Evaluation of VIDAS Salmonella (SLM) immunoassay method with Rappaport-Vassiliadis (RV) medium for detection of Salmonella in foods: collaborative study

A collaborative study was conducted to compare the VIDAS Salmonella (SLM) with Rappaport-Vassiliadis (RV) method for detection of Salmonella in foods to the current standard method presented in the U.S. Food and Drug Administration's Bacteriological Analytical Manual (BAM) and the culture method presented in AOAC's Official Methods of Analysis. The VIDAS SLM with RV method uses tetrathionate broth in combination with RV medium in place of selenite cystine broth for selective enrichment, thereby eliminating the hazardous waste issue for laboratories. Twenty five laboratories participated in the evaluation, each testing one or more of 8 test products: nonfat dry milk, dried egg, soy flour, lactic casein, milk chocolate, raw ground pork, raw ground turkey, and raw peeled shrimp. Results of the study showed no significant differences in the numbers of confirmed positive samples with the VIDAS SLM with RV procedure and the BAM/AOAC culture procedure. The VIDAS SLM with RV method was effective for rapid detection of Salmonella in foods. It is recommended that AOAC INTERNATIONAL modify the VIDAS Salmonella SLM procedure to include the RV method.     

Evaluation of the BAX system for detection of Salmonella in selected foods: collaborative study

A multilaboratory study was conducted to compare the automated BAX System to the standard cultural methods for detection of Salmonella in selected foods. Five food types--frankfurters, raw ground beef, mozzarella cheese, raw frozen tilapia fish, and orange juice--at 3 inoculation levels, were analyzed by each method. A sixth food type, raw ground chicken, was tested using 3 naturally contaminated lots. A total of 16 laboratories representing government and industry participated. In this study, 1386 samples were analyzed, of which 1188 were paired samples and 198 were unpaired samples. Of the 1188 paired samples, 461 were positive by both methods and 404 were negative by both methods. Thirty-seven samples were positive by the BAX System but negative by the standard reference method, and 11 samples were positive by standard cultural method and negative by the BAX System. Of the 198 unpaired samples, 106 were positive by the BAX System and 60 were positive by the standard cultural method. A Chi square analysis of each of the 6 food types, at the 3 inoculation levels tested, was performed. For all foods, the BAX System demonstrated results comparable to those of the standard reference methods based on the Chi square results.     

RapidChek SELECT Salmonella enteritidis test system for the detection of Salmonella enteritidis in poultry house drag swabs, shell egg pools, and chicken carcass rinsates

The RapidChek SELECT Salmonella Enteritidis Test System was validated for the detection of Salmonella Enteritidis (SE) in poultry house drag swabs, shell egg pools, and chicken carcass rinsates. The method utilizes RapidChek SELECT Salmonella (AOAC PTM License No. 080601) proprietary primary and secondary enrichment media. Following enrichment, an immunochromatographic test strip is inserted into the tube containing the secondary enrichment broth, developed for 10 min, and interpreted. Salmonella Enteritidis-inoculated samples (1-5 CFU SE/analytical unit) were tested by the test method as well as the appropriate cultural reference method U.S. Food and Drug Administration-Bacteriological Analytical Manual (drag swabs and egg pools) or U.S. Department of Agriculture-Food Safety and Inspection Service (chicken carcass rinsates). A total of 80 samples were tested by both methods in the study. Fifty-two samples were positive by the RapidChek SELECT Salmonella Enteritidis method and 38 were found positive by the respective reference method. The sensitivity of the method was 100% and the specificity was 100%. The accuracy of the test method was 137%, indicating that the method was more sensitive than the reference method. The RapidChek SELECT Salmonella Enteritidis method was tested with 82 Salmonella Group D1 strains including 63 Salmonella Enteritidis strains as well as 32 non-Salmonella Group D1 strains representing 10 bacteria genera. The test method detected all 82 Group D1 strains (100% sensitivity). None of the non-Salmonella Group D1 or other genera of bacteria were detected, indicating a specificity of 100%. The method was shown to be highly robust and stable under control and accelerated stability conditions.     

