International Journal 

of Food Microbiology 

ELSEVIER International Journal of Food Microbiology 37 (1997) 47-54 

Viability of Escherichia coli 0157:H7 in pepperoni during the 

manufacture of sticks and the subsequent storage of slices at 21, 4 

and - 20°C under air, vacuum and CO2 

Nancy G. Faith\ Nelly Parniere1.a, Trina Larson\ Timothy D. LorangU, 

John B. Luchanskya.b.* 

"Department o{ Food Microbiology and Toxicolog_v. Food Research Institute, 1925 Willow Drive. Madison WI 53706, USA 

"Department o{ Food Science, University o{ Wisconsin. Madison WI 53706, USA 

Received 13 January 1997; received in revised form 4 April I 997; accepted 23 April I 997 

*corresponding author. Tel.:+ I 608 2637777; fax : + 1 608 

2631114; e-mail: jbluchan@facstaff.wisc.edu 

'Present address: The National Institute of Agrifood Industries 

(INSFAJ, 65, Rue de Saint-Brieuc, 35042 Rennes, France. Tel.: 

+ 33 2 47665221; e-mail: parniere@france-cybermedia.com 

0168-1605/97/$17.00 © 1997 Elsevier Science BV. All rights reserved 

PII S0168-1605(97)00052-4 

Abstract 

A raw. pepperoni batter (75% pork:25% beef with a fat content of about 32%) was inoculated with a pediococcal starter 

culture (about 108 cfu/g) and a five-strain cocktail of Escherichia coli O157:H7 ( 2: 2 X 107 cfu/g), mixed with non-meat 

ingredients, and then hand-stuffed into 55 mm fibrous casings to form sticks. The numbers of the pathogen were determined 

before stuffing, after fermentation, after drying/slicing, and after periods of storage. For storage, slices were packaged under 

air, vacuum or CO2 and stored at - 20, 4 and 21°C. Sticks were fermented at 36°C and 85% relative humidity (RH) to 

-cc; pH 4.8 and then dried at I 3°C and 65% RH to a moisture/protein ratio (M/Pr) of -cc; 1.6: I. Fermentation and drying 

resulted in the numbers of the pathogen decreasing by about 2 log, 0 units. During storage, the temperature rather than the 

atmosphere had the greater effect on pathogen numbers. The greatest reductions in numbers were observed during storage at 

21°C, when numbers decreased to about 2 and 3.8 log 10 cfu/g within 14 days in product stored under air and vacuum, 

respectively, and a 5 log 111 reduction was observed for both atmospheres within 28 days. Regardless of the storage 

atmosphere, numbers did not decrease below 3.6 or 3.7 log, 0 cfu/g after 90 days of storage at - 20 or 4°C, respectively. 

These data confirm that fermentation and drying are sufficient to eliminate only about 2 log, 0 cfu/g of E.coli O157:H7 from 

fermented sausage, and that additional strategies, such as storage for at least 2 weeks at ambient temperature in air, are 

required to achieve a 5 to 6 log, 0 reduction in the numbers of the pathogen in sliced pepperoni. © 1997 Elsevier Science 

B.Y. 

Keywords: Escherichia coli O157:H7; Pepperoni; Pathogen; Fermentation; Survival 

1. Introduction 

There has been much concern over the association

of Escherichia coli 0157:H7 with meats, and considerable research initiated to determine how to control 
48 N.G. Faith et al. I lntemutional Jo11mul of" Food Microhiologv 37 ( /997) 47-54 

the pathogen in fermented meat products. Shortly 

after an outbreak of E. coli O 157: H7 infection which 

was linked to a dry, fermented pork/beef salami 

( Centers for Disease Control, 1995a), several laboratories initiated studies on the viability of this 

pathogen in a variety of fermented meats, to develop 

manufacturing processes that would reliably reduce 

pathogen numbers by 5 log 10 units. The impetus for 

these validation-type studies was the regulatory 

mandate from the United States Department of 

Agriculture (USDA) Food Safety and Inspection 

Service (FSIS) that manufacturers of dry and semidry fermented sausage demonstrate a 5-log 10 unit 

reduction of £. coli O157:H7 during processing 

(Reed, 1995). With the possible exception of beef 

jerky (Harrison and Harrison, 1996), the few reports 

published to date have revealed that fermentation and 

drying are only sufficient to effect a reduction of 

about 2 log 10 units in the numbers of this pathogen 

during the manufacture of pepperoni (Hinkens et al.. 

