Evaluation of Hot-Water Post-Packaging Pasteurization as a Post-Lethality 

Treatment against Listeria monocytogenes on Ready-to-Eat Beef Sticks 

and Natural Casing Wieners 

Steven C. Ingham, Melody A. Fanslau, Michael D. DeVita, Rishi K. Wadhera, 

and Dennis R. Buege 

University of Wisconsin-Madison 

ABSTRACT 

This study was conducted to evaluate hot-water post-packaging 

pasteurization (PPP) as a post-lethality treatment against Listeria 

monocytogenes on Ready-to-Eat beef snack sticks and natural casing 

wieners. Using a commercially available plastic packaging film specifically 

designed for PPP applications and 2.8 liters of boiling water in a sauce pan 

on a hot plate, an average reduction in L. monocytogenes numbers of at 

least 2 logs was obtained for three brands of beef snack sticks and three 

brands of natural casing wieners using heating times of 1.0 minute for 

individually packaged beef snack sticks and 4.0 minutes for seven-perpackage beef snack sticks. A treatment of 7.0 minutes for four-per-package 

natural casing wieners was also judged to be of potential commercial use. 

Cooked-out fat and moisture resulting from selected treatments ranged 

from 0.4 to 1.1% by weight for beef snack sticks and from 0.6 to 1.2% by 

weight for natural casing wieners. For all products tested in consumer 

sensory evaluation panels, the products treated by hot-water PPP (with 

cooked-out fat and moisture removed) were rated equal to or significantly 

better than corresponding untreated products. 

On June 6, 2003, the United States Department of Agriculture (USDA) published 

an interim final rule addressing the control of Listeria monocytogenes on readyto-eat (RTE) meat and poultry products (USDA, 2003). This rule went into effect 

October 6, 2003 and was intended to encourage processors of RTE products to 

take one or more specific steps to ensure the absence of L. monocytogenes on 

1 
their products. Possible steps range from using focused sanitation procedures, 

to adding ingredients or using processing treatments designed to kill L. 

monocytogenes or inhibit its growth. Under the regulations, the processor is also 

required to perform testing for L. monocytogenes or Listeria spp. on food contact 

surfaces in the area of the plant in which RTE products are handled after 

cooking. The amount of testing is related to the types of RTE products made, 

product ingredients, and how the products are processed and handled. In 

particular, the rule requires processors of RTE meat and poultry products to 

adopt one of three designated “Alternatives” to control L. monocytogenes on their 

products. The Alternatives involve varying levels of control and microbiological 

testing. In Alternative 1, the processor uses a post-lethality treatment that 

reduces or eliminates L. monocytogenes AND an antimicrobial agent or process 

that suppresses or limits L. monocytogenes growth throughout product shelf-life. 

In Alternative 2, the processor uses either a post-lethality treatment that reduces 

or eliminates L. monocytogenes OR an antimicrobial agent or process that 

suppresses or limits L. monocytogenes growth throughout product shelf-life. 

Under Alternative 3, only sanitation measures are relied upon to control L. 

monocytogenes. 

The objective of this work was to evaluate a potential post-lethality 

treatment, hot-water post-packaging pasteurization, for use by small and very 

small processors under Alternatives 1 or 2 of the USDA regulations. This postlethality treatment was studied for use with beef snack sticks and natural casing 

wieners, two products made by a large number of small and very small 

2 
processors. In the processing of beef snack sticks, the addition of salt, cooking, 

and smoking have already been shown to be effective antimicrobial processes by 

making the finished product unsuitable for L. monocytogenes growth (Ingham et 

al., 2004). Some processors also acidify beef snack sticks, either through 

fermentation or through the addition of an acidulant. Either means of acidification 

would provide further inhibition of L. monocytogenes growth. For beef snack 

sticks then, the adoption of hot-water post-packaging pasteurization as an 

effective post-lethality treatment could allow processors to operate under 

Alternative 1. The second product studied, natural casing wieners, has 

traditionally been made without the addition of any ingredients that effectively 

inhibit L. monocytogenes growth. Thus, conducting hot-water post-packaging 

pasteurization of this product would allow processors to operate under 

Alternative 2. However, many processors have begun adding an anti-microbial 

agent such as sodium lactate to their natural casing wiener recipes. For these 

processors, the use of hot-water post-packaging pasteurization would allow 

operation under Alternative 1. 

