Hot water post-process pasteurization of cook-in-bag turkey

breast treated with and without potassium lactate and sodium diacetate and acidified sodium chlorite for control of Listeria monocytogenes

JOHN B. LUCHANSKY¹*, GEORGE COCOMA², and JEFFREY E. CALL¹

¹Microbial Food Safety Research Unit, U.S. Department of Agriculture, Agricultural Research Service, Eastern Regional Research Center, 600 East Mermaid Lane, Wyndmoor, Pennsylvania 19038, USA, and ²Professional Resource Organization, Edmund, Oklahoma 73003, USA.

Key words: Listeria monocytogenes, turkey, post process pasteurization, hot water, potassium lactate, sodium diacetate, acidified sodium chlorite, meat, food safety.

*   Author for correspondence: Tel: 215-233-6620; Fax: 215-233-6581; E-mail: jluchansky@errc.ars.usda.gov.

^{^}    Mention of trade names or commercial products in this article is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture.

I  Portions of this research were presented at the Annual Meeting of the International Association for Food Protection, August 08-11, 2004, Phoenix, AZ (20).

ABSTRACT

Surface pasteurization and food grade chemicals were evaluated for post process control of listeriae on cook-in-bag turkey breasts (CIBTB). Individual CIBTB were obtained directly from a commercial manufacturer and surface inoculated (20 ml) with a 5-strain cocktail (ca. 7.0 log₁₀ total) of Listeria innocua. In each of two trials, the product was showered/submerged for up to 9 minutes with water heated to190°, 197°, or 205°F in a commercial pasteurization tunnel. Surviving listeriae were recovered from CIBTB by rinsing and then enumerated on MOX agar plates following incubation at 37°C for 48 hours. As expected, higher water temperatures and longer residence times resulted in a greater reduction of L. innocua. About a 2.0-log₁₀ reduction was achieved within 3 minutes at 205° and 197°F and within 7 minutes at 190°F. In related experiments, the following treatments were evaluated for control of Listeria monocytogenes on CIBTB: 1) potassium lactate/sodium diacetate solution (1.54% potassium lactate and 0.11% sodium diacetate) added to the formulation in the mixer and 150 ppm acidified sodium chlorite (ASC) applied to the surface with a pipette, or 2) potassium lactate/sodium diacetate solution only, or 3) no potassium lactate/sodium diacetate solution and no ASC. Each CIBTB was inoculated (20 ml) with about 5 log₁₀ CFU of a five-strain mixture of L. monocytogenes and then vacuum-sealed. In each of two trials, one-half of the CIBTB were exposed to 203°F water for 3 minutes in a pasteurization tunnel and the other one-half were not, and then all CIBTB were stored at 4°C for up to 60 days and L. monocytogenes were enumerated by direct plating onto MOX agar. Heating resulted in an initial reduction of about 2 log₁₀ CFU of L. monocytogenes per CIBTB. For heated CIBTB, L. monocytogenes increased by about 2 log₁₀ CFU per CIBTB in 28 (treatment 1), 28 (treatment 2), and 14 (treatment 3) days. Thereafter, pathogen levels reached about 7 log₁₀ CFU per CIBTB in 45, 45, and 21 days for treatments 1, 2, and 3, respectively. In contrast, for non‑heated CIBTB, L. monocytogenes levels increased from about 5 log₁₀ CFU per CIBTB to about 7 log₁₀ CFU per CIBTB in 28, 21, and 14 days for treatments 1, 2, and 3, respectively. Lastly, in each of three trials we tested the effect of hot water (203°F for 3 minutes) post process pasteurization of inoculated CIBTB on lethality towards L. monocytogenes and validated that it resulted in a 1.8-log₁₀ reduction in pathogen levels. Collectively, these data establish that hot water post-process pasteurization alone is effective in reducing L. monocytogenes on the surface of CIBTB. However, as used in this study, the potassium lactate/sodium diacetate solution and ASC were only somewhat effective at controlling the subsequent outgrowth of this pathogen during refrigerated storage.