1996), salami (Clavero and Beuchat. 1996; Glass et 

al., 1992; Nickelson et al., 1996 ). or summer sausage 

( Calicioglu et al., 1997 ). Thus. efforts have intensified to identify additional intervention strategies to 

obtain greater reductions of the numbers of this 

pathogen in fermented meats. 

As reported by Hinkens et al. (1996 ), heating 

pepperoni sticks, after fermentation at 36°C to :S pH 

5 and drying, to an internal temperature of 63°C 

instantaneous or to 53°C for 60 min. resulted in a 

2 5-log 10 unit decrease in numbers without visibly 

affecting the texture or appearance of the product. 

Similarly, fermentation at 41°C to pH 4.6 or pH 5 

and post-fermentation heating of sticks to an internal 

temperature of 54°C resulted in a > 5 log 1 0 decrease 

in counts of £. coli O I 57:H7 in summer sausage 

(Calicioglu et al., 1997). As another example, extended storage or fermentation of sausage sticks at 

32°C to pH 4.6 reduced counts of the pathogen by at 

least 4 log 10 cfu/g (Nickelson et al., 1996). Aside 

from the application of heat, relatively little else has 

been reported on other intervention strategies to 

reduce numbers of this pathogen in fermented meats. 

There are several categories and varieties of allbeef or beef/pork (55%/45%) fermented sausage 

that have a moisture/protein (M/Pr) ratio of 1.6: I 

and, as such. are called pepperoni (United States 

Department of Agriculture, Food Safety and Inspection Service, 1986). With about 115 000 000 kg used 

annually by pizzerias in the USA alone and with 

about 57% of customers for pizzas in a 2-store 

survey selecting pepperoni as their favorite topping, 

pepperoni is arguably the most popular variety of 

pizza topping in the United States (A. Willman, 

personal communication). About 90% of the pepperoni destined for use as a topping on frozen pizza 

is sold in stick form and is typically stored refrigerated under air or CO 2 for up to 30 days, the 

remainder being pre-sliced and typically stored refrigerated or frozen for up to 30 or 90 days, 

respectively (T. Lorang, unpublished data). Pepperoni pizza topping targeted for use by pizzerias is 

sliced in bulk and stored refrigerated or frozen with a 

shelf life of 30 or 90 days, respectively (T. Lorang. 

unpublished data). In a previous study (Hinkens et 

al., 1996), we reported that fermentation/drying of 

pepperoni sticks reduced the numbers of the pathogen by about 2 log 10 units. The present study 

expanded upon our previous research with pepperoni 

( Hinkens et al., 1996) and monitored the effect of 

different storage conditions on the fate of £. coli 

O157:H7 in pepperoni slices intended for use as 

pizza topping. 

2. Materials and methods 

2. /. Bacterial strains. 

Five strains of £. coli O157:H7 were used to 

inoculate pepperoni. The strains were EC5058, a 

beef isolate from the University of Wisconsin Food 

Research Institute; C7927, a human isolate from the 

Massachusetts apple cider outbreak of 1991 ( Besser 

et al.. 1993 ); F-90, a sausage isolate from the 

Washington/California dry-cured salami outbreak of 

1994 (Centers for Disease Control, 1995a); EC204P. 

a pork isolate from the University of Wisconsin Food 

Research Institute; and C9490, a human isolate from 

the Western States hamburger patty outbreak of 1993 

(Centers for Disease Control, 1993 ). The E. coli 

strains were maintained as recommended by the 

USDA/FSIS (Reed, 1995). A commercial Pediococrns acidilactici starter culture (Lactacel 115; Quest 

International. Sarasota. FL) was maintained and 

propagated according to the manufacturer's instructions. 
N.G. Faith et al. I International Journal of Food Microbiology 37 ( 1997) 47-54 49 

2.2. Preparation of E. coli O 157:H7 and P. 

acidilactici inocula. 

