Validation of Pepperoni Process for Control of Shiga Toxin–Producing Escherichia coli

2012 ◽  
Vol 75 (5) ◽  
pp. 838-846 ◽  
Author(s):  
KATHLEEN A. GLASS ◽  
CHARLES W. KASPAR ◽  
JEFFREY J. SINDELAR ◽  
ANDREW L. MILKOWSKI ◽  
BRIAN M. LOTZ ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Limit Of Detection ◽  
The Other ◽  
Direct Plating ◽  
E Coli ◽  
Log Reduction ◽  
Three Samples ◽  
Processing Steps

The objective of this study was to compare the survival of non-O157 Shiga toxin–producing Escherichia coli (STEC) with E. coli O157:H7 during pepperoni production. Pepperoni batter was inoculated with 7 log CFU/g of a seven-strain STEC mixture, including strains of serotypes O26, O45, O103, O111, O121, O145, and O157. Sausages were fermented to pH ≤4.8, heated at 53.3°C for 1 h, and dried for up to 20 days. STEC strains were enumerated at designated intervals on sorbitol MacConkey (SMAC) and Rainbow (RA) agars; enrichments were completed in modified EC (mEC) broth and nonselective tryptic soy broth (TSB). When plated on SMAC, total E. coli populations decreased 2.6 to 3.5 log after the 1-h heating step at 53.3°C, and a 4.9- to 5-log reduction was observed after 7 days of drying. RA was more sensitive in recovering survivors; log reductions on it were 1.9 to 2.6, 3.8 to 4.2, and 4.6 to 5.3 at the end of cook, and at day 7 and day 14 of drying, respectively. When numbers were less than the limit of detection by direct plating on days 14 and 20 of drying (representing a 5-log kill), no more than one of three samples in each experiment was positive by enrichment with mEC broth; however, STEC strains were recovered in TSB enrichment. Freezing the 7-day dried sausage for 2 to 3 weeks generated an additional 1- to 1.5-log kill. Confirmation by PCR revealed that O103 and O157 had the greatest survival during pepperoni productions, but all serotypes except O111 and O121 were occasionally recovered during drying. This study suggests that non-O157 STEC strains have comparable or less ability than E. coli O157 to survive the processing steps involved in the manufacture of pepperoni. Processes suitable for control of E. coli O157 will similarly inactivate the other STEC strains tested in this study.

Method for the Detection of Priority Shiga Toxin–Producing Escherichia coli in Beef Trim

2013 ◽  
Vol 76 (10) ◽  
pp. 1689-1696 ◽  
Author(s):  
GEORGE HUSZCZYNSKI ◽  
MARTINE GAUTHIER ◽  
SAM MOHAJER ◽  
ALEXANDER GILL ◽  
BURTON BLAIS
Keyword(s):  
Escherichia Coli ◽  
Multiplex Pcr ◽  
Shiga Toxin ◽  
Limit Of Detection ◽  
Turnaround Time ◽  
Direct Plating ◽  
E Coli ◽  
Agar Media ◽  
Pcr Techniques ◽  
Positive Results

A method has been developed for the detection in beef trim of priority Shiga toxin–producing E. coli (STEC) strains, defined as E. coli possessing the virulence factors stx1 and/or stx2 and intimin (eae), with O serogroups O26, O45, O103, O111, O121, O145, or O157. The method is based on recovery of the target bacteria by overnight enrichment in a broth optimized for recovery of O157 and non-O157 STEC, followed by screening using multiplex PCR techniques targeting (i) stx1, stx2, and eae (STE PCR) and (ii) gene sequences associated with the seven priority O serogroups (Poly O PCR), and then direct plating of broth samples positive in both STE and Poly O PCR onto Rainbow agar. Colonies on agar media were screened batchwise for STEC by the STE PCR, and presumptive isolates were characterized using a multiplex PCR and cloth-based hybridization array system targeting key virulence and O serogroup-specific markers. Using one representative strain of each priority O serogroup individually inoculated in beef trim samples, the method exhibited a limit of detection approaching 1 to 2 viable STEC cells per 65 g. None of the uninoculated trim samples produced positive results with either of the screening PCR procedures or on analysis of colonies recovered on plating media. STEC-negative samples were readily identified by screening PCR within 24 h, with a turnaround time of fewer than 4 days for confirmation of positives. The inclusivity and exclusivity characteristics of the screening PCR techniques were verified using a total of 65 different priority STEC strains: 24 nonpriority STEC, 15 non-STEC bacteria, and only those strains bearing the targeted characteristics produced screening PCR-positive results.