Salmonella in foods: new enrichment procedure for TECRA Salmonella Visual Immunoassay using a single rv(R10) only, TT only, or dual rv(R10) and TT selective enrichment broths (AOAC official method 998.09): collaborative study

A collaborative study was conducted to compare a new enrichment procedure for the TECRA Salmonella Visual Immunoassay (TSVIA) with the reference method given in the U.S. Food and Drug Administration's Bacteriological Analytical Manual (7th Ed.). Three food types (milk powder, pepper, and soy flour) were analyzed in Australia and 3 food types (milk chocolate, dried egg, and raw turkey) were analyzed in the United States. Thirty-eight collaborators participated in the study. The TECRA method was evaluated using both Rappaport-Vassiliadis R10 (RV(R10)) and tetrathionate (TT) broths for selective enrichment. M broth cultures arising from each of the 2 selective enrichment broths were tested in the TSVIA using 2 individual wells, one for each selective broth, and a single well to test the pooled selective enrichment broths. The results for the pooled enrichment broths were reported elsewhere. This study presents the results for the use of single enrichment broths, i.e., RV(R10) only or TT only, with the TSVIA. No significant differences (p > 0.05) were observed for the pairwise comparison of the proportion of positive samples for either RV(R10) or TT used as a single enrichment broth for the TSVIA with that for the reference method.     

Evaluation of VIDAS Immuno-Concentration Salmonella (ICS)/VIDAS Salmonella (SLM) immunoassay method for detection of Salmonella in selected foods (Method Modification 2001.09): collaborative study

A new method for detection of Salmonella in foods in a minimum of 24 h was adopted as an AOAC Official First Action Method for selected foods (2001.09) using both the VIDAS Immuno-Concentration Salmonella (ICS) and VIDAS Salmonella (SLM) methods.     

Reveal Salmonella 2.0 test for detection of Salmonella spp. in foods and environmental samples. Performance Tested Method 960801

Reveal Salmonella 2.0 is an improved version of the original Reveal Salmonella lateral flow immunoassay and is applicable to the detection of Salmonella enterica serogroups A-E in a variety of food and environmental samples. A Performance Tested Method validation study was conducted to compare performance of the Reveal 2.0 method with that of the U.S. Department of Agriculture-Food Safety and Inspection Service or U.S. Food and Drug Administration/Bacteriological Analytical Manual reference culture methods for detection of Salmonella spp. in chicken carcass rinse, raw ground turkey, raw ground beef, hot dogs, raw shrimp, a ready-to-eat meal product, dry pet food, ice cream, spinach, cantaloupe, peanut butter, stainless steel surface, and sprout irrigation water. In a total of 17 trials performed internally and four trials performed in an independent laboratory, there were no statistically significant differences in performance of the Reveal 2.0 and reference culture procedures as determined by Chi-square analysis, with the exception of one trial with stainless steel surface and one trial with sprout irrigation water where there were significantly more positive results by the Reveal 2.0 method. Considering all data generated in testing food samples using enrichment procedures specifically designed for the Reveal method, overall sensitivity of the Reveal method relative to the reference culture methods was 99%. In testing environmental samples, sensitivity of the Reveal method relative to the reference culture method was 164%. For select foods, use of the Reveal test in conjunction with reference method enrichment resulted in overall sensitivity of 92%. There were no unconfirmed positive results on uninoculated control samples in any trials for specificity of 100%. In inclusivity testing, 102 different Salmonella serovars belonging to serogroups A-E were tested and 99 were consistently positive in the Reveal test. In exclusivity testing of 33 strains of non-salmonellae representing 14 genera, 32 were negative when tested with Reveal following nonselective enrichment, and the remaining strain was found to be substantially inhibited by the enrichment media used with the Reveal method. Results of ruggedness testing showed that the Reveal test produces accurate results even with substantial deviation in sample volume or device development time.     

Evaluation of VIDas immuno-concentration Salmonella/VIDAS salmonella immunoassay method for detection of Salmonella in selected foods: collaborative study

The VIDAS Immuno-concentration Salmonella ICS)/VIDAS Salmonella (SLM) immunoassay method for the detection of Salmonella was compared to the Bacteriological Analytical Manual (BAM)/AOAC culture method in a collaborative study. Thirty-two laboratories participated in the evaluation. Each laboratory tested one or more of the 6 test products: milk chocolate, nonfat dry milk, dried whole egg, soy flour, ground black pepper, and ground raw turkey. The 2 methods were in agreement for 1,266 of the 1,440 samples. Of the 174 samples not in agreement, 69 were VIDAS CS/SLM-positive and BAM/AOAC-negative and 105 were VIDAS ICS/SLM-negative and BAM/AOAC-positive.     

