Determining the variability associated with testing shelled corn for aflatoxin using different analytical procedures in Louisiana in 1998.

The number of elevator facilities with laboratories to test shelled corn for aflatoxin on site is increasing. The inherent difficulty in accurately determining the true aflatoxin concentration of a lot of corn may have serious implications. Deviations from the true value are of even greater significance at busy locations where a high throughput is desired. This study was instituted to measure (1) the differences in aflatoxin test results between elevator laboratories and the Louisiana Agricultural Chemistry (LAC) laboratory and (2) the variability in aflatoxin test results associated with sampling, sample preparation, and analysis of shelled corn at such locations. One hundred lots of shelled corn from 10 elevators in Louisiana were analyzed for aflatoxin using the Aflatest method (at elevators and at the LAC laboratory) and high-performance column liquid chromatography (HPLC; LAC laboratory only). Mean aflatoxin levels determined at elevator laboratories were significantly (P < 0.05) lower from those obtained in the LAC laboratory using the Aflatest method. Overall, Aflatest method results were lower than those obtained by HPLC. This difference may be attributed to analyst technical dexterity, difficulty in providing careful attention to detail in a high throughput environment, and/or substandard facilities found at elevators. The total variance was partitioned into the combined sampling plus subsampling variance and analytical variance. The sampling and sample preparation steps accounted for about 91.5% of the total variability. When using the HPLC analytical method, the analytical step contributed only 8.5% to the total variance.

Feasibility of immunodiagnostic devices for the detection of ricin, amanitin, and T-2 toxin in food.

Qualitative and quantitative comparisons were conducted of commercially available immunodiagnostic devices for the detection of three select agents with oral LD50 values > or = 0.1 mg/kg of body weight. Ricin (oral LD50 > 1 mg/kg), amanitin (oral LD50 approximately 0.1 mg/kg), and T-2 toxin (oral LD50 > 1 mg/kg) were spiked into beverages, produce, dairy, and baked goods and assayed using commercially available enzyme-linked immunosorbent assays (ELISAs) and lateral flow devices. In all cases, the commercial diagnostic kits successfully detected all three select agents at concentrations below what might be a health concern. The considerable difference between the limit of detection of the immunodiagnostic devices employed (typically < or = 0.020 microg/g) and the amount of the select agent necessary to pose a health threat in a single serving of food facilitated the design of protocols for the high throughput screening of food samples. These protocols entailed simple extraction methods followed by sample dilution. Lateral flow devices and sandwich ELISAs for the detection of ricin had no significant background problems due to the food matrices. Competitive ELISAs, which typically have unacceptably high background reactions with food samples, successfully detected amanitin and T-2 toxin.

Performance Tested Method multiple laboratory validation study of ELISA-based assays for the detection of peanuts in food.

Performance Tested Method multiple laboratory validations for the detection of peanut protein in 4 different food matrixes were conducted under the auspices of the AOAC Research Institute. In this blind study, 3 commercially available ELISA test kits were validated: Neogen Veratox for Peanut, R-Biopharm RIDASCREEN FAST Peanut, and Tepnel BioKits for Peanut Assay. The food matrixes used were breakfast cereal, cookies, ice cream, and milk chocolate spiked at 0 and 5 ppm peanut. Analyses of the samples were conducted by laboratories representing industry and international and U.S governmental agencies. All 3 commercial test kits successfully identified spiked and peanut-free samples. The validation study required 60 analyses on test samples at the target level 5 microg peanut/g food and 60 analyses at a peanut-free level, which was designed to ensure that the lower 95% confidence limit for the sensitivity and specificity would not be <90%. The probability that a test sample contains an allergen given a prevalence rate of 5% and a positive test result using a single test kit analysis with 95% sensitivity and 95% specificity, which was demonstrated for these test kits, would be 50%. When 2 test kits are run simultaneously on all samples, the probability becomes 95%. It is therefore recommended that all field samples be analyzed with at least 2 of the validated kits.

Effect of processing on aflatoxin.

