Validation of a Modification to Performance-Tested MethodSM 070601: Reveal® Listeria Test for Detection of Listeria spp. in Selected Foods and Selected Environmental Samples

Abstract A modification to Performance-Tested MethodSM (PTM) 070601, Reveal® Listeria Test (Reveal), is described. The modified method uses a new media formulation, LESS enrichment broth, in single-step enrichment protocols for both foods and environmental sponge and swab samples. Food samples are enriched for 2730 h at 30C and environmental samples for 2448 h at 30C. Implementation of these abbreviated enrichment procedures allows test results to be obtained on a next-day basis. In testing of 14 food types in internal comparative studies with inoculated samples, there was a statistically significant difference in performance between the Reveal and reference culture U.S. Food and Drug Administration's Bacteriological Analytical Manual (FDA/BAM) or U.S. Department of Agriculture-Food Safety and Inspection Service (USDA-FSIS) methods for only a single food in one trial (pasteurized crab meat) at the 27 h enrichment time point, with more positive results obtained with the FDA/BAM reference method. No foods showed statistically significant differences in method performance at the 30 h time point. Independent laboratory testing of 3 foods again produced a statistically significant difference in results for crab meat at the 27 h time point; otherwise results of the Reveal and reference methods were statistically equivalent. Overall, considering both internal and independent laboratory trials, sensitivity of the Reveal method relative to the reference culture procedures in testing of foods was 85.9 at 27 h and 97.1 at 30 h. Results from 5 environmental surfaces inoculated with various strains of Listeria spp. showed that the Reveal method was more productive than the reference USDA-FSIS culture procedure for 3 surfaces (stainless steel, plastic, and cast iron), whereas results were statistically equivalent to the reference method for the other 2 surfaces (ceramic tile and sealed concrete). An independent laboratory trial with ceramic tile inoculated with L. monocytogenes confirmed the effectiveness of the Reveal method at the 24 h time point. Overall, sensitivity of the Reveal method at 24 h relative to that of the USDA-FSIS method was 153. The Reveal method exhibited extremely high specificity, with only a single false-positive result in all trials combined for overall specificity of 99.5.

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Validation of a Modification to Performance-Tested MethodSM 010403: Microwell DNA Hybridization Assay for Detection of Listeria spp. in Selected Foods and Selected Environmental Surfaces

Journal of AOAC International ◽

10.1093/jaoac/92.2.438 ◽

2009 ◽

Vol 92 (2) ◽

pp. 438-448 ◽

Cited By ~ 1

Author(s):

Susan Alles ◽

Linda X Peng ◽

Mark A Mozola

Keyword(s):

Ceramic Tile ◽

Reference Method ◽

High Specificity ◽

Single Step ◽

Method Performance ◽

Media Formulation ◽

Independent Laboratory ◽

Environmental Surfaces ◽

Crab Meat ◽

Time Point

Abstract A modification to Performance-Tested MethodSM 010403, GeneQuence<sup/> Listeria Test (DNAH method), is described. The modified method uses a new media formulation, LESS enrichment broth, in single-step enrichment protocols for both foods and environmental sponge and swab samples. Food samples are enriched for 2730 h at 30C, and environmental samples for 2448 h at 30C. Implementation of these abbreviated enrichment procedures allows test results to be obtained on a next-day basis. In testing of 14 food types in internal comparative studies with inoculated samples, there were statistically significant differences in method performance between the DNAH method and reference culture procedures for only 2 foods (pasteurized crab meat and lettuce) at the 27 h enrichment time point and for only a single food (pasteurized crab meat) in one trial at the 30 h enrichment time point. Independent laboratory testing with 3 foods showed statistical equivalence between the methods for all foods, and results support the findings of the internal trials. Overall, considering both internal and independent laboratory trials, sensitivity of the DNAH method relative to the reference culture procedures was 90.5. Results of testing 5 environmental surfaces inoculated with various strains of Listeria spp. showed that the DNAH method was more productive than the reference U.S. Department of Agriculture-Food Safety and Inspection Service (USDA-FSIS) culture procedure for 3 surfaces (stainless steel, plastic, and cast iron), whereas results were statistically equivalent to the reference method for the other 2 surfaces (ceramic tile and sealed concrete). An independent laboratory trial with ceramic tile inoculated with L. monocytogenes confirmed the effectiveness of the DNAH method at the 24 h time point. Overall, sensitivity of the DNAH method at 24 h relative to that of the USDA-FSIS method was 152. The DNAH method exhibited extremely high specificity, with only 1 false-positive reactions overall.