Each of the five strains of E. coli O157:H7 was 

grown separately in 250 ml of trypticase soy broth 

(TSB; Difeo Laboratories Inc., Detroit, MI) supplemented with 1% glucose, at 37°C, overnight, with 

shaking. The five cell suspensions were harvested 

and combined to achieve a final inoculum of about 

I X l O9 cfu / ml containing equal numbers of the 

strains, as previously described (Hinkens et al., 

1996). 6 ml of the thawed pediococcal starter culture 

were added to 42 ml of sterile dH 2O for addition to 

11.33 kg of batter. 

2.3. Manufacture of pepperoni. 

A flow diagram for pepperoni manufacture and 

storage is provided in Fig. 1. The raw meat supplied 

by a commercial manufacturer was maintained at 

about 4°C during batter preparation which was 

conducted essentially as described (Hinkens et al., 

1996). Raw batters were prepared in 46 kg batches, 

which were composed of 75% pork and 25% beef 

with a fat content of about 32% and contained 0.63o/c 

dextrose (A. E. Staley, Decatur, IL), 3% spice mix 

(Doskocil house blend; Doskocil Company, Inc., 

Jefferson, WI), and 2% cure mixture (Doskocil house 

Fermentation 

(at 36°C, 90% RH to pH 4.8) 

I 

Drying 

(for 18 days at 13"C, 65% RH) 

I 

Slicing 

I 

Packaging/Storage 

21°C 4"C -20°C 

I 

Air Vacuum Air Vacuum CO2 

Fig. I. Pepperoni manufacture. Flow diagram depicting the 

pepperoni processes evaluated. 

blend). The batter was inoculated with the cocktail of 

E. coli O157:H7 to obtain numbers of ::::- 2 X 10 7 

cfu/ g of batter, and with tke thawed pediococcal 

starter culture to achieve about 10 8 cfu / g of batter. A 

control batter without E. coli O 157:H7 was similarly 

prepared. The batter was ground using a commercial 

grinder (model 84142; Hobart Manufacturing Co., 

Troy, OH) through a 1/8" plate and then stuffed 

using a hand stuffer (Koch Supplies, Inc., Kansas 

City, MO) into 55 mm fibrous casings (TeePak, Inc., 

Westchester, IL), to a final length of 4 7 cm to give 

sticks which each weighed about 700 g. The sticks 

were transferred to a smokehouse (model 1000; 

Vortron, Inc., Beloit, WI) and fermentation was 

conducted as previously described (Hinkens et al., 

1996) to a final pH of <pH 4.8, which was 

achieved after 16 to 20 h. 

2.4. Drying, slicing and storage of pepperoni. 

The sticks were dried at l 3°C and 65% RH for 

about 18 days to a M/ Pr of :S 1.6: I in an environmentally controlled chamber (Biotron facility; 

University of Wisconsin, Madison, WI). The dried 

sticks were cut into slices of about 1.9 g each using a 

Globe slicing machine (Model 500; Stamford, CT). 

Portions, each of 55 slices, were placed into clear, 

polyethylene bags (Curion® Grade 863; 18 X 23 cm; 

0 2 < 1.0 CC per 100 inch2 per 24 hat 23°C and 9% 

RH with MVTR 0.5 gm H2O per 100 inch2 per 24 h 

at 38°C and 90% RH; Curwood Inc., Oshkosh, WI) 

for packaging under air, CO 2 ( :S I% residual 0 2 ), or 

vacuum. The latter two atmospheres were generated 

using a Multivac vacuum-packaging machine (Sepp 

Haggenmiiller KG, Germany), and the percent residual oxygen present was determined on representative bags packaged under CO 2 using a headspace 

analyzer (Model HS-750: Modem Controls, Inc., 

Minneapolis, MN). For slices stored under air, the 

bags were sealed about 0.5 cm from the top using an 

impulse heat sealer (model TISH-300; Electric Heating Equipment Co., Ltd., Taiwan, R.O.C.) to achieve 

an unoccupied volume of about 650 ml. Packages 

were stored at - 20, 4, or 21 °C for 1, 4, 7, 14, 28, 

60, or 90 days. At each sampling interval, three 

packages were removed from storage and a 25 g 

portion of pepperoni from each package was analyzed microbiologically. 
50 N.G. Faith et al. I lmemotiona/ Jo11ma/ of' Food Microhio/ogv 37 ( /997) 47-54 

2.5. Microbiological analyses of pepperoni hatters, 

sticks, and slices. 

