High-temperature, short-time pasteurization of milk is ineffective against spore-forming bacteria such as Bacillus anthracis (BA), but is lethal to its vegetative cells. Crossflow microfiltration ...(MF) using ceramic membranes with a pore size of 1.4μm has been shown to reject most microorganisms from skim milk; and, in combination with pasteurization, has been shown to extend its shelf life. The objectives of this study were to evaluate MF for its efficiency in removing spores of the attenuated Sterne strain of BA from milk; to evaluate the combined efficiency of MF using a 0.8-μm ceramic membrane, followed by pasteurization (72°C, 18.6s); and to monitor any residual BA in the permeates when stored at temperatures of 4, 10, and 25°C for up to 28 d. In each trial, 95 L of raw skim milk was inoculated with about 6.5 log₁₀ BA spores/mL of milk. It was then microfiltered in total recycle mode at 50°C using ceramic membranes with pore sizes of either 0.8μm or 1.4μm, at crossflow velocity of 6.2m/s and transmembrane pressure of 127.6kPa, conditions selected to exploit the selectivity of the membrane. Microfiltration using the 0.8-μm membrane removed 5.91±0.05 log₁₀ BA spores/mL of milk and the 1.4-μm membrane removed 4.50±0.35 log₁₀ BA spores/mL of milk. The 0.8-μm membrane showed efficient removal of the native microflora and both membranes showed near complete transmission of the casein proteins. Spore germination was evident in the permeates obtained at 10, 30, and 120min of MF time (0.8-μm membrane) but when stored at 4 or 10°C, spore levels were decreased to below detection levels (≤0.3 log₁₀ spores/mL) by d 7 or 3 of storage, respectively. Permeates stored at 25°C showed coagulation and were not evaluated further. Pasteurization of the permeate samples immediately after MF resulted in additional spore germination that was related to the length of MF time. Pasteurized permeates obtained at 10min of MF and stored at 4 or 10°C showed no growth of BA by d 7 and 3, respectively. Pasteurization of permeates obtained at 30 and 120min of MF resulted in spore germination of up to 2.42 log₁₀ BA spores/mL. Spore levels decreased over the length of the storage period at 4 or 10°C for the samples obtained at 30min of MF but not for the samples obtained at 120min of MF. This study confirms that MF using a 0.8-μm membrane before high-temperature, short-time pasteurization may improve the safety and quality of the fluid milk supply; however, the duration of MF should be limited to prevent spore germination following pasteurization.
For each of two trials, freshly ground beef of variable fat content (higher: 70:30 %lean:%fat; lower: 93:7 %lean:%fat) was separately inoculated with ca. 7.0 log CFU/g of a single strain of ...Escherichia coli serotypes O26:H11, O45:H2, O103:H2, O104:H4, O111:H-, O121:H19, O145:NM, and O157:H7. Next, ca. 3-g samples of inoculated beef were transferred into sterile filter bags and then flattened (ca. 1.0 mm thick) and vacuum sealed. For each temperature and sampling time, three bags of the inoculated wafers of beef were submerged in a thermostatically controlled water bath and heated to an internal temperature of 54.4°C (130°F) for up to 90 min, to 60°C (140°F) for up to 4 min, or to 65.6°C (150°F) for up to 0.26 min. In lower fat wafers, D-values ranged from 13.5 to 23.6 min, 0.6 to 1.2 min, and 0.05 to 0.08 min at 54.4, 60.0, and 65.6°C, respectively. Heating higher fat wafers to 54.4, 60.0, and 65.6°C generated D-values of 18.7 to 32.6, 0.7 to 1.1, and 0.05 to 0.2 min, respectively. In addition, we observed reductions of ca. 0.7 to 6.7 log CFU/g at 54.4°C after 90 min, ca. 1.1 to 6.1 log CFU/g at 60.0°C after 4 min, and 0.8 to 5.8 log CFU/g at 65.6°C after 0.26 min. Thus, cooking times and temperatures effective for inactivating a serotype O157:H7 strain of E. coli in ground beef were equally effective against the seven non-O157:H7 Shiga toxin–producing strains investigated herein.