There are a large number of variables involved in hot-water postpackaging pasteurization. These include packaging film composition and 

thickness, mass of product, volume of water, proportions of water and treated 

product, water temperature, and treatment time. In order to simplify potential 

techniques that small and very small processors might adopt, we focused our 

efforts by using a single type of packaging film, typical masses and packaging 

configurations of products, a single amount of hot water, and a single initial water 

3 
temperature. We varied the treatment time, keeping in mind that processors 

would prefer shorter treatments for reasons of economics and efficiency. 

We also paid considerable attention to whether effective hot-water postpackaging pasteurization treatments affected the sensory properties of beef 

snack sticks and natural casing wieners. Because these products would be 

subjected to a thermal process in a confining package, cook-out (almost entirely 

fat) was determined to be a potential drawback of this procedure. Therefore, we 

measured the amounts of cook-out and also conducted consumer sensory 

panels of treated and untreated products. 

Materials and Methods 

Meat Products 

Three brands each of beef snack sticks and natural casing wieners were 

purchased at local grocery stores and transported within 30 minutes to the 

laboratory. Samples were refrigerated (5°C+ 1°C) until use. A sample from a 

randomly chosen lot of each product was analyzed in the laboratory for water 

activity using a Decagon water activity meter (AquaLab Series 3 TE, Pullman, 

WA) and pH (sample homogenized in distilled water) using an Accumet AB15 pH 

meter (Fisher Scientific, Itasca, IL) and probe. Samples were also sent to a 

commercial laboratory to be analyzed for % moisture, % salt, and % fat using 

forced air oven determination of moisture – AOAC method 950.46Bb, 

potentiometric method for salt – AOAC method 980.25, and Soxhelet method for 

fat – AOAC method 960.39 (AOAC International, 1998). The Moisture: Protein 

ratio (MPR) and % water-phase salt (WPS) were calculated from these analytical 

4 
results. Representative products were measured to determine average length 

and diameter. Physical and chemical characteristics of the products are shown 

in Table 1. 

Bacterial Cultures and Preparation of Inoculum 

The L. monocytogenes strains used in this study were obtained from the 

laboratory of Dr. Eric Johnson at the Food Research Institute, University of 

Wisconsin-Madison. Strain Scott A was a clinical isolate, strains LM 101 and LM 

108 were isolated from hard salami, strain LM 310 was isolated from goat 

cheese, and strain V7 was isolated from raw milk. Stock cultures were 

maintained at -20°C in Brain Heart Infusion broth (BHIB; Difco, Becton-Dickinson, 

Sparks, MD) with 10% (w/v) glycerol (Fisher ) added. Working cultures, 

maintained at 4°C on Brain Heart Infusion agar (BHIA; Difco), were prepared 

monthly from frozen stock cultures. To obtain a working culture, a strain was 

cultured twice successively at 35°C for 18-24 h in BHIB, streaked to a BHIA 

plate, incubated at 35°C for 18-24 h and examined for purity, and then stored at 

4°C. Inoculation cultures were prepared for each strain by transferring a loopful 

of growth from the working culture plate to 9 ml of BHIB and incubating at 35°C 

for 20-24 h. To prepare the 5-strain inoculum cocktail, the BHIB cultures were 

combined into a 50-ml sterile plastic centrifuge tube, and centrifuged for 10 

minutes at 5,000 x g. The supernatant in the tube was decanted, and the pellet 

was resuspended to the original volume in Butterfield’s phosphate diluent (BPD; 

5 
Nelson Jameson, Marshfield, WI) to make an inoculum cocktail. The inoculum 

cocktail was serially diluted in BPD and plated to determine cell concentration. 

