Modification of the BAX®Salmonella Test Kit to Include a Hot Start Functionality (Modification of AOAC Official MethodSM 2003.09)

Abstract In 2010, the BAX® System PCR assay for Salmonella was modified to include a hot start functionality designed to keep the reaction enzyme inactive until PCR begins. To validate the assay's Official Methods of Analysis status to include this procedure modification, an evaluation was conducted on four food types that were simultaneously analyzed with the BAX System and either the U.S. Food and Drug Administration's Bacteriological Analytical Manual or the U.S. Department of Agriculture-Food Safety and Inspection Service Microbiology Laboratory Guidebook reference method for detecting Salmonella. Identical performance between the BAX System method and the reference methods was observed. Additionally, lysates were analyzed using both the BAX System Classic and BAX System Q7 instruments with identical results using both platforms for all samples tested. Of the 100 samples analyzed, 34 samples were positive for both the BAX System and reference methods, and 66 samples were negative by both the BAX System and reference methods, demonstrating 100% correlation. No instrument platform variation was observed. Additional inclusivity and exclusivity testing using the modified test kit demonstrated the test kit to be 100% accurate in evaluation of test panels of 352 Salmonella strains and 46 non-Salmonella strains.

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In-House Validation Study of the DuPont Qualicon BAX System Q7 Instrument with the BAX System PCR Assay for Salmonella (Modification of AOAC Official MethodSM 2003.09 and AOAC Research Institute Performance-Tested MethodSM 100201)

Journal of AOAC International ◽

10.1093/jaoac/92.3.989 ◽

2009 ◽

Vol 92 (3) ◽

pp. 989-994 ◽

Cited By ~ 1

Author(s):

George Tice ◽

Bridget Andaloro ◽

H Kirk White ◽

Lance Bolton ◽

Siqun Wang ◽

...

Keyword(s):

Validation Study ◽

Reference Method ◽

Pcr Assay ◽

Paired Samples ◽

Reference Methods ◽

Comparable Performance ◽

Aoac Official ◽

Inspection Service ◽

Positive Results ◽

The U.S

Abstract In 2006, DuPont Qualicon introduced the BAX<sup/> system Q7 instrument for use with its assays. To demonstrate the equivalence of the new and old instruments, a validation study was conducted using the BAX system PCR Assay for Salmonella, AOAC Official MethodSM 2003.09, on three food types. The foods were simultaneously analyzed with the BAX system Q7 instrument and either the U.S. Food and Drug Administration Bacteriological Analytical Manual or the U.S. Department of AgricultureFood Safety and Inspection Service Microbiology Laboratory Guidebook reference method for detecting Salmonella. Comparable performance between the BAX system and the reference methods was observed. Of the 75 paired samples analyzed, 39 samples were positive by both the BAX system and reference methods, and 36 samples were negative by both the BAX system and reference methods, demonstrating 100 correlation. Inclusivity and exclusivity for the BAX system Q7 instrument were also established by testing 50 Salmonella strains and 20 non-Salmonella isolates. All Salmonella strains returned positive results, and all non-Salmonella isolates returned a negative response.

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Evaluation of a Multiplex Real-Time PCR Method for Detecting Shiga Toxin–Producing Escherichia coli in Beef and Comparison to the U.S. Department of Agriculture Food Safety and Inspection Service Microbiology Laboratory Guidebook Method†

Journal of Food Protection ◽

10.4315/0362-028x.jfp-13-248 ◽

2014 ◽

Vol 77 (2) ◽

pp. 180-188 ◽

Cited By ~ 28

Author(s):

PINA M. FRATAMICO ◽

JAMIE L. WASILENKO ◽

BRADLEY GARMAN ◽

DANIEL R. DeMARCO ◽

STEPHEN VARKEY ◽

...