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An Analysis of Clostridium difficile Environmental Contamination During and After Treatment for C difficile Infection

Open Forum Infectious Diseases ◽

10.1093/ofid/ofaa362 ◽

2020 ◽

Vol 7 (11) ◽

Author(s):

Kerrie Davies ◽

Damian Mawer ◽

A Sarah Walker ◽

Claire Berry ◽

Timothy Planche ◽

...

Keyword(s):

Clostridium Difficile ◽

Environmental Contamination ◽

Environmental Samples ◽

Transmission Risk ◽

Fecal Samples ◽

Treatment Groups ◽

End Of Treatment ◽

Significant Difference ◽

Time Point

Abstract Background Lower Clostridium difficile spore counts in feces from C difficile infection (CDI) patients treated with fidaxomicin versus vancomycin have been observed. We aimed to determine whether environmental contamination is lower in patients treated with fidaxomicin compared with those treated with vancomycin/metronidazole. Methods The CDI cases were recruited at 4 UK hospitals (Leeds, Bradford, and London [2 centers]). Environmental samples (5 room sites) were taken pretreatment and at 2–3, 4–5, 6–8, and 9–12 days of treatment, end of treatment (EOT), and post-EOT. Fecal samples were collected at diagnosis and as often as produced thereafter. Swabs/feces were cultured for C difficile; percentage of C difficile-positive samples and C difficile bioburden were compared between different treatment arms at each time point. Results Pre-EOT (n = 244), there was a significant reduction in environmental contamination (≥1 site positive) around fidaxomicin versus vancomycin/metronidazole recipients at days 4–5 (30% vs 50% recipients, P = .04) and at days 9–12 (22% vs 49%, P = .005). This trend was consistently seen at all other timepoints, but it was not statistically significant. No differences were seen between treatment groups post-EOT (n = 76). Fidaxomicin-associated fecal positivity rates and colony counts were consistently lower than those for vancomycin/metronidazole from days 4 to 5 of treatment (including post-EOT); however, the only significant difference was in positivity rate at days 9–12 (15% vs 55%, P = .03). Conclusions There were significant reductions in C difficile recovery from both feces and the environment around fidaxomicin versus vancomycin/metronidazole recipients. Therefore, fidaxomicin treatment may lower the C difficile transmission risk by reducing excretion and environmental contamination.

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Comparative Evaluation of Veriflow®Listeria Species to USDA Culture-Based Method for the Detection of Listeria spp. in Food and Environmental Samples

Journal of AOAC International ◽

10.5740/jaoacint.17-0080 ◽

2017 ◽

Vol 100 (5) ◽

pp. 1434-1444 ◽

Cited By ~ 1

Author(s):

Adam C Joelsson ◽

Shawn P Terkhorn ◽

Ashley S Brown ◽

Amrita Puri ◽

Benjamin J Pascal ◽

...