Inoculation of Meat Products 

Two different methods were used to inoculate products. Natural casing wieners, 

and beef snack sticks to be packaged four-per-package or seven-per-package, 

were inoculated by placing 110 ml of a 1:10 dilution (in BPD) of inoculum cocktail 

in a large glass Petri dish and rolling an individual product piece through the 

inoculum for two complete revolutions. Each beef snack stick to be packaged 

individually was inoculated by placing 25 μl of the 1:10 dilution (BPD) of inoculum 

cocktail on each of 5 sites along the beef snack stick. The inoculum was then 

spread across the beef snack stick surface using a sterile plastic spreader. After 

inoculation, products were allowed to dry at room temperature (20-22°C) for 90 

minutes in a biosafety level 2 hood before packaging. Initial inoculum levels 

were typically 4.5 – 5.0 log CFU per sample (see below for description of 

sampling procedure) for beef snack sticks (range was 3.1 – 5.3) and 5.5 – 6.0 for 

natural casing wieners (range of 3.0 – 6.3). 

Packaging of Meat Products 

A commercial plastic film designed for use in hot-water/steam post-packaging 

pasteurization was obtained from Curwood, Inc. (Product SPP94, Material No. 

CPS302976; New London, WI). Bags were custom-sized for products using 

scissors and a heat-sealer on a commercial-type vacuum packaging machine. 

6 
Dry, inoculated products were placed in appropriate bags and vacuum-packaged 

(approx. 1 atm). Trials were done with one, four, and seven beef snack sticks 

per package and with four natural casing wieners per package. 

Hot-Water Post-Packaging Pasteurization Treatments 

Prior to pasteurization, 2.8 liters of tap water were placed in an aluminum sauce 

pan. Pan dimensions were 21 cm diameter and 12 cm height. A plastic test tube 

rack was submerged in the pan using a lead “donut” weight. This rack was used 

to hold the packaged product off of the pan bottom during heating and thus allow 

maximum water contact with the product. Either of two hot plates was used to 

heat the water to boiling, whereupon pasteurization trials were conducted. The 

hot plates had maximum power of 113 and 118 watts. A single package of 

inoculated product was placed between levels of the test tube rack in the boiling 

water. Although the water stopped boiling when packaged product was first 

added, it resumed boiling within a short time (< 30 seconds for the largest 

packages). After the prescribed time had elapsed, the package was immediately 

removed from the water and placed in ice-water slush. The next sample was not 

heated until after the water had again boiled. Additional water was added to the 

pan as needed to maintain water volume. A trial consisted of three packages of 

product, all inoculated with an inoculum cocktail from the same broth cultures, for 

each of several treatment times. Untreated control packages were handled 

identically to treated packages other than during submersion in the boiling water. 

7 
Enumeration of Inoculum Bacteria 

Each chilled package of product was removed from the ice-water slush, after 

which the package surface was patted dry with a paper towel and then sprayed 

with 70% ethanol. Following a 20-minute drying period, each package was 

aseptically opened. A composite sample, consisting of a 2.5 cm segment from 

the center of each beef snack stick or natural casing wiener in the package, was 

obtained from the package and transferred to a stomacher filter bag. The 

package, minus remaining product, was also transferred to the stomacher filter 

bag. A standard 198 ml volume of BPD was added to the stomacher bag and the 

contents were then manually massaged for 1 minute and manually shaken for 1 

minute. This procedure ensured thorough contact of BPD with product and 

package interior surfaces. From the initial sample dilution, further dilutions were 

made, as appropriate, using BPD. From the initial dilution, 1.0 ml was distributed 

for spread-plating among three plates (0.3, 0.3, and 0.4 ml) of Listeria Selective 

Agar (LSA; Oxoid, Ogdensburg, NY) with Listeria Selective Supplements (Oxford 

formulation; Oxoid). From the original dilution and each subsequent dilution, 0.1 

ml was spread on one LSA plate per dilution. Plates were incubated at 35°C for 

48 h and then examined for typical L. monocytogenes colonies (small-medium, 

brown-to-black colonies surrounded by a black precipitate zone), which were 

counted. For each product tested in each trial, one presumptive L. 

monocytogenes colony was selected for confirmation testing. The colony was 

transferred to BHIA and cultured, and then tested for Gram stain reaction, cellular 

morphology, oxidase activity, and biochemical characteristics (API Listeria kit, 

8 
bioMerieux, Hazelwood, MO). Throughout the study, all presumptive isolates 

were confirmed as L. monocytogenes. For each trial, the difference in log CFU 

between treated and untreated product was considered the treatment lethality. 

