Reveal®E. coli 2.0 Method for Detection of Escherichia coli O157:H7 in Raw Beef

2011 ◽  
Vol 94 (6) ◽  
pp. 1835-1845 ◽  
Author(s):  
Rebecca Hoerner ◽  
Jill Feldpausch ◽  
R Lucas Gray ◽  
Stephanie Curry ◽  
Paul Lewis ◽  
...  
Keyword(s):  
Test Performance ◽  
Test Procedure ◽  
Ground Beef ◽  
Test Sample ◽  
Sample Volume ◽  
Method Performance ◽  
E Coli ◽  
Test Specificity ◽  
Inspection Service ◽  
Positive Results

Abstract Reveal®E. coli 2.0 is a new lateral-flow immunodiagnostic test for detection of E. coli O157:H7 and O157:NM in raw beef trim and ground beef. Compared with the original Reveal E. coli O157:H7 assay, the new test utilizes a unique antibody combination resulting in improved test specificity. The device architecture and test procedure have also been modified, and a single enrichment protocol was developed which allows the test to be performed at any point during an enrichment period of 12 to 20 h. Results of inclusivity and exclusivity testing showed that the test is specific for E. coli serotypes O157:H7 and O157:NM, with the exception of two strains of O157:H38 and one strain of O157:H43 which produced positive reactions. In internal and independent laboratory trials comparing the Reveal 2.0 method to the U.S. Department of Agriculture–Food Safety and Inspection Service reference culture procedure for detection of E. coli O157:H7 in 65 and 375 g raw beef trim and ground beef samples, there were no statistically significant differences in method performance with the exception of a single internal trial with 375 g ground beef samples in which the Reveal method produced significantly more positive results. There were no unconfirmed positive results by the Reveal assay, for specificity of 100%. Results of ruggedness testing showed that the Reveal test produces accurate results even with substantial deviation in sample volume or device incubation time or temperature. However, addition of the promoter reagent to the test sample prior to introducing the test device is essential to proper test performance.

Validation of the ANSR®E. coli O157:H7 Method for Detection of E. coli O157:H7

2016 ◽  
Vol 99 (3) ◽  
pp. 705-716 ◽  
Author(s):  
Ryan Viator ◽  
Susan Alles ◽  
Quynh-Nhi Le ◽  
Edan Hosking ◽  
Evan Meister ◽  
...  
Keyword(s):  
Target Genes ◽  
Ground Beef ◽  
Reference Method ◽  
Single Step ◽  
Fluorescence Detector ◽  
Method Performance ◽  
E Coli ◽  
Reference Methods ◽  
Inspection Service ◽  
The U.S

Abstract A performance validation of the ANSR® for E. coli O157:H7 method was conducted in selected food matrixes. This assay uses selective nicking enzyme amplification technology to amplify target genes. Samples are enriched for 12–24 h and then lysed. The assay is completed within 40 min using real-time detection in a combination incubator/fluorescence detector and software. When 44 distinct strains of Escherichia coli O157:H7 and 6 strains of E. coli O157:NM were tested for inclusivity, all 50 strains produced positive results. In exclusivity testing, 57 strains representing 33 species of closely related Gram-negative bacteria belonging to the Enterobacteriaceae family, including 11 non-H7 O157 strains and shiga toxin-producing E. coli other than O157:H7, were evaluated. All 57 nontarget strains generated negative ANSR assay results. Using 80% lean ground beef and beef trim (approximately 20% fat), ANSR method performance was compared to the U.S. Department of Agriculture, Food Safety and Inspection Service Microbiology Laboratory Guidebook reference culture procedure. ANSR performance with baby spinach and sprout irrigation water was measured against the U.S. Food and Drug Administration Bacteriological Analytical Manual reference method. ANSR method performance was not statistically different to that of the reference methods using two different enrichment options. For ground beef and beef trim, the standard enrichment in modified Tryptone Soya Broth can be analyzed using the ANSR assay with a 1:10 dilution of the enrichment in phosphate-buffered saline and produces equivalent results to the reference method. Additionally, in most matrixes tested (exception is spinach which required 24 h enrichment) the assay offers great efficiency and flexibility over the reference method with a 12–24 h single-step enrichment. Equivalent results were observed at both time points (12 and 24 h) to reference methods. Small changes to the assay parameters minimally affected ANSR method performance. Finally, accelerated stability results from three independently manufactured lots support a shelf-life of 6 months when stored at 4°C.