Low Prevalence of Salmonella and Shiga Toxin–Producing Escherichia coli in Lymph Nodes of Australian Beef Cattle

2017 ◽  
Vol 80 (12) ◽  
pp. 2105-2111 ◽  
Author(s):  
Gavin Bailey ◽  
Long Huynh ◽  
Lachlan Govenlock ◽  
David Jordan ◽  
Ian Jenson
Keyword(s):  
Escherichia Coli ◽  
Lymph Nodes ◽  
Shiga Toxin ◽  
Limit Of Detection ◽  
Ground Beef ◽  
Plate Count ◽  
Salmonella Spp ◽  
Live Animal ◽  
E Coli ◽  
Low Prevalence

ABSTRACT Salmonella contamination of ground beef has been viewed as originating from the surface of carcasses. Recent studies have identified lymph nodes as a potential source of Salmonella contamination because these tissues play an active role in containment of pathogens in the live animal and because some lymph nodes are unavoidably present in manufacturing beef trimmings or primal cuts that may be incorporated into ground beef. A survey was conducted of the microbiological status of lymph nodes from Australian cattle at the time of slaughter to determine the prevalence of microbiological contamination. Sets of lymph nodes (n = 197), consisting of the superficial cervical (prescapular), prepectoral, axillary, presternal, popliteal, ischiatic, subiliac (precrural), coxalis, and iliofemoralis (deep inguinal), were collected from five geographically separated Australian abattoirs over a period of 14 months. Samples were tested for the presence of Salmonella spp. and Shiga toxin–producing Escherichia coli by BAX PCR assay. Aerobic plate count, E. coli, and coliforms were enumerated with a lower limit of detection of 80 CFU per node. The observed prevalence of Salmonella within peripheral lymph nodes was 0.48% (7 of 1,464). Two of the seven lymph nodes in which Salmonella organisms were detected came from the same animal. Grass-fed, grain-fed, and cull dairy cattle were all found to have detectable Salmonella in lymph nodes. All Salmonella detections occurred during cooler months of the year. No Shiga toxin–producing E. coli were detected. Aerobic microorganisms were detected above the limit of quantification in 3.2% of nodes (median count 2.24 log per node), and E. coli was detected in 0.8% of nodes (median count 3.05 log per node). The low prevalence of Salmonella and low concentration of aerobic microorganisms in Salmonella-positive lymph nodes of Australian cattle at the time of slaughter suggest that the likelihood of lymph nodes contributing significantly to the presence of Salmonella in ground beef is low.

scholarly journals Activation of Prophage eib Genes for Immunoglobulin-Binding Proteins by Genes from the IbrAB Genetic Island of Escherichia coli ECOR-9

2002 ◽  
Vol 184 (13) ◽  
pp. 3640-3648 ◽  
Author(s):  
Carol H. Sandt ◽  
James E. Hopper ◽  
Charles W. Hill
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Phylogenetic Group ◽  
The Other ◽  
Overlapping Genes ◽  
Left Boundary ◽  
E Coli ◽  
Genome Homology ◽  
K 12 ◽  
Immunoglobulin Binding