Listeria monocytogenes remains a significant foodborne pathogen, especially for ready-to-eat (RTE) meats. Evidence for this can be found in a risk assessment of 23 selected categories of RTE foods (http://www.foodsafety.gov/~dms/lmr2-toc.html) wherein deli foods and non-reheated frankfurters were identified as the highest risk foods on a per serving basis. Further evidence is provided by its recovery from commercially-prepared and retail RTE foods and by the continued documentation of product recalls and human illness. Surveys in the United States between 1990 and 2003 involving ~100,000 food samples estimated the prevalence of L. monocytogenes at 1.6% to 7.6% in meat, fish, and vegetable products, most of which were RTE foods (15, 18, 39). Also, Thomsen and McKenzie (38) reported that between 1982 and 1998 there were an average of 5 recalls per year due to contamination of foods with L. monocytogenes. Moreover, in recent years there have been several illnesses and deaths attributed to food borne listeriosis (8-11, 24). Thus, further research is warranted to evaluate post process interventions to enhance the safety of high volume RTE foods that may be associated with L. monocytogenes or that may support its growth.

Due to the severity of listeriosis and the number and magnitude of food recalls, regulatory agencies have established requirements for manufacturers to better control L. monocytogenes in meat and poultry. More specifically, the USDA Food Safety and Inspection Service (USDA/FSIS) has established new rules/guidelines for manufacturers of RTE meat and poultry products (1). In simple terms, this ruling provides manufacturers with the following three alternatives for determining the degree to which regulatory testing would be implemented: alternative 1, whereby a manufacturer would use a post process lethality step AND an antimicrobial to control outgrowth (lowest testing frequency); alternative 2, whereby a manufacturer would use either a post process lethality step OR an antimicrobial to control outgrowth (moderate/increased testing relative to alternative 1); or alternative 3, whereby a manufacturer would rely on sanitation alone (most testing). Although further clarification is expected regarding how much lethality is required or how little outgrowth is allowed, there is an immediate and critical need for identifying and implementing post process interventions for killing and/or inhibiting L. monocytogenes in RTE meats.

Various interventions are effective against L. monocytogenes, including some that have proven useful in controlling this pathogen in the event of post process contamination of RTE foods. Heat is arguably the most effective and most studied method for post process pasteurization. Several investigators have validated the use of heat to control L. monocytogenes associated with frankfurters, summer sausage, ground beef, ground pork, deli-type meat, and poultry products (5, 16, 29, 31, 32, 40, 42). Depending on the product, as well as the times and temperatures used for heating, reductions of L. monocytogenes ranging from 2.0 to 7.0 log₁₀ CFU have been reported. However, certain food grade chemicals, including organic acids such as potassium lactate and sodium diacetate, alone or in combination with other interventions, are also effective at controlling L. monocytogenes in RTE meats (3, 4, 6, 22, 30, 35, 36, 37, 41). In addition to use as a flavorant, the USDA/FSIS has approved the use of “lactates” at levels up to 4.8% of the total formulation to slow the growth of pathogens in fully cooked products. In addition, pH-altering inorganic compounds, such as acidified sodium chlorite (ASC), are also effective for controlling this pathogen, notably on fresh beef and on beef and broiler carcasses (7, 17). In the United States, ASC is approved at 500 to 1200 ppm at pH 2.3-2.9 for various applications, including for poultry and red meats, as well as processed, comminuted or formed meat products. For example, Mullerat et al. (25) reported reductions of L. monocytogenes, Salmonella spp., Escherichia coli O157:H7, Staphylococcus aureus, and Pseudomonas aeruginosa ranging from <1.0 to 5.0 log₁₀ after a 15-minute exposure to a solution containing 10 mM sodium chlorite. Although heat and food grade chemicals have antilisterial activity, a combination of interventions may be required to assist producers in satisfying current regulatory requirements for controlling L. monocytogenes in RTE meat products without adversely impacting product quality. Thus, the objective of the present study was to investigate the efficacy of using a combination of potassium lactate and sodium diacetate as ingredients and ASC as a surface-applied agent coupled with post-packaging hot water pasteurization to control L. monocytogenes on the surface of cook-in-bag turkey breasts (CIBTB).