scholarly journals Reveal 8-Hour Test System for Detection of Escherichia coli O157:H7 in Raw Ground Beef, Raw Beef Cubes, and Iceberg Lettuce Rinse: Collaborative Study

2001 ◽  
Vol 84 (3) ◽  
pp. 719-736 ◽  
Author(s):  
Charles B Bird ◽  
Rebecca J Hoerner ◽  
Lawrence Restaino ◽  
G Anderson ◽  
W Birbari ◽  
...  
Keyword(s):  
Collaborative Study ◽  
Ground Beef ◽  
Culture Method ◽  
The United States ◽  
Test System ◽  
Private Industry ◽  
E Coli ◽  
Inspection Service ◽  
High Level ◽  
The U.S

Abstract Five different food types were analyzed by the Reveal for E. coli O157:H7 8-Hour Test System (Reveal 8) and either the U.S. Food and Drug Administration's Bacteriological Analytical Manual (BAM) culture method or the U.S. Department of Agriculture Food Safety Inspection Service (FSIS) culture method for the presence of E. coli O157:H7. A total of 27 laboratories representing academia and private industry in the United States and Canada participated. Food types were inoculated with E. coli O157:H7 at 2 different levels: a high level where predominantly positive results were expected, and a low level where fractional recovery was anticipated. During this study, 1110 samples and controls were analyzed by both the Reveal 8 and by BAM or FSIS by each of the collaborators (2220 samples in total). For each set of samples, 740 were artificially inoculated with E. coli O157:H7, and 370 were uninoculated controls. The Reveal 8 detected 528 presumptive positives of which 487 were confirmed positive by the BAM culture method. In comparison, BAM and FSIS detected 489 of the 740 artificially contaminated samples as positive. In an additional in-house study performed only on chilled and frozen raw ground beef, 240 artificially inoculated samples were analyzed by both the Reveal 8 and by FSIS. The Reveal 8 detected and confirmed 104 samples as positive compared to 79 confirmed positive by FSIS.

scholarly journals Detex for Detection of Escherichia coli O157 in Raw Ground Beef and Raw Ground Poultry

2001 ◽  
Vol 84 (3) ◽  
pp. 752-760 ◽  
Author(s):  
Yvette M Henry ◽  
Nandini Natrajan ◽  
Wendy F Lauer
Keyword(s):  
Escherichia Coli ◽  
False Positive Rate ◽  
Ground Beef ◽  
Escherichia Coli O157 ◽  
Department Of Agriculture ◽  
E Coli ◽  
Metal Plating ◽  
Positive Rate ◽  
Meat Samples ◽  
Inspection Service

Abstract A method for detection of Escherichia coli O157 in beef and poultry is presented. The method is antibody-based and uses a patented antibody-specific metal-plating procedure for the detection of E. coli O157 in enriched meat samples. Both raw ground beef and raw ground poultry were tested as matrixes for the organism. The sensitivity and specificity of the assay were 98 and 90%, respectively. The accuracy of the assay was 96%. Overall, the method agreement between the E. coli O157 Detex assay and the U.S. Department of Agriculture/Food Safety Inspection Service method was 96%. Sample temperature upon loading of the apparatus was critical to the observed false-positive rate of the system.

Food Safety and Inspection Service Regulatory Testing Program for Escherichia coli O157:H7 in Raw Ground Beef

2005 ◽  
Vol 68 (3) ◽  
pp. 462-468 ◽  
Author(s):  
ALECIA LAREW NAUGLE ◽  
KRISTIN G. HOLT ◽  
PRISCILLA LEVINE ◽  
RON ECKEL
Keyword(s):  
Escherichia Coli ◽  
Food Safety ◽  
Ground Beef ◽  
Fiscal Year ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Inspection Service ◽  
Rate Ratios ◽  
Log Linear ◽  
Regulatory Actions