Modification of the BAX System PCR assay for detecting Salmonella in beef, produce, and soy protein isolate. Performance Tested Method 100201

The BAX System PCR assay for Salmonella detection in foods was previously validated as AOAC Research Institute (RI) Performance Tested Method (PTM) 100201. New studies were conducted on beef and produce using the same media and protocol currently approved for the BAX System PCR assay for E. coli O157:H7 multiplex (MP). Additionally, soy protein isolate was tested for matrix extension using the U.S. Food and Drug Administration-Bacteriological Analytical Manual (FDA-BAM) enrichment protocols. The studies compared the BAX System method to the U.S. Department of Agriculture culture method for detecting Salmonella in beef and the FDA-BAM culture method for detecting Salmonella in produce and soy protein isolate. Method comparison studies on low-level inoculates showed that the BAX System assay for Salmonella performed as well as or better than the reference method for detecting Salmonella in beef and produce in 8-24 h enrichment when the BAX System E. coli O157:H7 MP media was used, and soy protein isolate in 20 h enrichment with lactose broth followed by 3 h regrowth in brain heart infusion broth. An inclusivity panel of 104 Salmonella strains with diverse serotypes was tested by the BAX System using the proprietary BAX System media and returned all positive results. Ruggedness factors involved in the enrichment phase were also evaluated by testing outside the specified parameters, and none of the factors examined affected the performance of the assay.     

Salmonella in foods--a new enrichment procedure for use with the TECRA Salmonella visual immunoassay: collaborative study.

A collaborative study was conducted to compare a new enrichment procedure for the TECRA Salmonella Visual Immunoassay with the reference method given in the U.S. Food and Drug Administration's Bacteriological Analytical Manual (BAM 7th Ed.). Three food types (milk powder, black pepper, and soy flour) were analyzed in Australia, and 3 food types (milk chocolate, dried egg, and raw turkey) were analyzed in the United States. Thirty-eight collaborators participated in the study. No significant differences (p > 0.05) were observed for the pairwise comparison of the proportion of positive samples for the TECRA method with that for the reference method. The new enrichment procedure for the TECRA method has been adopted First Action by AOAC INTERNATIONAL.     

The effect of preenrichment and selective enrichment media on recovery of Salmonella Typhi from the tropical fruit mamey

The Bacteriological Analytical Manual (BAM) Salmonella culture method did not detect Salmonella Typhi in mamey, the tropical fruit that was implicated in a 1999 typhoid outbreak. The relative effectiveness of BAM's nonselective preenrichment and selective media for the recovery of S. Typhi from mamey was examined to determine if the BAM's preenrichment/selective enrichment strategy was the cause of the method's failure with this food. The preenrichment media were lactose broth, buffered peptone water (BPW), and universal preenrichment (UP) broth. The selective enrichment media were selenite cystine (SC) broth, tetrathionate (TT) broth, and Rappaport-Vassiliadis (RV) medium. UP broth was significantly more effective (P < 0.05) than either lactose broth or BPW for the recovery of 2 different S. Typhi strains from mamey. Of 120 test portions tested, 105 S. Typhi-positive test portions were recovered using UP broth, whereas only 1 S. Typhi-positive test portion was recovered using BPW, and no S. Typhi-positive test portions were recovered using lactose broth. SC and TT broths were both significantly more effective (P < 0.05) than RV medium. Of 105 S. Typhi-positive test portions, SC broth recovered 80, TT broth recovered 67, and RV medium recovered 9. After the above comparison, an incomplete UP (UPI) broth formulation was found to be significantly more effective (P < 0.05) than the complete formulation (UPC). Of 80 total positive test portions, UPI recovered 71, whereas UPC recovered only 48. The following UP broth formulations were compared to determine if any of the components of the UP broth formulation were inhibitory to S. Typhi: UPC, UP broth without sodium pyruvate (UPS), UP broth without ferric ammonium citrate (UPF), UP broth without MgSO4 (UPM), and UPI. It was found that none of the ingredients were inhibitory to S. Typhi and that, out of 140 total test portions, UPI and UPF, with 108 and 103 positive test portions, respectively, were significantly more effective (P < 0.05) than the other UP broth formulations (82 for UPC, 74 for UPS, and 60 for UPM). Rather than being inhibitory to the growth of S. Typhi, it appears that ferric ammonium citrate enhanced the growth of competitors which suppressed the growth of S. Typhi. These results demonstrate that UPI and UPF are effective for the recovery of S. Typhi from mamey.     