Microbiological testing was conducted for each of 

three trials using triplicate samples of meat and 

duplicate platings of each dilution. Batter, sticks, or 

slices were tested for viable E. coli O157:H7 by 

direct plating of macerated and diluted meat samples 

prior to stuffing, after fermentation, after drying/ 

slicing. and after storage at different temperatures 

and atmospheres. At each sampling interval, three 

25-g portions of the batter, or a 25-g cross section 

from the middle of each of three sticks, or a 25-g 

composite of slices were aseptically transferred to 

Stomacher bags (Seward Medical, London, UK) 

containing 225 ml of 0.1 % peptone and processed 

and plated onto MacConkey sorbitol agar (MSA; 

Difeo) as described previously (Hinkens el al., 

1996 ). When numbers of the pathogen decreased 

below 10 1 cfu / g, the presence/ absence of the pathogen was determined by enrichment as previously 

described ( Hinkens et al., 1996). The raw meat was 

tested for background levels of£. coli O157:H7 and 

other non-sorbitol-fermenting bacteria by spread 

plating macerated and diluted meat samples onto 

MSA plates and for total aerobic bacterial numbers 

by spread plating macerated and diluted meat samples onto trypticase soy agar (TSA; Difeo) plates. 

Immediately after stuffing. the sticks were also tested 

for viable pediococci and lactic acid bacteria (LAB) 

by spread plating macerated and diluted meat samples onto MRS (Difeo) agar plates. Plates were 

incubated at 42°C for 24 h to recover E. coli 

O157:H7 and at 37°C for 48 h to recover pediococci/ 

LAB. Representative isolates of each type were 

confirmed as previously described (Hinkens et al., 

1996). 

2.6. Chemical analyses of pepperoni sticks and 

organoleptic observations of slices. 

At each sampling time (before stuffing, after 

fermentation, after drying, and after storage), three 

control sticks were removed for chemical analyses. 

The control sticks were either transported on ice 

directly to a commercial testing laboratory, or were 

held at ~ 20°C for up to 7 days and then delivered 

for testing. Chemical analyses were performed on a 

composite from each of these three sticks following 

Association of Official Analytical Chemists (AOAC) 

procedures. Each composite was tested for fat 

(AOAC procedure 960.39), moisture (AOAC procedure 950.46 ), protein (AOAC procedure 928.06 ), 

and salt (AOAC procedure 935.47) as reported for 

meat products by McNeal ( 1990). The water activity 

(a") was determined using a Rotronic water activity 

meter (model DT; Huntington, NY). The pH and 

titratable acidity (TA) were determined as previously 

described (Luchansky et al., 1992). The TA was 

expressed as the percent lactic acid. Pepperoni slices 

were also evaluated for any changes in color or 

texture, or for the appearance of yeast/ molds, by 

visual inspection and palpation of slices. 

2. 7. Statistirnl analyses. 

Data were analyzed using version 5 of the Statistical Analysis System user's guide (SAS Institute, 

Cary. NC). 

3. Results 

3.1. Testing of raw meat. 

Analyses of the raw meat before inoculation with 

the serotype O 157: H7 cocktail and pediococcal 

starter culture revealed that none of the raw meat 

tested contained indigenous E. coli O 157: H7 by 

direct plating (data not shown). The raw meat 

produced total aerobic plate counts ranging from 

1.8 1 X I 0 to 5 X I 0° cfu/g of meat (average = 

1.7 X 10° cfu/g). The pediococci/LAB count of the 

raw meat ranged from 3 2 X 10 to 2.7 X 101 cfu/g of 

meat (average = 1.8 X 101 cfu/g). The pediococci/ 

LAB count after the addition of the starter culture to 

the batter ranged from 5 X 107 to 1.2 X IO 8 cfu / g of 

meat (average = 7.8 10 7 X cfu/g). The proximate 

composition of the pepperoni also displayed the 

expected levels for pH, a", salt, protein, moisture. 

and fat (Table I). 