We analyzed raw ground beef testing data to determine whether a decrease in the rate of Escherichia coli O157:H7–positive raw ground beef samples has occurred since the inception of Food Safety and Inspection Service (U.S. Department of Agriculture) regulatory actions and microbiological testing concerning this commodity and pathogen. A main effects log-linear Poisson regression model was constructed to evaluate the association between fiscal year and the rate of E. coli O157: H7–positive raw ground beef samples while controlling for the effect of season for the subset of test results obtained from fiscal year (FY)2000 through FY2003. Rate ratios were used to compare the rate of E. coli O157:H7–positive raw ground beef samples between sequential years to identify year-to-year differences. Of the 26,521 raw ground beef samples tested from FY2000 through FY2003, 189 (0.71%) tested positive for E. coli O157:H7. Year-to-year comparisons identified a 50% reduction in the rate of positive ground beef samples from FY2002 to FY2003 when controlling for season (95% CI, 10 to 72% decrease; P = 0.02). This decrease was the only significant year-to-year change in the rate of E. coli O157:H7–positive raw ground beef samples but was consistent in samples obtained from both federally inspected establishments and retail outlets. We believe this decrease is attributed to specific regulatory actions by Food Safety and Inspection Service and subsequent actions implemented by the industry, with the goal of reducing E. coli O157:H7 adulteration of raw ground beef. Continued monitoring is necessary to confirm that the decrease in the rate of E. coli O157:H7 in raw ground beef samples we observed here represents the beginning of a sustained trend.

Crystal Diagnostics MultiPath System™

2014 ◽  
Vol 97 (6) ◽  
pp. 1592-1600 ◽  
Author(s):  
Curtis H Stumpf ◽  
Weidong Zhao ◽  
Brian Bullard ◽  
Christine Ammons ◽  
Karl I Devlin ◽  
...  
Keyword(s):  
Liquid Crystal ◽  
Laboratory Tests ◽  
Ground Beef ◽  
Escherichia Coli O157 ◽  
Department Of Agriculture ◽  
E Coli ◽  
Independent Laboratory ◽  
Diagnostics System ◽  
Inspection Service ◽  
The U.S

Abstract The Crystal Diagnostics MultiPath System™ provides rapid detection of Escherichia coli O157 in fresh raw ground beef, raw beef trim, and spinach. The Crystal Diagnostics system combines patented Liquid Crystal technology with antibody-coated paramagnetic microspheres to selectively capture and detect E. coli O157 in food matrixes. This is the only liquid crystal-based biosensor commercially available for the detection of pathogens. The Crystal Diagnostics system expeditiously provides the sensitivity and accuracy of the U.S. Department of Agriculture Food Safety Inspection Service (USDA-FSIS) and the U.S. Food and Drug Administration Bacteriological Analytical Manual (FDA-BAM) methods for detecting as low as one CFU of E. coli O157 per 375 g of raw ground beef and raw beef trim, or 200 g of raw spinach. An internal inclusivity validation demonstrated detection of all 50 tested strains of E. coli O157. The internal and independent laboratory tests demonstrate that the method is rapid and sensitive for detecting of E. coli O157 in fresh raw ground beef, beef trim, and spinach.

Validation of a Modified ANSR® for E. coli O157:H7 Method for Detection of E. coli O157:H7 in Select Foods

2019 ◽  
Vol 102 (1) ◽  
pp. 96-107
Author(s):  
Susan Alles ◽  
Brooke Roman ◽  
Quynh-Nhi Le ◽  
Edan Hosking ◽  
Wesley Colangelo ◽  
...  
Keyword(s):  
Validation Study ◽  
Laboratory Testing ◽  
Ground Beef ◽  
Nucleic Acid Amplification ◽  
Method Performance ◽  
E Coli ◽  
Independent Laboratory ◽  
Accelerated Stability ◽  
Nucleic Acid Amplification Technology ◽  
The U.S

Abstract Background: The ANSR method is based on isothermal nucleic acid amplification technology. The modifications to assay components improve sensitivity of the assay and robustness of the internal positive control. Objective: A Performance Tested MethodSM validation study was conducted to assess performance of a modified version of the ANSR® for Escherichia coli O157:H7 method. Methods: The validation study included inclusivity/exclusivity, matrix, robustness, accelerated stability, and independent laboratory testing. Results: In inclusivity testing of 55 strains of E. coli O157:H7 and E. coli O157:NM variants, all strains produced positive results. In exclusivity testing of 41 strains including E. coli of other serotypes and bacteria of closely related genera, all strains produced negative results. In matrix testing of beef trim, raw ground beef, spinach, and sprout-irrigation water, ANSR method performance was compared with that of the U.S. Department of Agriculture Food Safety and Inspection Service Microbiology Laboratory Guidebook or the U.S. Food and Drug Administration Bacteriological Analytical Manual reference culture procedures. Conclusions: all trials, ANSR method performance was not statistically different from that of the reference methods. Results of independent laboratory testing of ground beef corroborated those of internal testing. Introducing modest changes to three assay operating parameters did not materially affect ANSR method performance. Finally, accelerated stability testing results of three independently manufactured lots of ANSR reagents support a shelf-life of 1 year when stored at 2–8°C.

