Effectiveness of Cross-Flow Microfiltration for Removal of Microorganisms Associated with Unpasteurized Liquid Egg White from Process Plant

ABSTRACT:  Thermal preservation is used by the egg industry to ensure the microbiological safety of liquid egg white (LEW); however, it does not eliminate all microorganisms and impairs some of the delicate functional properties of LEW. In this study, a pilot-scale cross-flow microfiltration (MF) process was designed to remove the natural microflora present in commercial LEW, obtained from a local egg-breaking plant, while maintaining the nutritional and functional properties of the LEW. LEW, containing approximately 10^{6 ± 1.7} colony forming units (CFU) per milliliter of total aerobic bacteria, was microfiltered using a ceramic membrane with a nominal pore size of 1.4 μm, at a cross-flow velocity of 6 m/s. To facilitate MF, LEW was screened, homogenized, and then diluted (1 : 2, w/w) with distilled water containing 0.5% sodium chloride. Homogenized LEW was found to have a threefold lower viscosity than unhomogenized LEW. Influence of MF temperature (25 and 40 °C) and pH (6 and 9) on permeate flux, transmission of egg white nutrients across the membrane, and microbial removal efficiency were evaluated. The pH had a significantly greater influence on permeate flux than temperature. Permeate flux increased by almost 148% when pH of LEW was adjusted from pH 9 to pH 6 at 40 °C. Influence of temperature on permeate flux, at a constant pH, however, was found to be inconclusive. Microbial removal efficiency was at least 5 log₁₀ CFU/mL. Total protein and SDS-PAGE analysis indicated that this MF process did not alter the protein composition of the permeate, compared to that of the feed LEW, and that the foaming properties of LEW were retained in the postfiltered samples.     

Inactivation of Shiga toxin-producing O157:H7 and non-O157:H7 Shiga toxin-producing Escherichia coli in brine-injected, gas-grilled steaks

We quantified translocation of Escherichia coli O157:H7 (ECOH) and non-O157:H7 verocytotoxigenic E. coli (STEC) into beef subprimals after brine injection and subsequently monitored their viability after cooking steaks cut therefrom. Beef subprimals were inoculated on the lean side with ca. 6.0 log CFU/g of a five-strain cocktail of rifampin-resistant ECOH or kanamycin-resistant STEC, and then passed once through an automatic brine-injector tenderizer, with the lean side facing upward. Brine solutions (9.9% ± 0.3% over fresh weight) consisted of 3.3% (wt/vol) of sodium tripolyphosphate and 3.3% (wt/vol) of sodium chloride, prepared both with (Lac(+), pH = 6.76) and without (Lac(-), pH = 8.02) a 25% (vol/vol) solution of a 60% potassium lactate-sodium diacetate syrup. For all samples injected with Lac(-) or Lac(+) brine, levels of ECOH or STEC recovered from the topmost 1 cm (i.e., segment 1) of a core sample obtained from tenderized subprimals ranged from ca. 4.7 to 6.3 log CFU/g; however, it was possible to recover ECOH or STEC from all six segments of all cores tested. Next, brine-injected steaks from tenderized subprimals were cooked on a commercial open-flame gas grill to internal endpoint temperatures of either 37.8 °C (100 °F), 48.8 °C (120 °F), 60 °C (140 °F), or 71.1 °C (160 °F). Regardless of brine formulation or temperature, cooking achieved reductions (expressed as log CFU per gram) of 0.3 to 4.1 of ECOH and 0.5 to 3.6 of STEC. However, fortuitous survivors were recovered even at 71.1 °C (160 °F) for ECOH and for STEC. Thus, ECOH and STEC behaved similarly, relative to translocation and thermal destruction: Tenderization via brine injection transferred both pathogens throughout subprimals and cooking highly contaminated, brine-injected steaks on a commercial gas grill at 71.1 °C (160 °F) did not kill all cells due, primarily, to nonuniform heating (i.e., cold spots) within the meat.     

