A collaborative study was conducted in 15 laboratories to evaluate 2 different techniques for enumerating Bacillus cereus in foods. A direct plating technique using mannitol-egg yolk-polymyxin agar and a most probable number (MPN) technique using trypticase-soy-polymyxin broth were compared for the enumeration of high and low populations of B. cereus in mashed potatoes. The collaborative results showed that the overall mean recovery obtained with the low population level was essentially the same by both techniques. However, the overall mean recovery was significantly higher by the direct plating technique at the high population level. A statistical evaluation of the data also showed that the direct plating technique had better repeatability and reproducibility than did the MFN technique at both the high and low population levels. These results suggest that the MPN technique is suitable for examining foods containing low populations of B. cereus, but that the direct plating technique is preferable for foods that contain a high population of this organism. The confirmatory technique used in the proposed method is reliable for presumptive identification of isolates as B. cereus. The method has been adopted as official first action.

Background: To properly clean and disinfect hospital mattresses, bed manufacturers recommend a 3- to 6-step process to remove all pathogenic bacteria. An alternative is to use a removable barrier on the mattress, which is laundered after each use. The current study was to determine efficacy of a commercial laundry process in eliminating Clostridioides difficile (C diff) spores, Mycobacterium terrae (M terrae), methicillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa (P aeruginosa), Klebsiella pneumoniae (K pneumoniae), and Escherichia coli (E coli) from a barrier. Methods: A test barrier received 3 unique microbial suspensions in separate locations, each suspension having a known quantity of specific microorganisms: C diff spores, M terrae, and a mixed suspension of MRSA, S aureus, P aeruginosa, K pneumoniae, and E coli. A wash load contained the test barrier and 11 additional ballast barriers. Various soils were spread onto the barriers to simulate heavy soiling that may occur in a wash load: Each barrier received a small amount of mixed soil, 50% received urine, 25% received blood, and 25% received a large amount of additional mixed soil. The load was laundered using 71°C (160°F) water, detergent, and chlorine bleach, with final drying at 71°C (160°F). After laundering, remaining colony-forming units (CFUs) of each microorganism were counted at the applied locations. Each test was replicated 3 times. Industry-accepted methods were used to produce suspensions, apply inoculum, and recover organisms after laundering. Results: Before laundering, test barriers contained at least 7.0 log10 cfu/mL of each microorganism distributed over 103 cm2. After laundering, in all cases, no residual CFUs were detected over the test area, resulting in greater than 6.0 log10 reductions for every organism. ( P < .05). Conclusions: Under extreme test conditions including the presence of soil, the laundry process removed all detectable pathogenic bacteria and spores from the barrier.

Background: The iQ-Check Real-Time PCR kits use PCR technology based on gene amplification and detection by a real-time PCR thermalcycler for the detection of target analytes in select food matrices. The iQ-Check E. coli O157:H7 [Performance Tested MethodSM (PTM) 020801] and STEC VirX and STEC SerO (combined PTM 121203) methods were previously validated for different matrices under different enrichment schemes. Objective: To modify the current iQ-Check E. coli O157:H7 Kit for the detection of Escherichia coli O157:H7 from 25 to 375 g for raw ground beef (17% fat), raw beef trim, and fresh spinach. In addition, a matrix extension was validated for iQ-Check E. coli O157:H7 for raw chicken breast without skin (25 g), raw chicken thigh with skin (25 g), mechanically separated chicken (25 g), and raw ground pork (25 g). The study also included the modification of the iQ-Check STEC VirX and SerO Kits for the detection of non-O157 Shiga toxin–producing E. coli (STEC) for raw ground beef (375 g), raw beef trim (375 g), and fresh spinach (375 g) from STEC Enrichment Broth to buffered peptone water (BPW). All tests were carried out at 8–22 h (10–22 h for fresh spinach). Methods: Ground beef, beef trim, and spinach were co-inoculated with E. coli O157:H7, non-O157 STECs, and Salmonella spp. and analyzed for E. coli O157:H7 and non-O157 STECs after an 8-22 h enrichment in BPW for the beef matrices and after a 10–22 h enrichment in BPW for spinach. The chicken matrices were inoculated with E. coli O157:H7 only and analyzed after an 8–22 h enrichment in BPW. The iQ-Check Free DNA Removal Solution workflow was utilized for all matrices. Confirmations at the 22 h time point and method comparisons were conducted with the appropriate reference method as outlined in the U.S. Food and Drug Administration Bacteriological Analytical Manual Chapter 4A or the U.S. Department of Agriculture Food Safety and Inspection Service Microbiology Laboratory Guidebook Chapters 5.09 and 5B.05. For the iQ-Check STEC VirX and STEC SerO Kits, inclusivity and exclusivity were also performed. Results: The two inclusivity and exclusivity evaluations indicated that the test methods can accurately detect the target analytes and correctly excluded nontarget organisms after 8 h of enrichment. In the method comparison study, the iQ-Check E. coli O157:H7 and STEC VirX and STEC SerO test kits demonstrated no statistically significant differences between candidate and reference method results or between presumptive and confirmed results for all food matrices analyzed and the two time points (8 or 10 and 22 h). Both time points produced the same results, with no discrepancies. Conclusions: The iQ-Check real-time PCR kits are effective methods for the detection of E. coli O157 and non-O157 STECs (both the virulence factors and the O groups) from raw ground beef, raw beef trim, and fresh spinach in 375 g samples enriched in BPW for 8–22 h (10–22 h for fresh spinach). In addition, the iQ-Check E. coli O157 Kit is effective in detecting E. coli O157 in 25 g samples of raw chicken breast without skin, raw chicken thigh with skin, mechanically separated chicken, and raw ground pork. The iQ-Check test kits allow the end user to pair enrichments for multiple target analytes, allowing the user to prepare a single enrichment and perform a single DNA extraction. The Free DNA Removal Solution removes free DNA from samples prior to PCR analysis, protecting DNA from intact and living cells. Highlights: The method modifications were granted based on the data collected.

Background: The MC-Media Pad™ Yeast and Mold (YM) is a ready-to use culture device that combines a test pad coated with medium and water-absorption polymers that is designed for the rapid quantification of yeast and mold in food products. Objective: The MC-Media Pad YM was compared with the U.S. Food and Drug Administration (FDA) Bacteriological Analytical Manual (BAM) for the enumeration of yeast and mold in frozen orange juice concentrate. Methods: The candidate method was evaluated using a paired study design in a multilaboratory collaborative study following the current AOAC Validation Guidelines. Three target contamination levels (low, 10–100 CFU/g; medium, 100–1000 CFU/g; and high 1000–10 000 CFU/g) and an uninoculated control level (0 CFU/g) were evaluated. MC-Media Pad YM devices were enumerated after 48 and 72 h of incubation. Results: Plate count obtained by both methods were log-transformed and the difference of means (including 95% confidence intervals), repeatability SD, and SD were determined for each contamination level. Conclusions: No statistical difference was observed between the MC-Media Pad YM (for both 48 and 72 h) and the FDA BAM for each contamination level. Highlights: The new method offers a convenient alternative to the reference method (FDA BAM) for detection of yeast and mold contamination in food products, yielding reliable and comparable results in 48 h compared to 5 days for the reference method.