Sample Size Determination for Food Sampling.

Industry and public health agencies sample and test food products for various purposes related to food safety and quality. Methods of sample selection and sample size determination are important in designing an optimal sampling plan. The appropriate sample size of a sampling plan depends on the objective. We examine the methods of sample size calculation for the following four objectives commonly associated with food sampling: (1) estimate prevalence (e.g., of contaminated products), (2) detect presence (e.g., of contaminated products), (3) estimate maximum prevalence, and (4) compare estimated prevalence with a specified value (e.g., a previous estimate or a threshold value). We illustrate these methods using examples and provide a web-based application (https://simple-sample.galaxytrakr.org/) written in R, using the shiny package, to help users with the application of each method.

On the Estimation of POD and LOD of Qualitative Microbiological Assays from a Multi-Laboratory Validation Study.

BACKGROUND: Jarvis et al. in 2019 (J. AOAC Int. 102: 1617-1623) estimated the mean laboratory effect (µ), standard deviation of laboratory effects (σ), probability of detection (POD), and level of detection (LOD) from a multi-laboratory validation study of qualitative microbiological assays using a random intercept complementary log-log model. Their approach estimated σ based on a Laplace approximation to the likelihood function of the model, but estimated µ from a fixed effectmodel due to a limitation in the MS Excel spreadsheet which was used by the authors to develop a calculation tool. OBJECTIVE: We compared the estimates of µ and σ from three approaches (the Laplace approximation that estimates µ and σ simultaneously from the random intercept model, adaptive Gauss-Hermite quadrature (AGHQ), and the method of Jarvis et al.) and introduced an R Shiny app to implement the AGHQ using the widely used "lme4" R package. METHODS: We conducted a simulation study to compare the accuracy of the estimates of µ and σ from the three approaches and compared the estimates of µ, σ, LOD, etc. between the R Shiny app and the spreadsheet calculation tool developed by Jarvis et al. for an example dataset. RESULTS: Our simulation study shows that, while the three approaches produce similar estimates of σ, the AGHQ has generally the best performance for estimating µ (and hence mean POD and LOD). The differences in the estimates between the R Shiny app and the spreadsheet were demonstrated using the example dataset. CONCLUSION: The AGHQ is the best method for estimating µ, POD, and LOD. HIGHLIGHTS: The user-friendly R Shiny app provides a better alternative to the spreadsheet.

Microbiome Population Dynamics of Cold-Smoked Sockeye Salmon during Refrigerated Storage and after Culture Enrichment.

ABSTRACT: Cold-smoked salmon is a ready-to-eat seafood product of high commercial importance. The processing and storage steps facilitate the introduction, growth, and persistence of foodborne pathogens and spoilage bacteria. The growth of commensal bacteria during storage and once the product is opened also influence the quality and safety of cold-smoked salmon. Here we investigated the microbial community through targeted 16S rRNA gene and shotgun metagenomic sequencing as means to better understand the interactions among bacteria in cold-smoked salmon. Cold-smoked salmon samples were tested over 30 days of aerobic storage at 4°C and cultured at each time point in a buffered Listeria enrichment broth (BLEB) commonly used to detect Listeria in foods. The microbiomes were composed of Firmicutes and Proteobacteria, namely, Carnobacterium, Brochothrix, Pseudomonas, Serratia, and Psychrobacter. Pseudomonas species were the most diverse species, with 181 taxa identified. In addition, we identified potential homologs to 10 classes of bacteriocins in microbiomes of cold-smoked salmon stored at 4°C and corresponding BLEB culture enrichments. The findings presented here contribute to our understanding of microbiome population dynamics in cold-smoked salmon, including changes in bacterial taxa during aerobic cold storage and after culture enrichment. This may facilitate improvements to pathogen detection and quality preservation of this food.

Dietary Supplement Adverse Event Report Data From the FDA Center for Food Safety and Applied Nutrition Adverse Event Reporting System (CAERS), 2004-2013.