Evaluation of the VIDAS Listeria (LIS) immunoassay for the detection of Listeria in foods using demi-Fraser and Fraser enrichment broths, as modification of AOAC Official Method 999.06 (AOAC Official Method 2004.06)

A multilaboratory study was conducted to compare the VIDAS LIS immunoassay with the standard cultural methods for the detection of Listeria in foods using an enrichment modification of AOAC Official Method 999.06. The modified enrichment protocol was implemented to harmonize the VIDAS LIS assay with the VIDAS LMO2 assay. Five food types--brie cheese, vanilla ice cream, frozen green beans, frozen raw tilapia fish, and cooked roast beef--at 3 inoculation levels, were analyzed by each method. A total of 15 laboratories representing government and industry participated. In this study, 1206 test portions were tested, of which 1170 were used in the statistical analysis. There were 433 positive by the VIDAS LIS assay and 396 positive by the standard culture methods. A Chi-square analysis of each of the 5 food types, at the 3 inoculation levels tested, was performed. The resulting average Chi square analysis, 0.42, indicated that, overall, there are no statistical differences between the VIDAS LIS assay and the standard methods at the 5% level of significance.     

Precollaborative study of the GeneQuence Salmonella assay using 24-hour enrichment protocols for detection of Salmonella spp. in select foods

New enrichment protocols are described for use with a DNA hybridization (DNAH) method for detection of Salmonella spp. in select foods. GeneQuence Salmonella, in its original version, utilized a 3-stage enrichment of minimum 42 h duration. New 2-stage procedures of 24-28 h duration are described for raw poultry, raw beef, pasteurized egg products, milk chocolate, and dry pet food. In the validation study described here, a total of 345 samples were tested by the abbreviated DNAH method in parallel with either the U.S. Food and Drug Administration's Bacteriological Analytical Manual (FDA/BAM) or U.S. Department of Agriculture-Food Safety and Inspection Service (USDA-FSIS) reference culture procedures. Results showed an overall sensitivity for the DNAH method of 97.1% (false-negative rate 2.9%). There were no false-positive results by the DNAH method; therefore the specificity was 100%. Overall agreement between the DNAH and reference culture methods was 98.5%. There were no significant differences in performance between the DNAH and reference methods for any of the foods tested as determined by Chi-square analysis. It is recommended that the DNAH method be subjected to AOAC collaborative study.     

Evaluation of VIDAs Immuno-concentration Salmonella assay Plus selective plate method (Hektoen enteric,bismuth sulfite,Salmonella identification) for detection of Salmonella in selected foods: collaborative study

The VIDAS Immuno-concentration Salmonella (ICS) plus selective plate method (Hektoen enteric, bismuth sulfite, Salmonella identification) method for the detection of Salmonella was compared to the Bacteriological Analytical Manual (BAM)/AOAC culture method in a collaborative study. Thirty-two laboratories participated in the evaluation. Each laboratory tested one or more of the 6 test products: milk chocolate, nonfat dry milk, dried whole egg, soy flour, ground black pepper, and ground raw turkey. The 2 methods were in agreement for 1,283 of the 1,440 test samples. Of the 157 test samples not in agreement, 82 were VIDAS ICS plus selective plate-positive and BAM/AOAC-negative, and 75 were VIDAS ICS plus selective plate-negative and BAM/AOAC-positive.     