Removal of Salmonella Enteritidis from commercial unpasteurized liquid egg white using pilot scale cross flow tangential microfiltration

Effectiveness of a cross flow microfiltration (MF) process for removal of a cocktail of Salmonella enterica serovar Enteritidis species from commercial unpasteurized liquid egg white (LEW) from a local egg breaking plant, while maintaining its functional properties was evaluated. To facilitate MF, LEW was wedge screened, homogenized and then diluted (1:2 w/w) with distilled water containing 0.5% sodium chloride. Diluted unpasteurized LEW was inoculated with five strains of S. Enteritidis (ATCC 4931, ATCC BAA-708, ATCC 49215, ATCC 49218, and ATCC BAA-1045) to a level of approximately 10⁷ CFU/mL of LEW and microfiltered using a ceramic membrane. Process parameters influencing egg white functional properties and pathogen removal efficiency were evaluated. Average permeates flux increased by almost 126% when pH of LEW was adjusted from pH 8 to pH 7 at 25 °C. Microbial removal efficiency was at least, on average, 6.8 Log₁₀ CFU/mL (limit of detection ≤ 0.5 Log₁₀ CFU/mL). Functional property analysis indicated that the MF process did not alter the foaming power of LEW.     

Control of Listeriamonocytogenes on commercially-produced frankfurters prepared with and without potassium lactate and sodium diacetate and surface treated with lauric arginate using the Sprayed Lethality in Container (SLIC®) delivery method

Viability of Listeriamonocytogenes was monitored on frankfurters formulated with or without potassium lactate and sodium diacetate at a ratio of ca. 7:1 and treated with lauric arginate (LAE; 22 or 44 ppm) using the Sprayed Lethality in Container (SLIC®) delivery method. Without antimicrobials, pathogen numbers remained relatively constant at ca. 3.3 log CFU/package for ca. 30 d, but then increased to ca. 8.4 log CFU/package over 120 d. Regardless of whether or not lactate and diacetate were included, when treated with LAE, pathogen numbers decreased from ca. 3.3 log CFU/package to ca. 1.5 log CFU/package within 2 h, but then increased to 7.3 and 6.7 log CFU/package, respectively, after 120 d. When frankfurters were formulated with lactate and diacetate and treated with LAE, pathogen numbers decreased by ca. 2.0 log CFU/package within 2 h and remained relatively unchanged over the 120 d. These data confirm that LAE provides an initial lethality towards L. monocytogenes and when used in combination with reduced levels/ratio of lactate and diacetate as an ingredient for frankfurters provides inhibition throughout shelf life.     

Validation of a commercial process for inactivation of Escherichia coli O157:H7, Salmonella typhimurium, and Listeria monocytogenes on the surface of whole muscle beef jerky

We validated the lethality of three time and temperature regimens for commercial processing of whole muscle beef jerky. A total of ca. 8.9 log CFU per strip of multiple-strain cocktails of Escherichia coli O157:H7, Salmonella Typhimurium, or Listeria monocytogenes were separately applied onto the surface of beef strips that were treated as follows: (i) inoculated but not marinated or (ii) inoculated and then marinated. A total of three beef strips for each treatment in each of three trials were separately inoculated with a cocktail of one of the three pathogens and placed on the top, middle, and bottom racks of a loading truck. The strips on the rack were loaded into a commercial smokehouse and cooked and dried for 1.5, 2.5, or 3.5 h at a target temperature of 180 degrees F (82.2 degrees C) with constant (natural hickory) smoking, but without the addition of humidity. Regardless of how the strips were treated or where the strips were placed on the loading rack, drying for 1.5, 2.5, or 3.5 h to a target temperature of 180 degrees F (average of 177.2 +/- 5.6 degrees F [80.7 +/- 3.1 degrees C]), with constant smoke at an initial average relative humidity of 63.1% to a final average relative humidity of 20.9% resulted in a decrease of > or = 7.3 log CFU per strip (> or = 6.9 log CFU/g) for each of the three pathogen cocktails. Of note, marinated strips that were cooked and dried for 2.5 and 3.5 h or nonmarinated strips cooked or dried for 3.5 h also satisfied the U.S. Food Safety and Inspection Service standard of identity (moisture-to-protein ratio < or = 0.75:1) and/or shelf-stability (water activity < or = 0.8) requirements for jerky.     