Keyword(s):

Real Time ◽

Real Time Pcr ◽

Virulence Genes ◽

Microbiology Laboratory ◽

Pcr Assay ◽

Department Of Agriculture ◽

E Coli ◽

System Method ◽

Inspection Service ◽

The U.S

The “top-six” non-O157 Shiga toxin–producing Escherichia coli (STEC) serogroups (O26, O45, O103, O111, O121, and O145) most frequently associated with outbreaks and cases of foodborne illnesses have been declared as adulterants in beef by the U.S. Department of Agriculture Food Safety and Inspection Service (FSIS). Regulatory testing in beef began in June 2012. The purpose of this study was to evaluate the DuPont BAX System method for detecting these top six STEC strains and strains of E. coli O157:H7. For STEC, the BAX System real-time STEC suite was evaluated, including a screening assay for the stx and eae virulence genes and two panel assays to identify the target serogroups: panel 1 detects O26, O111, and O121, and panel 2 detects O45, O103, O145. For E. coli O157:H7, the BAX System real-time PCR assay for this specific serotype was used. Sensitivity of each assay for the PCR targets was ≥1.23 × 103 CFU/ml in pure culture. Each assay was 100% inclusive for the strains tested (20 to 50 per assay), and no cross-reactivity with closely related strains was observed in any of the assays. The performance of the BAX System methods was compared with that of the FSIS Microbiology Laboratory Guidebook (MLG) methods for detection of the top six STEC and E. coli O157:H7 strains in ground beef and beef trim. Generally, results of the BAX System method were similar to those of the MLG methods for detecting non-O157 STEC and E. coli O157:H7. Reducing or eliminating novobiocin in modified tryptic soy broth (mTSB) may improve the detection of STEC O111 strains; one beef trim sample inoculated with STEC O111 produced a negative result when enriched in mTSB with 8 mg/liter novobiocin but was positive when enriched in mTSB without novobiocin. The results of this study indicate the feasibility of deploying a panel of real-time PCR assay configurations for the detection and monitoring of the top six STEC and E. coli O157:H7 strains in beef. The approach could easily be adapted for additional multiplex assays should regulations expand to include other O serogroups or virulence genes.

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Romer Labs RapidChek®Listeria monocytogenes Test System for the Detection of L. monocytogenes on Selected Foods and Environmental Surfaces

Journal of AOAC International ◽

10.5740/jaoacint.18-0035 ◽

2018 ◽

Vol 101 (5) ◽

pp. 1490-1507

Author(s):

Gregory Juck ◽

Verapaz Gonzalez ◽

Ann-Christine Olsson Allen ◽

Meredith Sutzko ◽

Kody Seward ◽

...

Keyword(s):

Listeria Monocytogenes ◽

Reference Method ◽

Test System ◽

Probability Of Detection ◽

Department Of Agriculture ◽

Detection Analysis ◽

Media System ◽

Reference Methods ◽

Environmental Surfaces ◽

Inspection Service

Abstract The Romer Labs RapidChek®Listeria monocytogenes test system (Performance Tested Method 011805) was validated against the U.S. Department of Agriculture-Food Safety and Inspection Service Microbiology Laboratory Guidebook (USDA-FSIS/MLG), U.S. Food and Drug Association Bacteriological Analytical Manual (FDA/BAM), and AOAC Official Methods of Analysis (AOAC/OMA) cultural reference methods for the detection of L. monocytogenes on selected foods including hot dogs, frozen cooked breaded chicken, frozen cooked shrimp, cured ham, and ice cream, and environmental surfaces including stainless steel and plastic in an unpaired study design. The RapidChek method uses a proprietary enrichment media system, a 44–48 h enrichment at 30 ± 1°C, and detects L. monocytogenes on an immunochromatographic lateral flow device within 10 min. Different L. monocytogenes strains were used to spike each of the matrixes. Samples were confirmed based on the reference method confirmations and an alternate confirmation method. A total of 140 low-level spiked samples were tested by the RapidChek method after enrichment for 44–48 h in parallel with the cultural reference method. There were 88 RapidChek presumptive positives. One of the presumptive positives was not confirmed culturally. Additionally, one of the culturally confirmed samples did not exhibit a presumptive positive. No difference between the alternate confirmation method and reference confirmation method was observed. The respective cultural reference methods (USDA-FSIS/MLG, FDA/BAM, and AOAC/OMA) produced a total of 63 confirmed positive results. Nonspiked samples from all foods were reported as negative for L. monocytogenes by all methods. Probability of detection analysis demonstrated no significant differences in the number of positive samples detected by the RapidChek method and the respective cultural reference method.