3.2. Microbiological testing of pepperoni sticks 

and slices during manufacture and storage. 

The raw batter contained 7.8±0.52 log 10 cfu/g of 

E.coli O157:H7. Thereafter, counts of the pathogen 

decreased to 6.8 ±0.40 log 10 cfu/ g after fermentation 
N.G. Faith et al. I International Journal of Food Microbiology 37 ( 1997) 47-54 51 

Table 1 

Chemical composition of pepperoni during manufacture and storage. 

Manufacturing step Chemical Analysis (value ::':: s.d; n = 3) 

pH TA(%)" M/Pr" a. Salt(%) Moisture(%) Protein(%) Fat(%) 

Batter 5.91 ::'::0.05 0.35::t::0.04 3.60::t::0.09 0.96::t::O.0O 3.28::t::0.06 47.93::t::0.80 13.32::t::0.14 33.29::t::0.25 

After fermentation 4.82::t::0.0l 0.60::t::0.06 3.54::t::0.08 0.96::'::0.00 3.24::t::0.09 46.80::':: 1.59 13.26::t::0.69 35.09::t:: 1.68 

After drying/slicing ' 4.77::t::0.0l 0.78::t::0.09 1.63 ::t::0.33 0.90::t::0.03 3.97::t::0.70 29.03::t::4.03 17.89::t:: 1.04 46.23::t::2.49 

d After storage 

(Temp., Atmos.) 

21°C Air 5.10 0.89 1.41 .883 4.46 27.65 19.56 48.04 

21°c Vacuum 4.76 0.95 1.40 .881 4.43 26.77 19.06 48.83 

4°C Air 4.77 0.96 1.43 .885 4.55 27.66 19.34 48.19 

4°C Vacuum 4.78 0.96 1.43 .882 4.46 27.11 18.98 48.88 

4°c co. 4.68 0.99 1.48 .881 4.35 27.83 18.83 48.06 

- 20°c co, 4.67 0.92 1.48 .885 4.62 27.61 18.65 46.78 

- 20°C Air 4.70 1.38 1.89 .900 4.12 31.82 16.86 45.13 

·' TA, titratable acidity as % lactic acid. 1

' M /Pr, moisture to protein ratio. 

' values are the averages of 3 trials. 

" values for 14 days of storage from trial # 2. 

Table 2 

E. coli O I 57:H7 counts in pepperoni slices during storage at different temperatures under different atmospheres. 

Temperature Atmosphere Time (days) of storage (log, 11 cfu g :1:: s.d.: 11 3) 

4 7 14 28 60 90 

21°c Air 5.51 :1::0.56 4.71 :1::0.67 (a)' 3.90:1::0.96 (a) < 1.88:1:: 1.55 (a) < 1.0:1::0.0 (a)" N.T.' N.T. 

Vacuum 5.59:1::0.67 4.93:1::0.63 (a) 4.39:1::0.78 (a) 3.77:1::0.88 (b) 1.90:1::0.51 la) < 1.0 (al"" < 1.0 (a)"" 

4°c Air 5.85:1::0.60 5.75:1::0.80 (b) 5.63:1:: 1.07 (b) 5.12:1:: 1.06 (c) 4.76:1::0.79 (b) 4.40:1::0.45 (bJ 3.71 :1:: 1.61 (b) 

Vacuum 5.95:1::0.76 6.20:1:: 1.01 (b) 5.56:1:: 1.07 (b) 5.29:1:: 1.03 (c) 5.07:1:: 1.12 (b) 4.77:1::0.78 (b) 4.29:1:: 1.26 (b) 

co, 5.83:1::0.81 5.87:1::0.96 (b) 5.54:1::0.74 (b) 5.81 :1::0.87 (c) 5.32:1::0.90 (b) 4.65::0.81 (bJ 4.13:1::l.46(b) 

-20°C Air'· 5.23:1::0.36 5.02:1::0.08 (a) 4.38:1::0. 17 (a) 4.41:1::0.31 (b) 3.43:1:: 1.04 (c) 4.13:1::0.71 (b) 3.58::0.64 (b) 

co, 5.08:1::0.65 4. 91 :1::0.85 (a) 4.16:1::0.14 (a) 4.45:1::0.96 (b) 3.74:1::0.71 (c) 3.91 :1::0.34 (b) 3.91 :1:: 1.90 (b) 

" Values in a column that are not followed by the same letter are significantly different (P -s 0.05). 

