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We present eight new radiocarbon dates for endemic and invasive rodents from Trouing Jérémie #5, a paleontologically-rich sink hole on the Tiburon Peninsula, Haiti. This includes new dates for two species that have been previously directly dated ( Isolobodon portoricensis and Brotomys voratus) as well as three endemic rodents which have no previous direct radiometric dates ( Plagiodontia velozi, Hexolobodon phenax, and Rhizoplagiodontia lemkei). The radiocarbon date for P. velozi, the largest of these species, is from the very early Holocene (10,995 ± 190 calendar years before present; Cal BP). Specimens of medium-bodied species, H. phenax and R. lemkei date to the mid-Holocene, while specimens of smaller-bodied I. portoricencis and B. voratus have dates falling in the Late-Holocene. These dates confirm that several of the extinct rodent species coexisted with the first humans, who arrived on the island ca. 6000 years ago. In contrast, murid Rattus specimens date to the last few centuries. Rats arrived with Europeans in the late 15th or early 16th Century and a radiocarbon date of ca. 500 Cal BP for one individual suggests that they likely spread quickly across the island. Collectively, these dates establish that vertebrate accumulations at Trouing Jérémie #5 span the Holocene. Remains from this site may provide a useful time sequence for future work examining ecological change across the Holocene as well as regional extirpation patterns.

Most researchers assume minimal impact of pretreatment on strontium isotope ratios (87Sr/86Sr) for bones and teeth, and methods vary tremendously. We compared 14 pretreatment methods, including no prep other than powdering enamel, ashing, soaking in water, an oxidizing agent (bleach or hydrogen peroxide) or acetic acid (0.1 M, 1.0 M, and 1.0 M buffered with calcium acetate), and a combination of these steps. We prepared and analyzed aliquots of powdered molar enamel from three proboscideans (one modern captive Indian elephant, Elephas maximus indicus; one Pleistocene mastodon, Mammut americanum; and one Miocene gomphothere, Afrochoerodon kisumuensis). Each pretreatment was performed in triplicate and we measured 87Sr/86Sr, Sr concentration, and uranium (U) concentration, using the same lab space and instrumentation for all samples. Variability in 87Sr/86Sr and Sr and U concentrations was considerable across pretreatments. Mean 87Sr/86Sr across methods ranged from 0.70999 to 0.71029 for the modern tooth, 0.71458 to 0.71502 for the Pleistocene tooth, and 0.70804 to 0.70817 for the Miocene tooth. The modern tooth contained the least Sr and negligible U. The Pleistocene tooth contained slightly more Sr and measurable amounts of U, and the Miocene tooth had approximately 5x more Sr and U than the Pleistocene tooth. For all three teeth, variance in 87Sr/86Sr, Sr concentrations, and U concentrations among replicates was statistically indistinguishable across pretreatments, but there were apparent differences among pretreatments for the modern and Pleistocene teeth. Both contained relatively little Sr, and it is possible that small amounts of exogenous Sr from reagents, building materials or dust affected some replicates for some pretreatments. For the modern tooth, median 87Sr/86Sr varied considerably (but statistically insignificantly) across pretreatments. For the Pleistocene tooth, variability in median 87Sr/86Sr was also considerable; some pretreatments were statistically distinct but there were no obvious patterns among methods. For the Miocene tooth, variability in median 87Sr/86Sr was much smaller, but there were significant differences among pretreatments. Most pretreatments yielded 87Sr/86Sr and Sr concentrations comparable to, or lower than, untreated powder, suggesting selective removal of exogenous material with high 87Sr/86Sr. Further evaluation of the mechanisms driving isotopic variability both within and among pretreatment methods is warranted. Researchers should clearly report their methods and avoid combining data obtained using different methods. Small differences in 87Sr/86Sr could impact data interpretations, especially in areas where isotopic variability is low.

With the advent of the COVID-19 pandemic, pharmacy students and educators experienced an abrupt shift as programmes that were previously taught exclusively in-person were then predominantly taught online. This sudden change provided little time for students to prepare for the new learning environment. The study objective was to explore pharmacy students' experiences of technology-enhanced learning during the COVID-19 pandemic. A cross-sectional survey was developed and distributed by email to all 3rd year (N = 76) and 4th year (N = 68) pharmacy students undertaking an MPharm programme in an Irish university. A total of 32 responses were collected, including 20 third year and 12 fourth year pharmacy students (response rates of 26.3% and 17.6%, respectively). The majority of respondents reported good or very good internet speed (71%) and stability (59%). Almost all were confident or very confident using Canvas (97%) prior to the onset of online learning. Respondents preferred engaging with other students in-person rather than online for coursework (68.8%) and learning new material (56.3%). Students favoured face-to-face delivery, with a recording of the session available online afterwards, for lectures (68.8%), workshops (50%) and tutorials (56.3%). Analysis of free-text comments indicates that respondents used recorded content to support exam revision and that a key drawback of online learning was social isolation. Pharmacy students favoured a blended learning approach, with in-person learning being recorded to support study and revision. Students' experience of TEL during the pandemic should be considered in the development and ongoing review of pharmacy programmes.

Background The Thermo Scientific™ SureTect™ Salmonella species PCR Assay utilizes Solaris™ reagents for performing PCR for the rapid and specific detection of Salmonella species in a broad range of foods and select environmental surfaces. Objective The aims were to demonstrate the reproducibility of the Thermo Scientific SureTect Salmonella species PCR Assay in a collaborative study using a challenging matrix, cocoa powder, and to extend the scope of the method. Method In the collaborative study, the candidate method was compared to the US Food and Drug Administration (FDA) Bacteriological Analytical Manual (BAM) Chapter 5 Salmonella reference method. The candidate method used two PCR thermocyclers, the Applied Biosystems™ QuantStudio™ 5 Real-Time PCR instrument (QS5) and the Applied Biosystems 7500 Fast Real-Time PCR instrument (7500 Fast). Fourteen participants from nine laboratories located within the United States and Europe were solicited for the collaborative study, with 12 participants submitting valid data. Three levels of contamination were evaluated for each matrix. Statistical analysis was conducted according to the probability of detection statistical model. In addition, 11 matrix studies were performed comparing the candidate method to the FDA/BAM Chapter 5 or US Department of Agriculture, Food Safety and Inspection Service, Microbiology Laboratory Guidebook 4.10 Isolation and Identification of Salmonella from Meat, Poultry, Pasteurized Egg, and Siluriformes (Fish) Products and Carcass and Environmental Sponges reference method. Nine of these matrices were also compared to the EN ISO 6579–1:2017/Amd.1:2020(E) Microbiology of the food chain—Horizontal method for the detection, enumeration and serotyping of Salmonella—Part 1: Detection of Salmonella spp.—AMENDMENT 1: Broader range of incubation temperatures, amendment to the status of Annex D, and correction of the composition of MSRV and SC reference method. Results In the collaborative study, the difference in laboratory results indicates equivalence between the candidate method and reference method for the matrix evaluated, and the method demonstrated acceptable interlaboratory reproducibility as determined in the collaborative evaluation. False-positive and false-negative rates were determined for the matrix and produced values of Conclusions Based on the data generated, the method demonstrated acceptable interlaboratory reproducibility data and statistical analysis. Highlights Due to the COVID-19 pandemic, some participants had to be trained remotely. Additionally, 375 g cocoa powder is known to be a challenging matrix for PCR methods. No unusual cross-contamination or false-positive/negative was reported, highlighting the ease of use, reproducibility, and robustness of the method.