MATERIALS AND METHODS

Bacterial strains. As described previously (30), approximately equal numbers of each of the following five strains of Listeria monocytogenes were used as a cocktail in this study: i) Scott A (serotype 4b, clinical isolate), ii) H7776 (serotype 4b, frankfurter isolate), iii) LM-101M (serotype 4b, beef and pork sausage isolate), iv) F6854 (serotype 1/2a, turkey frankfurter isolate), and v) MFS-2 (serotype 1/2a, environmental isolate from a pork processing plant). Likewise, approximately equal numbers of the following five strains of Listeria innocua were used as a cocktail in some experiments: i) 2428 (raw egg), ii) 2283 (turkey), iii) LA-1 (cheese), iv) S5-VJ-S (sausage), and v) LG2 (ground lamb). Isolates were passed twice in brain heart infusion (BHI; Difco Laboratories, Detroit, MI) broth at 37°C so that cells would be in the stationary phase for each experiment. Stock cultures were maintained by storage in BHI plus 10% (wt/vol) glycerol in 1.5-ml portions in cryovials and held at -80°C.

Inoculation of commercially-prepared cook-in-bag turkey breasts (CIBTB) with L. innocua for use in hot water post-process pasteurization studies. Whole turkey breasts (about 8 to 9 pounds) were de-boned, hand-formed, vacuum-sealed, cooked, and cooled to 4°C by a cooperating commercial processor. After processing, the uncured CIBTB were boxed, transported back to the laboratory, and stored at 4°C for up to 7 days before used. Each CIBTB was aseptically removed from its original packaging and re-packaged into a 12 × 18 inch post-pasteurization bag (CNP310T; Cryovac, Saddlebrook, NJ). Twenty milliliters of the L. innocua cocktail were added to each package to achieve a target level of about 7 log₁₀ CFU per CIBTB. Each package was then massaged by hand for about 2 minutes to distribute the inoculum and then vacuum sealed to 950 mBar using a Multivac A300/16 vacuum-packaging unit (Sepp Haggemüller KG, Wolfertschwenden, Germany). Next, the CIBTB were exposed to either 205°F water for residence times of 0, 1, 2, and 3 minutes, 197°F water for 0, 3, 5, and 7 minutes, or 190°F for 0, 5, 7, and 9 minutes in a pilot plant scale pasteurization tunnel (model #PT1030, Gal-Esh Stainless Steel Products Ltd., Holon, Israel). In brief, the CIBTB were transported through the pasteurization tunnel on a conveyer belt adjusted using the tunnel’s variable speed variator to achieve the targeted residence times, that being 1 to 9 minutes depending on the experiment. While in the tunnel, approximately one-half of each CIBTB was submerged in water heated to the target temperature and the other one-half was continuously showered with hot water heated to the target temperature. The temperature of the top and bottom surfaces of selected CIBTB were monitored during runs at all three temperatures using a stainless steel DeltaTRAK [model EBI-125A, (48 mm diameter X 28 mm thick), Delta Trak, Inc., Pleasanton, CA] data logger. For each CIBTB, a portion of the meat approximately the same diameter and thickness of the data logger was removed from the product and replaced with the data logger. For each temperature and for each trial, a data logger was placed on the top and on the bottom surface of a single CIBTB to monitor temperature during transit through the pasteurization tunnel. After heating, the CIBTB were quickly cooled by submersion in an ice water bath. For each of two trials, three CIBTB were analyzed per residence time at each temperature.

