Monocytogenes Cocktail Research Articles

Elimination of Listeria monocytogenes on Hotdogs by Infrared Surface Treatment

The objective of this research was to develop an infrared pasteurization process with automatic temperature control for inactivation of surface-contaminated Listeria monocytogenes on ready-to-eat meats such as hotdogs. The pasteurization system contained 4 basic elements: an infrared emitter, a hotdog roller, an infrared sensor, and a temperature controller. The infrared sensor was used to monitor the surface temperature of hotdogs while the infrared emitter, modulated by a power controller, was used as a heating source. The surface temperature of hotdogs was increased to set points (70, 75, 80, or 85 degrees C), and maintained for bacterial kill. The infrared surface pasteurization was evaluated using hotdogs that were surface-inoculated with a 4-strain L. monocytogenes cocktail to an average initial inoculum of 7.32 log (CFU/g). On the average 1.0, 2.1, 3.0, or 5.3 log-reduction in L. monocytogenes was observed after the surface temperature of hotdogs was increased to 70, 75, 80, or 85 degrees C, respectively. Holding the sample temperature led to additional bacterial inactivation. With a 3 min holding at 80 degrees C or 2 min at 85 degrees C, a total of 6.4 or 6.7 logs of L. monocytogenes were inactivated. This study demonstrated that the infrared surface pasteurization was effective in inactivating L. monocytogenes in RTE meats.

Impact of Dilution Ratios on Listeria monocytogenes Growth during University of Vermont Medium Enrichment of Deli Meats

In the U.S. Department of Agriculture (USDA) method for Listeria detection, a 25-g composite food sample is enriched in 225 ml of University of Vermont medium (UVM), giving a detection limit of 0.04 CFU/g. However, in a recent large-scale four-state deli meat survey for L. monocytogenes, 125-g samples enriched in 1,125 ml of UVM were requested to increase the detection limit to 0.008 CFU/g. To circumvent problems associated with large volumes of UVM, the impact on L. monocytogenes growth of lower dilution ratios used for enrichment and most-probable-number (MPN) detection was compared with the results obtained using the conventional 1:10 dilution. In this study, 125-g samples of cured turkey, uncured turkey, ham, and roast beef were inoculated with a six-strain L. monocytogenes cocktail to contain ∼1 × 103 CFU/g. This cocktail was then diluted 1:3, 1:5, or 1:10 in UVM, homogenized, enriched at 30°C, and periodically plated on modified Oxford agar to determine generation times during 24 h of incubation. The same enrichment protocol was also assessed in a three-tube MPN assay using 125-g samples inoculated with L. monocytogenes to contain ∼1 CFU/g. The effects of two homogenization methods, stomaching and pulsifying, on Listeria growth were compared using oven-roasted turkey breast diluted 1:3, 1:5, and 1:10 in UVM. Overall, the growth rates, generation times, and MPN values for each of the four selected deli meats were similar (P > 0.05) using UVM enrichment ratios of 1:3, 1:5, and 1:10, with no significant (P > 0.05) differences in L. monocytogenes growth rate or generation time between experiments using pulsifying and stomaching. These findings indicate that lower volumes of UVM can be used in the USDA procedure when examining deli meats without compromising Listeria recovery.

Inhibitory Activity of Colicin E1 against Listeria monocytogenes

Colicins are gram-negative bacteriocins produced by and effective against Escherichia coli and related species. Colicin E1 (ColE1) is composed of three functional domains, which collectively have a pore-forming effect on targeted bacteria. ColE1 binding and translocation domains are highly specific in contrast to the pore-forming domain, implying that ColE1 could be broadly effective. In this study, the activity of ColE1 against Listeria monocytogenes was evaluated in broth and on surfaces of ready-to-eat products. Individual strains of L. monocytogenes were examined in broth containing ColE1 at 0, 0.1, 1, or 10 μg/ml. Although strain differences in sensitivity to ColE1 existed, growth was significantly reduced in all strains at doses as low as 0.1 μg/ml. Sterilized ham slices were submerged in a five-strain L. monocytogenes cocktail (either 7 or 4 log CFU/ml) and placed in vacuum packages containing 0, 1, 5, 10, 25, or 50 μg of ColE1. Ham slices were then stored at 4 or 10°C, and samples were removed and examined for L. monocytogenes after 1, 3, 7, and 14 days. Reduction of L. monocytogenes by ColE1 was dependent on initial inoculum concentration and storage temperature. For slices stored at 4°C, treatment with 25 μg reduced Listeria growth below detection limits for the slices inoculated with 4 log CFU/ml for the entire 14 days, whereas for the 7-log CFU/ml slices, growth was detected at 7 days postinoculation. For slices stored at 10°C, 10 μg/ml ColE1 significantly inhibited growth of L. monocytogenes for up to 3 days for both inoculation groups. These data indicate that ColE1 is highly effective against Listeria.