Background The Assurance® GDS for Cronobacter Tq II assay is a nucleic acid amplification system for the qualitative detection of Cronobacter. The method uses an upfront concentration of the target organism from the enrichment by immunomagnetic separation (IMS) using the PickPen® device. Objective The Assurance GDS for Cronobacter Tq II method was evaluated for Official Methods of AnalysisSM certification. Method The matrix was compared to the ISO 22964:2017: Microbiology of the Food Chain—Horizontal Method for the Detection of Cronobacter spp. standard and using an alternative confirmation procedure. The alternative method was evaluated using 10 g test portions in an unpaired study design for powdered infant formula (milk based with iron and DHA) containing probiotics. Eleven technicians from eight laboratories located within the United States and Europe participated in the collaborative study. Statistical analysis was conducted according to the probability of detection (POD) statistical model as presented in the AOAC validation guidelines. The difference in laboratory POD (dLPODC) values with 95% confidence intervals across collaborators was calculated for each level between the candidate and reference method results and between the candidate presumptive and confirmed results. Results Results obtained for the low inoculum level test portions produced a dLPOD value with a 95% confidence interval of 0.03 (−0.18, 0.15). The dLPOD results indicate equivalence between the candidate method and reference method for the matrix evaluated. The method also demonstrated acceptable inter-laboratory reproducibility as determined in the collaborative evaluation. There were no false negative results; the false positive rate was determined and produced a value of Conclusions Based on the data generated, the method demonstrated Assurance GDS for Cronobacter Tq II assay produced acceptable interlaboratory reproducibility data and statistical analysis. Highlights The Assurance GDS for Cronobacter Tq II method is suitable for the rapid qualitative detection of Cronobacter in infant formulas, infant cereals, ingredients, and environmental samples.

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Testing milk for antibiotics before acceptance into dairies is required by the U.S. Pasteurized Milk Ordinance. Technological advances in tests have reduced screening times and improved detection accuracy. This work describes the validation of the Charm Rapid One Step Assay Beta-Lactam 30 Second Test according to the U.S. Food and Drug Administration Center for Veterinary Medicine protocol for raw commingled milk. Milk is added to the lateral flow test strip in an incubator/reader to deliver a 30 second result. Independent laboratory validation followed sensitivity, interference, and incurred residue protocols. Sensitivity, in parts per billion (ppb = µg/kg), using a probit curve determined 90% percent detection with 95% confidence, which met National Conference of Interstate Milk Shipments (NCIMS) specifications. Six U.S. approved beta-lactam drugs were detected below, but within 50% of, target/tolerance levels for penicillin G 2.9 ppb, ampicillin 5.9 ppb, amoxicillin 5.8 ppb, cephapirin 13 ppb, cloxacillin 8.1 ppb, and ceftiofur metabolites 73 ppb. No interferences were observed from 33 animal drugs at 100 ppb, somatic cells at 1.2 million/mL, or bacterial levels of >300 000 CFU/mL. Incurred residue detection levels were similar to levels determined with the spiked parent compound. The data support NCIMS that the BL30SEC method met U.S. criteria for testing milk for beta-lactams.

Background: The Bio-Rad iQ-Check Listeria spp. Kit uses real-time PCR technology for detection of Listeria species in select food matrixes and environmental surfaces. Objective: The iQ-Check Listeria spp. method was modified to reduce the enrichment medium volume for environmental sponges from 225 and 100 to 60 mL and to reduce the enrichment time for sponges and swabs from 25 ± 1 to as short as 18 h. The modified method was validated with stainless steel, polystyrene plastic, and sealed concrete using sponges or swabs with two different neutralizing buffers (Letheen Broth and HiCap™ Neutralizing Broth). In addition, the Bio-Rad Free DNA Removal Solution was used for all environmental samples. Methods: The iQ-Check Listeria spp. modified method was compared with the reference culture method in the U.S. Department of Agriculture Food Safety and Inspection Service Microbiology Laboratory Guidebook Chapter 8.10 using an unpaired study design. Results: In the method comparison study, the iQ-Check Listeria spp. modified method demonstrated no statistical difference in performance between candidate and reference method results or between presumptive and confirmed results for all environmental surfaces analyzed using HiCap Neutralizing Broth (World Bioproducts LLC) and Letheen broth. Conclusions: The modified iQ-Check Listeria spp. method is an effective method for the detection of Listeria species in environmental surfaces using both types of neutralizing buffer. Highlights: The method modification was granted based on the data collected.

Background: The Bruker MALDI Biotyper® method utilizes matrix-assisted laser desorption/ionization time-of-flight MS for the rapid and accurate identification and confirmation of Gram-negative bacteria from select media types. The alternative method was evaluated in a method extension study of AOAC INTERNATIONAL First Action Official MethodSM 2017.09 using nonselective and selective agars to identify Cronobacter spp., Salmonella spp., Campylobacter spp., and select Gram-negative bacteria. Results obtained by the Bruker MALDI Biotyper were compared to the traditional biochemical methods as prescribed in the appropriate reference methods. Methods: Two collaborative studies were organized, one in the United States focusing on Cronobacter spp. and other Gram-negative bacteria and one in Europe focusing on Salmonella spp. and other Gram-negative bacteria. Fourteen collaborators from seven laboratories located within the United States participated in the first collaborative study for Cronobacter spp. Fifteen collaborators from 15 service laboratories located within Europe participated in the second collaborative study for Salmonella spp. For each target organism (either Salmonella spp. or Cronobacter spp.), a total of 24 blind-coded isolates were evaluated. In each set of 24 organisms, there were 16 inclusivity organisms (Cronobacter spp. or Salmonella spp.) and 8 exclusivity organisms (non-Cronobacter spp. and non-Salmonella spp. closely related Gram-negative organisms). For the Campylobacter spp. method extension, 17 collaborators from eight laboratories located within the United States (seven laboratories) and Canada (one laboratory) participated in the collaborative study. A total of 24 blind-coded isolates were evaluated. In each set of 24 organisms, there were 16 inclusivity organisms (Campylobacter spp.) and 8 exclusivity organisms (non-Campylobacter spp. closely related Gram-negative organisms). Results: After testing was completed, the total percentage of correct identifications from each agar type for each strain was determined at a percentage of 100.0% to the genus level for the Cronobacter study and a percentage of 100.0% to the genus level for the Salmonella study. For the Campylobacter method extension, a correct identification and confirmation rate of 100.0% was obtained for the Campylobacter organisms at the species level. For non-Cronobacter, non-Salmonella, and non-Campylobacter organisms, 100.0% were correctly identified. Conclusions: The results indicated that the alternative method produced equivalent results when compared to the confirmatory procedures specified by each reference method. Highlights: The method extension can be modified to include the identification and confirmation of Campylobacter jejuni, Campylobacter coli, and Campylobacter lari.