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Roka Listeria Detection Method Using Transcription Mediated Amplification to Detect Listeria Species in Select Foods and Surfaces

Journal of AOAC International ◽

10.5740/jaoacint.12-063 ◽

2012 ◽

Vol 95 (6) ◽

pp. 1672-1688 ◽

Cited By ~ 3

Author(s):

Hua Yang ◽

Shannon Kaplan ◽

Michael Reshatoff ◽

Ernie Hu ◽

Alexis Zukowski ◽

...

Keyword(s):

Probability Of Detection ◽

Culture Methods ◽

Department Of Agriculture ◽

Detection Analysis ◽

Reference Methods ◽

Smoked Salmon ◽

Environmental Surfaces ◽

Detection Assay ◽

Inspection Service ◽

The U.S

Abstract The Roka Listeria Detection Assay was compared to the reference culture methods for nine select foods and three select surfaces. The Roka method used Half-Fraser Broth for enrichment at 35 ± 2°C for 24–28 h. Comparison of Roka's method to reference methods requires an unpaired approach. Each method had a total of 545 samples inoculated with a Listeria strain. Each food and surface was inoculated with a different strain of Listeria at two different levels per method. For the dairy products (Brie cheese, whole milk, and ice cream), our method was compared to AOAC Official MethodSM993.12. For the ready-to-eat meats (deli chicken, cured ham, chicken salad, and hot dogs) and environmental surfaces (sealed concrete, stainless steel, and plastic), these samples were compared to the U.S. Department of Agriculture/Food Safety and Inspection Service-Microbiology Laboratory Guidebook (USDA/FSIS-MLG) method MLG 8.07. Cold-smoked salmon and romaine lettuce were compared to the U.S. Food and Drug Administration/Bacteriological Analytical Manual, Chapter 10 (FDA/BAM) method. Roka's method had 358 positives out of 545 total inoculated samples compared to 332 positive for the reference methods. Overall the probability of detection analysis of the results showed better or equivalent performance compared to the reference methods.

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Detection of Listeria spp. Using ACTERO Listeria Enrichment Media

Journal of AOAC International ◽

10.5740/jaoacint.14-006 ◽

2014 ◽

Vol 97 (4) ◽

pp. 1127-1136

Author(s):

David Claveau ◽

Sergiy Olishevskyy ◽

Michael Giuffre ◽

Gabriela Martinez

Keyword(s):

Stainless Steel ◽

Environmental Samples ◽

Incubation Temperature ◽

Reference Method ◽

Selective Medium ◽

Single Step ◽

Department Of Agriculture ◽

Steel Surfaces ◽

Inspection Service ◽

The U.S

Abstract ACTERO™ Listeria Enrichment Media (ACTERO Listeria) is a selective medium developed for a single-step recovery and enrichment of Listeria spp. from environmental samples. Robustness testing of the ACTERO Listeria medium demonstrated good performance when minor changes were introduced to the incubation temperature and time. All 54 Listeria strains tested, representing the most frequently isolated Listeria species from food (L. monocytogenes, L. ivanovii, L. seeligeri, L. welshimeri, and L. grayi), were successfully enriched in ACTERO Listeria. None of the 30 nontarget strains tested in the exclusivity study was recovered after incubation in ACTERO Listeria. Recovery of Listeria was consistent across three independently produced lots of the ACTERO Listeria, and the prepared medium was stable for 45 days when stored at 4°C in the dark. Matrix studies performed with environmental sponge samples from plastic and stainless steel surfaces demonstrated similar recovery of Listeria spp. in a single-step enrichment using ACTERO Listeria from plastic, and significantly better recovery from stainless steel surfaces when compared to the U.S. Department of Agriculture-Food Safety and Inspection Service reference method. The results of this study prove that ACTERO Listeria Enrichment Media can be effectively used in replacement of the two-step enrichment suggested by the reference method without affecting the recovery of Listeria spp. from environmental samples.