Fate of Shiga toxin-producing O157:H7 and non-O157:H7 Escherichia coli cells within blade-tenderized beef steaks after cooking on a commercial open-flame gas grill

We compared the fate of cells of both Shiga toxin-producing Escherichia coli O157:H7 (ECOH) and Shiga toxin-producing non-O157:H7 E. coli (STEC) in blade-tenderized steaks after tenderization and cooking on a gas grill. In phase I, beef subprimal cuts were inoculated on the lean side with about 5.5 log CFU/g of a five-strain mixture of ECOH or STEC and then passed once through a mechanical blade tenderizer with the lean side facing up. In each of two trials, 10 core samples were removed from each of two tenderized subprimals and cut into six consecutive segments starting from the inoculated side. Ten total cores also were obtained from two nontenderized (control) subprimals, but only segment 1 (the topmost segment) was sampled. The levels of ECOH and STEC recovered from segment 1 were about 6.0 and 5.3 log CFU/g, respectively, for the control subprimals and about 5.7 and 5.0 log CFU/g, respectively, for the tenderized subprimals. However, both ECOH and STEC behaved similarly in terms of translocation, and cells of both pathogen cocktails were recovered from all six segments of the cores obtained from tenderized subprimals, albeit at lower levels in segments 2 to 6 than those found in segment 1. In phase II, steaks (2.54 and 3.81 cm thick) cut from tenderized subprimals were subsequently cooked (three steaks per treatment) on a commercial open-flame gas grill to internal temperatures of 48.9, 54.4, 60.0, 65.6, and 71.1°C. Regardless of temperature or thickness, we observed 2.0- to 4.1-log and 1.5- to 4.5-log reductions in ECOH and STEC levels, respectively. Both ECOH and STEC behaved similarly in response to heat, in that cooking eliminated significant numbers of both pathogen types; however, some survivors were recovered due, presumably, to uneven heating of the blade-tenderized steaks.     

Prevalence, characterization and sources of Listeria monocytogenes in blue crab (Callinectus sapidus) meat and blue crab processing plants

Seven blue crab processing plants were sampled to determine the prevalence and sources of Listeria spp. and Listeria monocytogenes for two years (2006–2007). A total of 488 raw crabs, 624 cooked crab meat (crab meat) and 624 environmental samples were tested by standard methods. Presumptive Listeria spp. were isolated from 19.5% of raw crabs, 10.8% of crab meat, and 69.5% of environmental samples. L. monocytogenes was isolated from 4.5% of raw crabs, 0.2% of crab meat, and 2.1% of environmental samples. Ninety-seven percent of the isolates were resistant to at least one of the ten antibiotics tested. Eight different serotypes were found among 76 L. monocytogenes isolates tested with the most common being 4b, 1/2b and 1/2a. Automated EcoRI ribotyping differentiated 11 ribotypes among the 106 L. monocytogenes isolates. Based on ribotyping analysis, the distribution of the ribotypes in each processing plant had a unique contamination pattern. A total of 92 ApaI and 88 AscI pulsotypes among the 106 L. monocytogenes isolates were found and distinct pulsotypes were observed in raw crab, crab meat and environmental samples. Ribotypes and serotypes recovered from crab processing plants included subtypes that have been associated with listeriosis cases in other food outbreaks. Our findings suggest that molecular methods may provide critical information about sources of L. monocytogenes in crab processing plants and will augment efforts to improve food safety control strategies such as targeting specific sources of contamination and use of aggressive detergents prior to sanitizing.     

Behavior of Escherichia coli O157:H7, Listeria monocytogenes, and Salmonella Typhimurium in teewurst, a raw spreadable sausage

The fate of Listeria monocytogenes, Salmonella Typhimurium, or Escherichia coli O157:H7 were separately monitored both in and on teewurst, a traditional raw and spreadable sausage of Germanic origin. Multi-strain cocktails of each pathogen (ca. 5.0 log CFU/g) were used to separately inoculate teewurst that was subsequently stored at 1.5, 4, 10, and 21 °C. When inoculated into commercially-prepared batter just prior to stuffing, in general, the higher the storage temperature, the greater the lethality. Depending on the storage temperature, pathogen levels in the batter decreased by 2.3 to 3.4, ca. 3.8, and 2.2 to 3.6 log CFU/g for E. coli O157:H7, S. Typhimurium, and L. monocytogenes, respectively, during storage for 30 days. When inoculated onto both the top and bottom faces of sliced commercially-prepared finished product, the results for all four temperatures showed a decrease of 0.9 to 1.4, 1.4 to 1.8, and 2.2 to 3.0 log CFU/g for E. coli O157:H7, S. Typhimurium, and L. monocytogenes, respectively, over the course of 21 days. With the possible exceptions for salt and carbohydrate levels, chemical analyses of teewurst purchased from five commercial manufacturers revealed only subtle differences in proximate composition for this product type. Our data establish that teewurst does not provide a favourable environment for the survival of E. coli O157:H7, S. Typhimurium, or L. monocytogenes inoculated either into or onto the product.     