Keyword(s):

Stainless Steel ◽

Ceramic Tile ◽

Pcr Amplification ◽

Reference Method ◽

Probability Of Detection ◽

Complex Data ◽

Listeria Species ◽

Environmental Surfaces ◽

Study Results ◽

Significant Difference

Abstract Veriflow®Listeria species (Veriflow LS) is a molecular-based assay for the presumptive detection of Listeria spp. from environmental surfaces (stainless steel, sealed concrete, plastic, and ceramic tile) and ready-to-eat (RTE) food matrixes (hot dogs and deli meat). The assay utilizes a PCRdetection method coupled with a rapid, visual, flow-based assay that develops in 3 min post-PCR amplification and requires only a 24 h enrichment for maximum sensitivity. The Veriflow LS system eliminates the need for sample purification, gel electrophoresis, or fluorophore-based detection of target amplification and does not require complex data analysis. This Performance Tested MethodSM validation study demonstrated the ability of the Veriflow LS assayto detect low levels of artificially inoculated Listeria spp. in six distinct environmental and food matrixes. In each unpaired reference comparison study, probability of detection analysis indicated that there was no significant difference between the Veriflow LS method and the U.S. Department of Agriculture Food Safety and Inspection Service Microbiology Laboratory Guide Chapter 8.08 reference method. Fifty-one strains of various Listeria spp. were detected in the inclusivity study, and 35 nonspecific organisms went undetected in the exclusivity study. The study results show that the Veriflow LS is a sensitive, selective, and robust assay for the presumptive detection of Listeria spp. sampled from environmental surfaces (stainless steel, sealed concrete, plastic, and ceramic tile) and RTE food matrixes (hot dogs and delimeat).

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Validation of the ANSR® for Campylobacter Method for Detection of Thermophilic Campylobacter spp. in Chicken Carcass Rinse and Turkey Sponge Samples

Journal of AOAC International ◽

10.5740/jaoacint.16-0241 ◽

2016 ◽

Vol 99 (6) ◽

pp. 1555-1564 ◽

Cited By ~ 2

Author(s):

Ryan Viator ◽

Susan Alles ◽

Quynh-Nhi Le ◽

Edan Hosking ◽

Preetha Biswas ◽

...

Keyword(s):

Target Genes ◽

Reference Method ◽

Common Species ◽

Single Step ◽

Fluorescence Detector ◽

Method Performance ◽

Chicken Carcass ◽

Inspection Service ◽

Nicking Enzyme ◽

Performance Validation

Abstract A performance validation of the ANSR® for Campylobacter method was conducted in selected matrixes. This assay used selective nicking enzyme amplification technology to amplify target genes. Samples were enriched for 20 to 24 h and then lysed. The assay was completed within 50 min using real-time detection in a combination incubator/fluorescence detector and software. When 50 distinct strains of Campylobacter jejuni, C. lari, or C. coli were tested for inclusivity, all 50 strains produced positive results. In exclusivity testing, 31 strains of related organisms, including seven nontarget Campylobacter strains and other common species, were evaluated. All 31 species generated negative ANSR assay results, including the nontarget Campylobacter strains. The ANSR for Campylobacter method was compared to the U.S. Department of Agriculture, Food Safety and Inspection Service Microbiology Laboratory Guidebook reference method using naturally contaminated chicken carcass rinse or turkey carcass sponge samples. ANSR method performance was not statistically different from the reference method using two different enrichment options. Equivalent results were observed at both time points (20 and 24 h) and in both atmospheres (microaerobic and aerobic) to reference methods. Method performance with chicken carcass rinse was confirmed in an independent laboratory study. Additionally, in robustness testing, small, deliberate changes to the assay parameters minimally affected ANSR method performance. Finally, accelerated stability results from three independently manufactured lots supported a shelf life of 6 months when stored at 4°C. The ANSR assay offered greater efficiency and flexibility when compared to the reference method with a 20–24 h single-step enrichment in a microaerobic or an aerobic atmosphere.

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Evaluation of VIDAS® UP Salmonella (SPT) Assay for the Detection of Salmonella in a Variety of Foods and Environmental Samples: Collaborative Study

Journal of AOAC International ◽

10.5740/jaoacint.cs2013_01 ◽

2013 ◽

Vol 96 (4) ◽

pp. 808-821 ◽

Cited By ~ 3

Author(s):

Patrick Bird ◽

Kiel Fisher ◽

Megan Boyle ◽

Travis Huffman ◽

Marc Juenger ◽

...