            Inoculation of commercially-prepared CIBTB with L. monocytogenes for use in post-process validation studies. A single batch of chopped/formed turkey breasts was prepared by the same commercial manufacturer as above. For these studies, as needed, 2.75% of a 60% solution of potassium lactate and sodium diacetate (K-lactate/Na-diacetate, a 56% potassium lactate, 4% sodium diacetate solution in dH₂O; Opti-Form® 4, Purac America, Inc., Lincolnshire, IL) was added to a ribbon mixer. The K-lactate/Na-diacetate solution was added to achieve a target concentration of potassium lactate and sodium diacetate of about 1.54 and 0.11%, respectively. After processing, the uncured CIBTB were boxed, transported back to the laboratory, and stored at 4°C for up to 7 days before used. Each CIBTB was aseptically removed from the original vacuum packaging and re-packaged as described above. As needed, a stock solution of acidified sodium chlorite (ASC, a 3.35% solution of sodium chlorite; Bio-Cide International, Norman, OK) was prepared and activated as follows: 2.84 g of citric acid (Sigma Chemical Co., St. Louis, MO) was added to 7.1 ml of the 3.35% sodium chlorite solution and allowed to “activate” for 10 minutes at room temperature. The “activated” or acidified sodium chlorite was then diluted in 947 ml of sterile distilled water to achieve a final concentration of 250 ppm. The pH of the ASC stock solution of ASC was ca. pH 2.5. Next, 15 ml of the 250 ppm stock solution was applied to the surface of the K-lactate/Na-diacetate containing CIBTB with the aid of a pipette to achieve a final concentration of ca.150 ppm when combined with the L. monocytogenes inoculum. This level was suggested by the supplier of ASC to compliment the types/levels of the other ingredients. Twenty milliliters of the L. monocytogenes cocktail were added to each package to achieve a target level of about 7 log₁₀ CFU per CIBTB. Each package was then massaged by hand for about 2 minutes to distribute the ASC and the inoculum before being vacuum sealed as described above. The CIBTB were exposed to 203°F water for residence times of 0, 1, 3, 5, and 7 minutes in the above mentioned pasteurization tunnel and the temperature of the top and bottom surfaces of selected CIBTB were monitored as described above. After heating, CIBTB were quickly cooled by submersion in an ice water bath. For each of three trials, three CIBTB were analyzed per each residence time at 203°F.

            Inoculation of commercially-prepared CIBTB with L. monocytogenes for use in hot water post-process pasteurization and shelf life studies. A single batch of CIBTB, with and without K-lactate/Na-diacetate, was obtained from the above mentioned commercial processor. The CIBTB were treated as follows: i) K-lactate/Na-diacetate plus ASC; ii) K-lactate/Na-diacetate only; and iii) no K-lactate/Na-diacetate, no ASC. Each CIBTB was re-packaged in pasteurization bags and 20 ml of the L. monocytogenes cocktail were added to each package to achieve a target level of about 5 log₁₀ CFU per CIBTB. For each of these three treatments, one portion of the inoculated CIBTB was not heated, whereas another portion of the CIBTB was exposed to 203°F water for 3 minutes in the pasteurization tunnel. All CIBTB were stored at 4°C and analyzed 0, 7, 14, 21, 28, 45, and/or 60 days post-inoculation. Control samples that were not inoculated with the pathogen were analyzed on days 1, 45, and 60 post-inoculation. For both trials, three CIBTB, heated and non-heated, were analyzed per sampling time point for each of the three treatments.

            Microbiological analyses. Surviving listeriae were enumerated using the USDA/ARS package rinse method (21) and spread-plating 250 ul of the resulting rinse fluid or dilutions thereof onto duplicate modified Oxford (MOX; 12) agar plates using a spiral plater (Autoplate 4000, Spiral Biotech, Gaithersburg, MD) and incubating for 48 hours at 37°C. Bacterial numbers were expressed as log₁₀ CFU per package with each package containing a single CIBTB. Select colonies were randomly selected and confirmed as L. monocytogenes following standard USDA/FSIS methods (12).

            Chemical analyses. The proximate composition of CIBTB was determined using methods approved and described by the Association of Official Analytical Chemists (23) as conducted by a commercial testing laboratory. For day 0 for one trial of the shelf life study proximate analyses were performed on two CIBTB treated with K-lactate/Na-diacetate and ASC and 2 CIBTB that were not treated with either of these food grade chemicals.