ABSTRACT Four distinct Escherichia coli immunoglobulin-binding (eib) genes, each of which encodes a surface-exposed protein that binds immunoglobulins in a nonimmune manner, are carried by separate prophages in E. coli reference (ECOR) strain ECOR-9. Each eib gene was transferred to test E. coli strains, both in the form of multicopy recombinant plasmids and as lysogenized prophage. The derived lysogens express little or no Eib protein, in sharp contrast to the parental lysogen, suggesting that ECOR-9 has an expression-enhancing activity that the derived lysogens lack. Supporting this hypothesis, we cloned from ECOR-9 overlapping genes, ibrA and ibrB (designation is derived from “immunoglobulin-binding regulator”), which together activated eib expression in the derived lysogens. The proteins encoded by ibrA and ibrB are very similar to uncharacterized proteins encoded by genes of Salmonella enterica serovar Typhi and E. coli O157:H7 (in a prophage-like element of the Sakai strain and in two O islands of strain EDL933). The genomic segment containing ibrA and ibrB has been designated the IbrAB island. It contains regions of homology to the Shiga toxin-converting prophage, Stx2, as well as genes homologous to phage antirepressor genes. The left boundary between the IbrAB island and the chromosomal framework is located near min 35.8 of the E. coli K-12 genome. Homology to IbrAB was found in certain other ECOR strains, including the other five eib-positive strains and most strains of the phylogenetic group B2. Sequencing of a 1.1-kb portion of ibrAB revealed that the other eib-positive strains diverge by ≤0.1% from ECOR-9, whereas eib-negative ECOR-47 diverges by 16%.

scholarly journals Variation in Acid Resistance among Shiga Toxin-Producing Clones of Pathogenic Escherichia coli

2005 ◽  
Vol 71 (5) ◽  
pp. 2493-2500 ◽  
Author(s):  
Teresa M. Large ◽  
Seth T. Walk ◽  
Thomas S. Whittam
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Survival Rates ◽  
Acid Resistance ◽  
Acid Decarboxylase ◽  
Infectious Dose ◽  
Clonal Group ◽  
E Coli ◽  
Log Reduction ◽  
Oxidative System

ABSTRACT Pathogenic strains of Escherichia coli, such as E. coli O157:H7, have a low infectious dose and an ability to survive in acidic foods. These bacteria have evolved at least three distinct mechanisms of acid resistance (AR), including two amino acid decarboxylase-dependent systems (arginine and glutamate) and a glucose catabolite-repressed system. We quantified the survival rates for each AR mechanism separately in clinical isolates representing three groups of Shiga toxin-producing E. coli (STEC) clones (O157:H7, O26:H11/O111:H8, and O121:H19) and six commensal strains from ECOR group A. Members of the STEC clones were not significantly more acid resistant than the commensal strains when analyzed using any individual AR mechanism. The glutamate system provided the best protection in a highly acidic environment for all groups of isolates (<0.1 log reduction in CFU/ml per hour at pH 2.0). Under these conditions, there was notable variation in survival rates among the 30 O157:H7 strains, which depended in part on Mg2+ concentration. The arginine system provided better protection at pH 2.5, with a range of 0.03 to 0.41 log reduction per hour, compared to the oxidative system, with a range of 0.13 to 0.64 log reduction per hour. The average survival rate for the O157:H7 clonal group was significantly less than that of the other STEC clones in the glutamate and arginine systems and significantly less than that of the O26/O111 clone in the oxidative system, indicating that this clonal group is not exceptionally acid resistant with these specific mechanisms.

Evaluating the Efficacy of Three U.S. Department of Agriculture–Approved Antimicrobial Sprays for Reducing Shiga Toxin–Producing Escherichia coli Surrogate Populations on Bob Veal Carcasses

2016 ◽  
Vol 79 (6) ◽  
pp. 956-962 ◽  
Author(s):  
N. J. SEVART ◽  
N. BAUMANN ◽  
H. THIPPAREDDI ◽  
T. A. HOUSER ◽  
J. B. LUCHANSKY ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Control Point ◽  
Spray Application ◽  
Department Of Agriculture ◽  
E Coli ◽  
Log Reduction ◽  
Instrumental Color ◽  
Antimicrobial Intervention ◽  
Spray Treatment