Background: The MC-Media Pad™ Yeast and Mold (YM) is a ready-to use culture device that combines a test pad coated with medium and water-absorption polymers that is designed for the rapid quantification of yeast and mold in food products. Objective: The MC-Media Pad YM was compared with the U.S. Food and Drug Administration (FDA) Bacteriological Analytical Manual (BAM) for the enumeration of yeast and mold in frozen orange juice concentrate. Methods: The candidate method was evaluated using a paired study design in a multilaboratory collaborative study following the current AOAC Validation Guidelines. Three target contamination levels (low, 10–100 CFU/g; medium, 100–1000 CFU/g; and high 1000–10 000 CFU/g) and an uninoculated control level (0 CFU/g) were evaluated. MC-Media Pad YM devices were enumerated after 48 and 72 h of incubation. Results: Plate count obtained by both methods were log-transformed and the difference of means (including 95% confidence intervals), repeatability SD, and SD were determined for each contamination level. Conclusions: No statistical difference was observed between the MC-Media Pad YM (for both 48 and 72 h) and the FDA BAM for each contamination level. Highlights: The new method offers a convenient alternative to the reference method (FDA BAM) for detection of yeast and mold contamination in food products, yielding reliable and comparable results in 48 h compared to 5 days for the reference method.

The 3M™ Molecular Detection Assay (MDA) 2 – Cronobacter combines the use of loop-mediated isothermal amplification to rapidly amplify nucleic acid sequences while using bioluminescence to detect the amplification. Using a paired study design, the MDA 2 – Cronobacter was compared with ISO 22964:2017 for the detection of Cronobacter species in powdered infant formula containing probiotics. Technicians from 11 laboratories from the United States, Mexico, and Croatia participated. Collaborators received test portions with three levels of contamination. Statistical analysis was conducted according to the probability of detection (POD) statistical model. Results obtained for the low-inoculum level test portions produced a difference in POD values obtained from combining all valid collaborator POD data values with 95% confidence intervals of –0.01, –0.12, and 0.10, indicating that the difference between methods was not statistically significant at the 0.05 probability level.

The AOAC Research Institute Performance Tested MethodsSM Program certified Sample6 DETECT/L™ in April 2014 (Certification No. 041401) for the detection of Listeria species (L. monocytogenes, L. innocua, L. ivanovii, L. seeligeri, L. marthii, L. welshimeri) on stainless steel environmental surfaces. A modification was approved in January 2016, increasing the concentration of sanitizer-neutralizing reagents in detection reagents, increasing the number of phage in the detection solution, and increasing the sample test volume. Moreover, changes to reduce the number of negative controls and add compatibility with polyurethane sponges were also approved. In this modification, to ensure that DETECT/L continues to meet performance expectations, Sample6 evaluated workflow changes to enhance sensitivity and the ease-of-use of the assay. Changes to the phage concentration and detection threshold, plus the inclusion of a confirmation step (DETECT Check), were validated to obtain better accuracy and optimize assay performance. Inclusivity, exclusivity, and robustness testing were conducted by Sample6 to evaluate the changes. A third-party laboratory compared the DETECT/L assay and the U.S. Department of Agriculture reference method in a stainless steel environmental surface matrix study. The data presented in this report demonstrate that the changes proposed to the DETECT/L assay meet or exceed the performance in the current configuration.

Background Peel PlateTM Enterobacteriaceae Bacteria (EB) is dried selective media on a 47 mm plastic plate that produces enzyme substrate colored colonies on rehydration and incubation for 24 h and up to 48 h at 37 ± 1°C. Purpose The method validation compared quantification of EB to reference methods ISO 21528:2017 Parts 1 and 2. Methods Matrixes compared were whole milk, skim powdered milk, vanilla ice cream, butter, infant formulas (soy- and dairy-based), infant cereals ± probiotic, environmental sponge swab of stainless steel surface, and poultry carcass rinse with two different peptone buffers. Results In inclusivity and exclusivity studies, the method detected 54 of 54 EB strains and did not detect 30 of 30 non-EB strains. In matrix studies, the claimed foods were tested at three contamination levels using paired analysis between the reference and Peel Plate EB methods. Colony-forming units per gram or mL [CFU/g (mL)] were log10 transformed for statistical analysis. The candidate method and reference method were shown to be equivalent by the performance requirement of all 95% confidence intervals on mean difference falling between −0.5 and +0.5 log10 CFU/g (mL). An international collaborative study with dried infant formula spiked with Cronobacter sakazakii at log10 CFU/g (mL) 1.05, 2.31, and 3.21 levels, produced method differences −0.16, 0.15, and 0.18 log10 CFU/g (mL) with repeatabilities (r) = 0.33, 0.20, and 0.12 log10 CFU/g (mL) and reproducibilities (R) = 0.45, 0.26, and 0.18 log10 CFU/g (mL). Conclusions Based on these evaluations, the candidate method is considered equivalent to the reference methods at both the 24 h and 48 h incubation periods at 37 ± 1°C. Highlights Ready to use Enterobacteriaceae method equivalent to ISO-21528:2017 Parts 1 and 2; EB test colored colonies at 37°C for 24 h are equivalent at 48 h incubation; Singlet determined CFU/mL are statistically the same as duplicate average results; EB test validated for infant formula and dairy products including with probiotics; EB test for environmental surfaces and poultry carcass rinses using peptone buffers.

Background The Solus One Salmonella immunoassay utilizes Salmonella specific selective media and automated liquid handling, for the rapid and specific detection of Salmonella species in select food types. Objective The candidate method was evaluated using 375 g test portions in an unpaired study design for a single matrix, instant non-fat dry milk (NFDM) powder. Method The matrix was compared to the United States Food and Drug Administration/Bacteriological Analytical Manual (FDA/BAM) Chapter 5 Salmonella reference method. Eleven participants from 10 laboratories within academia and industry, located within the United States, Mexico, South Africa, Germany, and the United Kingdom, contributed data for the collaborative study. Three levels of contamination were evaluated for each matrix: an uninoculated control level [0 colony forming units (CFU)/test portion], a low inoculum level (0.2–2 CFU/test portion) and a high inoculum level (2–5 CFU/test portion). Statistical analysis was conducted according to the Probability of Detection (POD) statistical model. Results Results obtained for the low inoculum level test portions produced a dLPOD value with a 95% confidence interval between the candidate method confirmed (both alternative and conventional confirmation procedures) and the reference method of 0.07 (−0.02, 0.15). Conclusions The dLPOD results indicate equivalence between the candidate method and the reference method for the matrix evaluated and the method demonstrated acceptable inter-laboratory reproducibility as determined in the collaborative evaluation. False positive and false negative rates were determined for the matrix and produce values of Highlights Based on the data generated, the method demonstrated acceptable inter-laboratory reproducibility data and statistical analysis.

Background The MC-Media Pad® Rapid Aerobic Count (RAC) is a ready-to-use culture device combining a test pad coated with medium and water absorption polymers that are designed for the rapid quantification of total aerobic bacteria in food products. Objective The MC-Media Pad RAC was compared to the U.S. Department of Agriculture Food Safety and Inspection Service Microbiology Laboratory Guidebook, Chapter 3.02: Quantitative Analysis of Bacteria in Foods as Sanitary Indicators for raw ground pork and the Standard Methods for the Examination of Dairy Products, Chapter 6: Microbial Count Methods for yogurt drink. Method The candidate method was evaluated against the reference methods using a paired study design in a multi-collaborator study, following the current AOAC INTERNATIONAL Official Methods of AnalysisSM Appendix J guidelines. Three target contamination levels (low, medium, and high) were evaluated. MC-Media Pad RAC devices were enumerated after 24 and 48 h of incubation. Results Plate counts obtained by both methods were log10-transformed and the difference of means (including 95% confidence intervals), repeatability SD, and reproducibility SD were determined for each contamination level. All 95% confidence intervals for mean difference fell easily within ±0.10, the performance requirement being ±0.5. Conclusion The MC-Media Pad RAC (for both 24 and 48 h) and both reference methods for each contamination level were therefore shown to be equivalent, with 97.5% confidence. Highlights The new method offers a convenient alternative to the reference methods for detection of aerobic plate count in food products, yielding reliable and comparable results in 24 or 48 h compared to 48 h for the reference methods.