            Statistical analyses. Data were analyzed using version 8.0 of the SAS statistical package (SAS Institute, Inc., Cary, NC). The data were compared by analysis of covariance and F-tests to evaluate the effect of time, temperature, and food grade antimicrobials on the viability of listeriae in packages of CIBTB during extended storage at 4°C for the shelf life component of this study and to evaluate the effect of chemicals on the thermal stability of the pathogen after passage through the pasteurization tunnel in the validation component of this study.

RESULTS

Proximate composition and temperature profile of CIBTB: A representative sample, consisting of two CIBTB treated with and two CIBTB treated without K-lactate/Na-diacetate and ASC collected from a single trial of the shelf life study, was forwarded to a private testing laboratory for proximate composition analyses (Table 1). Chemical analyses revealed significant differences (P > 0.05) among NaCl, fat, carbohydrate, and lactic acid levels in CIBTB treated with and without K-lactate/Na-diacetate and ASC but did not reveal appreciable differences in levels of the other chemicals assayed. Although there were significant differences in the levels of some of the chemicals tested, by empirical analyses these differences did not have an appreciable effect on the outgrowth of L. monocytogenes in CIBTB during storage. Results also revealed no untoward effects on the texture or appearance of CIBTB under such conditions (data not shown). Also, sampling of non-inoculated packages of CIBTB revealed the absence (i.e., <20 CFU per package) of indigenous L. monocytogenes by direct plating (data not shown).

To address the issue of heat transfer/delivery we measured the temperature achieved for both the top and bottom surfaces of CIBTB during exposure to water heated to 190°, 197°, 203°, and 205°F. The maximum top surface temperatures obtained for the hot water post-process pasteurization studies using L. innocua were 194°, 191°, and 176°F when the water in the pasteurization tunnel was heated to 205°, 197°, and 190°F, respectively. The maximum bottom surface temperatures obtained for these same studies were 195°, 185°, and 186°F when the water in the pasteurization tunnel was heated to 205°, 197°, and 190°F, respectively. During post-processing pasteurization validation studies at 203°F for 3 minutes using CIBTB containing K-lactate/Na-diacetate and ASC and inoculated with L. monocytogenes, the maximum top and bottom surface temperatures achieved were 182° and 192°F, respectively. In comparison, the maximum top and bottom surface temperatures achieved in CIBTB not treated with food grade chemicals were 183° and 192°F, respectively, following treatment at 203°F for 3 minutes. These data indicate that the temperature of the water was higher than the temperature of the product and that in general the temperatures at the bottom were higher than the top probably because the bottom surface was submerged and this allowed for better heat transfer.

            Post-packaging surface pasteurization of CIBTB inoculated with L. innocua. We used a L. innocua cocktail as a surrogate for L. monocytogenes to gain insight on the fate of listeriae on CIBTB following transit through the pasteurization tunnel. These data were then used in subsequent inoculated package experiments to validate the lethality of the hot water pasteurization step on L. monocytogenes. As expected, the higher the temperature and the longer the residence time, the greater the reduction in the population of L. innocua (Table 2). For each of the three temperatures tested, we observed at least a 2.0-log₁₀ reduction within 7 minutes at 190°F, 3 minutes at 197°F, and 3 minutes at 205°F. The greatest reduction, that being 2.1 and 2.55 log_10, was achieved within 3 minutes at 205°F and within 7 minutes at 197°F, respectively. Visual inspection confirmed that these time and temperature combinations had little or no effect on the texture or appearance of CIBTB (data not shown).

            Validation of a defined time/temperature combination for post-packaging pasteurization of CIBTB to control L. monocytogenes. Based on our results from hot water surface pasteurization of L. innocua on CIBTB and on input from an industry partner, we validated the lethality towards L. monocytogenes of processing CIBTB in a pasteurization tunnel using water heated to 203°F (Table 3). One portion of CIBTB was formulated with K-lactate/Na-diacetate and surface treated with ASC (treatment 1), whereas another portion was formulated without K-lactate/Na-diacetate and ASC (treatment 2). Based on the average of three independent trials, with the possible exception of heating for 1 minute wherein a 1.5-log₁₀ reduction was achieved for CIBTB without antimicrobials (i.e., treatment 2) and a 0.92-log₁₀ reduction was achieved for CIBTB containing antimicrobials (i.e., treatment 1), there was essentially no difference in lethality relative to formulation for CIBTB processed in the pasteurization tunnel using water heated to 203°F for 3, 5, or 7 minutes. These data validate that processing for 3 minutes with water heated to 203°F would result in about a 1.8-log₁₀ reduction in levels of L. monocytogenes using the conditions, product, and pasteurization tunnel tested herein.