Naturally occurring toxicant contamination of foods with mycotoxins is unavoidable and unpredictable and poses a unique challenge to food safety. Aflatoxins are toxic mold metabolites produced by toxigenic strains of Aspergillus species. Primary commodities susceptible to aflatoxin contamination include corn, peanuts and cottonseed and animal-derived foods such as milk when the animal is fed aflatoxin-contaminated feed. Risks associated with aflatoxin-contaminated foods can be reduced through the use of specific processing and decontamination procedures. Factors, which influence the effectiveness of a specific process or procedure, include the chemical stability of the mycotoxin(s), nature of the process, type and interaction with the food/feed matrix and interaction with multiple mycotoxins if present. Practical decontamination procedures must: 1) inactivate, destroy, or remove the toxin, 2) not produce or leave toxic residues in the food/feed, 3) retain the nutritive value of the food/feed, 4) not alter the acceptability or the technological properties of the product, and, if possible, 5) destroy fungal spores. For aflatoxins, multiple processing and/or decontamination schemes have been successful in reducing aflatoxin concentrations to acceptable levels. Physical cleaning and separation procedures, where the mold-damaged kernel/seed/nut is removed from the intact commodity, can result in 40-80% reduction in aflatoxins levels. Processes such as dry and wet milling result in the distribution of aflatoxin residues into less utilized fractions of the commodity. The ammoniation of aflatoxin-contaminated commodities has altered the concentrations as well as toxic and carcinogenic effects of aflatoxin by greater than 99%. Nonbiological materials such as selected anticaking agents covalently bind aflatoxins from aqueous suspensions, diminish aflatoxin uptake by animals, prevent acute aflatoxicosis, and decrease aflatoxin residues in milk. Ultimately, the best processing or decontamination process is one that is approved by regulatory agencies, cost-effective, and reduces the mycotoxin concentration to acceptable levels.

U.S. perspective on mycotoxin regulatory issues.

Control programs set up by the Food and Drug Administration (FDA) for aflatoxin, an unavoidable natural contaminant produced by specific molds that invade a number of feedstuffs and basic foods, provide an example of forces that affect risk assessment and management strategies by a regulatory agency. More recently, on an international scale, efforts to establish international food standards for fumonisin, deoxynivalenol, ochratoxin A, zearalenone, and patulin, as well as for aflatoxin, demonstrate the complexity of developing regulations and/or standards designed to protect consumer health and ensure fair trade practices on a global scale. Current FDA regulations for aflatoxins address public health concerns for potential contamination in basic foods, residues in milk, and animal feeds for numerous commodities and applications. Regulatory limits, sampling and analytical procedures, decontamination and/or diversion to less risk uses for contaminated product are components of mycotoxin control programs. Current efforts by FDA to establish regulatory controls for deoxynivalenol, fumonisin, and patulin add further insight on the role that safety and risk assessment procedures play in the development of action levels and advisories for mycotoxins.

Dairy Herd Mastitis Quality Control Program.

Milk from 34 dairy herds was tested over a 12-month period using a mastitis evaluation program in which Streptococcus agalactiae , Staphylococcus aureus and leucocytes were counted. There was a significant decrease in the total mastitic bacterial count ( S. agalactiae plus S. aureus ) over the testing period; however, the curve was bimodal, showing high points in the winter and summer months. The leucocyte count alone was not a good indicator of the mastitic condition of the herd. In approximately 12% of the test results, there was a high bacterial count with a low leucocyte count or a high leucocyte count with a low bacterial count.

Direct Application of a New Hypochlorite Sanitizer for Reducing Bacterial Contamination on Foods.