Sexual and reproductive health (SRH) issues are a common yet frequently underreported problem in adolescents and adults with cystic fibrosis (CF). As the CF population continues to experience overall health gains in lung function, nutrition, and life expectancy, it is likely that SRH will become a more prevalent issue, and more research will be needed to better understand the needs of the CF population. The focus of this review is to provide an overview of the current state of the literature regarding SRH in the CF population.

The Bruker MALDI Biotyper® method utilizes matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS for the rapid and accurate confirmation and identification of Gram-positive bacteria from select media types. This alternative method was evaluated using nonselective and selective agar plates to identify and confirm Listeria monocytogenes, Listeria species, and select Gram-positive bacteria. Results obtained by the Bruker MALDI Biotyper were compared with the traditional biochemical methods as prescribed in the appropriate reference method standards. Sixteen collaborators from 16 different laboratories located within the European Union participated in the collaborative study. A total of 36 blind-coded isolates were evaluated by each collaborator. In each set of 36 organisms, there were 16 L. monocytogenes strains, 12 non-monocytogenes Listeria species strains, and 8 additional Gram-positive exclusivity strains. After testing was completed, the total percentage of correct identifications (to both genus and species level) and confirmation from each agar type for each strain was determined at a percentage of 99.9% to the genus level and 98.8% to the species level. The results indicated that the alternative method produced equivalent results when compared with the confirmatory procedures specified by each reference method.

The iQ-Check Salmonella II Real-Time PCR test kit utilizes Salmonella-specific oligonucleotide probes and primers for the rapid and specific detection of Salmonella species in select food types. The alternative method was evaluated by using 375 g test portions in an unpaired study design for two matrices, milk chocolate and dry dog food. Each matrix was compared with the U.S. Food and Drug Administration Chapter 5 Salmonella reference method. Fourteen technicians from 12 laboratories, including academia and industry, located within the United States and Canada participated in the collaborative study. Three levels of contamination were evaluated for each matrix: 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). The statistical analysis was conducted according to the Probability of Detection (POD) statistical model. The results obtained for the low inoculum level test portions produced a difference in the candidate presumptive and confirmatory results (dLPOD) value with a 95% confidence interval of −0.05, (−0.15, 0.06) for the milk chocolate and 0.10, (−0.01, 0.21) for the dry dog food. The dLPOD results indicate an equivalence between the candidate method and reference method for the matrices evaluated, and the method demonstrated acceptable interlaboratory reproducibility as determined in the collaborative evaluation. False positive and false negative rates were determined for each matrix and produce values of

QIAGEN mericon Escherichia coli O157 Screen Plus and mericon E. coli Shiga toxin-producing E. coli (STEC) O-Type Pathogen Detection Assays use Real-Time PCR technology for the rapid, accurate detection of E. coli O157 and the “big six” (O26, O45, O103, O111, O121, O145) (non-O157 STEC) in select food types. Using a paired study design, the assays were compared with the U.S. Department of Agriculture, Food Safety Inspection Service Microbiology Laboratory Guidebook Chapter 5.09 reference method for the detection of E. coli O157:H7 in raw ground beef. Both mericon assays were evaluated using the manual and an automated DNA extraction method. Thirteen technicians from five laboratories located within the continental United States participated in the collaborative study. Three levels of contamination were evaluated. Statistical analysis was conducted according to the probability of detection (POD) statistical model. Results obtained for the low-inoculum level test portions produced a difference between laboratories POD (dLPOD) value with a 95% confidence interval of 0.00 (–0.12, 0.12) for the mericon E. coli O157 Screen Plus with manual and automated extraction and mericon E. coli STEC O-Type with manual extraction and –0.01 (–0.13, 0.10) for the mericon E. coli STEC O-Type with automated extraction. The dLPOD results indicate equivalence between the candidate methods and the reference method.

OBJECTIVE: This study evaluates the roles of medical and social complexity in health care use outcomes in cystic fibrosis (CF) after transfer from pediatric to adult care. METHODS: Retrospective cohort design included patients with CF who were transitioned into adult care at Indiana University from 2005 to 2015. Predictor variables included demographic and comorbidity data, age at transition, treatment complexity score (TCS), and an objective scoring measure of their social complexity (Bob’s Level of Social Support, BLSS). Outcome variables included outpatient visit rates and hospitalization rates. Pearson’s correlations and linear regression were used to analyze the data. RESULTS: The median age of the patients (N=133) at the time of transition was 20 (IQR 19–23) years. The mean FEV(1) % predicted at transition was 69 ± 24%. TCS correlated with outpatient visit rates (r = 0.3, p = 0.003), as well as hospitalization rates (r = 0.4, p < 0.001); while the BLSS only correlated with hospitalization rates (r = 0.7, p < 0.001). After adjusting for covariates, the strongest predictors of post-transfer hospitalizations are BLSS (p

BACKGROUND Several nongenetic factors, such as socioeconomic status, environmental exposures, and adherence have been described to have an impact on outcomes in cystic fibrosis (CF). OBJECTIVE To determine the effect of social complexity on the % predicted forced expiratory volume (ppFEV1 ) before and after transfer to adult care among adolescents with CF. METHODS Retrospective, single center, cohort study included all patients with CF who were transitioned into adult care between 2005 and 2015 at Indiana University. Social complexity (Bob's level of social support, [BLSS]) was collected at transfer. Linear mixed regression models assessed the relationship between ppFEV1 decline over time and BLSS with other covariates. RESULTS The median age of the patients (N = 133) at the time of transfer was 20 years (interquartile range: 19-23). Overall, there was a decline in lung function over time in our population (ppFEV1 at 24 months pretransfer 77 ± 20%, ppFEV1 at 24 months, posttransfer 66 ± 24%; P

The 3M™ Molecular Detection Assay (MDA) 2 – Listeria monocytogenes uses loop-mediated isothermal amplification of unique DNA target sequences combined with bioluminescence to rapidly detect Listeria monocytogenes in a broad range of food types and on environmental surfaces. Using an unpaired study design, technicians from 13 laboratories located in the United States and Canada compared the 3M MDA 2 – Listeria monocytogenes to the U.S. Department of Agriculture Food Safety Inspection Service Microbiology Laboratory Guidebook Chapter 8.09 “Isolation and Identification of Listeria monocytogenes from Red Meat, Poultry, and Egg Products, and Environmental Samples” reference method for the detection of L. monocytogenes in deli turkey and raw chicken breast fillet. Each matrix was evaluated at three levels of contamination: 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). Statistical analysis was conducted according to the probability of detection (POD) statistical model. Results obtained for the low inoculum level test portions produced a difference in the collaborating laboratory POD (dLPOD) value of 0.04 with a 95% confidence interval of (−0.08, 0.17) for deli turkey, indicating that the difference between methods was not statistically significant at the 0.05 probability level. For raw chicken breast fillet, a dLPOD value of 0.16 with a 95% confidence interval of (0.04, 0.28) indicated a statistically significant difference, with an observed higher proportion of positive results by the candidate method compared to the reference method.