The lethality of each trial for each treatment is shown in Table 2, along with mean 

lethality (and standard deviation when n = 3) for each treatment. 

Sensory Quality of Treated Products 

The percent of product weight lost as cooked-out fat and water was determined 

for single trials of uninoculated beef snack sticks heated for 3 and 4 minutes (four 

beef snack sticks per package) and for 4 and 5 minutes (seven beef snack sticks 

per package), and for single trials of uninoculated natural casing wieners heated 

5, 6, and 7 minutes. Results of these trials are shown in Table 3. Unscreened 

panelists (n from 178-192) also evaluated treated (4 minutes for beef snack 

sticks packaged seven-per-package and 7 minutes for natural casing wieners 

packaged four-per-package) and untreated beef snack sticks or natural casing 

wieners, which were presented in random order in cups coded with a three-digit 

random number. The treated and control natural casing wieners were separately 

warmed by being placed under boiling water for 4 minutes and then held warm in 

an insulated container prior to serving. The panelist ballot contained two 

structured seven-point hedonic scales with possible scores ranging from 1 = 

dislike very much to 7 = like very much. Mean scores for treated and untreated 

products were calculated and an analysis of variance appropriate for a 

randomized complete block design (Steel and Torrie, 1960) was conducted. 

9 
Results of sensory evaluation and subsequent statistical analysis of the sensory 

data are shown in Table 4 and 5. 

Results and Discussion 

Compliance guidelines from USDA state that an effective post-lethality treatment 

must reduce numbers of L. monocytogenes by at least 1.0 log (USDA, 2004). 

However, a higher standard of lethality must be met in order to reduce regulatory 

sampling frequency. In the latter case, L. monocytogenes numbers must be 

reduced by at least 2.0 logs. All of the PPP treatments tested had some lethality 

against L. monocytogenes on vacuum-packaged beef snack sticks and natural 

casing wieners (Table 2). With the compliance guidelines in mind, we can 

recommend hot-water PPP treatments of at least 1.0 minute for individually 

packaged beef snack sticks, at least 4.0 minutes for four-per-package and 

seven-per-package beef snack sticks, and at least 7.0 minutes for four-perpackage natural casing wieners as having potential commercial utility (Table 2). 

Treating individual beef snack sticks for 1.0 minute often eliminated all 

inoculum L. monocytogenes cells. Results from these trials would support 

validation of the 1.0 minute treatment for commercial use. Treating one-perpackage beef snack sticks for 1.5 minutes and seven-per-package beef snack 

sticks for at least 4.0 minutes also resulted in complete elimination of inoculum L. 

monocytogenes. The seemingly high variability in some of the latter trials was 

solely the result of trial-to-trial differences in the number of cells in the inoculum. 

Thus, these results would support validation of a 1.5 minute treatment time for 

10 
individually packaged beef snack sticks and a 4.0 minute treatment time for 

seven-per-package beef snack sticks. Trials with four-per-package beef snack 

sticks were less extensive, but the results showed that a treatment time of 4.0 

minutes would consistently come close to or exceed a 2.0 log reduction. 

Although a treatment time of 5.0 minutes was only tested on four-per-package 

beef snack sticks from one processor, it is likely that this treatment time could be 

validated for commercial use because this treatment time was consistently 

effective on seven-per-package beef snack sticks. 

Lethality of hot-water PPP against L. monocytogenes on natural casing 

wieners was lower than that seen for beef snack sticks (Table 2). This difference 

is likely caused by the wieners having a larger mass than the beef snack sticks 

and thus taking longer to heat. A treatment time of 7.0 minutes appeared to 

result in sufficient lethality to meet the guideline for effectiveness, but did not 

reach the lethality level needed to reduce regulatory sampling frequency. 

Variability in results for wieners and beef snack sticks was probably due to 

slight differences in product size, product surface topography, heat transfer 

through adjoining pieces, and tightness of vacuum-packaging. Increasing the 

PPP treatment time beyond those times studied here would undoubtedly 

decrease the variability of results and increase lethality, but would probably not 

be economically feasible for processors. 