Keyword(s):

Confidence Intervals ◽

Environmental Samples ◽

Collaborative Study ◽

Reference Method ◽

The United States ◽

Probability Of Detection ◽

Inoculum Level ◽

Test Portion ◽

Low Level ◽

Significant Difference

Abstract The VIDAS® UP Salmonella (SPT) uses recombinant phage proteins to detect Salmonella species in human and animal food products and production environmental samples after 18–26 h of enrichment. The VIDAS SPT assay is performed with the automated VIDAS or mini-VIDAS instruments. The VIDAS SPT method was compared in a multilaboratory collaborative study to the U.S. Department of Agriculture/Food Safety and Inspection Service-Microbiology Laboratory Guidebook (USDA/FSIS-MLG) 4.05 (2011) Isolation and Identification of Salmonella from Meat, Poultry, Pasteurized Egg and Catfish Products reference method following the current AOAC guidelines. A total of 15 laboratories representing government, academia, and industry throughout the United States participated. One matrix, raw ground beef, was analyzed using two different test portion sizes, 25 and 375 g. Each test portion was artificially contaminated with Salmonella at three inoculation levels, an uninoculated control level (0 CFU/test portion), a low inoculum level (0.2–2 CFU/test portion), and a high inoculum level (2–5 CFU/test portion). In this study, 1656 unpaired replicate samples were analyzed. Of those unpaired replicates, 476 were presumptive positive by the VIDAS method, with 475 confirmed positive by the traditional confirmation procedures and 476 confirmed positive by an alternative confirmation procedure. There were 411 confirmed positive replicates by the USDA/FSIS-MLG reference method. Statistical analysis was conducted according to the probability of detection (POD). For the low-level 375 g test portions, the following dLPOD values, with 95% confidence intervals, were obtained: 0.01 (−0.12, +0.15) for samples confirmed following the traditional confirmation; 0.02 (−0.18, +0.2) for samples confirmed following traditional confirmation on IBISA and ASAP; and 0.03 (−0.18, +0.24) for samples confirmed following the alternative confirmation on IBISA and ASAP. For the low-level 25 g test portions, the following dLPOD values, with 95% confidence intervals, were obtained: 0.41, (0.32, +0.49) for samples confirmed following the traditional confirmation, the traditional confirmation on IBISA and ASAP, and the alternative confirmation on IBISA and ASAP. With 0.0 within the confidence intervals for the 375 g test portions, there was no statistically significant difference in the number of positive samples detected by the VIDAS SPT method and the USDA/FSIS-MLG method at the 0.05 level. For the 25 g test portions, a statistically significant difference was observed between the VIDAS SPT method and the reference method for the low inoculum level, where the VIDAS SPT method recovered a higher number of positive results than the reference method. It is recommended that the VIDAS SPT method with the optional ASAP and IBISA agar confirmation method be adopted for Official First Action status for the detection of Salmonella in a variety of foods and environmental samples.

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Validation of the ANSR® Listeria monocytogenes Method for Detection of Listeria monocytogenes in Selected Food and Environmental Samples

Journal of AOAC International ◽

10.5740/jaoacint.15-0200 ◽

2016 ◽

Vol 99 (1) ◽

pp. 112-123

Author(s):

Oscar Caballero ◽

Susan Alles ◽

Quynh-Nhi Le ◽

R Lucas Gray ◽

Edan Hosking ◽

...