3M Molecular Detection Assay (MDA) 2–Listeria uses loop-mediated isothermal amplification and bioluminescence detection to rapidly detect Listeria species in a broad range of food types and environmental surfaces. Using an unpaired study design, MDA 2–Listeria was compared with the U.S. Department of Agriculture, Food Safety and Inspection Service's Microbiology Laboratory Guidebook Chapter 8.09 “Isolation and identification of Listeria monocytogenes from red meat, poultry and egg products, and environmental samples” reference method for the detection of Listeria in deli turkey and raw chicken breast fillet. Technicians from 13 laboratories located within the continental United States and Canada participated in the collaborative study. Each matrix was evaluated at three levels of contamination: uninoculated control (0 CFU/test portion), low inoculum (0.2–2 CFU/test portion), and high inoculum (2–5 CFU/test portion). Statistical analysis was conducted according to the probability of detection (POD) statistical model. Results obtained for the low-inoculum-level test portions produced a difference between two laboratory POD values (dLPOD) with 95% confidence intervals of 0.04 (–0.08, 0.17) for deli turkey, indicating the difference between the methods was not statistically significant at the P = 0.05. For raw chicken breast fillet, a dLPOD value with 95% confidence interval of 0.16 (0.04, 0.28) indicated a statistically significant difference between the two methods, with an observed higher proportion of positive results by the candidate method than the reference method.

The Applied Biosystems™ RapidFinder™ STEC Detection Workflow (Thermo Fisher Scientific) is a complete protocol for the rapid qualitative detection of Escherichia coli (E. coli) O157:H7 and the “Big 6” non-O157 Shiga-like toxin-producing E. coli (STEC) serotypes (defined as serogroups: O26, O45, O103, O111, O121, and O145). The RapidFinder STEC Detection Workflow makes use of either the automated preparation of PCR-ready DNA using the Applied Biosystems PrepSEQ™ Nucleic Acid Extraction Kit in conjunction with the Applied Biosystems MagMAX™ Express 96-well magnetic particle processor or the Applied Biosystems PrepSEQ Rapid Spin kit for manual preparation of PCR-ready DNA. Two separate assays comprise the RapidFinder STEC Detection Workflow, the Applied Biosystems RapidFinder STEC Screening Assay and the Applied Biosystems RapidFinder STEC Confirmation Assay. The RapidFinder STEC Screening Assay includes primers and probes to detect the presence of stx1 (Shiga toxin 1), stx2 (Shiga toxin 2), eae (intimin), and E. coli O157 gene targets. The RapidFinder STEC Confirmation Assay includes primers and probes for the “Big 6” non-O157 STEC and E. coli O157:H7. The use of these two assays in tandem allows a user to detect accurately the presence of the “Big 6” STECs and E. coli O157:H7. The performance of the RapidFinder STEC Detection Workflow was evaluated in a method comparison study, in inclusivity and exclusivity studies, and in a robustness evaluation. The assays were compared to the U.S. Department of Agriculture (USDA), Food Safety and Inspection Service (FSIS) Microbiology Laboratory Guidebook (MLG) 5.09: Detection, Isolation and Identification of Escherichia coli O157:H7 from Meat Products and Carcass and Environmental Sponges for raw ground beef (73% lean) and USDA/FSIS-MLG 5B.05: Detection, Isolation and Identification of Escherichia coli non-O157:H7 from Meat Products and Carcass and Environmental Sponges for raw beef trim. No statistically significant differences were observed between the reference method and the individual or combined kits forming the candidate assay using either of the DNA preparation kits (manual or automated extraction). For the inclusivity and exclusivity evaluation, the RapidFinder STEC Detection Workflow, comprising both RapidFinder STEC screening and confirmation kits, correctly identified all 50 target organism isolates and correctly excluded all 30 nontarget strains for both of the assays evaluated. The results of these studies demonstrate the sensitivity and selectivity of the RapidFinder STEC Detection Workflow for the detection of E. coli O157:H7 and the “Big 6” STEC serotypes in both raw ground beef and beef trim. The robustness testing demonstrated that minor variations in the method parameters did not impact the accuracy of the assay and highlighted the importance of following the correct incubation temperatures

Background: The iQ-Check Real-Time PCR kits use PCR technology based on gene amplification and detection by a real-time PCR thermalcycler for the detection of target analytes in select food matrices. The iQ-Check E. coli O157:H7 [Performance Tested MethodSM (PTM) 020801] and STEC VirX and STEC SerO (combined PTM 121203) methods were previously validated for different matrices under different enrichment schemes. Objective: To modify the current iQ-Check E. coli O157:H7 Kit for the detection of Escherichia coli O157:H7 from 25 to 375 g for raw ground beef (17% fat), raw beef trim, and fresh spinach. In addition, a matrix extension was validated for iQ-Check E. coli O157:H7 for raw chicken breast without skin (25 g), raw chicken thigh with skin (25 g), mechanically separated chicken (25 g), and raw ground pork (25 g). The study also included the modification of the iQ-Check STEC VirX and SerO Kits for the detection of non-O157 Shiga toxin–producing E. coli (STEC) for raw ground beef (375 g), raw beef trim (375 g), and fresh spinach (375 g) from STEC Enrichment Broth to buffered peptone water (BPW). All tests were carried out at 8–22 h (10–22 h for fresh spinach). Methods: Ground beef, beef trim, and spinach were co-inoculated with E. coli O157:H7, non-O157 STECs, and Salmonella spp. and analyzed for E. coli O157:H7 and non-O157 STECs after an 8-22 h enrichment in BPW for the beef matrices and after a 10–22 h enrichment in BPW for spinach. The chicken matrices were inoculated with E. coli O157:H7 only and analyzed after an 8–22 h enrichment in BPW. The iQ-Check Free DNA Removal Solution workflow was utilized for all matrices. Confirmations at the 22 h time point and method comparisons were conducted with the appropriate reference method as outlined in the U.S. Food and Drug Administration Bacteriological Analytical Manual Chapter 4A or the U.S. Department of Agriculture Food Safety and Inspection Service Microbiology Laboratory Guidebook Chapters 5.09 and 5B.05. For the iQ-Check STEC VirX and STEC SerO Kits, inclusivity and exclusivity were also performed. Results: The two inclusivity and exclusivity evaluations indicated that the test methods can accurately detect the target analytes and correctly excluded nontarget organisms after 8 h of enrichment. In the method comparison study, the iQ-Check E. coli O157:H7 and STEC VirX and STEC SerO test kits demonstrated no statistically significant differences between candidate and reference method results or between presumptive and confirmed results for all food matrices analyzed and the two time points (8 or 10 and 22 h). Both time points produced the same results, with no discrepancies. Conclusions: The iQ-Check real-time PCR kits are effective methods for the detection of E. coli O157 and non-O157 STECs (both the virulence factors and the O groups) from raw ground beef, raw beef trim, and fresh spinach in 375 g samples enriched in BPW for 8–22 h (10–22 h for fresh spinach). In addition, the iQ-Check E. coli O157 Kit is effective in detecting E. coli O157 in 25 g samples of raw chicken breast without skin, raw chicken thigh with skin, mechanically separated chicken, and raw ground pork. The iQ-Check test kits allow the end user to pair enrichments for multiple target analytes, allowing the user to prepare a single enrichment and perform a single DNA extraction. The Free DNA Removal Solution removes free DNA from samples prior to PCR analysis, protecting DNA from intact and living cells. Highlights: The method modifications were granted based on the data collected.