Visual immunoprecipitate assay eight hour method for detection of enterohemorrhagic Escherichia coli O157:H7 in raw and cooked beef (modification of AOAC Official Method 996.09): collaborative study

AOAC Official Method 996.09, Visual Immunoprecipitate Assay (VIP) for Escherichia coli O157:H7, was modified to incorporate a new enrichment protocol using BioControl EHEC8 medium for testing raw and cooked beef. Foods were tested by VIP assay and the U.S. Department of Agriculture/Food Safety and Inspection Service (USDA/FSIS) enrichment procedure and the FDA Bacteriological Analytical Manual (BAM) isolation and confirmation techniques. A total of 15 collaborators participated. Raw and cooked ground beef were inoculated with E. coli O157:H7 at 2 different levels: a high level, where predominantly positive results were expected, and a low level where fractional recovery was anticipated. Collaborators tested 396 test portions and controls by both methods, for a total of 792 test portions. Of the 396 paired test portions, 75 were positive and 230 were negative by both the VIP and culture methods. Eleven test portions were presumptively positive by VIP and could not be confirmed culturally; 32 were negative by VIP, but confirmed positive by culture; and 65 were negative by the culture method, but confirmed positive by the VIP method. There was no statistical difference between results obtained with the VIP for EHEC 8 h method and the culture method except for cooked beef, where the VIP had significantly higher recovery for one inoculation level.     

Modification of the BAX Salmonella test kit to include a hot start functionality (modification of AOAC Official Method 2003.09)

In 2010, the BAX System PCR assay for Salmonella was modified to include a hot start functionality designed to keep the reaction enzyme inactive until PCR begins. To validate the assay's Official Methods of Analysis status to include this procedure modification, an evaluation was conducted on four food types that were simultaneously analyzed with the BAX System and either the U.S. Food and Drug Administration's Bacteriological Analytical Manual or the U.S. Department of Agriculture-Food Safety and Inspection Service Microbiology Laboratory Guidebook reference method for detecting Salmonella. Identical performance between the BAX System method and the reference methods was observed. Additionally, lysates were analyzed using both the BAX System Classic and BAX System Q7 instruments with identical results using both platforms for all samples tested. Of the 100 samples analyzed, 34 samples were positive for both the BAX System and reference methods, and 66 samples were negative by both the BAX System and reference methods, demonstrating 100% correlation. No instrument platform variation was observed. Additional inclusivity and exclusivity testing using the modified test kit demonstrated the test kit to be 100% accurate in evaluation of test panels of 352 Salmonella strains and 46 non-Salmonella strains.     

Evaluation of the GeneQuence DNA hybridization method in conjunction with 24-hour enrichment protocols for detection of Salmonella spp. in select foods: collaborative study

A multilaboratory study was conducted to compare performance of the GeneQuence DNA hybridization (DNAH) method incorporating new 24 h enrichment protocols and reference culture procedures for detection of Salmonella spp. in select foods. Six food types (raw ground turkey, raw ground beef, dried whole egg, milk chocolate, walnuts, and dry pet food) were tested by the DNAH method and by the culture methods of either the U.S. Department of Agriculture-Food Safety and Inspection Service (USDA-FSIS) or the U.S. Food and Drug Administration's Bacteriological Analytical Manual (FDA/BAM). Fifteen laboratories participated in the study. Four of the foods tested (raw ground turkey, dried whole egg, milk chocolate, and dry pet food), showed no statistically significant differences in performance between the DNAH method and the reference procedure as determined by Chi square analysis. Sensitivity rates for the DNAH method ranged from 92 to 100%. The DNAH method, with the specific enrichment protocol evaluated, was found to be ineffective for detection of Salmonella spp. in walnuts. For raw ground beef, results from one trial showed a statistically significant difference in performance, with more positives obtained by the reference method. However, evidence suggests that the difference in the number of positives was likely due to lack of homogeneity of the test samples rather than to DNAH method performance.     

Validation study to demonstrate the equivalence of a minor modification (TECRA ULTIMA protocol) to AOAC Method 998.09 (TECRA Salmonella Visual Immunoassay) with the cultural reference method