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Validation Study of a Rapid ELISA for Detection of Deoxynivalenol in Wheat, Barley, Malted Barley, Corn, Oats, and Rice

Journal of AOAC International ◽

10.1093/jaoac/93.2.600 ◽

2010 ◽

Vol 93 (2) ◽

pp. 600-610 ◽

Cited By ~ 8

Author(s):

Anthony Lupo ◽

Chris Roebuck ◽

Ken Settimo ◽

Anna Quain ◽

Justina Kennedy ◽

...

Keyword(s):

Rapid Method ◽

Electron Capture ◽

Validation Study ◽

Reference Method ◽

Elisa Test ◽

Electron Capture Detection ◽

Reference Methods ◽

Test Kit ◽

Aoac Official ◽

Research Institute

Abstract Neogen Corp. developed the Veratox DON test kit for the detection of deoxynivalenol (DON). The purpose of this study was to validate the method under the requirements of the AOAC Research Institute Performance Tested MethodsSM (PTM) program. There are two AOAC Official MethodsSM for DON detection: 986.17 and 986.18, the first of which is a TLC method and the second a GC method. A rapid method (PTM 000701) has also been performance tested by the AOAC Research Institute. One of the most widely used reference methods; however, is a GC method with electron capture detection that is referred to as the reference method in this paper. Although considered the reference method, the GC procedure is complicated and requires the investment of both expensive equipment and a highly skilled technician. A rapid (e.g., ELISA) test kit needs to be validated by the AOAC Research Institute. The Veratox 2/3 method is highly reproducible with average CV values <10, and is very accurate, showing >97 correlation to reference methods.

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Validation of a Microwell DNA Probe Assay for Detection of Listeria spp. in Foods

Journal of AOAC International ◽

10.1093/jaoac/89.3.651 ◽

2006 ◽

Vol 89 (3) ◽

pp. 651-668 ◽

Cited By ~ 2

Author(s):

Omar A Oyarzabal ◽

Nicole M Behnke ◽

Mark A Mozola ◽

Wallace H Andrews ◽

Elliot T Ryser ◽

...

Keyword(s):

Phosphate Buffer Solution ◽

Food Samples ◽

Negative Control ◽

Buffer Solution ◽

Department Of Agriculture ◽

Reference Methods ◽

Probe Assay ◽

Food Commodities ◽

Inspection Service ◽

The U.S

Abstract A new DNA hybridization assay in microwell format for detection of Listeria spp. in foods and environmental samples was developed. This assay uses Listeria-specific oligonucleotide probes labeled with horseradish peroxidase and a photometrically determined end point. Validation studies with 15 different food commodities and a variety of environmental sample types were conducted to compare the performance of this alternative test versus reference methods. Meats, seafood, dairy products, and vegetables comprised the categories of food tested. Food samples were inoculated at 2 levels and refrigerated or frozen for at least 72 h. Uninoculated (negative) control samples were included in each trial. Samples were enriched according to the procedure recommended by either the U.S. Food and Drug Administration (FDA) or the U.S. Department of Agriculture (USDA), Food Safety and Inspection Service (FSIS). Samples enriched for 24 h were transferred to Oxford agar plates and incubated for 24 h. The surface of the plateswas then swabbed and any growth present was transferred to phosphate buffer solution for the performance of the DNA assay. A standard confirmation procedure was used to compare the number of positive samples obtained with the DNA method versus reference methods. Statistical analyses of the results indicate that the proposed alternative method performs equally to cultural referencemethods. TheDNAassay is able to detect as lowas 1 colony-forming unit of Listeria in a 25 g food sample, with results available as early as 48 h after the start of sample enrichment.

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Evaluation of the Thermo Scientific RapidFinder™ Salmonella Species, Typhimurium, and Enteritidis Multiplex PCR Kit

Journal of AOAC International ◽

10.5740/jaoacint.17-0344 ◽

2018 ◽

Vol 101 (4) ◽

pp. 1059-1100 ◽

Cited By ~ 1

Author(s):

Emma Scopes ◽

Jessica Screen ◽

Katharine Evans ◽

David Crabtree ◽

Annette Hughes ◽

...