ABSTRACT Effective antimicrobial intervention strategies to reduce Shiga toxin–producing Escherichia coli (STEC) risks associated with veal are needed. This study evaluated the efficacy of lactic acid (4.5%, pH 2.0), Citrilow (pH 1.2), and Beefxide (2.25%, pH 2.3) for reducing STEC surrogates on prerigor and chilled bob veal carcasses and monitored the effects of these interventions on chilled carcass color. Dehided bob veal carcasses were inoculated with a five-strain cocktail of rifampin-resistant, surrogate E. coli bacteria. E. coli surrogates were enumerated after inoculation, after water wash, after prechill carcass antimicrobial spray application, after chilling for 24 h, and after postchill carcass antimicrobial spray application; carcass color was measured throughout the process. A standard carcass water wash (~50°C) reduced the STEC surrogate population by 0.9 log CFU/cm2 (P ≤ 0.05). All three antimicrobial sprays applied to prerigor carcasses delivered an additional ~0.5-log reduction (P ≤ 0.05) of the surrogates. Chilling of carcasses for 24 h reduced (P ≤ 0.05) the surrogate population by an additional ~0.4 log cycles. The postchill application of the antimicrobial sprays provided no further reductions. Carcass L*, a*, and b* color values were not different (P &gt; 0.05) among carcass treatments. Generally, the types and concentrations of the antimicrobial sprays evaluated herein did not negatively impact visual or instrumental color of chilled veal carcasses. This study demonstrates that warm water washing, followed by a prechill spray treatment with a low-pH chemical intervention, can effectively reduce STEC risks associated with veal carcasses; this provides processors a validated control point in slaughter operations.

scholarly journals Persistent Colonization of Sheep byEscherichia coli O157:H7 and Other E. coliPathotypes

2000 ◽  
Vol 66 (11) ◽  
pp. 4926-4934 ◽  
Author(s):  
N. A. Cornick ◽  
S. L. Booher ◽  
T. A. Casey ◽  
H. W. Moon
Keyword(s):  
Escherichia Coli ◽  
Young Adult ◽  
Shiga Toxin ◽  
The Other ◽  
E Coli ◽  
Adult Sheep ◽  
Food Borne ◽  
Food Borne Illness ◽  
Byescherichia Coli ◽  
Frequency Magnitude

ABSTRACT Shiga toxin-producing Escherichia coli (STEC) is an important cause of food-borne illness in humans. Ruminants appear to be more frequently colonized by STEC than are other animals, but the reason(s) for this is unknown. We compared the frequency, magnitude, duration, and transmissibility of colonization of sheep by E. coli O157:H7 to that by other pathotypes of E. coli. Young adult sheep were simultaneously inoculated with a cocktail consisting of two strains of E. coli O157:H7, two strains of enterotoxigenic E. coli (ETEC), and one strain of enteropathogenic E. coli. Both STEC strains and ETEC 2041 were given at either 107 or 1010CFU/strain/animal. The other strains were given only at 1010 CFU/strain. We found no consistent differences among pathotypes in the frequency, magnitude, and transmissibility of colonization. However, the STEC strains tended to persist to 2 weeks and 2 months postinoculation more frequently than did the other pathotypes. The tendency for persistence of the STEC strains was apparent following an inoculation dose of either 107 or 1010 CFU. One of the ETEC strains also persisted when inoculated at 1010 CFU. However, in contrast to the STEC strains, it did not persist when inoculated at 107 CFU. These results support the hypothesis that STEC is better adapted to persist in the alimentary tracts of sheep than are other pathotypes ofE. coli.

Thermal Inactivation of Escherichia coli O157:H7 and Non-O157 Shiga Toxin–Producing Escherichia coli Cells in Mechanically Tenderized Veal†

2014 ◽  
Vol 77 (7) ◽  
pp. 1201-1206 ◽  
Author(s):  
JOHN B. LUCHANSKY ◽  
ANNA C. S. PORTO-FETT ◽  
BRADLEY A. SHOYER ◽  
HARSHAVARDHAN THIPPAREDDI ◽  
JESUS R. AMAYA ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Thermal Inactivation ◽  
Escherichia Coli O157 ◽  
Internal Temperature ◽  
E Coli ◽  
Log Reduction ◽  
Cooking Oil ◽  
Escherichia Coli Cells ◽  
Cooking Times

