Concentration of Listeria monocytogenes in skim milk and soft cheese through microplate immunocapture.

Microplate immunocapture is an inexpensive method for the concentration of foodborne pathogens using an antibody-coated microplate. The objective of this study was to determine the efficacy of microplate immunocapture as an alternative to traditional enrichment for concentrating Listeria monocytogenes to levels detectable with selective plating or real-time PCR. L. monocytogenes isolates serologically characterized as Type 1 (1/2a) and Type 4 (untypeable) were grown overnight and diluted to 100 to 106 colony-forming units (CFU)/mL. The isolates were used to optimize microplate immunocapture in tryptic soy broth with 0.6% yeast extract (TSBYE), skim milk, and queso fresco samples. Following microplate immunocapture, the bacteria were streaked onto polymyxin-acriflavine-LiCl-ceftazidime-aesculin-mannitol (PALCAM) agar, followed by incubation at 37 °C for 24 ±â€¯2 h. The bacteria also underwent real-time polymerase chain reaction (PCR). The optimized microplate immunocapture method was tested in triplicate for its ability to capture L. monocytogenes in broth and food samples. Overall recovery rates for L. monocytogenes in food samples at cell populations of 100, 102, and 104 CFU/25 g using microplate immunocapture with real-time PCR were 88.9%, 94.4%, and 100%, respectively. Recovery in these matrices using microplate immunocapture with selective plating was comparatively lower, at 0%, 44.4%, and 100%, respectively. Conventional culture method showed 100% detection at each inoculation level. Microplate immunocapture combined with real-time PCR shows high potential to reduce the time required for detection, with concentration of L. monocytogenes to detectable levels within 1-4 h. The incorporation of a short enrichment step may improve recovery rates at low cell levels.

Microbial safety and quality of fresh herbs from Los Angeles, Orange County and Seattle farmers' markets.

BACKGROUND: Farmers' markets have been growing in popularity in the United States, but the microbial quality and safety of the food sold at these markets is currently unknown. The purpose of this study was to assess the microbial safety and quality of fresh basil, parsley and cilantro sold at farmers' markets in the Los Angeles, Orange County and greater Seattle areas. RESULTS: A total of 133 samples (52 basil, 41 cilantro and 40 parsley) were collected from 13 different farmers' markets and tested for Salmonella and generic Escherichia coli. One sample (parsley) was confirmed positive for Salmonella and 24.1% of samples were positive for generic E. coli, with a range of 0.70-3.15 log CFU g(-1) . Among the herbs tested, basil showed the highest percentage of samples with generic E. coli (26.9%), followed by cilantro (24.4%) and then parsley (20.0%). For 12% of samples, the levels of generic E. coli exceeded guidelines established by the Public Health Laboratory Service for microbiological quality of ready-to-eat foods. CONCLUSION: Overall, this study indicates the presence of Salmonella and generic E. coli in fresh herbs sold at farmers' markets; however, additional studies are needed to determine the sources and extent of contamination.

Cleaning frequency and the microbial load in ice-cream.

This study investigates the efficacy of a 62 h cleaning frequency in the manufacturing of ice-cream. Various product and product contact surfaces were sampled progressively throughout the time period between cleaning cycles, and analyzed for microbial growth. The coliform and standard plate counts (SPC) of these samples did not vary significantly over time after 0, 24, 48, or 62 h from Cleaning in Place (CiP). Data for product contact surfaces were significant for the SPC representing sample locations. Some of the variables in cleaning practices had significant influence on microbial loads. An increase in the number of flavors manufactured caused a decrease in SPC within the 24 h interval, but by the 48 h interval the SPC increased. More washouts within the first 24 h interval were favorable, as indicated by decreased SPC. The more frequently the liquefier was sanitized within the 62 h interval, the lower the SPC. This study indicates that food safety was not compromised and safety practices were effectively implemented throughout the process.