In addition to the challenges inherent in achieving consistently adequate 

lethality in an economical manner, processors adopting PPP would face the 

challenge of maintaining desirable sensory properties in the treated product. The 

11 
cook-out in beef snack sticks treated by PPP ranged from 0.2 to 1.1% by weight 

for beef snack sticks and from 0.4 to 1.2% by weight for natural casing wieners. 

Particularly for beef snack sticks, cooked-out fat will be visible in packaged 

product and may be unacceptable to consumers. When PPP-treated beef snack 

sticks (with cooked-out fat and moisture removed) were served alongside 

untreated beef snack sticks to untrained consumer panelists, no negative effect 

of the PPP treatment was detected (Table 4). The hot-water PPP-treated beef 

snack sticks from processor B were actually evaluated as significantly better than 

the untreated control. Similar results were obtained for the natural casing 

wieners (Table 5). 

In summary, we believe that it is possible to meet USDA compliance 

guidelines for an effective post-lethality treatment by treating beef snack sticks 

and natural casing wieners with the relatively simple technique of hot-water PPP. 

Before PPP can be widely adopted by small and very small processors of these 

products, however, further validation studies are necessary. In addition, 

processors must determine whether hot-water PPP treatments would result in 

product characteristics that are acceptable to their customers. 

12 
References 

1. AOAC International. 1998. Official methods of analysis of AOAC 

International. 16th edition, 4th revision, Volume II. P. Cunniff (ed.). AOAC 

International, Gaithersburg, MD. 

2. Ingham, S.C., D.R. Buege, B.K. Dropp, and J.A. Losinski. 2004. Survival 

of Listeria monocytogenes during storage of ready-to-eat meat products 

processed by drying, fermentation, and/or smoking. J. Food Prot. In 

Press. 

3. Steel, R.G.D. and J.H. Torrie. 1960. Principles and procedures of 

statistics. New York: McGraw-Hill. 

4. United States Department of Agriculture, Food Safety and Inspection 

Service. 2004. Compliance guidelines to control Listeria monocytogenes 

in post-lethality exposed ready-to-eat meat and poultry products. 

Available at http://www.fsis.usda.gov/OPDE/rdad/FRPubs/97-

013F/CompGuidelines.pdf. Accessed 19 April 2004. 

5. United States Department of Agriculture, Food Safety and Inspection 

Service. 2003. Control of Listeria monocytogenes in ready-to-eat meat 

and poultry products; final rule. Fed. Regist. 68: 34207-34254. 

13 
Table 1. Physical and chemical characteristics of commercial beef snack sticks and natural casing wie

water post-packaging pasteurization. Product Beef Snack Sticks Natural Casing WienProcessor A B C A B 

Average Pre-heat Length (cm) 20 12.6 15.1 12 13.2 Average pre-heat Diameter (cm) 1.3 1.6 1.4 2.1 2.4 

pH 4.5 5.0 5.0 5.9 6.1 

Water activity (a

w) 0.91 0.86 0.89 0.95 0.97 

Moisture (%) 36.0 29.1 34.6 53.6 52.3 Protein (%) 20.6 17.2 20.0 12.3 12.8 

Moisture:Protein 1.7 1.7 1.7 4.3 4.1 

Water-phase 9.6 8.8 10.1 3.6 3.8 

Salt (%) Fat (%) 35.6 46.4 36.6 27.1 31.6 
Table 2. Lethality of hot-water post-packaging pasteurization treatments against Listeria monocytogen

beef snack sticks and natural casing wieners. A single package of product was submerged in 2.8 liters the time indicated, then chilled immediately thereafter in ice-water slush. Product Treatment Decrease (log CUF/piece) in Listeria monocytog