The TECRA Salmonella Visual Immunoassay (VIA) using Rappaport-Vassiliadis RV[R10] as a single selective enrichment broth has Final Action approval (AOAC Method 998.09). TECRA has recently developed a protocol (TECRA ULTIMA), which involves the addition of a new additive to a 1 mL aliquot of the RV[R10] broth, prior to the heat-killing step, thereby allowing the RV[R10] broth to be tested directly in the kit and thus eliminating the need for the 2 h post-enrichment in M broth. An in-house validation study was conducted to compare the modified AOAC Method 998.09 to the reference culture method. Three foods were used in the study: Naturally contaminated raw ground poultry at high (10-50 cells/25 g), and low (1-5 cells/25 g) levels; and milk powder and peanut butter, artificially inoculated at low and high levels with Salmonella bovismorbificans and S. enterica Mbandaka, respectively. Twenty test portions were analyzed for each level with 10 uninoculated control samples per food. Overall, no significant differences (p <0.05) were observed when the proportion of positive test portions for the modified VIA were compared with that for the reference method. This minor modification, which employs the additive (provided in the TECRA ULTIMA SALMONELLA Test Kit) to permit the direct analysis of RV[R10] broth has demonstrated the utility of the TECRA ULTIMA SALMONELLA protocol. It is recommended that the minor modification to Method 998.09 be approved First Action as an additional option within the method.     

TECRA Unique test for rapid detection of Salmonella in food: collaborative study

The TECRA Unique Salmonella test uses the principle of immunoenrichment to allow rapid detection of Salmonellae in food. A collaborative study was conducted to compare the TECRA Salmonella Unique test with the reference culture method given in the U.S. Food and Drug Administration's Bacteriological Analytical Manual. Three food types (milk powder, pepper, and soy flour) were analyzed in Australia and 2 food types (milk chocolate and dried egg) were analyzed in the United States. Forty-one collaborators participated in the study. For each of the 5 foods at each of the 3 levels, a comparison showed no significant differences (p > or = 0.05) in the proportion of positive test samples for Unique and that for the reference method using the Chi-square test for independence with continuity correction.     

Reveal Salmonella enteritidis test for detection of Salmonella enteritidis in shell eggs and environmental samples

Reveal Salmonella Enteritidis (SE) is a lateral flow-based immunodiagnostic assay used for rapid detection of Salmonella enterica serovar Enteritidis from pooled shell eggs and environmental samples. This assay uses highly specific antibodies to accurately detect S. Enteritidis. Studies were conducted to compare the performance of this test against reference procedures for detection of S. Enteritidis from both pooled shell eggs and environmental samples. Pooled shell eggs were inoculated with low levels ofS. Enteritidis and were enriched according to the procedure prescribed by the U.S. Food and Drug Administration. Uninoculated samples were included in each trial. Reveal SE exhibited 100% sensitivity and 100% specificity in comparison to the reference method in all trials. An abbreviated 48 h/(no hold) enrichment procedure was also developed and validated for detection ofS. Enteritidis from pooled shell egg samples. This shortened enrichment procedure can be used in conjunction with the Reveal SE test and offers a significant enrichment time savings of 96 h. Chi-square analysis revealed that there was no significant difference between the abbreviated Reveal method and the reference procedure for detection ofS. Enteritidis from pooled shell egg samples. Out of 245 natural drag swabs screened internally, only three samples tested Reveal SE positive and were confirmed by the reference procedure, resulting in 100% sensitivity and 100% specificity. An external laboratory screened 147 poultry house environmental samples and obtained 35 Reveal SE confirmed positives for Reveal SE sensitivity of 100% and specificity of 90%. Inoculation trials with drag swabs resulted in 96% sensitivity and 100% specificity. Thus, these data demonstrate that Reveal SE is a highly sensitive and specific assay for the detection of S. Enteritidis from both pooled shell eggs and environmental samples.     

Salmonella in selected foods by VIDAS immuno-concentration Salmonella plus selective plate method (Hektoen enteric,xylose lysine desoxycholate,bismuth sulfite): collaborative study

The VIDAS Immuno-concentration Salmonella (ICS) plus selective plate method (Hektoen enteric, xylose lysine desoxycholate, bismuth sulfite) method for the detection of Salmonella was compared to the Bacteriological Analytical Manual (BAM)/AOAC culture method in a collaborative study. Thirty-two laboratories participated in the evaluation. Each laboratory tested one or more of the 6 test products: milk chocolate, nonfat dry milk, dried whole egg, soy flour, ground black pepper, and ground raw turkey. The 2 methods were in agreement for 1,297 of the 1,455 samples. Of the 158 samples not in agreement, 82 were VIDAS ICS plus selective plate-positive and BAM/AOAC-negative, and 76 were VIDAS ICS plus selective plate-negative and BAM/AOAC-positive.