Detection of Salmonella in onion and garlic powders: collaborative study.

The relative efficiency of trypticase soy broth with added 0.5% K2SO3 and lactose broth as pre-enrichment media for recovering Salmonella from onion powder and garlic powder was collaboratively studied. For each spice, 13 collaborators each received 5 duplicate samples; 4 of the 5 replicate samples were inoculated with 1 of 4 levels of S. thompson ranging, at initiation of analysis, from greater than 3 to 93 organisms/g onion powder and greater than 3 to 43 organisms/g garlic powder. Salmonella growth was inhibited in each of these spices as evidenced by a rapid decline of most probable number values in samples determined immediately after and 7 days following inoculation. Collaborative results of cultural analyses demonstrated superiority of the modified tripticase soy broth for recovering Salmonella in each of the 2 spices. The improved method of detecting Salmonella in onion and garlic powders has been adopted as official first action.     

Multitest system for biochemical identification of Salmonella, Escherichia coli, and other Enterobacteriaceae isolated from foods: collaborative study

Nine laboratories evaluated the ability of the MICRO-ID test system to correctly identify members of the Enterobacteriaceae. A total of 78 isolates representing 11 genera of enterics that had been previously isolated from foods were used in the collaborative study. The collaborators streaked each isolate on plate count agar and incubated the plates overnight at 35 degrees C to check purity of the isolates. Then they proceeded with the method in which an isolated colony is transferred to a plate count agar slant and is incubated 18-24 h at 35 degrees C. Growth from the slant is emulsified in 3.5 mL physiological saline to a density comparable to a McFarland No. 2 tube and is then used to inoculate the test (MICRO-ID) strip. The strip is incubated 4 h at 35 degrees C and the reactions are read and recorded. Isolates are identified by using an octal code and the test kit manual. The system correctly identified 98.8% of the Salmonella isolates, 97.7% of the E. coli isolates, and 84.6% of the other 9 enteric genera tested. The system has been approved interim official first action as an alternative to conventional biochemical tests (1) for presumptive generic identification of food-borne Salmonella and for screening and elimination of non-Salmonella isolates; (2) for identification of E. coli from foods; and (3) for presumptive generic identification of other Enterobacteriaceae isolated from foods.     

Visual immunoassay for detection of Salmonella in foods: collaborative study

A collaborative study was performed in 13 laboratories to validate a visual enzyme immunoassay (EIA) procedure, TECRA, for rapid detection of Salmonella in foods. The EIA method was compared with the standard culture procedure for detection of Salmonella in 6 food types: ground black pepper, soy flour, dried whole eggs, milk chocolate, nonfat dry milk, and raw deboned turkey. Uninoculated and inoculated samples were included in each food group analyzed. There was no significant difference in the productivity of the EIA and culture procedures at the 5% level for any of the 6 foods. The enzyme immunoassay screening method has been approved interim official first action.     

Enzyme immunoassay for detection of Salmonella in foods: collaborative study

A collaborative study was performed in 25 laboratories to validate an enzyme immunoassay (EIA) procedure utilizing 2 specific monoclonal antibodies for rapid detection of Salmonella in foods. The EIA was compared with the standard culture procedure for detection of Salmonella in 6 food types: ground black pepper, soy isolate, dried whole eggs, milk chocolate, nonfat dry milk, and raw deboned turkey. Uninoculated and inoculated samples were included in each food group analyzed, with the exception of poultry which was naturally contaminated. There was no significant difference in the productivity of the EIA and culture procedures at the 5% level for any of the 6 foods. The enzyme immunoassay screening method has been adopted official first action.     

DNA hybridization assay for detection of Salmonella in foods: collaborative study

A collaborative study was performed in 11 laboratories to validate a DNA hybridization (DNAH) procedure for detection of Salmonella in foods. The DNAH procedure was compared to the standard culture method for detection of Salmonella in 6 foods: ground pepper, soy flour, dry whole egg, milk chocolate, nonfat dry milk, and raw deboned turkey. With the exception of turkey which was naturally contaminated, uninoculated and inoculated samples of each food group were analyzed. Results for the DNAH method were significantly better than for the standard culture method at the 5% probability level for the detection of Salmonella in turkey. There was no significant difference between the methods for the other 5 foods. The method has been adopted official first action.     