Use of One-Ply Composite Tissues in an Automated Optical Assay for Recovery of Listeria from Food Contact Surfaces and Poultry-Processing Environments

A novel one-ply composite tissue (CT) method using the Soleris (formerly BioSys) optical analysis system was compared with the conventional U.S. Department of Agriculture (USDA) environmental sponge enrichment method for recovery of Listeria from food contact surfaces and poultry-processing environments. Stainless steel and high-density polyethylene plates were inoculated to contain a six-strain L. monocytogenes cocktail at 104, 102, and 10 CFU per plate, whereas samples from naturally contaminated surfaces and floor drains from a poultry-processing facility were collected with CTs and environmental sponges. CT samples were transferred into Soleris system vials, and presumptive-positive samples were further confirmed. Sponge samples were processed for Listeria using the USDA culture method. L. monocytogenes recovery rates from inoculated stainless steel and polyethylene surfaces were then compared for the two methods in terms of sensitivity, specificity, and positive and negative predictive values. No significant differences (P > 0.05) were found between the two methods for recovery of L. monocytogenes from any of the inoculated stainless steel and polyethylene surfaces or environmental samples. Sensitivity, specificity, and overall accuracy of the CT-Soleris for recovery of Listeria from environmental samples were 83, 97, and 95%, respectively. Listeria was detected 2 to 3 days sooner with the CT-Soleris method than with the USDA culture method, thus supporting the increased efficacy of this new protocol for environmental sampling.

Transfer of Listeria monocytogenes during Slicing of Turkey Breast, Bologna, and Salami with Simulated Kitchen Knives

In response to continued concerns regarding Listeria cross-contamination during the slicing of deli meats, a series of specially prepared grade 304 and 316 stainless steel kitchen knife blades was inoculated with a six-strain Listeria monocytogenes cocktail (108, 105, and 103 CFU per blade) composed of two weak, two medium, and two strong biofilm-forming strains. The blades were then attached to an Instron 5565 electromechanical compression analyzer and used to slice whole chubs of delicatessen turkey breast, bologna, and salami to entirety (30 slices) at a cutting speed of 8.3 mm/s. Homogenates of the slices in University of Vermont Medium were surface or pour plated with modified Oxford agar and then enriched. Listeria transfer from knife blades inoculated at 108 CFU per blade was logarithmic, with a 2-log decrease seen after 8 to 12 slices and direct counts obtained thereafter out to 30 slices. However, blades containing 105 and 103 CFU per blade typically yielded direct counts out to only 20 and 5 slices, respectively. Normalizing data on a log scale for the first 10 slices resulted in significantly greater Listeria transfer and “tailing” from grade 304 as opposed to grade 316 stainless (P < 0.05) for all three products. After 1 year of use, surface roughness values as determined by surface profilometry were significantly greater (P < 0.001) for grade 304 than for grade 316 stainless blades. Cutting force and blade sharpness were not significantly different (P > 0.05) within stainless steel grade (P < 0.05) for each product. However, significant differences in cutting force were seen between salami and turkey (P < 0.05) for grades 304 and 316 stainless, respectively. In addition to compositional differences in the deli meats and knife blades, wear and scoring on the blade likely affected Listeria transfer during slicing.