"Positive for£. coli O157:H7 by enrichment. 

'N.T. = not tested due to mold contamination. 

"Values from trial #3. 

' Values from trials #2 and #3. 

and to about 5. 9 :±: 1.10 log 10 cfu I g after drying. 

Within 14 days counts of the pathogen were reduced 

by about an additional 2 (vacuum) and 4 (air) log 10 

during storage at 21 °C compared to a 1 to 2 log 10 

decrease at 4°C and about a 1.5 log 10 decrease at 

- 20°C. Whereas pathogen numbers decreased to 

below detection by direct plating within 28 (air) and 

60 (vacuum) days during storage at 21 °C, the greatest 

reductions during storage at 4°C and - 20°C, regardless of atmosphere, were approximately 4 log 10 after 

90 days. However, it was still possible to recover the 

pathogen by enrichment during storage at 21 °C on 

days 28 (air), 60 (vacuum), and 90 (vacuum). Differences among treatments were also observed that 

were attributed to the storage atmosphere, but such 

differences were not as significant as the differences 

observed with storage temperature. In general, storage under air was more detrimental to the serotype 

O157:H7 cocktail than storage under vacuum or 

under CO2 (Table 2). At day 14, numbers of the 

pathogen after storage under air at 21 °C were 

significantly (P < 0.05) lower than observed for 

other treatments at day 14. 

There was no appreciable difference in "the proximate composition of sticks/ slices related to storage 

temperature or atmosphere (Table 1). However, after 

= 
52 N.G. Faith et al. I /11ternatio11al Journal of" Food Microhiologv 37 ( /997) ../7-54 

storage of slices in air at 21 °C for 28 days the 

product changed from bright orange to grayish 

brown and yeast and mold growth was observed on 

slices from several packages (data not shown). 

Storage of slices at lower temperatures and/ or at 

other atmospheres did not result in any visible 

changes in color and did not support mold or yeast 

growth (data not shown). 

4. Discussion 

Escherichia coli O I 57:H7 has caused considerable 

illness and death during the 1990s (Griffin, 1995). 

Notable examples include foodborne outbreaks associated with undercooked hamburger patties from a 

fast food restaurant chain (Centers for Disease 

Control, 1993 ), apple cider prepared using fallen 

apples contaminated with deer feces ( Besser et al., 

1993 ), unpasteurized apple juice or juice mixtures 

containing apple juice (Centers for Disease Control, 

1996), and fermented sausage with an unconfirmed 

source of contamination (Centers for Disease Control, 1995a). There was also an outbreak in Japan 

with an unconfirmed source involving > 9,000 individuals (Anonymous, I 996) and an outbreak in 

Australia from uncooked, semidry fermented sausage 

contaminated with another enterohemorrhagic 

serotype, E. coli O I I I : NM ( Centers for Disease 

Control, 1995b). In hindsight, the relative tolerance 

of E. coli O157:H7 to both acid (Arnold and Kaspar, 

1995: Benjamin and Datta, 1995; Miller and Kaspar, 

1994) and heat (Jackson et al., 1995) may partly 

explain the ability of this pathogen to persist and 

cause illness in certain foods previously regarded as 

low risk. There may also be significant variations in 

the degree of acid and/ or heat tolerance from strainto-strain, and a subpopulation of cells of a given 

strain of O157:H7 may also be more or less resistant 

to heat and/or acid than are the majority of cells 

within the same population (Todd et al., 1993: C. W. 

Kaspar and J. B. Luchansky, unpublished data). 

Largely as a result of the Washington/California 

salami outbreak of I 994 (Centers for Disease Control, 1995a), the USDA/FSIS developed guidelines 

for sausage manufacturers to validate processes to 

ensure a 5 log reduction in counts of E. coli 

O157:H7 in fermented meats (Reed, 1995). 

