4. AOACRIMicroMethods-2018Awards

194

1426  C rowley et al . : J ournal of AOAC I nternational V ol . 95, N o . 5, 2012

the isolates. Participants were asked to provide their own Gram stain reagents due to challenges associated with federal regulations in the transportation of the reagents.

and TSAB cultures were 95, 96, and 97%, respectively. The VITEK 2 GP reagents were stable over 18-month time studies and provided highly reproducible results when compared between lots. Assay ruggedness was demonstrated for three critical parameters, including age of culture, inoculum density ,and inoculum age. Overall results indicate that the VITEK 2 GP method is an acceptable automated method for the identification of selected Gram-positive organisms. The method was awarded PTM certification No. 120702 on December 10, 2007. The collaborative study evaluated the ability of the VITEK 2 GP identification method to correctly identify 720 inclusivity strains representative of 12 different claimed organisms. A total of 120 exclusivity strains representative of six different nonclaimed organisms were also evaluated. All isolates analyzed were obtained from the three recommended subculture culture media: TSA, CBA, and TSAB (6). Table 1. Inclusivity (claimed) isolates used in VITEK 2 GP collaborative study Organisms ID No. Source a Origin Listeria grayi 125274 BMX Industry Listeria innocua 12524 BMX Fish Listeria ivanovii ssp. Ivanovii 12913 BMX Reference lab Listeria ivanovii ssp . Londoniensis 12914 BMX Clinical Listeria monocytogenes 12502 BMX Shellfish Listeria seeligeri 6217 BMX Creamer Listeria welshimeri 12517 BMX Beef Staphylococcus hyicus 13889 BMX Reference lab Staphylococcus intermedius 16409 BMX Veterinary Staphylococcus aureus 8819 BMX Reference lab Staphylococcus aureus 8852 BMX Reference lab Staphylococcus epidermidis 8890 BMX Reference lab a  BMX = bioMérieux culture collection. A total of 20 laboratories participated in the collaborative study, consisting of industry, government, and private testing laboratories involved in the testing of food products. Qualified laboratories currently using the VITEK 2 system were invited to act as collaborators. Each participating laboratory received 18 isolates on Brain Heart Infusion (BHI) agar slants. The identity of each isolate had been previously confirmed by traditional biochemical confirmations and/or Certificate of Analysis. There were 12 claimed isolates (target inclusivity organisms; Table 1) and six isolates of exclusivity organisms (nontarget organisms; Table 2). A detailed collaborative study packet outlining all necessary information related to the study, including isolate preparation and documentation of results, was sent to each collaborating laboratory prior to the initiation of the study. All participating laboratories were provided with GP cards, prepared culture media plates, and any additional supplies needed to identify Collaborative Study Study Design

Preparation of Isolates

Pure isolates of each organism from the bioMérieux (BMX) culture collection were sent on BHI agar slants to each of the participating laboratories. Each isolate used had been well characterized. Microscopic and plate morphology, standard biochemicals, at least two phenotypic methods, and, when needed, molecular sequencing were performed. To prepare for distribution to the collaborators, growth from each strain was transferred to BHI agar slants from growth that had been subcultured onto CBA and incubated for 18–24 h at 35–37°C. Following incubation, slants were labeled and prepared for shipment. All isolates were labeled with a randomized, blind-coded 3-digit number affixed to the sample vial. Isolates were shipped in leak-proof, insulated containers on a Monday via overnight delivery according to the Category B Dangerous Goods shipment regulations set forth by the International Air Transportation Association. Upon receipt on Tuesday, isolates were streaked onto CBA to check for purity. Collaborators were instructed to hold the original BHI agar slants in the refrigerator at 2–5°C for the duration of the study. All isolates were shipped at ambient temperature. Collaborators received all 18 isolates during the same week of testing. Participants were instructed to inspect the isolates upon receipt of the package and document that they were received in good condition on the Sample Receipt Confirmation form provided in the shipment. Forms were then faxed or emailed to the study director. Upon receipt of the shipment, collaborators were instructed to streak each isolate received onto CBA to check for purity and incubate for 18–24 h at 35–37°C. Following incubation, a Gram stain was conducted on one isolated colony from each strain according to the U.S. Food and Drug Administration’s Bacteriological Analytical Manual (FDA/BAM) method (7) and results were recorded. Those isolates that were identified as characteristic Gram-positive organisms were streaked to the three recommended culture media, TSA, CBA, and TSAB, and Isolate Distribution Analysis of Isolates

Table 2. Exclusivity isolates used in VITEK 2 GP collaborative study Organisms ID No. Source a

Origin

202608 110125 112380 111663 304232 305527

BMX Tomato paste BMX Hospital BMX Beef hide BMX Reference lab BMX Clinical BMX Reference lab

Bacillus coagulans Citrobacter freundii

Escherichia coli

Serratia marcescens Aspergillus niger Candida albicans

a  BMX = bioMérieux culture collection.

03/10/2019