The efficacy of the buffered sodium hypochlorite solution, Bionox, in controlling bacterial contamination was evaluated on fresh-cut poultry (chicken, light and dark meat), fish fillets, fruit, and vegetables. Food products were immersed in a nutrient broth suspension of Salmonella enteritidis (ATCC #13067) for 30 s and allowed to drip/air dry for 10 s prior to exposure to the sanitizing agent. Food products were then each vigorously washed with 100 ml of buffered peptone which was plated in serial dilution on XLD-N agar and incubated at 37°C for 24 h. Typical Salmonella colonies as well as non- Salmonella colonies growing on the XLD-N plates were counted and identified. Results showed the sanitizing solution to be effective in reducing S. enteritidis on all test foodstuffs. Count reductions of 4, 3, and 2 logs per gram on chicken, vegetables, and fruit, respectively, were achieved. Salmonella reductions of two logs were also achieved on fish fillets, but the sanitizer performance depended to some extent on the background bacterial flora present prior to the addition of the Salmonella test organism. The effect of the sanitizing solution on protein functionality, lipid oxidation, and starch degradation was determined using protein dispersibility and solubility assays, peroxide and iodine values, and changes in reducing sugars levels, respectively. Results showed no adverse effects on these parameters after exposure of the food products to the sanitizing solution.

Comparison of the Microbial Quality of Raw Shrimp from China, Ecuador, or Mexico at Both Wholesale and Retail Levels.

Aerobic plate counts (APC), Listeria spp., and Vibrio spp. and antibiotic resistance patterns of Vibrio spp. were determined for imported shrimp from China, Ecuador, and Mexico obtained from wholesale/frozen and retail/previously frozen markets. Statistically significant differences in APC were observed among source countries and wholesale/frozen versus retail/previously frozen samples. Wholesale/frozen shrimp products were consistently excellent quality with respect to APC; higher contamination levels were observed in retail/previously frozen samples. Listeria spp. and L. monocytogenes were isolated from 16.7 and 6.7% of shrimp samples, respectively. Vibrio spp. were present in 63.3% of the samples, more often isolated from shrimp from Mexico or China than Ecuador. The majority of isolates were identified as V. parahaemolyticus (36.7%), V. alginolyticus (26.7%), or V. vulniftcus (16.7%), and 53.7% were resistant to at least one antibiotic. These data reveal that important differences in microbial quality occur in raw shrimp products as a function of source and between retail and Wholesale products.

Preparation of 14C-Labeled Penicillic Acid with High Specific Activity and Yield.

14C-Labeled penicillic acid was produced by stationary culture incubation of Penicillium cyclopium (NRRL 1888) on a modified Raulin-Thom broth medium containing 14C-labeled acetate. Approximately 1.2 g of radioactive compound, with a specific activity of 23.0 µCi/mmole, was produced in 9 days in 1500 ml of the broth. Incorporation of the isotope into penicillic acid was 11. 9%. Production of the radiolabeled compound with high specific activity was achieved by correlating the monitoring of expired 14C-CO2 with production of penicillic acid during the fermentation. The effects of various growth substrates, pH, and incubation times on production of non-labeled penicillic acid also were investigated. Results show that sterile rice is an excellent substrate, that among liquid media examined, higher yields were obtained in stationary rather than in shake cultures, and that higher yields of penicillic acid were obtained at pH 3.5 or lower. Simultaneous monitoring of penicillic acid production and 14C-label incorporation is essential to detect and isolate a high yield of labeled compound with high specific activity.

Reduction of Mutagenic Potentials in Milk: Effects of Ammonia Treatment on Aflatoxin-Contaminated Cottonseed.

Milks obtained from cows fed rations containing aflatoxin-contaminated cottonseed, ammonia-treated aflatoxin-contaminated cottonseed, and uncontaminated cottonseed were tested for mutagenic potential using the Salmonella /mammalian microsome mutagenicity assay using Salmonella typhimurium strains TA98 and TA100. Standard assay protocol was used with S-9 liver homogenate added. Samples including whole milk, nonfat dry milk powder, cream, and reconstituted whole milk were applied directly to the plates in triplicate. As a control, samples of whole milk, reconstituted whole milk, and nonfat dry milk powder from cows fed uncontaminated feed were spiked with aflatoxin B1 and tested for mutagenic activity. High levels of mutagenic activity were observed in all samples from cows exposed to aflatoxin-contaminated cottonseed and the aflatoxin-spiked milks. This high activity was not evident in whole milk and whole milk component samples from cows fed the ammonia-treated aflatoxin-contaminated cottonseed or nonaflatoxin containing cottonseed. A low level of mutagenic potential was evident in whole milk from the ammonia treated group using TA100 tester strain.