Several factors in the formulation and preparation 

of pepperoni may affect viability of serotype 

O157:H7 strains of E. coli. Previous studies indicated that indigenous bacteria, the spice/ cure mix, 

and smoking do not appreciably influence numbers 

of E. coli O157:H7 in pepperoni (Hinkens et al., 

1996). Amongst the other factors that could influence 

the viability of serotype O I 57: H7 strains in a 

fermented meat, the combined effects of time, temperature, and pH/ acid type influence the survival of 

the pathogen. For example, extended time at :S pH 

4.6 and/or 2: 32°C as well as extensive drying at 

elevated temperature, such as 60°C for IO h, resulted 

in an appreciable decrease in pathogen numbers 

( Harrison and Harrison, I 996; Nickelson et al., 

1996 ). However, it is unlikely that these parameters 

could be adjusted sufficiently to deliver a 5-D kill of 

E. coli O I 57:H7 in pepperoni. Similar results were 

also observed in synthetic media. For example, £. 

coli O157:H7 was more sensitive to acid conditions 

at abusive than at refrigeration temperatures (AbdulRaouf et al., 1993; Besser et al., 1993: Miller and 

Kaspar, 1994). Although post-fermentation heating 

of pepperoni sticks and/ or manufacture of sticks at 

reduced pH levels delivered the mandated reduction 

of 5 log cfu g I in counts of E. coli O 157: H7 

(Hinkens et al., 1996: Nickelson et al., 1996: 

Calicioglu et al., 1997), some sausage varieties 

cannot be so treated without causing undesirable 

sensory or textural changes in the product and/or 

without substantially increasing processing costs. 

Heating may also cause ·greasing out' and may yield 

a product more prone to cupping when used as a 

pizza topping (S. Seideman, personal communication). If heating regimes appropriate to the product 

cannot be identified, then storage of pepperoni slices 

in air at ambient temperature may provide a practicable safety measure. 

In the present study, pathogen numbers were 

monitored during storage of pepperoni slices at 

different temperatures and under different atmospheres. During storage, the numbers of the pathogen 

were reduced to a greater extent at ambient temperature than at chiller or freezing temperatures. The data 

revealed that storing pepperoni slices at 21 °C under 

air or vacuum were sufficient to deliver a 5 log 

reduction in pathogen numbers. The data also revealed that refrigerated or frozen storage resulted in 

an additional about 1.5 to 2.5 log reduction over the 

about 2 log reduction achieved via fermentation and 
N.G. Faith et al. I International Journal t!{ Food Microhiolorrv 37 ( 1997) 47-54 53 

drying. Although storage at ambient temperature 

resulted in the greatest reduction in pathogen numbers, this treatment produced some undesirable 

changes in product appearance. Studies are underway 

to quantify the effect of cooking pizza containing 

pepperoni slices spiked with E. coli O157:H7 on 

pathogen viability and to evaluate other cost-effective intervention strategies such as food grade (bio)preservatives and both pre- and post-fermentation 

pasteurization which might be used to reduce the 

likelihood of foodborne illness due to E. coli 

O157:H7. 

Acknowledgments 

The technical assistance of Patrick Krakar and 

Ann Roering is gratefully acknowledged. Our appreciation is also extended to Chuck Baum (UWMadison Biotron), Dennis Buege (UW-Madison, 

Muscle Biology Laboratory), Steven Devcich 

(TeePak), Gerry Durnell and Alys Willman (National 

Association of Pizza Operators), Larry Hand (Diversitech), Chuck Kaspar (UW-Madison Food Research Institute), Pat Podratz (Modern Controls), and 

Bev Traver and Steve Seideman (Doskocil) for 

supplies, service, equipment, and/ or consultations. 

We also thank the following individuals for supplying cultures; Jay H. Lewis (Washington State 

Public Health Lab) for strain F-90, and Joy G. Wells 

and Timothy J. Barrett (Centers for Disease Control 

and Prevention) for strain C9490. In addition, we 

greatly appreciate the secretarial assistance of Barbara Cochrane.This work was supported by monies 

from the National Cattlemen's Beef Association and 

contributions to the Food Research Institute from the 

food industry. Portions of this research will be 

presented at the Annual Meeting of the Institute of 

Food Technologists, June 14-18, 1997, in Orlando, 

FL. 

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