Intragastric Inoculation with a Cocktail of Listeria monocytogenes Strains Does Not Potentiate the Severity of Infection in A/J Mice Compared to Inoculation with the Individual Strains Comprising the Cocktail

Although multistrain cocktails of Listeria monocytogenes are used in food inoculation experiments, no studies, to our knowledge, have been reported that use these cocktails in an intragastric mouse model. In this study, we used a five-strain L. monocytogenes cocktail consisting of strains Scott A, MFS108, 101M, V7, and 310 and a four-strain L. monocytogenes cocktail containing strains Scott A, EGD, H7738, and F2365. Here, we report that intragastric inoculation of anesthetized mice with approximately 106 CFU of a cocktail of L. monocytogenes strains does not result (P > 0.05) in a more severe infection (on the basis of the CFU of Listeria spp. recovered from the spleen, liver, and blood) than inoculation of mice with similar numbers of the individual strains comprising the cocktail. Nor did we observe any consistent relationship between susceptibility of L. monocytogenes strains to inactivation in synthetic gastric fluid in vitro and virulence in mice.

Inhibition of Listeria monocytogenes on the Surface of Individually Packaged Hot Dogs with a Packaging Film Coating Containing Nisin

The objective of this study was to determine the effectiveness of packaging films coated with a methylcellulose/hydroxypropyl methylcellulose–based solution containing 10,000, 7,500, 2,500, or 156.3 IU/ml nisin for controlling Listeria monocytogenes on the surfaces of vacuum-packaged hot dogs. Barrier film coated with a methylcellulose/hydroxypropyl methyl-cellulose–based solution containing nisin or no nisin (control) was heat sealed to form individual pouches. Hot dogs were placed in control and nisin-containing pouches and inoculated with a five-strain L. monocytogenes cocktail (approximately 5 log CFU per package), vacuum sealed, and stored for intervals of 2 h and 7, 15, 21, 28, and 60 d at 4°C. After storage, hot dogs and packages were rinsed with 0.1% peptone water. Diluent was spiral plated on modified oxford agar and tryptic soy agar and incubated to obtain counts (CFU per package). L. monocytogenes counts on hot dogs packaged in films coated with 156.3 IU/ml nisin decreased slightly (~0.5-log reduction) through day 15 of refrigerated storage but was statistically the same (P > 0.05) as hot dogs packaged in films without nisin after 60 d of storage. Packaging films coated with a cellulose-based solution containing 10,000 and 7,500 IU/ml nisin significantly decreased (P < 0.05) L. monocytogenes populations on the surface of hot dogs by greater than 2 log CFU per package throughout the 60-d study. Similar results were observed for hot dogs packaged in films coated with 2,500 IU/ml nisin; however, L. monocytogenes populations were observed to be approximately 4 log CFU per package after 60 d of refrigerated storage from plate counts on tryptic soy and modified oxford agars.

Antilisterial Effects of a Sorbate-Nisin Combination In Vitro and on Packaged Beef at Refrigeration Temperature

The antilisterial effects of a sorbate-nisin combination were assessed in vitro and on beef at refrigeration temperature. Three hemolytic pathogenic strains of Listeria monocytogenes, reference strain NCTC 7973, food strain L70, and clinical strain L94, were stored at 4°C in phosphate-buffered saline, pH 5.5, containing a combination of sorbate (0.2% wt/vol) and nisin (40 IU/ml). After 4 weeks, hemolysin production by the strains had ceased, their subsequent lag phases at 37°C were extended from an initial 1.23 to 1.32 h to a final 7.13 to 8.06 h and their pathogenicity for chick embryos had decreased from an initial 93.3 to 95.5% to a final 43.3 to 60.0%. Sterile beef steaks of normal pH (5.4 to 5.5) were inoculated with a cocktail of the three strains at approximately 5 log CFU/cm2 and the surface of half the steaks was treated with the antimicrobial solution 1.0% sorbate plus 1,000 IU of nisin per ml. The meat was packaged under vacuum or 100% carbon dioxide and stored at 4°C for 4 weeks. On untreated meat, L. monocytogenes grew by 1.79 log cycles in vacuum packages, but in CO2 packages the initial population decreased by 0.54 log cycle. On treated vacuum-packaged meat, L. monocytogenes decreased during storage to the extent that 96.5% of the initial pathogen load was eliminated, but the lag phase of the remaining cells at 37°C was unaffected. On treated CO2-packaged meat L. monocytogenes decreased during storage to the extent that 89.3% of the initial pathogen load was eliminated, and for surviving cells the lag phase at 37°C was extended. Treatment with the sorbate-nisin combination did not significantly affect pathogenicity of the L. monocytogenes cocktail recovered from vacuum- or carbon dioxide-packages after storage, in contrast to the in vitro study, where pathogenicity was clearly attenuated. The reason for this difference is unknown.