            Effect of post-processing surface pasteurization and formulation of CIBTB for control of L. monocytogenes during refrigerated storage. In addition to verifying the initial/direct reduction achieved using post processing surface pasteurization at 203°F, we also evaluated the effect of heat and food grade chemicals for precluding outgrowth of L. monocytogenes over the expected refrigerated shelf life of the product. The CIBTB were formulated as follows: K-lactate/Na-diacetate as an ingredient (treatment 1), K-lactate/Na-diacetate as an ingredient and surface applied ASC (treatment 2), and no K-lactate/Na-diacetate and no ASC (treatment 3). In general, levels of L. monocytogenes were significantly reduced (P > 0.05) on the surface of CIBTB that were subjected to 203°F water for 3 minutes, regardless of treatment type, compared with CIBTB that were not heated (Table 4). Specifically, pathogen numbers were reduced by 2.2, 2.4, and 2.2 log₁₀ CFU for treatments 1, 2, and 3, respectively, for heated CIBTB. These results compare favorably with results detailed in the previous section on processing CIBTB for 3 minutes at 203°F for post process pasteurization wherein we observed a 1.8-log₁₀ reduction in levels of L. monocytogenes.

During subsequent storage at 4°C (Table 4), for heated CIBTB, L. monocytogenes levels increased by about 2 log₁₀ CFU per CIBTB in 28 (treatment 1), 28 (treatment 2), and 14 (treatment 3) days. Thereafter, pathogen levels reached about 7 log₁₀ CFU per CIBTB in 45, 45, and 21 days for treatments 1, 2, and 3, respectively. In contrast, in non-heated CIBTB, the pathogen numbers increased from about 5 log₁₀ CFU per CIBTB to about 7 log₁₀ CFU per CIBTB in 28, 21, and 14 days for treatments 1, 2, and 3, respectively. Thereafter, for all treatments, pathogen levels continued to increase to about 10⁸ to 10¹⁰ CFU per CIBTB. In general, inclusion of K-lactate/Na-diacetate and/or ASC had no apparent effect on controlling the subsequent outgrowth of the pathogen during refrigerated storage. These data indicate that use of K-lactate/Na-diacetate as an ingredient alone or in combination with surface applied ASC was not totally effective at controlling outgrowth of L. monocytogenes during extended refrigerated storage.

DISCUSSION

The number and magnitude of meat and poultry recalls in recent years due to contamination of RTE foods with L. monocytogenes is staggering. Between 1999 and 2002 there were 5 major recalls that alone resulted in the voluntary recall of some 130 million pounds of RTE foods. Of particular relevance to the present study was a recall in 2002 of some 30 million pounds of deli turkey products linked to 43 illnesses and 13 deaths (11). The various recalls and epidemiologically-linked illnesses associated with RTE foods have most likely been a major factor in the implementation of a recent USDA/FSIS ruling that requires processors of post-lethality exposed RTE foods to control the contamination of such products with L. monocytogenes using a Hazard Analysis Critical Control Point (HACCP) plan or Sanitation Standard Operating Procedure (SSOP). Manufacturers are also required to formulate/process RTE meat and poultry products to achieve some level of post-process lethality and/or to prevent subsequent outgrowth of the pathogen during shelf life. Thus, along with several other investigators, we evaluated processes aimed at eliminating any cells of L. monocytogenes that may be acquired post process as incidental contaminants of RTE meat and poultry products. In the present study we tested different temperatures and residence times for processing CIBTB in a commercial hot water pasteurization tunnel for post-process lethality against L. monocytogenes. We also evaluated food grade chemicals for the ability to inhibit L. monocytogenes during extended refrigerated storage. Several investigators have confirmed that these treatments are effective when used singly, but there have been far fewer studies evaluating their efficacy to control L. monocytogenes when used in combination for RTE poultry during refrigerated storage.