The Bruker MALDI Biotyper® method utilizes matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS for the rapid and accurate identification and confirmation of Gram-negative bacteria from select media types. The alternative method was evaluated using nonselective and selective agars to identify Cronobacter spp., Salmonella spp., and select Gram-negative bacteria. Results obtained by the Bruker MALDI Biotyper were compared to the traditional biochemical methods as prescribed in the appropriate reference methods. Two collaborative studies were organized, one in the United States focusing on Cronobacter spp. and other Gram-negative bacteria, and one in Europe focusing on Salmonella spp. and other Gram-negative bacteria. Fourteen collaborators from seven laboratories located within the United States participated in the first collaborative study for Cronobacter spp. Fifteen collaborators from 15 service laboratories located within Europe participated in the second collaborative study for Salmonella spp. For each target organism (either Salmonella spp. or Cronobacter spp.), a total of 24 blind-coded isolates were evaluated. In each set of 24 organisms, there were 16 inclusivity organisms (Cronobacter spp. or Salmonella spp.) and 8 exclusivity organisms (closely related non-Cronobacter spp. and non-Salmonella spp. Gram-negative organisms). After testing was completed, the total percentage of correct identifications from each agar type for each strain was determined at a percentage of 100.0% to the genus level for the Cronobacter study and a percentage of 100.0% to the genus level for the Salmonella study. For both non-Cronobacter and non-Salmonella organisms, a percentage of 100.0% was correctly identified. The results indicated that the alternative method produced equivalent results when compared to the confirmatory procedures specified by each reference method.

Background The 3M™ Petrifilm™ Rapid E. coli/Coliform Count Plate is a selective and differential sample-ready-culture medium designed for the rapid enumeration of Escherichia coli (E. coli) and coliforms in the food and beverage industries. Objective The 3M Petrifilm Rapid E. coli/Coliform Count Plate was compared to the U.S. Food and Drug Administration Bacteriological Analytical Manual (BAM) Chapter 4 Enumeration of Escherichia coli and the Coliform Bacteria, the International Organization of Standards (ISO) 4832:2006 Microbiology of food and animal feeding stuffs — Horizontal method for the enumeration of coliforms—Colony-count technique, and ISO 16649-2:2017 Microbiology of food and animal feeding stuffs—Horizontal method for the enumeration of beta-glucuronidase-positive Escherichia coli—Part 2 Colony-count technique at 44 degrees C using bromo-4-chloro-3- indolyl beta-D-glucuronide methods for the enumeration of E. coli and coliforms in dry dog kibble. Method The candidate method was evaluated using two diluents, Butterfield's phosphate buffered diluent and peptone salt solution, in a paired study design with each reference method in a multi-laboratory collaborative study following the current AOAC Validation Guidelines. Three target contamination levels and an uninoculated control level were evaluated. Results The candidate and reference methods were not statistically different at each contamination level. Reproducibility values obtained during the collaborative study were similar between the candidate and reference methods. Conclusion These results demonstrate that the candidate method is equivalent to the reference methods. Highlight 3M Petrifilm Rapid E. coli/Coliform Count Plate was recommended for Official First Action status for enumeration of E. coli and coliforms in a broad range of foods and environmental surfaces.

Peel Plate™ EC is a low-profile plastic, 47 mm culture dish with an adhesive top that contains a dried medium with Gram-negative selective agents and with enzyme substrate indicators for β-galactosidase (coliform) and β-glucuronidase (Escherichia coli). The method provides a conventional quantitative coliform (red) and E. coli (blue/purple/black) count with simple rehydration and incubation for 24 ± 2 h at 35 ± 1°C, while providing a total coliform result, sum of E. coli, and coliform without color differential in dairy products at 32 ± 1°C for 24 ± 2 h. Dairy matrixes claimed and supported with total coliform data are whole milk, skim milk, chocolate milk (2% fat), heavy cream (35% fat), pasteurized whole goat milk, ultra-high-temperature pasteurized milk, powdered milk, lactose-reduced milk, strawberry milk, shredded cheddar cheese, raw cow milk, raw goat milk, raw sheep milk, sour cream, condensed milk, eggnog, vanilla ice cream, condensed whey, yogurt, and cottage cheese. Matrixes claimed for E. coli and total coliform detection are raw ground beef, mixed cellulose 0.45 μm filtered bottled water, environmental sponge of stainless steel, raw ground turkey, dry dog food, liquid whole pasteurized eggs, milk chocolate, leafy green (mixed greens) rinse/flume water, irrigation water, poultry carcass rinse, and large animal carcass sponge. The method has been independently evaluated for total coliform in whole milk, skim milk, chocolate milk, and heavy cream. The method was also independently evaluated for E. coli and coliform in ground beef, filtered bottled water, and sponge rinse from stainless steel surfaces. In inclusivity and exclusivity studies, the method detected 57 of 58 different strains of coliform and E. coli at 32 ± 1°C and 35 ± 1°C in and excluded 31 of 32 different noncoliform strains consisting of Gram-negative and Gram-positive bacteria. In the matrix study, each matrix was assessed separately at each contamination level in comparison to an appropriate reference method. Colony counts were determined for each level and then log10 transformed. The transformed data were evaluated for repeatability, log-mean comparison between methods with 95% confidence interval, and r2. A 95% confidence interval range of −0.5 to 0.5 on the mean difference was used as the acceptance criterion to establish significant statistical difference between methods. The evaluations demonstrate that the Peel Plate EC method provides no statistical differences across most of the matrixes. The coliform r2 values were greater than 0.9 except in the case of skim milk (r2 = 0.77 and 0.69), sheep milk (0.84), and chocolate (0.81). In the case of skim milk, the three highest concentrations were significantly biased low compared with the reference method, whereas in the case of chocolate, the highest concentration was significantly biased high. The E. coli r2 values were greater than 0.9 except in the case of hog rinse (0.89), flume water (0.82), and chocolate (0.77). The lower values were generally from only a 1 log difference between highest and lowest concentrations except in the case of chocolate, in which the highest concentration was biased high compared with the reference method. Within-method repeatability of Peel Plate EC was similar to the reference method, with relative SDs generally less than 5% when log10 means were ≥1.5. QC data support that the Peel Plate EC is stable for 1 year when refrigerated. Incubation temperature ranges, 30–36°C, and times, 22–26 and 48 h for yogurt, were not significantly different in paired t-test comparison. The method is selective without the need for confirmation, although confirmation of coliform and E. coli was performed as part of the validation work.