Viability of
Listeria
monocytogenes was monitored on frankfurters formulated with or without potassium lactate and sodium diacetate at a ratio of ca. 7:1 and treated with lauric arginate (LAE; 22 or ...44
ppm) using the Sprayed Lethality in Container (SLIC®) delivery method. Without antimicrobials, pathogen numbers remained relatively constant at ca. 3.3
log
CFU/package for ca. 30
d, but then increased to ca. 8.4
log
CFU/package over 120
d. Regardless of whether or not lactate and diacetate were included, when treated with LAE, pathogen numbers decreased from ca. 3.3
log
CFU/package to ca. 1.5
log
CFU/package within 2
h, but then increased to 7.3 and 6.7
log
CFU/package, respectively, after 120
d. When frankfurters were formulated with lactate and diacetate and treated with LAE, pathogen numbers decreased by ca. 2.0
log
CFU/package within 2
h and remained relatively unchanged over the 120
d. These data confirm that LAE provides an initial lethality towards
L. monocytogenes and when used in combination with reduced levels/ratio of lactate and diacetate as an ingredient for frankfurters provides inhibition throughout shelf life.
In phase I, beef subprimals were inoculated on the lean side with ca. 0.5 to 3.5 log CFU/g of a rifampin-resistant (rif(r)) cocktail of Escherichia coli O157:H7 and passed once, lean side up, through ...a mechanical blade tenderizer. Inoculated subprimals that were not tenderized served as controls. Ten core samples were removed from each subprimal and cut into six consecutive segments: segments 1 to 4 comprised the top 4 cm and segments 5 and 6 the deepest 4 cm. Levels of E. coli O157:H7 recovered from segment 1 of control subprimals when inoculated with ca. 0.5, 1.5, 2.5, or 3.5 log CFU/g were 0.6, 1.46, 2.5, and 3.19 log CFU/g, respectively. Following tenderization, pathogen levels recovered from segment 1 inoculated with 0.5 to 3.5 log CFU/g were 0.22, 1.06, 2.04, and 2.7 log CFU/g, respectively. Levels recovered in segment 2 were 7- to 34-fold lower than levels recovered from segment 1. Next, in phase II, the translocation of ca. 4 log CFU of the pathogen per g was assessed for lean-side-inoculated subprimals passed either once (LS) or twice (LD) through the tenderizer and for fatside-inoculated subprimals passed either once (FS) or twice (FD) through the tenderizer. Levels in segment 1 for LS, LD, FS, and FD tenderized subprimals were 3.63, 3.52, 2.85, and 3.55 log CFU/g, respectively. The levels recovered in segment 2 were 14- to 50-fold lower than levels recovered in segment 1 for LS, LD, FS, and FD subprimals. Thus, blade tenderization transfers E. coli O157:H7 primarily into the topmost 1 cm, but also into the deeper tissues of beef subprimals.
The effect of high-hydrostatic-pressure processing (HPP) on the survival of a 5-strain rifampicin-resistant cocktail of Listeria monocytogenes in Queso Fresco (QF) was evaluated as a postpackaging ...intervention. Queso Fresco was made using pasteurized, homogenized milk, and was starter-free and not pressed. In phase 1, QF slices (12.7×7.6×1cm), weighing from 52 to 66g, were surface inoculated with L. monocytogenes (ca. 5.0 log10 cfu/g) and individually double vacuum packaged. The slices were then warmed to either 20 or 40°C and HPP treated at 200, 400, and 600MPa for hold times of 5, 10, 15, or 20min. Treatment at 600MPa was most effective in reducing L. monocytogenes to below the detection level of 0.91 log10 cfu/g at all hold times and temperatures. High-hydrostatic-pressure processing at 40°C, 400MPa, and hold time ≥15min was effective but resulted in wheying-off and textural changes. In phase 2, L. monocytogenes was inoculated either on the slices (ca. 5.0 log10 cfu/g; ON) or in the curds (ca. 7.0 log10 cfu/g; IN) before the cheese block was formed and sliced. The slices were treated at 20°C and 600MPa at hold times of 3, 10, and 20min, and then stored at 4 and 10°C for 60d. For both treatments, L. monocytogenes became less resistant to pressure as hold time increased, with greater percentages of injured cells at 3 and 10min than at 20min, at which the lethality of the process increased. For the IN treatment, with hold times of 3 and 10min, growth of L. monocytogenes increased the first week of storage, but was delayed for 1 wk, with a hold time of 20min. Longer lag times in growth of L. monocytogenes during storage at 4°C were observed for the ON treatment at hold times of 10 and 20min, indicating that the IN treatment may have provided a more protective environment with less injury to the cells than the ON treatment. Similarly, HPP treatment for 10min followed by storage at 4°C was the best method for suppressing the growth of the endogenous microflora with bacterial counts remaining below the level of detection for 2 out of the 3 QF samples for up to 84d. Lag times in growth were not observed during storage of QF at 10°C. Although HPP reduced L. monocytogenes immediately after processing, a second preservation technique is necessary to control growth of L. monocytogenes during cold storage. However, the results also showed that HPP would be effective for slowing the growth of microorganisms that can shorten the shelf life of QF.