Since heat is the most common and probably the most effective intervention, prefatory experiments were conducted to evaluate a range of time and temperature combinations likely to be used by manufacturers as a post-processing lethality treatment for RTE poultry products. These experiments used a 5-strain cocktail of L. innocua as a surrogate for L. monocytogenes and CIBTB that did not contain any lactates. The times and temperatures evaluated in this portion of the study resulted in an appreciable reduction in pathogen levels, that being about a 2.5-log₁₀ decrease per CIBTB, but did not result in any undesirable effects on the product per se, that being no discoloration or foul odors, etc. Moreover, these results compare favorably with those already published.

There have been several studies undertaken to evaluate both pre- and post-process surface pasteurization of RTE meat and poultry products to control L. monocytogenes. For example, Muriana and colleagues reported significant reductions of L. monocytogenes on deli-type turkey following post process submersion in hot water and/or in a radiant oven (14, 26, 27). Similar results were obtained by Murphy et al. (28) on turkey breasts using submersion heating to reduce levels of L. monocytogenes. Conditions for heating should also be adjusted to accommodate differences in lethality toward L. monocytogenes due to the formulation/type of CIBTB and manufacturing conditions (27). Regardless, our findings validated that hot water post-process pasteurization delivered at least a 1.8-log₁₀ reduction of L. monocytogenes per CIBTB after passage for 3 minutes in a pasteurization tunnel while showered/submerged with water heated to 203°F. In practice, it would be desirable for cost, product quality, and pathogen lethality to process CIBTB through the pasteurization tunnel in <3 minutes using water heated to 203°F. The challenge is to apply a sufficient amount of heat for a sufficient length of time to achieve the desired reduction of the target pathogen without causing undesirable effects on the product.

The efficacy of potassium lactate and sodium diacetate for controlling L. monocytogenes in RTE meats has been well demonstrated by a number of researchers (3, 4, 30, 33, 34, 41). In a previous study, we also showed that inclusion of 2% potassium lactate as an ingredient in frankfurters was quite effective at preventing outgrowth of the pathogen during refrigerated storage, that being <1.0 log₁₀ increase over 90 days (30). However, potassium lactate did not have an appreciable effect on thermal inactivation of the pathogen when present on the surface of frankfurters that were subsequently heated at near boiling temperatures, that is similar results were obtained for frankfurters formulated with and without potassium lactate (31). We did report that re-heating at near boiling temperatures for about 36 seconds was sufficient to achieve a 5‑log₁₀ reduction in pathogen numbers (31). The results of our previous studies are in agreement with the present study relative to thermal inactivation of the pathogen, but are not in agreement relative to outgrowth of the pathogen during refrigerated storage. This may be due, in part, to the use of lower levels of potassium lactate in the present study (ca. 1.5%) compared to our previous study (ca. 2.0%) and/or the fact that frankfurters and CIBTB differ in their formulation, shape, surface area, and/or topography. However, the present study also used sodium diacetate (ca. 0.11%) as an ingredient in combination with potassium lactate and we were anticipating a synergistic antilisterial effect. Further experiments must be conducted to optimize the levels/types of antimicrobials for controlling listeriae on CIBTB and to evaluate the most effective method to deliver antimicrobials to RTE products.

Although the effectiveness of ASC has not been thoroughly demonstrated for RTE meats, a recent study revealed that 0.12% of ASC reduced levels of L. monocytogenes on fresh beef by about 1.8 log₁₀ within 2 weeks at 4°C, albeit while negatively affecting the color of the beef (19). Frank et al. (13) reported that 10 minute exposures of a biofilm of L. monocytogenes on stainless steel to a combination of alkali cleaning and ASC were effective at achieving about a 6.0-log₁₀ reduction of the pathogen. Previous studies have also evaluated higher levels (500 to 1200 ppm) of ASC than what was used in the present study (150 ppm). Regardless, in the present study ASC did not have an appreciable effect on inhibiting the outgrowth of L. monocytogenes, presumably because far less ASC was applied than what is recommended/typical. Further work is warranted to establish if higher levels and/or different points of delivery and/or different delivery methods would yield a greater antilisterial response with ASC or other food grade chemicals.