Use of the Petrifilm™ Test Kit-HEC Direct Blot Enzyme Immunoassay Method without Swab Pre-Enrichment to Screen for Low Levels of Escherichia coli O157 : H7 on Beef Carcasses by Surface Swabbing.

A total of 750 beef carcasses were assayed to detect E. coli O157:H7 by surface swabbing. Each swab represented a 200-cm2 composite surface area. Escherichia coli O157:H7 was not detected in any of the carcass samples. Assays were conducted from carcass swab suspensions by direct plating on Petrifilm™ E. coli Count plates followed by an E. coli O157:H7 assay using the Petrifilm™ Test Kit-HEC direct blot enzyme immunoassay without sample pre-enrichment, and by enrichment using a modified U.S. Department of Agriculture (USDA) Petrifilm™ E. coli O157:H7 procedure. Additionally, beef slabs were inoculated with E. coli O157 :H7 to verify that recovery by the direct swab technique was at least as efficient as enrichment of excised tissue samples, which is the usual USDA method. Escherichia coli O157:H7 inoculation of 4,32, and 180 CFU/25 cm2 were designated low, medium, and high surface contamination levels, respectively. The percentage of recovery of E. coli O157:H7 using the direct swab technique without sample pre-enrichment was 60, 80, and 100% for low, medium, and high surface inoculation levels, respectively. In comparison, recovery from 25-g enrichments of ground excised samples from beef slabs containing 1.4,9.6, and 50 CFU of E. coli O157:H7 was 0, 20, and 73%, respectively. Similarly, the percent recovery of this organism from nonground excised samples containing 56 CFU/25 g enrichment was 68%, versus 100% by direct swab technique without sample pre-enrichment from beef containing 190 CFU/25 cm2 Direct surface swab tests with the 3M Petrifilm™ Test Kit-HEC without sample pre-enrichment proved to be a sensitive, nondestructive, alternate means of evaluating beef carcasses for the presence of E. coli O157:H7.

Direct Enumeration of Escherichia coli O157:H7 from Petrifilm™ EC Count Plates Using the Petrifilm™ Test Kit - HEC Without Sample Pre-Enrichment.

Enumeration of Escherichia coli O157:H7 from Petrifilm™ E. coli (PEC) Count plates was studied incorporating the Petrifilm™ -HEC kit without sample pre-enrichment. Samples containing various E. coli O157:H7 inoculum levels were plated according to PEC package insert instructions. Plates were incubated at 32°C for dairy products and at 35°C for other samples. Total coliform, E. coli and E. coli O157:H7 levels were quantified in both control and inoculated samples. The average recovery from various food matrices was 74.8%. Food samples included retail soft cheese, clams, ground beef, chicken, salad mix, apple juice, cantaloupe and raw milk. The overall recovery was also determined on surface samples including cutting boards and preparation tables, at 116.3%. Results suggest that enumeration of E. coli O157:H7 from PEC Count plates is possible for products containing background coliforms as high as 40,000 colony forming units (CFU)/g, and with minimal background coliforms at <10 CFU/g. This method is a fast and efficient means of quantifying levels of presumptive positive E. coli O157:H7 from PEC Count plates.

Selective Enumeration of Bifidobacterium bifidum , Enterococcus faecium , and Streptomycin-Resistant Lactobacillus acidophilus from a Mixed Probiotic Product.

Modified VF-Bouillon agar with 0.5 mg/ml lithium chloride, 20 µg/ml sodium lauryl sulfate, 5 mg/ml sodium propionate, and 10 µg/ml neomycin sulfate was used with a triple-layer diffusion technique to selectively enumerate Bifidobacterium bifidum . Modified Brigg's agar was used to enumerate Enterococcus faecium . Modified Brigg's agar with 1,200 µg/ml streptomycin sulfate was used in a double-layer diffusion technique to selectively enumerate a streptomycin-resistant strain of Lactobacillus acidophilus . Selective enumeration of the individual bacterial components was compared to the mixture with an average 99% recovery of each component.