Keyword(s):

Listeria Monocytogenes ◽

Storage Temperature ◽

Reference Method ◽

Single Step ◽

Stainless Steel Surface ◽

Method Performance ◽

Inspection Service ◽

Nicking Enzyme ◽

Performance Results ◽

The U.S

Abstract Work was conducted to validate performance of the ANSR® for Listeria monocytogenes method in selected food and environmental matrixes. This DNA-based assay involves amplification of nucleic acid via an isothermal reaction based on nicking enzyme amplification technology. Following single-step sample enrichment for 16–24 h for most matrixes, the assay is completed in 40 min using only simple instrumentation. When 50 distinct strains of L. monocytogenes were tested for inclusivity, 48 produced positive results, the exceptions being two strains confirmed by PCR to lack the assay target gene. Forty-seven nontarget strains (30 species), including multiple non-monocytogenes Listeria species as well as non-Listeria, Gram-positive bacteria, were tested, and all generated negative ANSR assay results. Performance of the ANSR method was compared with that of the U.S. Department of Agriculture, Food Safety and Inspection Service Microbiology Laboratory Guidebook reference culture procedure for detection of L. monocytogenes in hot dogs, pasteurized liquid egg, and sponge samples taken from an inoculated stainless steel surface. In addition, ANSR performance was measured against the U.S. Food and Drug Administration Bacteriological Analytical Manual reference method for detection of L. monocytogenes in Mexican-style cheese, cantaloupe, sprout irrigation water, and guacamole. With the single exception of pasteurized liquid egg at 16 h, ANSR method performance as quantified by the number of positives obtained was not statistically different from that of the reference methods. Robustness trials demonstrated that deliberate introduction of small deviations to the normal assay parameters did not affect ANSR method performance. Results of accelerated stability testing conducted using two manufactured lots of reagents predicts stability at the specified storage temperature of 4°C of more than 1 year.

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Validation of the One Broth One Plate for Salmonella Method for Detection of Salmonella Spp. in Select Food and Environmental Samples: AOAC Performance Tested MethodSM 102002

Journal of AOAC International ◽

10.1093/jaoacint/qsaa149 ◽

2020 ◽

Author(s):

Susan Alles ◽

Brooke Roman ◽

Quynh-Nhi Le ◽

Magdalena Kurteu ◽

Ezzeddine Elmerhebi ◽

...

Keyword(s):

Validation Study ◽

Environmental Samples ◽

Reference Method ◽

Black Pepper ◽

Single Step ◽

Probability Of Detection ◽

Stainless Steel Surface ◽

Salmonella Spp ◽

Simple Method ◽

The U.S

Abstract Background One Broth One Plate for Salmonella (OBOP Salmonella) is a rapid and simple method for detection of Salmonella spp. in food and environmental samples using traditional culture methodology. The method utilizes single-step enrichment followed by plating to a selective/differential, chromogenic agar. Objective The purpose of the validation study was to measure the effectiveness of the OBOP Salmonella method in comparison to reference culture procedures. Methods Performance of the OBOP Salmonella method was compared to that of the U.S. Food and Drug Administration Bacteriological Analytical Manual Chapter 5 reference method for queso fresco, smoked salmon, cantaloupe, chocolate, black pepper, chili powder, dry pet food, and sponge samples from a stainless steel surface, or to that of the U.S. Department of Agriculture Microbiology Laboratory Guidebook Chapter 4.10 method for raw ground turkey, chicken carcass rinse, and pasteurized liquid egg. Inclusivity/exclusivity, robustness, and stability/lot-to-lot consistency testing was also performed. Results In the matrix study, there were no statistically significant differences in performance between the OBOP Salmonella and reference methods, as determined by probability of detection analysis (P < 0.05), for any of the matrixes examined. All 104 Salmonella spp. strains produced positive results in inclusivity testing, and all 33 non-salmonellae exclusivity strains tested negative with the OBOP Salmonella method. Conclusions Results of the validation study show that the OBOP Salmonella method is a reliable procedure for detection of Salmonella spp. in select matrixes. The method is simple to perform, requires no specialized equipment, and produces results in as little as 37 h.