Preflattened veal cutlets (ca. 71.5 g, ca. 0.32 cm thick) were surface inoculated with ca. 6.8 log CFU/g of a multistrain cocktail of Escherichia coli O157:H7 (ECOH) or a cocktail made of single strains of serogroups O26, O45, O103, O104, O111, O121, and O145 of Shiga toxin–producing E. coli (STEC) cells and then were mechanically tenderized by passing once through a “Sir Steak” tenderizer. For each cooking time, in each of at least three trials, three inoculated and tenderized cutlets, with and without breading, were individually cooked in 15 or 30 ml of canola oil for 0.0, 0.75, 1.0, 1.25, 1.5, 1.75, or 2.25 min per side on an electric skillet set at 191.5°C. The temperatures of the meat and of the skillet were monitored and recorded using a type J thermocouple. Regardless of the breading or volume of oil used to cook the meat, the longer the cooking times, the higher was the internal temperature of the meat, along with a greater reduction of both ECOH and STEC. The average final internal temperature of the meat at the approximate geometric center ranged from 56.8 to 93.1°C. Microbial reductions of ca. 2.0 to 6.7 log CFU/g and ca. 2.6 to 6.2 log CFU/g were achieved for ECOH and STEC, respectively. Our data also revealed no differences in thermal inactivation of ECOH relative to the volume of oil used to cook nonbreaded cutlets. However, when cooking breaded cutlets, the use of more (30 ml) compared with less (15 ml) cooking oil resulted in greater reductions in pathogen numbers. To deliver about a 5.0-log reduction of ECOH and STEC, and to achieve the recommended internal temperature of 71.1°C, it was necessary to cook mechanically tenderized veal cutlets for at least 1.5 min per side on a preheated electric skillet set at 191.5°C and containing 15 ml of cooking oil. These data also established that cooking times and temperatures effective for inactivating serotype O157:H7 strains of E. coli in tenderized veal are equally effective against the additional six non-O157 Shiga toxin–producing strains investigated herein.

Specificity Analysis of a Novel Phage-Derived Ligand in an Enzyme-Linked Fluorescent Assay for the Detection of Escherichia coli O157:H7

2009 ◽  
Vol 72 (5) ◽  
pp. 1078-1081 ◽  
Author(s):  
CHRISTINE ROZAND ◽  
PETER C. H. FENG
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Cross Reactivity ◽  
The Other ◽  
Antibody Detection ◽  
Escherichia Coli O157 ◽  
Citrobacter Freundii ◽  
Fluorescent Assay ◽  
E Coli ◽  
Rough Strain

An assay using a phage-derived ligand to capture Escherichia coli O157:H7 prior to antibody detection was evaluated for assay specificity. Analysis of 200 strains showed that the assay was highly specific for the O157 serogroup. It detected all the O157:H7 strains including Shiga toxin–producing O157 nonmotile strains as well as O157 non-H7 strains. In addition, the assay detected various O157:H7 phenotypic variants that are not easily detected by routine analytical methods, as well as a rough strain that did not express O157 antigen and therefore is undetectable serologically. The phage ligand assay showed no cross-reactivity to the other E. coli serotypes. Isolates of Salmonella group N and a few Citrobacter freundii strains that cross-reacted with anti-O157 sera also showed cross-reactivity with the phage ligand. However, other strains that cross-reacted serologically with anti-O157 sera were correctly identified as negative with the phage ligand assay, including several strains of E. coli that nonspecifically autoagglutinate latex reagents.

Inactivation of Shiga Toxin–Producing Escherichia coli in Single-Strength Lemon and Lime Juices Containing Preservatives

2011 ◽  
Vol 74 (10) ◽  
pp. 1746-1750 ◽  
Author(s):  
AI KATAOKA ◽  
ELENA ENACHE ◽  
MARIA SOHAIL ◽  
PHILIP H. ELLIOTT ◽  
D. GLENN BLACK
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Food Products ◽  
Storage Conditions ◽  
E Coli ◽  
Log Reduction ◽  
Validation Tests ◽  
Shelf Stable

The objective of this study was to determine the inactivation of non-O157 Shiga toxin–producing Escherichia coli (STEC) serotypes in comparison with O157 STEC in commercially produced, shelf-stable lemon and lime juices. The present validation tests confirmed that storage of the juices containing preservatives at room temperatures (22°C) for 3 days (72 h) ensures a &gt;6-log reduction of O26, O45, O103, O111, O121, O145, and O157 STEC. These results demonstrate that non-O157 STEC had survival abilities comparable to those of E. coli O157:H7 strains in acidic food products such as lemon and lime juices (pH 2.5 ± 0.1); therefore, the storage conditions deemed to inactivate E. coli O157:H7 similarly inactivate the non-O157 serotypes.