BACKGROUND: The Food and Drug Administration (FDA)'s Center for Food Safety and Applied Nutrition (CFSAN) oversees the safety of the nation's foods, dietary supplements, and cosmetic products. OBJECTIVE: To present a descriptive analysis of the 2004-2013 dietary supplement adverse event report (AER) data from CAERS and evaluate the 2006 Dietary Supplements and Nonprescription Drug Consumer Protection Act as pertaining to dietary supplements adverse events reporting. METHODS: We queried CAERS for data from the 2004-2013 AERs specifying at least 1 suspected dietary supplement product. We extracted the product name(s), the symptom(s) reported, age, sex, and serious adverse event outcomes. We examined time trends for mandatory and voluntary reporting and performed analysis using SAS v9.4 and R v3.3.0 software. RESULTS: Of the total AERs (n = 15 430) received from January 1, 2004, through December 31, 2013, indicating at least 1 suspected dietary supplement product, 66.9% were mandatory, 32.2% were voluntary, and 0.9% were both mandatory and voluntary. Reported serious outcomes included death, life-threatening conditions, hospitalizations, congenital anomalies/birth defects and events requiring interventions to prevent permanent impairments (5.1%). The dietary supplement adverse event reporting rate in the United States was estimated at ~2% based on CAERS data. CONCLUSIONS: This study characterizes CAERS dietary supplement adverse event data for the 2004-2013 period and estimates a reporting rate of 2% for dietary supplement adverse events based on CAERS data. The findings show that the 2006 Dietary Supplements and Nonprescription Drug Consumer Protection Act had a substantial impact on the reporting of adverse events.

Contribution of ionic silver to genotoxic potential of nanosilver in human liver HepG2 and colon Caco2 cells evaluated by the cytokinesis-block micronucleus assay.

Extensive human exposure to food- and cosmetics-related consumer products containing nanosilver is of public concern because of the lack of information about their safety. Genotoxicity is an important endpoint for the safety and health hazard assessment of regulated products including nanomaterials. The in vitro cytokinesis-block micronucleus (CBMN) assay is a very useful test for predictive genotoxicity testing. Recently, we have reported the genotoxicity of 20 nm nanosilver in human liver HepG2 and colon Caco2 cells evaluated using the CBMN assay. The objective of our present study was three-fold: (i) to evaluate if HepG2 and Caco2 cells are valuable in vitro models for rapid genotoxicity screening of nanosilver; (ii) to test the hypothesis that the nanoparticle size and cell types are critical determinants of its genotoxicity; and (iii) to determine if ionic silver contributes to the nanosilver genotoxicity. With these objectives in mind, we evaluated the genotoxic potential of 50 nm nanosilver of the same shape, composition, surface charge, obtained from the same commercial source, under the same experimental conditions and the same genotoxic CBMN endpoint used for the previously tested 20 nm silver. The ionic silver (silver acetate) was also evaluated under the same conditions. Results of our study show that up to the concentrations tested in these cell types, the smaller (20 nm) nanosilver induces micronucleus formation in both the cell types but the larger (50 nm) nanosilver and the ionic silver provide a much weaker response compared with controls under the same conditions.

Flow cytometric evaluation of the contribution of ionic silver to genotoxic potential of nanosilver in human liver HepG2 and colon Caco2 cells.

Exposure to nanosilver found in food- and cosmetics-related consumer products is of public concern because of the lack of information about its safety. In this study, two widely used in vitro cell culture models, human liver HepG2 and colon Caco2 cells, and the flow cytometric micronucleus (FCMN) assay were evaluated as tools for rapid predictive screening of the potential genotoxicity of nanosilver. Recently, we reported the genotoxicity of 20 nm nanosilver using these systems. In the current study presented here, we tested the hypothesis that the nanoparticle size and cell types were critical determinants of its genotoxicity. To test this hypothesis, we used the FCMN assay to evaluate the genotoxic potential of 50 nm nanosilver of the same shape, composition, surface charge and obtained from the same commercial source using the same experimental conditions and in vitro models (HepG2 and Caco2) as previously tested for the 20 nm silver. Results of our study show that up to the concentrations tested in these cultured cell test systems, the smaller (20 nm) nanoparticle is genotoxic to both the cell types by inducing micronucleus (MN). However, the larger (50 nm) nanosilver induces MN only in HepG2 cells, but not in Caco2 cells. Also in this study, we evaluated the contribution of ionic silver to the genotoxic potential of nanosilver using silver acetate as the representative ionic silver. The MN frequencies in HepG2 and Caco2 cells exposed to the ionic silver in the concentration range tested are not statistically significant from the control values except at the top concentrations for both the cell types. Therefore, our results indicate that the ionic silver may not contribute to the MN-forming ability of nanosilver in HepG2 and Caco2 cells. Also our results suggest that the HepG2 and Caco2 cell cultures and the FCMN assay are useful tools for rapid predictive screening of a genotoxic potential of food- and cosmetics-related chemicals including nanosilver.

