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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

Background Actero™ Salmonella Enrichment Media1 (Actero™ Salmonella) is a culture broth developed to recover Salmonella spp. from foods and environmental surfaces. Performance of Actero™ Salmonella broth has already been assessed and validated (AOAC Performance Tested MethodSM 041403) for the detection of Salmonella spp. in various foods, feeds and environmental samples. Objective This study aimed to validate the performance of a modified version of Actero™ Salmonella broth by incorporating one of the two liquid supplements into the powdered formula. Methods Inclusivity, exclusivity, stability, and lot-to-lot studies were carried out. Raw ground beef, chicken carcass rinse, dry pet food and stainless steel samples were enriched for 14–20 h at 35–39°C and analyzed using real-time PCR assay as well as by direct plating. Results The Probability of Detection assay confirmed the equivalent performance of the alternative methods as compared to the reference methods. All Salmonella strains, except Salmonella II : 57: z29:-, were able to grow in Actero™ Salmonella broth. One-half of the non-target strains did not grow in Actero™ Salmonella broth, whereas the atypical for Salmonella growth was observed for other non-target microorganisms subsequently plated onto selective and differential agars. Lot-to-lot consistency was demonstrated for three consecutively manufactured lots of the broth. The liquid broth was proven to be stable at 4°C for up to 9 weeks of storage. Conclusions and Highlights The incorporation of one of the two specific supplements into a powdered formula of Actero™ Salmonella broth made it more convenient to use without compromising the performance and accuracy.

The microbiological standard method for the detection of Salmonella spp. is time-consuming, and consequently there is a need for an alternative rapid methodology for its detection. In this study, two open-formula real-time PCR methods for detecting the inv A and ttr gene Salmonella spp. have been successfully optimized and in-house validated. Different DNA extraction procedures were used (boiling, Chelex resin and standard I - Biorad). The relative sensitivity and false positive ratio for the alternative methods were 100% and 0.0%, respectively. The relative trueness was in range from 96.8% to 99.2%. No false negative results were detected. The lowest Ct values were obtained using the protocol for detecting the ttr gene after DNA extraction by the Chelex. The results were compared in a large number of food and environmental samples to those of the SRPS EN ISO 6579:2008 standard method and commercial kit (IQ check® Salmonella II kit, Bio-Rad, USA). Practical applications The whole procedure of real-time PCR methods in this study was approximately 20 hr, in contrast to 4–5 days of analysis time for the standard method (SRPS EN ISO 6579:2008). The real-time PCR method also provides a high level of sensitivity and specificity with a low risk of cross-contamination. The implementation of the method for routine analysis will help improve safety in the food production chain, by providing results compatible with the ISO standard, but more rapidly.

The capacity to detect low levels of healthy and sub-lethally injured Salmonella enterica cells in chocolate by two alternative rapid detection methods iQ-Check(TM)Salmonella II real-time PCR (Bio-Rad) and VIDAS® Easy SLM (BioMérieux) was assessed and compared with ISO 6579:2005. Chocolate, a low moisture food known to support the survival of Salmonella, was challenged as food matrix. Buffered peptone water (BPW) did not support the recovery of low levels of sub-lethally injured S. enterica independent of the detection method, while BPW supplemented with milk powder enabled detection by the three examined methods. However, inhibition of real-time PCR was observed since for one out of three repetitions of chocolate inoculated with a low number of sub-lethally injured S. enterica cells, no PCR signal was obtained. Therefore, attention should be paid to the enrichment step to avoid false negative results due to the presence of especially sub-lethally injured Salmonella cells in chocolate. An appropriate sample preparation (such as enrichment media and conditions for incubation) remains the key factor for reliable detection including sub-lethally injured cells and should be evaluated, if necessary optimized, for each detection assay.

iQ-Check Salmonella II is a real-time PCR kit for detection of Salmonella in foods. Specific oligonucleotide probes are used to detect target DNA during the amplification, by hybridizing to the amplicons. These probes are linked to a fluorophore, which fluoresces only when hybridized to the target sequence. As part of an Emergency Response Validation due to a massive outbreak and subsequent recall, peanut butter was tested to compare the performance of iQ-Check Salmonella II to the U.S. Food and Drug Administration's Bacteriological Analytical Manual (FDA-BAM) reference method for detection of Salmonella. A single enrichment in buffered peptone water was used for a reduced enrichment time of 21 1 h over the 48 h reference method. There was no significant difference in the performance of the iQ-Check kit when compared to the FDA-BAM method, as determined by Chi-square analysis. All samples identified as positive by iQ-Check were confirmed by reference method protocol.

When the mutants of Salmonella lacking UDPGal-4-epimerase were grown in the presence of galactose, they exhibited marked bacteriolysis in ordinary medium and were convested to soheroplasts in hypertomic medium. Lysis was not observed in non-growing cells. Synthesis of cell wall does not seem to be inhibited by galactose at least up to 5–10 min before lysis, but cell wall synthesized in the presence of galactose had a sugar composition quite different from that formed in its absence. In the light of these findings, the mechanism of lysis was discussed, considering the metabolic consequences of the absence of epimerase.

Bacteriological investigations were performed with 31 samples of feces from reptiles, mainly from snakes of a snake exhibition in St. Gall. 27 Salmonella and Arizona serotypes were recovered from 20 specimens (64.5%). Seven of the Salmonella isolates belonged to the sub-genus I, two to the sub-genus II, while 14 could be classified as belonging to the sub-genus III (Arizona). Single infections have occurred in 13, and double infections in 7 of the cases under study. One Salmonella serotype of the sub-genus II – Salmonella II 13,23,36:1, Z281,5 – and three Arizona serotypes – Arizona 7a,7b:23:31, Arizona 23:29:21 and Arizona 24:26:30 – could be declared as new types.

Populations derived from Salmonella typhimurium surviving 1, 5, or 10 exposures to 0.5 Mrad did not show any increase in resistance to antibiotic discs containing 5 μg of tetracycline, 5 μg of chloromycetin, or 50 units of polymyxin. The frequency of resistance to 3 μg of tetracycline, 10 units of ampicillin, 120 μg of chloromycetin, or 30 μg of streptomycin per milliliter of agar, did not increase among progeny derived from either of two strains of S. typhimurium surviving 5–10 exposures to 0.5 Mrad. So, too, the virulence and radiation resistance of three strains of S. typhimurium exposed to radiation-growth cycling remained essentially unchanged or decreased.A mixture which originally contained 10 different serotypes of Salmonella was cycled at 0.23 Mrad. After 10 exposures the frequency of resistance of the progeny to ampicillin, chloromycetin, and streptomycin increased 87-, 55- and 13-fold respectively. The LD50 was essentially the same, but the radiation resistance (D value) increased from 0.053 Mrad in controls to 0.074 Mrad.The evidence presented seems to indicate that mutants are attained more readily from Salmonella cultures recycled at 0.25 Mrad or less than from those recycled at 0.5 Mrad. The pathogenicity of either one was usually essentially unchanged or reduced.