Enzyme immunoassay for detection of Salmonella in low-moisture foods: collaborative study

A collaborative study was performed in 15 laboratories to evaluate a modification of the enzyme immunoassay (EIA) method for detection of Salmonella in foods (46.B21-46.B29). The modified EIA requires 18-24 h pre-enrichment, 6-8 h selective enrichment, and 14-18 h M-broth post-enrichment prior to performing the assay, which requires 1-2 h. Total assay time is 40-52 h. The modified method was compared with the standard culture method for detection of Salmonella in 5 low-moisture foods: nonfat dry milk, milk chocolate, meat and bone meal, dry whole egg, and ground pepper. The modified method has been adopted official first action for use with low-moisture foods.     

Fluorescent enzyme immunoassay for rapid screening of Salmonella in foods: collaborative study

A collaborative study was performed in 13 laboratories to validate an enzyme immunoassay (EIA) procedure for rapid detection of Salmonella in foods. The EIA was compared with the standard culture procedure for detection of Salmonella in 6 food types: ground black pepper, soy flour, dried whole eggs, milk chocolate, nonfat dry milk, and raw deboned turkey. Uninoculated and inoculated samples were included in each food group analyzed. There was no significant difference in the proportion of samples positive by the EIA and culture procedures at the 5% level for any of the 6 foods. The enzyme immunoassay screening method has been adopted official first action as a rapid screening method for detection of Salmonella.     

Immunodiffusion screening method for detection of motile Salmonella in foods: collaborative study

A collaborative study was performed to validate the performance of the 1-2 TEST for detection of motile salmonellae in foods. Detection is based on observation of an immobilized band of cells. Twenty-three laboratories participated in the study. The 1-2 TEST (immunodiffusion test) was compared with the standard culture procedure (BAM/AOAC; FDA Bacteriological Analytical Manual) for detection of Salmonella in 6 food types: ground black pepper, soy flour, dried whole egg, milk chocolate, nonfat dry milk, and raw deboned turkey. Uninoculated and inoculated samples were included in each food group analyzed. After the tests on the 6 foods were completed, analysis of the data for turkey and soy flour showed that certain collaborators obtained data inconsistent with the data from the majority of collaborators. No specific method deviations to account for the inconsistencies were reported by those collaborators, so the collaborative testing of these 2 foods was repeated. Analysis of data for pepper, chocolate, nonfat dry milk, dried whole egg, and the second set of soy flour and turkey indicated 96.1% agreement between the BAM/AOAC and immunodiffusion test methods. The false negative rates for the immunodiffusion test and BAM/AOAC methods were 3.6 and 1.7%, respectively. There was no significant difference in the productivity of the immunodiffusion test and BAM/AOAC method at the 5% level for any of the 6 foods. The immunodiffusion screening method has been approved official first action for detection of motile Salmonella in foods.     

Colorimetric deoxyribonucleic acid hybridization assay for rapid screening of Salmonella in foods: collaborative study

A collaborative study was performed in 11 laboratories to validate a colorimetric DNA hybridization (DNAH) method for rapid detection of Salmonella in foods. The method was compared to the standard culture method for detection of Salmonella in nonfat dry milk, milk chocolate, soy isolate, dried whole egg, ground black pepper, and raw ground turkey. Samples inoculated with high (0.4-2 cells/g) and low (0.04-0.2 cells/g) levels of Salmonella and uninoculated control samples were included in each food group analyzed. There was no significant difference in the proportion of samples positive by DNAH and culture procedure for any of the 6 foods. The colorimetric DNA hybridization assay screening method has been adopted official first action as a rapid screening method for detection of Salmonella in all foods.     

Dry rehydratable film for enumeration of total aerobic bacteria in foods: collaborative study

A rehydratable dry-film plating procedure for aerobic plate counts has been compared to the standard agar plate method (966.23B and C, 15th ed.; 46.014-46.015, 14th ed.) in a collaborative study by 12 laboratories. Each laboratory analyzed the normal microflora of 3 samples in duplicate for 6 products. The aerobic plate counts ranged from 1.0 x 10(3) to 1.0 x 10(8) cfu/g. The products were flour, nuts, frozen raw shrimp, spice, frozen raw ground turkey, and frozen and refrigerated vegetables. Repeatability standard deviations of the 2 methods did not differ significantly for 13 of 18 test samples. For 1 shrimp and 2 turkey samples, the dry-film method had lower repeatability variances (P less than 0.05) and for 1 spice sample the agar method had lower repeatability variances (P less than 0.05). Relative standard deviations of repeatability were between 1.7 and 15.5% for the dry-film method and 1.2 and 16.0% for the agar method. Relative standard deviations of reproducibility ranged from 2.4 to 23.4% for the dry-film method and 2.3 to 18.8% for the agar method. The dry rehydratable film method has been adopted official first action for determination of the aerobic plate count.     