Comparative cytotoxicity of nanosilver in human liver HepG2 and colon Caco2 cells in culture.

The use of silver nanoparticles in food, food contact materials, dietary supplements and cosmetics has increased significantly owing to their antibacterial and antifungal properties. As a consequence, the need for validated rapid screening methods to assess their toxicity is necessary to ensure consumer safety. This study evaluated two widely used in vitro cell culture models, human liver HepG2 cells and human colon Caco2 cells, as tools for assessing the potential cytotoxicity of food- and cosmetic-related nanoparticles. The two cell culture models were utilized to compare the potential cytotoxicity of 20-nm silver. The average size of the silver nanoparticle determined by our transmission electron microscopy (TEM) analysis was 20.4 nm. The dynamic light scattering (DLS) analysis showed no large agglomeration of the silver nanoparticles. The concentration of the 20-nm silver solution determined by our inductively coupled plasma-mass spectrometry (ICP-MS) analysis was 0.962 mg ml(-1) . Our ICP-MS and TEM analysis demonstrated the uptake of 20-nm silver by both HepG2 and Caco2 cells. Cytotoxicity, determined by the Alamar Blue reduction assay, was evaluated in the nanosilver concentration range of 0.1 to 20 µg ml(-1) . Significant concentration-dependent cytotoxicity of the nanosilver in HepG2 cells was observed in the concentration range of 1 to 20 µg ml(-1) and at a higher concentration range of 10 to 20 µg ml(-1) in Caco2 cells compared with the vehicle control. A concentration-dependent decrease in dsDNA content was observed in both cell types exposed to nanosilver but not controls, suggesting an increase in DNA damage. The DNA damage was observed in the concentration range of 1 to 20 µg ml(-1) . Nanosilver-exposed HepG2 and Caco2 cells showed no cellular oxidative stress, determined by the dichlorofluorescein assay, compared with the vehicle control in the concentration range used in this study. A concentration-dependent decrease in mitochondria membrane potential in both nanosilver exposed cell types suggested increased mitochondria injury compared with the vehicle control. The mitochondrial injury in HepG2 cells was significant in the concentration range of 1 to 20 µg ml(-1) , but in Caco2 cells it was significant at a higher concentration range of 10 to 20 µg ml(-1) . These results indicated that HepG2 cells were more sensitive to nanosilver exposure than Caco2 cells. It is generally believed that cellular oxidative stress induces cytotoxicity of nanoparticles. However, in this study we did not detect any nanosilver-induced oxidative stress in either cell type at the concentration range used in this study. Our results suggest that cellular oxidative stress did not play a major role in the observed cytotoxicity of nanosilver in HepG2 and Caco2 cells and that a different mechanism of nanosilver-induced mitochondrial injury leads to the cytotoxicity. The HepG2 and Caco2 cells used this study appear to be targets for silver nanoparticles. The results of this study suggest that the differences in the mechanisms of toxicity induced by nanosilver may be largely as a consequence of the type of cells used. This differential rather than universal response of different cell types exposed to nanoparticles may play an important role in the mechanism of their toxicity. In summary, the results of this study indicate that the widely used in vitro models, HepG2 and Caco2 cells in culture, are excellent systems for screening cytotoxicity of silver nanoparticles. These long established cell culture models and simple assays used in this study can provide useful toxicity and mechanistic information that can help to better inform safety assessments of food- and cosmetic-related silver nanoparticles.