Enterobacteriaceae In Foods Research Articles

Validation of the Soleris®Enterobacteriaceae Method for Detection of Enterobacteriaceae in Dried Cannabis Flower: AOAC Performance Tested MethodSM 121901.

The Soleris®Enterobacteriaceae vial is a growth-based, automated method for detection of bacteria of the family Enterobacteriaceae in foods and other sample types including nutraceuticals and cosmetics. The Soleris method is used in a "dilute-to-specification" or threshold manner, in which a result is scored as positive or negative around a predetermined cutoff (in CFU/g) established by the dilution and volume of sample homogenate tested. The Soleris method was granted AOAC Performance Tested MethodSM (PTM) status for select foods after successful completion of a validation study (PTM 121901). The objective of this study was to validate the method for the detection of Enterobacteriaceae in dried cannabis flower [>0.3% delta-9-tetrahydrocannabinol (THC)]. The matrix study included comparison of Soleris method presumptive results to confirmation from the Soleris vials, and comparison of the Soleris confirmed results to those of the ISO 21528-2:2017 colony count method. Test materials at four different levels of contamination ranging from 7.8 to 3500 CFU/g were tested at three dilutions, corresponding to test thresholds. Probability of detection analysis at P < 0.05 showed there were no significant differences between Soleris presumptive and confirmed results, and no significant differences between Soleris confirmed and ISO 21528-2:2017 results. The results provided evidence that the Soleris Enterobacteriaceae test is an accurate method for detection of Enterobacteriaceae in dried cannabis flower. The Soleris Enterobacteriaceae method provides cannabis industry QC personnel with an effective method for analysis of dried cannabis flower and produces results in 20-24 h.

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

BackgroundSoleris®Enterobacteriaceae is a growth-based, automated method for detection of Enterobacteriaceae in food.ObjectiveA 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.MethodsInclusivity 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.ResultsInclusivity 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.ConclusionsSoleris 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.HighlightsThe Soleris method offers the advantages of labor savings and results within 18 h.

Comparison of In-house and Commercial Real-time PCR Systems for the Detection of Enterobacteriaceae and their Evaluation Within an Interlaboratory Study Using Infant Formula Samples

Traditional detection methods for Enterobacteriaceae in foods are time-consuming and laborious. The current study assessed the specificity of three real-time PCR primer sets. Set A (IEC primers) targeted the conserved flanking regions of the 16S rRNA, the 16S-ITS-23S gene region. Set B (ENT primers) annealed to Escherichia coli 16S ribosomal RNA gene. The third set (C) used a D-LUX™ (Light Upon eXtension) single FAM-labelled forward primer and a corresponding unlabeled primer. Set A was specific for E. coli and for some non-Enterobacteriaceae. SYBR Green-based real-time PCR confirmed the specificity of set B for the Enterobacteriaceae but also detected Vibrionaceae. In contrast, set C was poorly specific. However, set D including the forward LUX™ primer from set C and the reverse primer from set B had a specificity comparable to that of set B, but with higher sensitivity. This combined set was successfully applied to detect Enterobacteriaceae in infant milk formula and compared favourably with a commercial real-time PCR kit.

Validation of an Alternative Method for Counting Enterobacteriaceae in Foods Based on Accuracy Profile

Several years ago, microbiologists developed a harmonized standard for the validation of alternative methods against the reference method, namely International Organization for Standardization (ISO) 16140:2003. Since then a new and better statistical approach known as the accuracy profile concept has been proposed for interpretation of data collected during a validation study. This approach defines acceptability limits that are used to verify that an alternative analytical method can produce results acceptable for a defined analytical objective. This paper demonstrates how the accuracy profile approach can be efficiently applied to any data set collected according to ISO guidelines and how to interpret the final calculations to decide if a method is valid or not. The procedure was applied to data obtained in the validation of a commercial kit for the enumeration of Enterobacteriaceae in foods. A comparison of several statistical outputs derived from application of the accuracy profile demonstrated that this simple numerical method can be applied despite the non-normal distribution of bacterial colony counts.

Development of a PCR Assay for Detection of Enterobacteriaceae in Foods

A broad-range PCR assay for the detection of bacteria belonging to the Enterobacteriaceae family was developed in this study. Primers targeting the bacterial 16S rRNA gene were newly designed and used in this PCR assay. To determine the specificity of the assay, 72 different bacterial species (of 49 genera), 2 fungi, 3 animals, and 4 plants were tested. Results were positive for every tested Vibrioaceae or Enterobacteriaceae strain except Proteus mirabilis. For all other bacterial strains and eukaryotes tested, results were negative. Bacterial DNA for PCR was prepared by a simple procedure with the use of Chelex 100 resin from culture after growth in brain heart infusion medium. To test this PCR assay for the monitoring of the Enterobacteriaceae family, either Escherichia coli or Salmonella Enteritidis was inoculated into various foods as an indicator. Prior to the PCR, the inoculation of 10 to 40 CFU of bacteria per g of food was followed by a 5-h enrichment culture step, and the PCR assay allowed the detection of bacterial cells. When actual examinations of the contamination of 15 noodle foods with Enterobacteriaceae by this PCR assay were conducted, 33% (5 of 15) of the samples tested positive. These results agreed with those of the Petri film Enterobacteriaceae Count Plate assay. Including the enrichment culture step, the entire PCR detection process can be completed within 7 h.