A Novel Selective Medium for Simultaneous Enrichment of Shiga Toxin–Producing Escherichia coli and Salmonella in Ground Beef

2018 ◽  
Vol 81 (8) ◽  
pp. 1252-1257 ◽  
Author(s):  
JOSEPH EGGERS ◽  
JOELLEN M. FEIRTAG ◽  
ALAN D. OLSTEIN ◽  
JOSEPH M. BOSILEVAC
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Ground Beef ◽  
Performance Standards ◽  
Selective Medium ◽  
Microbiological Analysis ◽  
Department Of Agriculture ◽  
Selective Enrichment ◽  
E Coli ◽  
Inspection Service

ABSTRACT Microbiological analysis of ground beef for contamination by both Salmonella and Shiga toxin–producing Escherichia coli (STEC) is performed by the U.S. Department of Agriculture, Food Safety Inspection Service (FSIS), as part of its Performance Standards Verification Testing program. FSIS has established a zero tolerance for STEC serotype O157:H7 and serogroups O26, O45, O103, O111, O121, and O145 because they are regarded as adulterants. The detection and isolation of these specific serogroups presents a technical challenge necessitating time-consuming and costly laboratory procedures that often exceed the technical capabilities of many small internal and reference laboratories. We describe here a method using a novel STEC and Salmonella selective (SSS) broth that allows for simultaneous selective enrichment of STEC and Salmonella sp., providing isolation and detection from the same broth. The method only involves direct plating from beef enrichments to detect suspect isolates that can be easily confirmed by using immunoassays or PCR, rendering the isolation simpler and less costly than the current described methods. In a side-by-side comparison with modified tryptic soy broth (mTSB), the use of SSS broth resulted in primarily isolating STEC and Salmonella sp., while substantially suppressing the growth of other gram-negative Enterobacteriacae by 90%. Significantly more (χ2 < 3.84) samples containing E. coli O157:H7 and STEC O26, O111, O121, and O145 and a nondifferent (χ2 > 3.84) number of samples containing STEC O103 and O45 were identified when enriching in SSS broth. Coenrichment using six different Salmonella serovars showed numerically greater but not significant (χ2 < 3.84) positive samples by using SSS broth compared with mTSB for a majority of serotypes.

Methods Used for Detection and Recovery of Escherichia coli O157:H7 Associated with a Food-Borne Disease Outbreak†

1995 ◽  
Vol 58 (6) ◽  
pp. 597-603 ◽  
Author(s):  
J. L.JOHNSON ◽  
B. E. ROSE ◽  
A. K. SHARAR ◽  
G. M. RANSOM ◽  
C. P. LATTUADA ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Environmental Samples ◽  
Disease Outbreak ◽  
Screening Methods ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Food Borne ◽  
Quantitative Analyses ◽  
Inspection Service ◽  
Positive Results

The current Food Safety and Inspection Service method for detection and recovery of Escherichia coli O157:H7, (including modified EC broth with novobiocin (mEC+n) and a direct blot ELISA). was used to analyze beef and environmental samples during an investigation of a food-borne disease outbreak attributed to consumption of undercooked hamburger patties. Double-modified trypticase soy broth (dmTSB) and a commercially available dipstick immunoassay were also used to improve detection/recovery of E. coli O157:H7. A total of 1,115 beef and environmental samples was screened with the direct blot ELISA and the dipstick immunoassay; 178 presumptive-positive samples (by either or both of the screening methods) were subjected to recovery/isolation procedures. Toxigenic E. coli O157:H7 was recovered from 45 samples: 40 hamburger-patty samples produced on the epidemiologically identified date, 3 hamburger-patty samples produced on another date, and 2 beef briskets. The organism was not recovered from environmental samples. Limited quantitative analyses indicated that contaminated hamburger patties contained fewer than 4.3 CFU of E. coli O157:H7 per g. Atypical, toxigenic ornithine decarboxylase–negative E. coli O157:H7 and nontoxigenic sorbitol-positive E. coli O157:H29 were also recovered. Both enrichment broths gave strong positive reactions with the two immunoassay screening methods, but E. coli O157:H7 was recovered more often from mEC+n broth than from dmTSB. Both screening methods gave positive results for 44 of the 45 beef samples found to contain E. coli O157:H7. False-positive results were frequently observed with both screening methods.