The 3M™ Molecular Detection Assay (MDA) 2

VereBeef™ Detection Kit, incorporating both multiplex PCR and microarray technologies on a lab-on-chip platform, is intended for qualitative detection and differentiation of Escherichia coli O157:H7, E. coli O26, E. coli O45, E. coli O103, E. coli O111, E. coli O121, E. coli O145, Shiga toxin-producing E. coli (STEC) virulence factors (stx1A, stx2A, eae), and Salmonella species in one test using raw beef trim samples. This product underwent extensive evaluations, including inclusivity-exclusivity, method comparison, robustness, lot-to-lot variability, and stability studies. The inclusivity/exclusivity study demonstrated that VereBeef Detection Kit specifically detects and identifies target analytes without occurrence of false-positive and false-negative detection. In the method comparison study, the performance of the VereBeef Detection Kit was compared with U.S. Department of Agriculture Food Safety and Inspection Service Microbiology Laboratory Guidebook’s methods for target organism detection in raw beef trim using E. coli O157:H7 single inoculation and Salmonella and non-O157 STEC dual inoculation. Data demonstrated equivalence in both methods. The robustness study showed that changesin the test parameters do not impact assay performance. Collectively, VereBeef Detection Kit is able to detect target pathogens in raw beef trim with a minimum enrichment time of 8 h for E. coli O157:H7 detection and 10 h for Salmonella and non-O157 STEC detection.

The iQ-Check Salmonella II Real-Time PCR test kit utilizes Salmonella-specific oligonucleotide probes and primers for the rapid and specific detection of Salmonella species in select food types. The alternative method was evaluated by using 375 g test portions in an unpaired study design for two matrices, milk chocolate and dry dog food. Each matrix was compared with the U.S. Food and Drug Administration Chapter 5 Salmonella reference method. Fourteen technicians from 12 laboratories, including academia and industry, located within the United States and Canada participated in the collaborative study. Three levels of contamination were evaluated for each matrix: 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). The statistical analysis was conducted according to the Probability of Detection (POD) statistical model. The results obtained for the low inoculum level test portions produced a difference in the candidate presumptive and confirmatory results (dLPOD) value with a 95% confidence interval of -0.05, (-0.15, 0.06) for the milk chocolate and 0.10, (-0.01, 0.21) for the dry dog food. The dLPOD results indicate an equivalence between the candidate method and reference method for the matrices evaluated, and the method demonstrated acceptable interlaboratory reproducibility as determined in the collaborative evaluation. False positive and false negative rates were determined for each matrix and produce values of2%. Based on the data generated, the method demonstrated acceptable interlaboratory reproducibility data and statistical analysis.

The 3M™ Petrifilm™ Rapid Yeast and Mold (RYM) Count Plate is a simple, ready-to-use chromogenic culture method for the rapid detection and enumeration of yeast and mold in food products. The 3M Petrifilm RYM Count Plate method was compared to the U. S. Food and Drug Administration Bacteriological Analytical Manual (FDA BAM) Chapter 18, Yeasts, Molds and Mycotoxins and the ISO 21527:2008 Microbiology of Food and Animal Feeding Stuffs—Horizontal Method for the Enumeration for Yeast and Molds – Part 1: Colony Count Technique in Products with Water Activity Greater Than 0.95 and Part 2: Colony Count Technique in Products with Water Activity Less Than or Equal to 0.95 reference methods for raw almonds and raw frozen ground beef patties (77% lean). The 3M Petrifilm RYM Count Plate method was evaluated using a paired study design in a multi-laboratory collaborative study following the current AOAC Validation Guidelines. Three target contamination levels (low, 10–100 CFU/g; medium, 100–1000 CFU/g; high 1000–10 000 CFU/g) as well as an uninoculated control level (0 CFU/g) were evaluated for each matrix. Samples evaluated by the 3M Petrifilm RYM Count Plate method were prepared in duplicate and incubated at both 25°C and 28°C. Plates at both temperatures were enumerated after 48 and 60 h of incubation. No significant difference was observed between the 3M Petrifilm RYM Count Plate method and the FDA BAM or ISO 21527 reference methods for each contamination level. No statistical differences were observed between samples analyzed by the 3M Petrifilm RYM Count Plate method (at either 25°C or 28°C) and the reference methods. No statistical significant differences were observed between enumeration of colonies at 48 and 60 h on the 3M Petrifilm RYM Count Plate method and the reference methods.

The 3M™ Petrifilm™Salmonella Express (SALX) System is a simple, ready-to-use chromogenic culture medium system for the rapid qualitative detection and biochemical confirmation of Salmonella spp. in food and food process environmental samples. The 3M Petrifilm SALX System was compared using an unpaired study design in a multilaboratory collaborative study to the U.S. Department of Agriculture/Food Safety and Inspection Service (USDA/FSIS) Microbiology Laboratory Guidebook (MLG) 4.07 (2013) Isolation and Identification of Salmonella from Meat, Poultry, Pasteurized Egg and Catfish Products and Carcass and Environmental Sponges for raw ground beef and the U.S. Food and Drug Administration Bacteriological Analytical Manual (FDA/BAM) Chapter 5, Salmonella (2011) reference method for dry dog food following the current AOAC validation guidelines. For this study, a total of 17 laboratories located throughout the continental United States evaluated 1872 test portions. For the 3M Petrifilm SALX System, raw ground beef was analyzed using 25 g test portions, and dry dog food was analyzed using 375 g test portions. For the reference methods, 25 g test portions of each matrix were analyzed. The two matrices were 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). Each inoculation level was statistically analyzed using the probability of detection statistical model. For the raw ground beef and dry dog food test portions, no significant differences at the 95% confidence interval were observed in the number of positive samples detected by the 3M Petrifilm SALX System versus either the USDA/FSIS-MLG or FDA/BAM methods.

QIAGEN mericon Escherichia coli O157 Screen Plus and mericon E. coli Shiga toxin-producing E. coli (STEC) O-Type Pathogen Detection Assays use Real-Time PCR technology for the rapid, accurate detection of E. coli O157 and the "big six" (O26, O45, O103, O111, O121, O145) (non-O157 STEC) in select food types. Using a paired study design, the assays were compared with the U.S. Department of Agriculture, Food Safety Inspection Service Microbiology Laboratory Guidebook Chapter 5.09 reference method for the detection of E. coli O157:H7 in raw ground beef. Both mericon assays were evaluated using the manual and an automated DNA extraction method. Thirteen technicians from five laboratories located within the continental United States participated in the collaborative study. Three levels of contamination were evaluated. Statistical analysis was conducted according to the probability of detection (POD) statistical model. Results obtained for the low-inoculum level test portions produced a difference between laboratories POD (dLPOD) value with a 95% confidence interval of 0.00 (-0.12, 0.12) for the mericon E. coli O157 Screen Plus with manual and automated extraction and mericon E. coli STEC O-Type with manual extraction and -0.01 (-0.13, 0.10) for the mericon E. coli STEC O-Type with automated extraction. The dLPOD results indicate equivalence between the candidate methods and the reference method.