 Time (min.) Processor Trial Values Mean Std. De

Beef Snack Sticks 1 per pkg. 0.5 A 0.7 2.3 b 0.3 1.1 1.0 

B 2.4 1.7 1.3 1.8 0.6 C 2.5 3.5 b 2.9 3.0 0.5 

1.0 A 1.0 2.3 b 2.7b 2.0 0.9 

B 3.7 b 2.5 2.9 b 3.0 0.6 

C 3.6 b 3.2 3.6 3.5 0.2 

2.3 b 1.5 A 4.4 b 2.7 b 3.1 1.1 

B 3.7 b 3.5 b 2.9 b 3.4 0.4 C 3.6 b 3.5 b 3.9 b 3.7 0.2 

Beef snack sticks 4 per pkg. 3 B 1.4 1.5 0.2 1.0 0.7 

C 1.4 1.4 0.9 1.2 0.3 

4 A 3.9 3.9 b 4.0 b 3.9 0.1 

B 2.7 3.9 b 1.4 b 2.7 1.3 C 2.1 1.9 2.5 2.2 0.3 

5 A 4.0 b 3.9 b 4.0 b 4.0 0.1 
Beef snack sticks, 

7 per pkg. 3 B 2.5 2.7 1.5 2.2 0.6 

C 2.1 1.8 0.4 1.4 0.9 

4 B 4.0 b 2.8 b 4.3 b 3.7 0.8 

C 2.2 b 2.8 b 2.8 b 2.6 0.3 

5 A 2.3 b 2.1 b 4.1 b 2.8 1.1 

C 2.2 b 2.8 b 2.8 b 2.6 0.3 

6 A 2.3 b 2.1 b 4.1 b 2.8 1.1 

Natural Casing Wieners 4 per pkg. 

5 A 1.3 0.9 

B 1.0 1.0 1.1 1.0 0.1 D 1.3

 6 A 2.0 1.2 1.5 1.6 0.4 

B 1.1 1.1 1.4 1.2 0.2 

D 1.1 1.4 

7 A 4.3 1.1 1.9 2.4 1.7 

B 1.4 3.4 D 1.6 1.8 

a Each value is the result from an individual trial. Three samples were analyzed for each trial. b Indicates that no surviving cells were detected. For such a trial, a value of [Log (zero time) - Log (Detection Limit)] + 0.1 

decrease in cells. 
Table 3. Cook-out loss during hot-water post-packaging pasteurization of beef snack sticks and natura

Product Treatment Time Processor % weight lost in cook-outa

 (min.) 

Beef Snack Sticks 4 per package 3 

4

7 per package 4 

5

A 

B

 C

 A

 B C 

A 

B

 C

 A

 B C 

0.9 

 0.2

 0.8 0.9 0.6 0.6 

1.1 

 0.4

 0.7 0.8 0.7 0.7 

Natural Casing Wieners 4 per package 5 A 

D

 0.4 

 0.4

 6 A

 D

 0.8 0.5

 7

aEach value is for a single package of product. A

 B D

 1.2 1.0 0.6 
Table 4. Consumer acceptance of beef snack sticks (seven per package) treated for 4.0 minutes with 

packaging pasteurization. “Trtmt” = treated product, “Ctrl” = untreated product. Processor A B C

 Trtmt. Ctrl Trtmt Ctrl Trtmt 

Assigned Descriptor Numerical score Number of responses Like very much = 7 64 65 52 49 Like moderately = 6 71 69 84 66 Like slightly = 5 33 35 32 33 

Neither like nor dislike = 4 6 7 8 11 

76 

61 41 

7 

Dislike slightly = 3 5 4 7 18 

Dislike moderately = 2 2 2 2 7 

Dislike very much = 1 1 0 1 2 

Mean score 5.95 5.98 5.84 5.47 

4 2 

1 

5.98 

Trtmt significantly different? NO YES NO 
Table 5. Consumer acceptance of natural casing wieners (four per package) treated for 7 minutes with 

packaging pasteurization. “Trtmt” = treated product, “Ctrl” = untreated product. Processor A B D

 Trtmt. Ctrl Trtmt Ctrl Trtmt 

Assigned Descriptor Numerical score Number of responses Like very much = 7 53 60 66 45 Like moderately = 6 79 81 68 79 Like slightly = 5 35 24 29 26 

Neither like nor dislike = 4 5 9 12 14 

56 

71 33 

7 

Dislike slightly = 3 6 6 6 10 

Dislike moderately = 2 3 3 2 6 

Dislike very much = 1 2 0 1 4 

Mean score 5.83 5.93 5.90 5.55 

9 2 

0 

5.85 

Trtmt significantly different? NO YES NO