scholarly journals USDA FSIS, FDA BAM, and ISO Culture Methods BD BBL CHROMagar O157 Media

2009 ◽  
Vol 92 (4) ◽  
pp. 1118-1127 ◽  
Author(s):  
Vicki Ritter ◽  
Susan Kircher ◽  
Nancy Dick
Keyword(s):  
Ground Beef ◽  
Reference Method ◽  
Food Samples ◽  
Culture Methods ◽  
Chi Square ◽  
Apple Cider ◽  
E Coli ◽  
Inspection Service ◽  
Chi Square Analysis ◽  
Method Agreement

Abstract BBL CHROMagar O157 media (CO) was evaluated for detection of Escherichia coli O157:H7 in raw ground beef and unpasteurized apple cider. The recovery of E. coli O157:H7 on CO was compared to the U.S. Food and Drug Administration (FDA) Bacteriological Analytical Manual (BAM), U.S. Department of Agriculture (USDA) Food Safety and Inspection Service (FSIS), and International Organization for Standardization (ISO) reference-plated media using the recommended enrichment broths. Of the 180 food samples tested, 45 were tested using BAM, 45 using the USDA method, and 90 using the ISO method. CO produced comparable results with the reference methods on all matrixes with a sensitivity of 100 and a specificity of 100. No false negatives were found in testing the food matrixes. There was no statistical difference in recovery based on Chi-square analysis. Method agreement for raw ground beef was 85 for the USDA FSIS method and 95 for the ISO method. Method agreement for unpasteurized apple cider was 100 for the ISO and FDA BAM methods. In all cases where method agreement was <100, CO detected more positives than the reference method media. Evaluation of known isolates on CO in inclusivity and exclusivity testing had a sensitivity and specificity of 100. The results of this study demonstrate that CO is an effective medium for the recovery and detection of E. coli O157:H7 in raw ground beef and unpasteurized apple cider using FDA BAM, USDA FSIS, and ISO methods.

Validation of Modifications to the ANSR® Listeria Method for Improved Ease of Use and Performance

2016 ◽  
Vol 99 (1) ◽  
pp. 98-111
Author(s):  
Oscar Caballero ◽  
Susan Alles ◽  
Quynh-Nhi Le ◽  
R Lucas Gray ◽  
Edan Hosking ◽  
...  
Keyword(s):  
Sample Volume ◽  
Ease Of Use ◽  
Stainless Steel Surface ◽  
Second Phase ◽  
Department Of Agriculture ◽  
Method Performance ◽  
Lysis Time ◽  
Accelerated Stability ◽  
And Performance ◽  
Inspection Service

Abstract A study was conducted to validate minor reagent formulation, enrichment, and procedural changes to the ANSR® Listeria method, Performance-Tested MethodSM 101202. In order to improve ease of use and diminish risk of amplicon contamination, the lyophilized reagent components were reformulated for increased solubility, thus eliminating the need to mix by pipetting. In the alternative procedure, an aliquot of the lysate is added to lyophilized ANSR reagents, immediately capped, and briefly mixed by vortexing. When three foods (hot dogs, Mexican-style cheese, and cantaloupe) and sponge samples taken from a stainless steel surface were tested, significant differences in performance between the ANSR and U.S. Food and Drug Administration Bacteriological Analytical Manual or U.S. Department of Agriculture, Food Safety and Inspection Service Microbiology Laboratory Guidebook reference culture procedures were seen with hot dogs and Mexican-style cheese after 16 h enrichment, with the reference methods producing more positive results. After 24 h enrichment, however, there were no significant differences in method performance for any of the four matrixes tested. Robustness testing was also conducted, with variations to lysis buffer volume, lysis time, and sample volume having no demonstrable effect on assay results. Accelerated stability testing was carried out over a 10-week period and showed no diminishment in assay performance. A second phase of the study examined performance of the ANSR assay following enrichment in a new medium, LESS Plus broth, designed for use with all food and environmental sample types. With the alternative LESS Plus broth, there were no significant differences in performance between the ANSR method and the reference culture procedures for any of the matrixes tested after either 16 or 24 h enrichment, although 24 h enrichment is recommended for hot dogs due to higher sensitivity. Results of inclusivity and exclusivity testing using LESS Plus broth showed that the ANSR assay is highly specific, with 100% expected results for target and nontarget bacteria.

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