            We initiated this study in response to the voluntarily recall of some 30 million pounds of poultry products by 2 processors in the Northeastern portion of the United States due to contamination with L. monocytogenes (11). It was critical to optimize and implement interventions to eliminate any L. monocytogenes that might become associated with CIBTB after cooking, but prior to final packaging, and then to preclude any residual cells from growing to potentially dangerous levels during refrigerated storage. In addition to demonstrating the effectiveness of hot water post process pasteurization of CIBTB for achieving “alternative 2” status, the further optimization of food grade chemicals to prevent outgrowth of L. monocytogenes may allow manufacturers of such products to reformulate and achieve “alternative 1” status. This may also be significant if regulatory agencies soften the current “zero tolerance” policy and allow a certain level, for example 100 CFU per gram, of L. monocytogenes in products that do not support its growth (2). Our findings establish that hot water post-process pasteurization is effective in reducing initial levels of L. monocytogenes by about 2.0-log₁₀ CFU on CIBTB and that K-lactate/Na-diacetate added as an ingredient and ASC applied to the surface of CIBTB were not effective at controlling the subsequent outgrowth of L. monocytogenes during refrigerated storage. Studies are ongoing to investigate in more detail the optimal time and temperature parameters required to enhance the thermal inactivation of L. monocytogenes in CIBTB, as well as to identify effective food grade chemicals and delivery methods for antimicrobials to prevent outgrowth of this pathogen in RTE meat and poultry products.

ACKNOWLEDGMENTS

We acknowledge the assistance/input of Jenny Bumanlag, Brian Dirks, Vijay Juneja, Wendy Kramer, and Joseph Sites of the USDA, Agricultural Research Service, Eastern Regional Research Center, Wyndmoor, PA.

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Proximate composition analyses of cook-in-bag turkey breasts (CIBTB) treated with and without K-lactate/Na-diacetate and acidified sodium chlorite (ASC)*

* Two CIBTB’s were analyzed per treatment. All CIBTB were obtained from day 0 of the shelf life study experiments.

** Indicates that values/levels are significantly different (P>0.05) between CIBTB treatments.

Effect of post-process hot water pasteurization treatments on CIBTB* inoculated with L. innocua (N = 2 trials, n = 3 CIBTB per sampling point).

* CIBTB were not treated with K-lactate/Na-diacetate or ASC for these experiments.

** log₁₀ CFU/package.

*** standard deviations (log₁₀).

N.D. = not determined.

Validation of post-process hot water pasteurization (203°F for 3 minutes) treatment of CIBTB inoculated with L. monocytogenes (N = 3 trials, n = 3 CIBTB per sampling point).

* CIBTB treated with K-lactate/Na-diacetate and ASC.

** CIBTB not treated with K-lactate/Na-diacetate or ASC.

***log₁₀ CFU/package.

**** Standard Deviations (log₁₀).

Effect of post-process hot water pasteurization treatments, K-lactate/Na-diacetate, and acidified sodium chlorite (ASC) on the viability of L. monocytogenes inoculated onto the surface of CIBTB during storage at 4°C (N = 2 trials, n = 3 CIBTB per sampling point).

*Treatment 1 = CIBTB treated with K-lactate/Na-diacetate and ASC. Treatment 2 = CIBTB treated with K-lactate/Na-diacetate only. Treatment 3 = CIBTB not treated with K-lactate/Na-diacetate or ASC. Initial levels of L. monocytogenes are lower for treatments that were heated compared to treatments that were not heated due to exposure of CIBTB to water heated at 203°F for 3 minutes prior to storage.

**log₁₀ CFU/package.

*** log₁₀ CFU/package values with the same letter are statistically similar.

****Standard Deviations (log₁₀).

N.D. = not determined.