mucin or bovine mucus did not inhibit bacterial lysis by phage SH1 or KH1. A phage treatment protocol was optimized using a mouse model of E. coli O157:H7 intestinal carriage. Oral treatment with SH1 or a mixture of SH1 and KH1 at phage/bacterium ratios ≥102 terminated the presence of fecal E. coli O157:H7 within 2 to 6 days after phage treatment. Untreated control mice remained culture positive for >10 days. To optimize bacterial carriage and phage delivery in cattle, E. coli O157:H7 was applied rectally to Holstein steers 7 days before the administration of 1010 PFU SH1 and KH1. Phages were applied directly to the rectoanal junction mucosa at phage/bacterium ratios calculated to be ≥102. In addition, phages were maintained at 106 PFU/ml in the drinking water of the phage treatment group. This phage therapy reduced the average number of E. coli O157:H7 CFU among phage-treated steers compared to control steers (P < 0.05); however, it did not eliminate the bacteria from the majority of steers.

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