Dry rehydratable films for enumeration of coliforms and aerobic bacteria in dairy products: collaborative study

A collaborative study was conducted to compare proposed dry-film plating methods, using aerobic count plates and coliform count plates, to standard agar plating methods for quantifying aerobic bacteria and coliforms in dairy products. In this study, 5 food products (chocolate milk, pasteurized cheese, nonfat dry milk, evaporated milk, and vanilla ice cream), selected as representative dairy products, were analyzed by 11 collaborating laboratories. The results indicate that the dry-film plating methods are equivalent to or better than the agar plating methods. The aerobic count and coliform count dry-film plating methods have been adopted official first action.     

Gram-negative identification card for identification of Salmonella, Escherichia coli, and other Enterobacteriaceae isolated from foods: collaborative study

Twelve laboratories evaluated the Gram-Negative Identification (GNI) Card to identify members of the Enterobacteriaceae. Eighty-four isolates, previously isolated from foods, were used in the collaborative study; the isolates represented 12 genera within the Enterobacteriaceae group. Each collaborator streaked each isolate on tryptic soy agar plates for purity. In the method, plates are incubated 18-24 h at 35 degrees C. Isolated colonies are then subcultured to tryptic soy agar slants and incubated 18-24 h at 35 degrees C. An emulsion is made from the growth on the slant in 1.8 mL 0.45% sodium chloride solution. The GNI Card is filled and placed in a reader/incubator. Isolates are identified and an identification is printed. The Vitek System correctly identified 96.7% of Salmonella sp., 97.0% of Escherichia coli, and an average of 93.8% of the other enteric genera. The method using the Vitek System and GNI Card has been approved interim official first action by AOAC as a screening method for the presumptive identification of Salmonella sp., E. coli, and other Enterobacteriaceae isolated from foods.     

Enumeration of total bacteria and coliforms in milk by dry rehydratable film methods: collaborative study

Eleven laboratories participated in a collaborative study to compare the dry rehydratable film (Petrifilm SM and Petrifilm VRB) methods, respectively, to the standard plate count (SPC) and violet red bile agar (VRBA) standard methods for estimation of total bacteria and coliform counts in raw and homogenized pasteurized milk. Each laboratory analyzed 16 samples (8 different samples in blind duplicate) for total count by both the SPC and Petrifilm SM methods. A second set of 16 samples was analyzed by the VRBA and Petrifilm VRB methods. The repeatability standard deviations (the square root of the between-replicates variance) of the SPC, Petrifilm SM, VRBA, and Petrifilm VRB methods were 0.05104, 0.0444, 0.14606, and 0.13806, respectively; the reproducibility standard deviations were 0.7197, 0.06380, 0.15326, and 0.13806, respectively. The difference between the mean log10 SPC and the mean log10 Petrifilm SM results was 0.027. For the VRBA and Petrifilm VRB methods, the mean log10 difference was 0.013. These results generally indicate the suitability of the dry rehydratable film methods as alternatives to the SPC and VRBA methods for milk samples. The methods have been adopted official first action.     

Direct plating method for enumeration of Staphylococcus aureus: collaborative study.

Considerable evidence has been published regarding the adverse effect of sodium chloride on physiologically impaired cells of Staphylococcus aureus, such as are to be expected in processed foods. A direct plating method for enumeration of S. aureus eliminating the use of sodium chloride was devised and subjected to collaborative study by 16 analysts. Results obtained by the direct plating method were compared to those obtained by the AOAC official first action method (46.036--46.040). Participating analysts examined duplicate samples at population levels of 91, 34, and 20 S. aureus/g. Coefficients of variation among analysts were considerably lower for the direct plating method (31, 81, and 48%, respectively) than for method 46.040 (59, 156, and 150%, respectively) at all 3 population levels. High coefficients of variation for the direct plating method at 2 of the 3 levels were due principally to low populations of S. aureus. The direct plating method has been adopted as official first action for general purpose use and use of method 46.036--46.040 has been restricted to raw food ingredients and nonprocessed foods.     

Semiautomated method for niacin and niacinamide in food products: collaborative study.

A collaborative study was conducted comparing a semiautomated colorimetric niacin method with a manual colorimetric and a microbiological method for 10 food products. Seven laboratories used the microbiological method, 7 laboratories used the manual colorimetric method, and 6 laboratories used the semiautomated method. The semiautomated method was more repeatable within a laboratory and more reproducible between laboratories than was either of the other methods. The semiautomated method results compared favorably with both the microbiological and manual colorimetric method results.     

Determination of amprolium in feeds: collaborative study.