Effect of high-pressure processing on reduction of Listeria monocytogenes in packaged Queso Fresco

The effect of high-hydrostatic-pressure processing (HPP) on the survival of a 5-strain rifampicin-resistant cocktail of Listeria monocytogenes in Queso Fresco (QF) was evaluated as a postpackaging intervention. Queso Fresco was made using pasteurized, homogenized milk, and was starter-free and not pressed. In phase 1, QF slices (12.7 × 7.6 × 1 cm), weighing from 52 to 66 g, were surface inoculated with L. monocytogenes (ca. 5.0 log₁₀ cfu/g) and individually double vacuum packaged. The slices were then warmed to either 20 or 40°C and HPP treated at 200, 400, and 600 MPa for hold times of 5, 10, 15, or 20 min. Treatment at 600 MPa was most effective in reducing L. monocytogenes to below the detection level of 0.91 log₁₀ cfu/g at all hold times and temperatures. High-hydrostatic-pressure processing at 40°C, 400 MPa, and hold time ≥15 min was effective but resulted in wheying-off and textural changes. In phase 2, L. monocytogenes was inoculated either on the slices (ca. 5.0 log₁₀ cfu/g; ON) or in the curds (ca. 7.0 log₁₀ cfu/g; IN) before the cheese block was formed and sliced. The slices were treated at 20°C and 600 MPa at hold times of 3, 10, and 20 min, and then stored at 4 and 10°C for 60 d. For both treatments, L. monocytogenes became less resistant to pressure as hold time increased, with greater percentages of injured cells at 3 and 10 min than at 20 min, at which the lethality of the process increased. For the IN treatment, with hold times of 3 and 10 min, growth of L. monocytogenes increased the first week of storage, but was delayed for 1 wk, with a hold time of 20 min. Longer lag times in growth of L. monocytogenes during storage at 4°C were observed for the ON treatment at hold times of 10 and 20 min, indicating that the IN treatment may have provided a more protective environment with less injury to the cells than the ON treatment. Similarly, HPP treatment for 10 min followed by storage at 4°C was the best method for suppressing the growth of the endogenous microflora with bacterial counts remaining below the level of detection for 2 out of the 3 QF samples for up to 84 d. Lag times in growth were not observed during storage of QF at 10°C. Although HPP reduced L. monocytogenes immediately after processing, a second preservation technique is necessary to control growth of L. monocytogenes during cold storage. However, the results also showed that HPP would be effective for slowing the growth of microorganisms that can shorten the shelf life of QF.     

Viability of multi-strain mixtures of Listeria monocytogenes, Salmonella typhimurium, or Escherichia coli O157:H7 inoculated into the batter or onto the surface of a soudjouk-style fermented semi-dry sausage

The fate of Listeria monocytogenes, Salmonella typhimurium, or Escherichia coli O157:H7 were separately monitored both in and on soudjouk. Fermentation and drying alone reduced numbers of L. monocytogenes by 0.07 and 0.74 log₁₀ CFU/g for sausages fermented to pH 5.3 and 4.8, respectively, whereas numbers of S. typhimurium and E. coli O157:H7 were reduced by 1.52 and 3.51 log₁₀ CFU/g and 0.03 and 1.11 log₁₀ CFU/g, respectively. When sausages fermented to pH 5.3 or 4.8 were stored at 4, 10, or 21 °C, numbers of L. monocytogenes, S. typhimurium, and E. coli O157:H7 decreased by an additional 0.08–1.80, 0.88–3.74, and 0.68–3.17 log₁₀ CFU/g, respectively, within 30 days. Storage for 90 days of commercially manufactured soudjouk that was sliced and then surface inoculated with L. monocytogenes, S. typhimurium, and E. coli O157:H7 generated average D-values of ca. 10.1, 7.6, and 5.9 days at 4 °C; 6.4, 4.3, and 2.9 days at 10 °C; 1.4, 0.9, and 1.6 days at 21 °C; and 0.9, 1.4, and 0.25 days at 30 °C. Overall, fermentation to pH 4.8 and storage at 21 °C was the most effective treatment for reducing numbers of L. monocytogenes (2.54 log₁₀ CFU/g reduction), S. typhimurium (5.23 log₁₀ CFU/g reduction), and E. coli O157:H7 (3.48 log₁₀ CFU/g reduction). In summary, soudjouk-style sausage does not provide a favorable environment for outgrowth/survival of these three pathogens.