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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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Sensitivity and Specificity of the BAX® for Screening/Listeria monocytogenes Assay: Internal Validation and Independent Laboratory Study

Journal of AOAC International ◽

10.1093/jaoac/84.4.1087 ◽

2001 ◽

Vol 84 (4) ◽

pp. 1087-1098 ◽

Cited By ~ 4

Author(s):

Alan M Hochberg ◽

Ann Roering ◽

Vidhya Gangar ◽

Michael Curiale ◽

W Mark Barbour ◽

...

Keyword(s):

Listeria Monocytogenes ◽

Sensitivity And Specificity ◽

Operating Parameters ◽

Culture Methods ◽

Internal Validation ◽

Independent Laboratory ◽

Significant Difference ◽

Standard Culture ◽

Crab Meat ◽

Ruggedness Testing

Abstract In this study to certify the BAX® for Screening/Listeria monocytogenes assay (DuPont Qualicon, Wilmington, DE), an internal evaluation was conducted on 16 food types that were simultaneously analyzed with the BAX system (BAX), and the ISO method for the detection of L. monocytogenes (ISO). No statistically significant difference in performance between the BAX and ISO methods was observed. Inclusivity/exclusivity testing showed that the BAX system was able to detect 97 of 97 (100%) of L. monocytogenes strains tested. None of 56 other Listeria species or non-Listeria tested gave a reproducible positive BAX result. Ruggedness testing demonstrated that performance of the assay was not affected by reasonable variability in the operating parameters. BAX was then submitted for independent laboratory validation. In this phase, BAX was compared with standard culture methods for the detection of L. monocytogenes in chicken (USDA-FSIS), crab meat (BAM), and milk (AOAC). This study validated product claims of sensitivity and specificity >98% in accordance with AOAC Performance Tested MethodSM requirements.

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Soleris®Enterobacteriaceae for the Detection of Enterobacteriaceae in Select Foods: AOAC Performance Tested MethodSM 121901

Journal of AOAC International ◽

10.1093/jaoacint/qsaa001 ◽

2020 ◽

Vol 103 (4) ◽

pp. 1081-1089

Author(s):

Susan Alles ◽

Brooke Roman ◽

Gail Betts ◽

Suzanne Jordan ◽

Linda Everis ◽

...

Keyword(s):

Reference Method ◽

Probability Of Detection ◽

Mozzarella Cheese ◽

Bacterial Strains ◽

Method Performance ◽

Dog Food ◽

Independent Laboratory ◽

Detection Analysis ◽

Automated Method ◽

Positive Results

Abstract Background Soleris®Enterobacteriaceae is a growth-based, automated method for detection of Enterobacteriaceae in food. Objective A study was conducted to validate the Soleris method for detection of Enterobacteriaceae in select foods (pasteurized milk, yogurt, mozzarella cheese, ice cream, dried milk, pasteurized liquid egg, frozen cooked chicken, deli ham, lettuce, and dry dog food) at a threshold of ≥ 10 CFU/g of product. Methods Inclusivity and exclusivity of the Soleris method were assessed by testing 55 and 38 target and non-target bacterial strains, respectively. Matrix testing was performed with one naturally contaminated and nine inoculated foods. Efficacy of the Soleris method was compared to that of the ISO 21528-2:2017 direct plating reference method using probability of detection analysis. Independent laboratory testing was conducted to verify method performance in two matrixes (yogurt and deli ham). Method robustness, stability, and lot-to-lot consistency of the Soleris reagents were also assessed. Results Inclusivity of the Soleris test was 91% and exclusivity was 100%. In matrix testing, there were no significant differences in the number of positive results obtained with the Soleris and reference methods for any of the matrixes examined. Overall, of 370 test portions, there were 176 positive results by the Soleris method and 177 positive results by the reference procedure. Conclusions Soleris Enterobacteriaceae is an effective method for detection of Enterobacteriaceae in the foods evaluated, with performance equivalent to that of the ISO 21528-2:2017 reference method. Highlights The Soleris method offers the advantages of labor savings and results within 18 h.

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