The fate of
Listeria monocytogenes,
Salmonella typhimurium, or
Escherichia coli O157:H7 were separately monitored both in and on soudjouk. Fermentation and drying alone reduced numbers of
L. ...monocytogenes by 0.07 and 0.74
log
10
CFU/g for sausages fermented to pH 5.3 and 4.8, respectively, whereas numbers of
S. typhimurium and
E. coli O157:H7 were reduced by 1.52 and 3.51
log
10
CFU/g and 0.03 and 1.11
log
10
CFU/g, respectively. When sausages fermented to pH 5.3 or 4.8 were stored at 4, 10, or 21
°C, numbers of
L. monocytogenes,
S. typhimurium, and
E. coli O157:H7 decreased by an additional 0.08–1.80, 0.88–3.74, and 0.68–3.17
log
10
CFU/g, respectively, within 30 days. Storage for 90 days of commercially manufactured soudjouk that was sliced and then surface inoculated with
L. monocytogenes,
S. typhimurium, and
E. coli O157:H7 generated average
D-values of ca. 10.1, 7.6, and 5.9 days at 4
°C; 6.4, 4.3, and 2.9 days at 10
°C; 1.4, 0.9, and 1.6 days at 21
°C; and 0.9, 1.4, and 0.25 days at 30
°C. Overall, fermentation to pH 4.8 and storage at 21
°C was the most effective treatment for reducing numbers of
L. monocytogenes (2.54
log
10
CFU/g reduction),
S. typhimurium (⩾5.23
log
10
CFU/g reduction), and
E. coli O157:H7 (3.48
log
10
CFU/g reduction). In summary, soudjouk-style sausage does not provide a favorable environment for outgrowth/survival of these three pathogens.
We demonstrated the effectiveness of delivering an antimicrobial purge/fluid into shrink-wrap bags immediately prior to introducing the product and vacuum sealing, namely the “Sprayed Lethality In ...Container” (SLIC™) intervention delivery method. The pathogen was
Listeria monocytogenes, the antimicrobials were acidic calcium sulfate (ACS; calcium sulfate plus lactic acid; 1:1 or 1:2 in dH
2O) and lauric arginate (LAE; Ethyl-
N-dodecanoyl-
l-arginate hydrochloride; 5% or 10% in dH
2O), and the product was commercially prepared “table brown” ham (ca. 3 pounds each). Hams were surface inoculated with a five-strain cocktail of
L. monocytogenes (ca. 7.0 log
10 CFU per ham), added to shrink-wrap bags that already contained ACS or LAE, vacuum-sealed, and stored at 4
°C for 24
h. Pathogen levels decreased by 1.2, 1.6, 2.4, and 3.1 log
10 CFU/ham and 0.7, 1.6, 2.2, and 2.6 log
10 CFU/ham in samples treated with 2, 4, 6, and 8
mL of a 1:1 and 1:2 solution of ACS, respectively. In samples treated with 2, 4, 6, and 8
mL of a 5% solution of LAE, pathogen levels decreased by 3.3, 6.5, 5.6, and 6.5 log
10 CFU/ham, whereas when treated with a 10% solution of LAE pathogen levels decreased ca. 6.5 log
10 CFU/ham for all application volumes tested. The efficacy of ACS and LAE were further evaluated in shelf-life studies wherein hams were surface inoculated with either ca. 3.0 or 7.0 log
10 CFU of
L. monocytogenes, added to shrink-wrap bags that contained 0, 4, 6, or 8
mL of either a 1:2 solution of ACS or a 5% solution of LAE, vacuum-sealed, and stored at 4
°C for 60 days. For hams inoculated with 7.0 log
10 CFU,
L. monocytogenes levels decreased by ca.1.2, 1.5, and 2.0 log
10 CFU/ham and 5.1, 5.4, and 5.5 log
10 CFU/ham within 24
h at 4
°C in samples treated with 4, 6, and 8
mL of a 1:2 solution of ACS and a 5% solution of LAE, respectively, compared to control hams that were not treated with either antimicrobial. Thereafter, pathogen levels remained relatively unchanged (±1.0 log
10 CFU/ham ) after 60 days at 4
°C in hams treated with 4, 6, and 8
mL of a 1:2 solution of ACS and increased by ca. 2.0–5.0 log
10 CFU/ham in samples treated with 4, 6, and 8
mL of a 5% solution of LAE. For hams inoculated with 3.0 log
10 CFU,
L. monocytogenes levels decreased by 1.3, 1.9, and 1.8 log
10 CFU/ham within 24
h at 4
°C in samples treated with 4, 6, and 8
mL of a 1:2 solution of ACS, respectively, compared to control hams that were not treated. Likewise, levels of the pathogen were reduced to below the limit of detection (i.e., 1.48 log
10 CFU/ham) in the presence of 4, 6, and 8
mL of a 5% solution of LAE within 24
h at 4
°C. After 60 days at 4
°C, pathogen levels remained relatively unchanged (±0.3 log
10 CFU/ham) in hams treated with 4, 6, and 8
mL of a 1:2 solution of ACS. However, levels of
L. monocytogenes increased by ca. 2.0 log
10 CFU/ham in samples treated with 4 and 6
mL of a 5% LAE solution within 60 days but remained below the detection limit on samples treated with 8
mL of this antimicrobial. These data confirmed that application via SLIC™ of both ACS and LAE, at the concentrations and volumes used in this study, appreciably reduced levels of
L. monocytogenes on the surface of hams within 24
h at 4
°C and showed potential for controlling outgrowth of the pathogen over 60 days of refrigerated storage.