3M™ Petrifilm™ Enterobacteriaceae Count Plate Method for Enumeration of Enterobacteriaceae in Selected Foods: Collaborative Study

The practice of detecting and enumerating all oxidase-negative, glucose-fermenting-Gram-negative rods (i.e., the family Enterobacteriaceae) is used to indicate unsanitary or inadequate food processing conditions. The objective of this interlaboratory collaborative study was to evaluate and compare the methods described in Standard Methods for the Examination of Dairy Products (SMEDP) and the Compendium of Methods for the Microbiological Examination of Foods (Compendium) with a commercial product, the 3M Petrifilm Enterobacteriaceae Count Plate, for the recovery of Enterobacteriaceae in foods. Six foods--cheddar cheese, milk, flour, frozen prepared meals, frozen broccoli, and nut pieces--were analyzed for Enterobacteriaceae by 12 collaborating laboratories. For each food tested, the collaborators received 8 blind test portions consisting of a control test portion and 3 levels of inoculated test portion, each in duplicate. Each test portion was tested by the Petrifilm Enterobacteriaceae Count Plate method as well as the SMEDP or Compendium methods. The precision estimates (repeatability or within-laboratory variation, and reproducibility or between-laboratory variation) were calculated with standard statistical techniques.

Evaluation of the 3M Petrifilm Enterobacteriaceae Count Plate Method for the Enumeration of Enterobacteriaceae in Foods

Results of the 3M Petrifilm Enterobacteriaceae Count (EB) plate method were compared with those of the standard violet red bile glucose agar (VRBG) method for the detection and enumeration of Enterobacteriaceae. Studies involving 107 bacterial strains demonstrated that the Petrifilm EB plate method is as sensitive as and more selective than the VRBG method. Sixty of the 62 pure Enterobacteriaceae cultures were recovered by both methods. In addition, 38 of the 45 non-Enterobacteriaceae organisms did not grow on the Petrifilm EB plate, while 28 of the 45 non-Enterobacteriaceae organisms did not grow on the VRBG plate. Colony counts from 174 naturally contaminated and 120 artificially inoculated dairy and nondairy food samples showed that the Petrifilm EB plate method performed as well as or better than the standard VRBG method for the enumeration of Enterobacteriaceae.

Comparison of the VITEK Gram-Negative Identification Card and an Enhanced Version for Identification of Salmonella, Escherichia coli, and Other Enterobacteriaceae:. Method Modification

The VITEK Gram-Negative Identification Card (GNI) was compared to an enhanced version of the test kit, the GNI + Card. The GNI Card is an official AOAC method (991.13) for identification of Salmonella, Escherichia coli, and other Enterobacteriaceae in foods. In this comparison 5 replicates of 124 gram-negative stock culture strains were evaluated to determine equivalency. Isolates were obtained primarily from food sources, with 108 of the isolates representing 15 genera within the family Enterobacteriaceae. Overall agreement between the GNI Card and the GNI + Card was 98.2%. Of the 23 strains of Salmonella tested, the longest identification time for the GNI Card was 10 h, with 43.5% of the strains identifying in 4 h or less. The longest identification time of the same Salmonella strains tested with the GNI + Card was 8 h, with 95.7% of the strains identifying within 4 h. The GNI + Card provided increased speed of identification while retaining the accuracy of the GNI Card. The GNI method for identifying Salmonella, Escherichia coli, and other Enterobacteriaceae was approved as a method modification by AOAC INTERNATIONAL.

Identification of Enterobacteriaceae in Foods with the AutoMicrobic System

Stock cultures (136) and fresh isolates (163) of Enterobacteriaceae from ground beef, processed chickens, frozen pot pies and commercial poultry feeds were identified to species with the AutoMicrobic System (AMS). All stock cultures and fresh isolates were also concurrently tested with two other identification systems (Micro-ID and API), previously evaluated and proven accurate for identification of Enterobacteriaceae. The AMS correctly identified to species 135/136 (99.3%) of the stock cultures and 160/163 (98.2%) of the fresh isolates. All Salmonella cultures tested (74) were correctly identified by AMS.

The scope and limitations of four methods for the rapid identification of Enterobacteriaceae in foods.

Four methods for the identification of Enterobacteriaceae in cooked frozen meat have been appraised. Although these methods could be modified to improve efficiency they could be clearly classified into two categories: (i) those that allow identification to genus level and which often cannot discriminate between two genera without the help of additional reactions; (ii) those that allow identification to species level and which only occasionally require additional reactions to differentiate one species from another. A different evaluation criterion was followed for each of these categories. In the first, as methods lead to final identification by means of data furnished by the manufacturer or via a number code which does not indicate probability, reactions were compared with those obtained by classical methods. In the second group, every number code admits three possibilities and indicates a probable biotype for each, thus reducing the risk of faulty interpretation on the part of the operator while also compensating for deficiencies of certain reactions. Evaluation was therefore based on the percentage of correctly identified cases per species. Efficiencies of the main biochemical reactions are also discussed.

The presumptive enumeration of lactose negative as well as lactose positive Enterobacteriaceae in foods.

The presumptive enumeration of lactose negative as well as lactose positive Enterobacteriaceae in foods.

The Presumptive Enumeration of Lactose Negative as well as Lactose Positive Enterobacteriaceae in Foods

The Presumptive Enumeration of Lactose Negative as well as Lactose Positive Enterobacteriaceae in Foods