The performances of two DuPont BAX System PCR assays for detecting Salmonella on a variety of low-moisture soy ingredients were evaluated against the U. S. Food and Drug Administration's Bacteriological Analytical Manual (FDA BAM) method or the International Organization for Standardization (ISO) 6579 reference method. These evaluations were conducted as a single laboratory validation at an ISO 17025 accredited third-party laboratory. Validations were conducted on five soy ingredients: isolated soy protein (ISP), soy fiber, fluid soy lecithin, deoiled soy lecithin, and soy nuggets, using a paired-study design. The ISP was analyzed as both 25- and 375-g composite test portions, whereas all other sample matrices were analyzed as 375-g composite test portions. To evaluate 25-g test portions of ISP, the test material was inoculated using Salmonella enterica subsp. enterica serovar Mbandaka (Q Laboratories isolate 11031.1). Salmonella enterica subsp. enterica serovar Tennessee (Q Laboratories isolate 11031.3) was used for all other trials. For each trial of the method comparison, 25 samples were analyzed for each matrix: 5 uninoculated controls and 20 samples inoculated at low levels (0.2 to 2 CFU per test portion) that were targeted to achieve fractionally positive results (25 to 75%). Using McNemar's chi-square analysis, no significant difference at P ≥ 0.05 (χ(2) ≤ 3.84) was observed between the number of positives obtained by the BAX System and the reference methods for all five test matrices evaluated. These studies indicate that the BAX System PCR assays, in combination with the single buffered peptone water primary enrichment and subsequent brain heart infusion regrowth step, demonstrate equivalent sensitivity and robustness compared with the FDA BAM and ISO reference methods for both 25- and 375-g composite samples. Moreover, there was no observed reduction of sensitivity in the larger 375-g composite samples for all five matrices.

The 3M™ Molecular Detection Assay (MDA) Salmonella utilizes isothermal amplification of nucleic acid sequences with high specificity, efficiency, rapidity and bioluminescence to detect amplification of Salmonella spp. in food, food-related, and environmental samples after enrichment. A method modification and matrix extension study of the previously approved AOAC Official MethodSM 2013.09 was conducted, and approval of the modification was received on March 20, 2014. Using an unpaired study design in a multilaboratory collaborative study, the 3M MDA Salmonella method was compared to the U.S. Department of Agriculture/Food Safety and Inspection Service (USDA/FSIS) Microbiology Laboratory Guidebook (MLG) 4.05 (2011), Isolation and Identification of Salmonella from Meat, Poultry, Pasteurized Egg, and Catfish Products for raw ground beef and the U.S. Food and Drug Administration (FDA)/Bacteriological Analytical Manual (BAM) Chapter 5, Salmonella reference method for wet dog food following the current AOAC guidelines. A total of 20 laboratories participated. For the 3M MDA Salmonella method, raw ground beef was analyzed using 25 g test portions, and wet dog food was analyzed using 375 g test portions. For the reference methods, 25 g test portions of each matrix were analyzed. Each matrix 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, 1512 unpaired replicate samples were analyzed. Statistical analysis was conducted according to the probability of detection (POD). For the low-level raw ground beef test portions, the following dLPOD (difference between the LPODs of the reference and candidate method) values with 95% confidence intervals were obtained: –0.01 (–0.14, +0.12). For the low-level wet dog food test portions, the following dLPOD with 95% confidence intervals were obtained: –0.04 (–0.16, +0.09). No significant differences were observed in the number of positive samples detected by the 3M MDA Salmonella method versus either the USDA/FSIS-MLG or FDA/BAM methods.

The DNAble Salmonella detection assay utilizes single overnight culture enrichment, user-friendly sample preparation, and isothermal DNA amplification for Salmonella detection. This report describes studies performed in support of AOAC Research Institute Performance Tested MethodSM certification of the DNAble assay. Selectivity (inclusivity and exclusivity) studies were performed in the sponsor's laboratory. DNAble detected 119 out of 120 Salmonella isolates, representing 100 Salmonella serovars, in the inclusivity study while none of the 35 diverse non-Salmonella strains (32 species) tested was detected in the exclusivity study. Consistency (lot-to-lot and stability), instrument variation, and robustness studies were also conducted by the sponsor. Statistically equivalent assay performance was observed in these studies demonstrating robust assay manufacture and performance despite variation of multiple parameters in these challenges. Matrix studies, performed in an independent laboratory, evaluated DNAble assay performance in dry pet food, on stainless steel surfaces, and poultry environmental drag swab samples. Two sample sizes (25 and 375 g) and two culture volumes (9:1 and 3:1, v/w) were evaluated in separate matrix studies for dry pet food to provide multiple certified testing options for assay users. DNAble assay performance for dry pet food and stainless steel was compared to the procedures described in the U.S. Food and Drug Administration (FDA) Bacteriological Analytical Manual, Chapter 5, Salmonella. Assay performance for drag swabs was compared to protocols dictated in the FDA Environmental Sampling and Detection of Salmonella in Poultry Houses guidelines. Matrix study results demonstrated statistically equivalent DNAble assay performance compared to these reference methods, ensuring that the DNAble assay provides results comparable to those of the reference methods.

A multilaboratory study was conducted to evaluate the ability of the DuPont™ BAX® System Real-Time PCR Assay for Salmonella to detect the target species in a variety of foods and environmental surfaces. Internal validation studies were performed by DuPont Nutrition & Health on 24 different sample types to demonstrate the reliability of the test method among a wide variety of sample types. Two of these matrixes—pork and turkey frankfurters and pasteurized, not-from-concentrate orange juice without pulp—were each evaluated in 14 independent laboratories as part of the collaborative study to demonstrate repeatability and reproducibility of the internal laboratory results independent of the end user. Frankfurter samples were evaluated against the U. S. Department of Agriculture, Food Safety and Inspection Service reference method as a paired study, while orange juice samples were evaluated against the U. S. Food and Drug Administration reference method as an unpaired study, using a proprietary media for the test method. Samples tested in this study were artificially inoculated with a Salmonella strain at levels expected to produce low (0.2–2.0 CFU/test portion) or high (5 CFU/test portion) spike levels on the day of analysis. For each matrix, the collaborative study failed to show a statistically significant difference between the candidate method and the reference method using the probability of detection statistical model.

The 3M™ Molecular Detection Assay (MDA) Salmonella is used with the 3M™ Molecular Detection System for the detection of Salmonella spp. in food, food-related, and environmental samples after enrichment. The assay utilizes loop-mediated isothermal amplification to rapidly amplify Salmonella target DNA with high specificity and sensitivity, combined with bioluminescence to detect the amplification. The 3M MDA Salmonella method was compared using an unpaired study design 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), Isolation and Identification of Salmonella from Meat, Poultry, Pasteurized Egg and Catfish Products for raw ground beef and the U.S. Food and Drug Administration/Bacteriological Analytical Manual (FDA/BAM) Chapter 5 Salmonella reference method for wet dog food following the current AOAC guidelines. A total of 20 laboratories participated. For the 3M MDA Salmonella method, raw ground beef was analyzed using 25 g test portions, and wet dog food was analyzed using 375 g test portions. For the reference methods, 25 g test portions of each matrix were analyzed. Each matrix 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, 1512 unpaired replicate samples were analyzed. Statistical analysis was conducted according to the probability of detection (POD). For the low-level raw ground beef test portions, the following dLPOD (difference between the POD of the reference and candidate method) values with 95% confidence intervals were obtained: –0.01 (–0.14, +0.12). For the low-level wet dog food test portions, the following dLPOD with 95% confidence intervals were obtained: –0.04 (–0.16, +0.09). No significant differences were observed in the number of positive samples detected by the 3M MDA Salmonella method versus either the USDA/FSIS-MLG or FDA/BAM methods.