Keyword(s):

Multiplex Pcr ◽

Probability Of Detection ◽

Specific Method ◽

Salmonella Spp ◽

Department Of Agriculture ◽

Reference Methods ◽

Thermo Fisher Scientific ◽

Reliable Performance ◽

Inspection Service ◽

The U.S

Abstract The Thermo Scientific RapidFinder™ Salmonella Species, Typhimurium, and Enteritidis Multiplex PCR Kit (candidate method) is a real-time PCR assay for the detection and differentiation of Salmonella spp., and the serovars S. Typhimurium, and S. Enteritidis from poultry, pork, and environmental samples. The method was validated in comparison to the U.S. Department of Agriculture Food Safety and Inspection Service and the U.S. Food and Drug Administration reference methods. Thermo Fisher Scientific (Basingstoke, United Kingdom) tested all matrixes. In addition, two matrixes were analyzed independently by Q Laboratories, Inc. (Cincinnati, OH). Few statistically significant differences were found between the candidate and reference methods when analyzed by probability of detection. When differences were observed, these were in favor of the candidate method. All 200 inclusivity strains and none of the 45 exclusivity strains were detected, which demonstrated that the RapidFinder Salmonella Species, Typhimurium, and Enteritidis Multiplex PCR Kit was able to detect all the major groups of Salmonella, the less common subspecies of S. enterica, and the rarely encountered S. bongori. None of the exclusivity isolates analyzed were detected. Robustness testing demonstrated that the assay gave reliable performance, with specific method deviations outside the recommended parameters. Accelerated stability testing was conducted, validating the assay shelf life.

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

Journal of AOAC International ◽

10.5740/jaoacint.15-0309 ◽

2016 ◽

Vol 99 (3) ◽

pp. 705-716 ◽

Cited By ~ 1

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.

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Validation of iQ-Check E. coli O157:H7 Real-Time PCR Test Kit for Detection of Escherichia coli O157:H7 in Selected Foods

Journal of AOAC International ◽

10.1093/jaoac/92.4.1095 ◽

2009 ◽

Vol 92 (4) ◽

pp. 1095-1104 ◽

Cited By ~ 1

Author(s):

Wendy F Lauer ◽

Sylvie Tymciu ◽

Caroline D Sidi ◽

Pierre Sonigo

Keyword(s):

Real Time ◽

Real Time Pcr ◽

Ground Beef ◽

Reference Method ◽

Target Sequence ◽

Apple Cider ◽

E Coli ◽

Test Kit ◽

Significant Difference ◽

The U.S

Abstract iQ-Check E. coli O157:H7 (Bio-Rad Laboratories, Hercules, CA) is a real-time PCR kit for detection of E. coli O157:H7 from selected foods. Specific fluorescent oligonucleotide probes are used to detect target DNA during the amplification, by hybridizing to the amplicons. These fluorescent probes are linked to a fluorophore which fluoresces only when hybridized to the target sequence. Three foods (ground beef, apple cider, fresh spinach) were selected to compare the performance of iQ-Check E. coli O157:H7 to the U.S. Department of Agriculture-Food Safety and Inspection Service Microbiology Laboratory Guidebook (MLG) reference method for ground beef and the U.S. Food and Drug Administration/Bacteriological Analytical Manual reference method for apple cider and fresh spinach. Three protocols were tested in this study: a shortened 8 h primary enrichment in buffered peptone water (BPW), a 24 h enrichment in BPW, and an enrichment in appropriate reference method enrichment broth. The iQ-Check E. coli O157:H7 method was able to identify more true/confirmed positive samples than the reference method. Inclusivity and exclusivity rates of the method were 100. iQ-Check E. coli O157:H7 performed as expected when minor procedural variations were introduced, validating the ruggedness of the method. There was no significant difference observed in performance over the shelf life of the kit.

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