A survey was conducted between March and October of 1994 to determine the prevalence and identify the sources of serotype O157:H7 isolates of Escherichia coli in Wisconsin dairy herds. A stratified ...sample of 400 farms was identified, and 70 farms with weaned calves less than 4 months old were included in the study During the prevalence study, 5 of the 70 farms (herd prevalence, 7.1 +/- 4.5%) and fecal samples from 10 of 560 calves (animal prevalence, 1.8%) tested positive for serotype O157:H7. In a follow-up study, the five O157:H7-positive farms and seven of the O157:H7-negative farms identified in the prevalence study were visited again. An additional 517 fecal samples from cattle of various ages were tested, and a total of 15 animals from four of the five herds that were previously positive and 4 animals from two of seven herds that were previously negative tested positive for E. coli O157:H7. Observations made during the follow-up study suggested that horizontal transmission was an important means of E. coli O157:H7 dissemination on the farms. A total of 302 environmental samples, were examined, and 2 animal drinking water samples from one previously negative farm and 1 animal drinking water sample from a previously positive farm contained E. coli O157:H7. Analyses by the pulsed-field gel electrophoresis technique of contour-clamped homogeneous electric field electrophoresis revealed that isolates from the same farm displayed identical or very similar XbaI restriction endonuclease digestion profiles (REDP), whereas isolates from different farms typically displayed different REDP. However, more than one REDP was usually observed for a given herd over the 8-month sampling period. Analyses of multiple isolates from an animal revealed that some animals harbored O157:H7 strains that had different REDP, although the REDP of isolates obtained from the same fecal sample were very similar
In the first part of this study, samples were collected from farms, cheese processing plants (CPPs), and retail markets located in various geographical areas of Sonora, Mexico, over a 12-month period ...during the summer of 2004 and winter of 2005. Four (all Queso Fresco QF from retail markets) of 349 total samples tested positive for Listeria monocytogenes (Lm). Of these four positive samples, three were collected in the northern region and one in the southern region of Sonora. Additionally, two were collected during the winter months, and two were collected during the summer months. For the second part of the study, a total of 39 samples from a farm, a CPP, and retail markets were collected and processed according to a combination of the Norma Oficial Mexicana NOM-143-SSA1-1995.10 method (NOM) and the U.S. Food and Drug Administration (FDA) Bacteriological Analytical Manual method, and 27 samples from these same locations were collected and processed according to the U.S. Department of Agriculture Food Safety and Inspection Service method (USDA-FSIS). The NOM-FDA method recovered the pathogen from 6 (15%) of 39 samples (one cheese and five product contact surfaces), while the USDA-FSIS method recovered the pathogen from 5 (18.5%) of 27 samples (all product contact surfaces). In addition, the 40 isolates recovered from the 15 total samples that tested positive for Lm grouped into five distinct pulsotypes that were ca. 60% related, as determined by pulsed-field gel electrophoresis analysis. The results of this study confirmed a 3.4% prevalence of Lm in QF collected from retail markets located in Sonora and no appreciable difference in the effectiveness of either the NOM-FDA or USDA-FSIS method to recover the pathogen from cheese or environmental samples.
Escherichia coli O157:H7 was recovered from colon fecal samples of pigs. Polymerase chain reaction confirmed two genotypes: isolates harboring the eaeA, stx1, and stx2 genes and isolates harboring ...the eaeA, stx1, and hly933 genes. We demonstrate that swine in the United States can harbor potentially pathogenic E. coli O157:H7.
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