Microbiologique Microfilm™ EBEc - a Rapid Method for the Simultaneous Enumeration of Enterobacteriaceae and Escherichia coli.

BACKGROUND: The Enterobacteriaceae and generic Escherichia coli are routinely enumerated in foods as part of product release criteria, or in the case of swabs, for environmental monitoring. OBJECTIVE: Microbiologique Microfilm™ EBEc is intended to provide a rapid and easy-to-use method for simultaneous enumeration of Enterobacteriaceae and E. coli on foods and environmental surfaces. Methods: This study evaluated the performance of Microfilm™ EBEc against ISO methods (ISO 21528-2:2017 for Enterobacteriaceae and ISO 16649-2: 2001 for E. coli) in 20 food matrixes and two environmental surfaces. Inclusivity, exclusivity, lot-to-lot reproducibility, ruggedness and stability studies were also performed on Microfilm™ EBEc. RESULTS: No significant differences and high correlation coefficients (R2) were observed between the Microfilm™ EBEc and the corresponding ISO methods in spiked food matrixes and environmental samples. Inclusivity studies showed expected results for all the E. coli and Enterobacteriaceae strains tested. In terms of exclusivity testing, all the strains tested failed to grow on Microfilm™ EBEc. A lot-to-lot study showed no significant differences in mean difference (log10) counts among the three lots of the Microfilm™ EBEc. Ruggedness studies showed no significant differences in mean difference (log10) counts at varying incubation temperatures and times. Stability studies on the Microfilm™ EBEc showed that it is stable at 2-25°C for 12 months, and at 45°C for 6 weeks. CONCLUSIONS: The results of this study indicate that the Microfilm™ EBEc is equivalent to the corresponding ISO methods for enumeration of Enterobacteriaceae and E. coli. Highlights: Microfilm™ EBEc offers a convenient and relatively fast test method for simultaneous enumeration of Enterobacteriaceae and E. coli in 24 h and has an advantage over the corresponding ISO methods that require two assays on the same sample for enumeration of Enterobacteriaceae and E. coli Gram-negative indicator groups.

Performance Validation of the Microbiologique MicrofilmTM Test System for AOAC Research Institute Performance Tested MethodSMCertification.

The Microfilm™ Test System is intended for quantitative microbiology and consists of three types of Microfilms for aerobic plate count (Microfilm APC), total coliform and Escherichia coli count (Microfilm TCEc), and yeast and mold count (Microfilm YMC). This study evaluated the performance of the Microfilm Test System against International Organization for Standardization (ISO) methods on 20 food matrixes and 2 environmental surfaces. Ruggedness, robustness, and stability were also determined, while inclusivity and exclusivity studies were performed on Microfilm TCEc and YMC. An independent laboratory evaluated the performance on four food matrixes and one environmental surface. No significant differences and high correlation coefficients were observed between the Microfilm Test System and the corresponding ISO methods (ISO 4833-1:2013 for APC, ISO 4832:2006 for total coliform count, ISO 16649-2: 2001 for E. coli, and ISO 21527 Part 1 and Part 2 for YMC) in spiked food matrixes and environmental samples. These results were corroborated by the independent laboratory. Inclusivity and exclusivity studies for Microfilm TCEc showed expected results for all the E. coli strains tested (blue-violet or violet color), while the related coliforms showed the expected blue-green colonies on the Microfilm. Similarly, all 100 fungal strains tested showed typical growth on Microfilm YMC. Exclusivity testing on Microfilm TCEc and YMC showed no growth of nontarget organisms. Robustness and ruggedness studies showed no significant differences in mean difference counts at varying incubation temperatures and times. Stability studies on three lots of the Microfilm Test System showed that it is stable at 2-25°C for 12 months and at 45°C for 6 weeks.

Effect of temperature and growth media on the attachment of Listeria monocytogenes to stainless steel.

Listeria monocytogenes ATCC 19111 cultivated in nutrient-rich medium (brain heart infusion, BHI) or starved in minimal medium (10% filter sterilized pond water and 90% sterilized distilled water) were investigated for their initial attachment to austenitic stainless steel No. 4 with satin finish at 4 degrees C, 20 degrees C, 30 degrees C, 37 degrees C, or 42 degrees C. A droplet (10 microl) containing approximately 10(7) CFU/ml of L. monocytogenes suspended in BHI or minimal medium was placed on the stainless steel surface. After holding in saturated humidity for 3 h at the desired temperature the surface was washed and prepared for scanning electron microscopy (SEM). Using SEM, attachment of L. monocytogenes was determined by counting cells remaining on the surface. When L. monocytogenes cultivated in BHI were used, with the exception of the number of attached cells being lower at 42 degrees C than at 37 degrees C and 30 degrees C, the number of attached cells increased with increasing temperature (P<0.05). When L. monocytogenes starved in minimal medium were used, the number of attached cells also increased with increasing attachment temperature (P<0.05), but the number of attached cells at 42 degrees C was lower than that at the other temperatures. The attachment of L. monocytogenes to stainless steel surface was greater when cultivated in rich medium of BHI vs starved in the minimal medium.

Attachment of Listeria monocytogenes to an austenitic stainless steel after welding and accelerated corrosion treatments.

Austenitic stainless steels, widely used in food processing, undergo microstructural changes during welding, resulting in three distinctive zones: weld metal, heat-affected zone, and base metal. This research was conducted to determine the attachment of Listeria monocytogenes in these three zones before and after exposure to a corrosive environment. All experiments were done with tungsten inert gas welding of type 304 stainless steel. The four welding treatments were large or small beads with high or low heat. After welding, all surfaces were polished to an equivalent surface finish. A 10-microl droplet of an L. monocytogenes suspension was placed on the test surfaces. After 3 h at 23 degrees C, the surfaces were washed and prepared for scanning electron microscopy, which was used to determine attachment of L. monocytogenes by counting cells remaining on each test surface. In general, bacteria were randomly distributed on each surface type. However, differences in surface area of inoculum due to differences in interfacial energy (as manifested by the contact angle) were apparent and required normalization of bacterial count data. There were no differences (P > 0.05) in numbers of bacteria on the three surface zones. However, after exposure to the corrosive medium, numbers of bacteria on the three zones were higher (P < 0.05) than those on the corresponding zones of noncorroded surfaces. For the corroded surfaces, bacterial counts on the base metal were lower (P < 0.05) than those on heat-affected and weld zones.