Viability of Escherichia coli O157:H7 in Fermented Semidry Low-Temperature-Cooked Beef Summer Sausage

1997 ◽  
Vol 60 (10) ◽  
pp. 1158-1162 ◽  
Author(s):  
MEHMET CALICIOGLU ◽  
NANCY G. FAITH ◽  
DENNIS R. BUEGE ◽  
JOHN B. LUCHANSKY
Keyword(s):  
Escherichia Coli ◽  
Low Temperature ◽  
Cold Water ◽  
Starter Culture ◽  
Escherichia Coli O157 ◽  
Internal Temperature ◽  
Direct Plating ◽  
E Coli ◽  
Log Reduction ◽  
Viable Cells

The population of inoculated Escherichia coli O157:H7 was monitored during the manufacture and storage of a semidry beef summer sausage processed by fermentation and cooking at a low temperature by heating to an internal temperature of 130°F (54°C). The all-beef batter (11% fat and nonmeat ingredients) was inoculated with the commercial starter culture Pediococcus acidilactici HP (≥8.6 log CFU/g of batter) and a five-strain mixture of E. coli O157:H7 (≥7 log CFU/g) and then hand stuffed into 2.5-inch (64-mm) diameter fibrous casings. The sausages were fermented at an initial temperature of 85°F (29°C) to a final temperature of 105°F (41°C) over ca. 13 h at 80% relative humidity (RH) to pH 4.6 or pH 5.0. After fermentation to pH 4.6, the internal temperature of the chubs was raised to 130°F (54°C) instantaneous over 3.6 h at 60% RH. After fermentation to pH 5.0, the internal temperature of the chubs was raised to 130°F (54°C) over 3.6 h at 60% RH and the chubs were maintained under these conditions for 0, 30, or 60 min. he chubs were cold water showered for 15 min and then chilled at 39°F (4°C) for 6 h before being vacuum packaged and stored at 39°F (4°C) or 77°F (25°C) for 7 days. Regardless of the target pH, fermentation alone resulted in only a 1.39-log CFU/g decrease in pathogen numbers. However, fermentation to pH 4.6 and heating to an internal temperature of 130°F (54°C) instantaneous reduced counts of E. coli O157:H7 by ≥7.0 log units to below detection levels (&lt;10 CFU/g). Pathogen numbers remained below levels detectable by direct plating, but viable E. coli O157:H7 cells were recovered by enrichment of samples during sausage storage at either refrigeration or abuse temperatures. In contrast, fermentation to pH 5.0 and heating to an internal temperature of 130°F (54°C) instantaneous resulted in a 3.2-log-unit decrease in counts of E. coli O157:H7. No appreciable reductions in pathogen numbers were observed thereafter following storage at either 39°F (4°C) or 77°F (25°C) for 7 days. Fermentation to pH 5.0 and heating to an internal temperature of 130°F (54°C) instantaneous followed by holding for 30 or 60 min resulted in about a 5- or 7-log reduction, respectively, in pathogen numbers. For chubs held for 30 min at 130°F (54°C), pathogen numbers decreased to 2.02 and &lt;1.0 log CFU/g at 39°F (4°C) and 77°F (25°C), respectively, after 7 days; viable cells were only observed by enrichment after storage at 77°F (25°C). For chubs held for 60 min at 130°F (54°C), pathogen numbers remained below levels detectable by direct plating, but viable cells were recoverable by enrichment after 7 days at both storage temperatures. These data will be useful guidelines to manufacturers for developing processing conditions to further ensure the safety of this category of fermented sausages relative to food-borne pathogens such as serotype O157:H7 strains of E. coli.

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