The official final action method, 42.028--42.032, for determining amprolium in feeds was modified by a change in the preparation of aluminum oxide for chromatography. A premix containing 0.5% amprolium was collaboratively studied by the modified and the official methods. Compared with the modified method, 87.7% of the drug was recovered from the premix by using the official method. The modification makes possible the assay of premixes as well as finished feeds. The official final action method has been modified to incorporate this change.     

Determination of zearalenone in corn: collaborative study.

Corn samples spiked at levels of 100, 300, 1000, and 2000 mug zearalenone/kg were sent to 22 collaborators for analysis by the Eppley method. All samples were yellow corn except one white corn sample spiked at 2000 mug/kg. Results from 16 collaborators were statistically analyzed. Only 4 of 16 collaborators detected zearalenone in the sample containing 100 mu/kg, but 11 detected the toxin in the sample containing 300 mug/kg. Average recoveries from all samples were 129% at 300 mug/kg, 101% at 1000 mug/kg, and 88% at 2000 mug/kg. The between-laboratory coefficients of variation were 53.0% at 300 mug/kg, 38.2% at 1000 mug/kg, and 27.0% at 2000 mug/kg. Five naturally contaminated corn samples, one in triplicate, were also provided. The mean level of zearalenone in the naturally contaminated samples ranged from 431 to 7622 mug/kg. The mean coefficient of variation for all samples was 40.5%. Two collaborators measured quantities of zearalenone on thin layer chromatographic plates densitometrically. Their results were not included in the statistical analysis, but the results indicated that densitometric measurement, given proper dilutions of solutions, could be used. The method has been adopted as official first action.     

Recovery of Salmonella species from nonfat dry milk rehydrated under rapid and reduced pre-enrichment conditions: collaborative study

A collaborative study was conducted to compare the relative efficiency of the AOAC rapid rehydration method with the reduced rehydration soak method for the recovery of Salmonella species from nonfat dry milk (NFDM). In the AOAC method, a 25 g sample of NFDM is rapidly rehydrated at a 1:9 sample/water ratio and mixed by swirling. After 60 min, the flask contents are adjusted to a pH of 6.8, and 0.45 mL of 1% aqueous brilliant green dye solution is added. The flasks are then incubated at 35 degrees C. In the soak method, a 25 g sample of NFDM is gently added to the sterile brilliant green (BG) water at a 1:9 sample/BG water ratio and allowed to soak undisturbed for 60 min at room temperature before incubation. Twelve collaborators analyzed 3 shipments of samples with the following results for the AOAC and soak methods: shipment 1-31 and 46 positive samples, respectively, with a 48% increase in detection by the soak method; shipment 3-45 and 66 positive samples, respectively, with a 47% increase in detection by the soak method; shipment 2--no significant difference in recovery of Salmonella species by the 2 methods. It is recommended that the official final action method for the detection of Salmonella species, 46.054-46.067, be revised to use the soak method for the analysis of nonfat dry milk.     

Microbiological method for assaying lincomycin in animal feed: collaborative study.

A microbiological assay for determining lincomycin in swine feed, supplement, and a vitamin-mineral premix was studied collaboratively in 16 laboratories. The design of the study involved a complete feed, feed supplement, and a vitamin-mineral premix covering a range of fortification from 20 to 80 g/ton and 80 to 2600 g/ton. Two methods of sample preparation were used depending on the concentration of lincomycin in the sample. Statistical evaluation of the results from the 2 methods indicated that 10 and 11 collaborators, respectively, had mean recoveries which were not significantly different from one another. Ten laboratories obtained a mean recovery of 112.2% (range 102.3--123.5%) for the lower level, and 11 laboratories obtained a mean recovery of 104.4% (range 100.0--107.7%) for the higher level. The method has been adopted as official first action.     

Improved hydrophobic grid membrane filter method, using EF-18 agar, for detection of Salmonella in foods: collaborative study

A collaborative study was carried out in 30 laboratories to validate improvements to the official final action hydrophobic grid membrane filter (HGMF) screening method for Salmonella in foods, 985.42, by comparing the performance of the improved HGMF method against that of the AOAC/BAM conventional culture method. Six products were included in the collaborative study: milk chocolate, raw deboned poultry meat, black pepper, soy flour, egg yolk powder, and nonfat dry milk. The raw deboned poultry meat was naturally contaminated with Salmonella, and the remaining 5 products were each inoculated in advance with low levels of individual Salmonella serotypes. The AOAC/BAM method produced 11 false negative results and the improved HGMF method produced 18 false negative results. The improved HGMF Salmonella method has been approved interim official first action for all foods to replace the HGMF official final action method, 985.42.