Presence of Escherichia coli O157:H7 in ground beef and ground baby beef meat

A total of 114 beef and baby beef samples were examined. The samples included ground baby beef, mixed ground baby beef and pork, and chopped and shaped meat. The samples were analyzed from 30 different grocery stores in Zagreb, Croatia. The object of this study was to evaluate the prevalence of Escherichia coli O157:H7 in the samples that can enhance the potential risk of outbreaks of hemorrhagic colitis and hemolytic uremic syndrome. The results in all tested samples of E. coli O157:H7 were negative. A single sample was positive in a latex agglutination test using antiserum to O157:H7. It was identified as Proteus vulgaris at the Pasteur Institute, Paris, France. This result correlates positively with cross-contamination with Yersinia enterocolitica 09, Brucella abortus, Salmonella type N, and Pseudomonas maltophila.     

Detection of freeze-injured Escherichia coli O157:H7 cells from foods by resuscitation prior to selective enrichment

We tried to detect Escherichia coli O157:H7 in food samples artificially contaminated with freeze-injured E. coli O157:H7 using an enrichment method with modified EC broth supplemented with novobiocin (mEC + n). When the samples were cultured for enrichment immediately after inoculation of freeze-injured cells, E. coli O157:H7 was not detected in 13 out of 18 samples. However, allowing the food samples to stand for 3 h at room temperature prior to enrichment in mEC + n remarkably improved recovery of E. coli O157:H7 except for some acidic foods. E. coli O157:H7 was detected in the acidic foods by introducing a resuscitation step of 3-h of incubation in a non-selective broth at room temperature prior to enrichment with mEC + n.     

Evaluation of standard enrichment broths for recovery of healthy and chlorine-injured Escherichia coli O157:H7 cells in kimchi

Food Science and Biotechnology - This study aimed to evaluate three standard enrichment broth preparations for the recovery of healthy and chlorine-injured E. coli O157:H7 cells in kimchi. The...     

Inactivation of Escherichia coli O157:H7 in packaged ground beef by allyl isothiocyanate

Commercial allyl isothiocyanate (AIT) was examined for its ability to reduce numbers of Escherichia coli O157:H7 inoculated in fresh ground beef packaged under nitrogen and stored refrigerated or frozen. A five-strain cocktail of E. coli O157:H7 containing 3 or 6 log10 cfu/g was inoculated into 100 g ground beef and formed into 10x1-cm patties. A 10-cm diameter filter paper disk treated with AIT suspended in sterile corn oil was placed on top of a single patty. One patty and paper disk were placed in a bag of Nylon/EVOH/PE with O2 permeability of 2.3 cm3 m(-2) 24 h atm at 23 degrees C. The bags were back-flushed with 100% nitrogen, heat-sealed and stored at 10, 4 and -18 degrees C for 8, 21 or 35 days, respectively. During storage, the AIT levels in the package headspaces were determined by gas liquid chromatography, and mesophilic bacteria and E. coli O157:H7 were counted. The mesophilic aerobic bacteria in ground beef patties were largely unaffected by the addition of AIT. At an initial population of 3 log10 cfu/g, E. coli O157:H7 was reduced by AIT to undetectable levels after 18 days at 4 degrees C or 10 days at -18 degrees C. In samples inoculated with 6 log10 cfu/g, a >3 log10 reduction of E. coli O157:H7 was observed after 21 days at 4 degrees C, while a 1 log10 reduction was observed after 8 and 35 days at 10 and -18 degrees C, respectively. The final AIT concentrations in the headspaces after storage at 10, 4, and -18 degrees C were 444, 456, and 112 microg/ml at 8, 21, and 35 days, respectively. Results showed that AIT can substantially reduce numbers of E. coli O157:H7 in fresh ground beef during refrigerated or frozen storage.     

Quantitative detection of Escherichia coli O157:H7 in ground beef by the polymerase chain reaction

A method for quantitative detection of Escherichia coli O157:H7 based on the polymerase chain reaction (PCR) was developed. The method used the NIH Image 1·61 software program to quantitatively analyse the intensity of the fluorescent image of the amplified PCR product. Based on the PCR with SLT1 and SLT2 primers used separately, a log-linear relationship between the numbers of cfu of E. coli O157:H7 inoculated into ground beef and the intensity of the PCR products was achieved with and without enrichment. Without enrichment, 150 cfu of E. coli O157:H7 per gram of ground beef were detected. In contrast, the detection limit decreased to 1·2 cfu g⁻¹ of ground beef using SLT1 and SLT2 primers after 4·5 h of enrichment using modified EC broth with 20 μg ml⁻¹ of novobiocin.     

Repair and growth of heat- and freeze-injured Escherichia coli O157:H7 in selective enrichment broths

Two Escherichia coli O157:H7 strains, ATCC 35150 and 43894, were heat injured in a beef infusion at 53°C for 40 and 50 min, respectively (1· 5–2·0 log₁₀cfu ml⁻¹of injury) and freeze injured at −25°C for 30 days (1 log₁₀cfu ml⁻¹of injury) as determined by plating on MacConkey agar with 0·60% bile salts #3 (Mac-BS) as the selective medium and on Brain Heart Infusion agar (BHIA) as the non-selective medium. Repair of injury was measured in five selective enrichment broths [buffered peptone water supplemented with vancomycin, cefsulodin, and cefixime (BPW-VCC), modified EC broth with novobiocin (mEC+n), enterohaemorrhagic E. coli enrichment broth (EEB), double modified TSB (dmTSB), and BCM^®E. coli enrichment broth (BCM^®-EB)] versus TSB as the non-selective control broth over 3 h incubation at 37°C and 42°C. Repair was measured as the increase in cfu ml⁻¹enumerated on Mac-BS with time vs the total cfu ml⁻¹(injured and uninjured cells) enumerated on BHIA. In mEC+n, EEB, and dmTSB some death of both heat- and freeze-injured cells occurred immediately during the 3 h incubation (decrease on BHIA plates), and there was either minimal or no repair of the injured cells at both temperatures. Efficient repair of heat injury was obtained with both BPW-VCC and BCM^®-EB, but the latter produced a growth rate and final cell concentration closer to TSB. In freeze-injury repair however, BPW-VCC gave poor results while repair in BCM^®-EB was equal to TSB. Both BCM^®-EB and BPW-VCC inhibited the growth of all Gram-positive and a select number of Gram-negative bacteria tested. The ability of the selective enrichment broth BCM^®-EB to resuscitate heat- and freeze-injured E. coli O157:H7 efficiently within 3 h, warrants further testing with other types of stress in both artificially and naturally contaminated foods.     

Magnetic nanoparticle-antibody conjugates for the separation of Escherichia coli O157:H7 in ground beef

The immunomagnetic separation with magnetic nanoparticle-antibody conjugates (MNCs) was investigated and evaluated for the detection of Escherichia coli O157:H7 in ground beef samples. MNCs were prepared by immobilizing biotin-labeled polyclonal goat anti-E. coli antibodies onto streptavidin-coated magnetic nanoparticles. For bacterial separation, MNCs were mixed with inoculated ground beef samples, then nanoparticle-antibody-E. coli O157:H7 complexes were separated from food matrix with a magnet, washed, and surface plated for microbial enumeration. The capture efficiency was determined by plating cells bound to nanoparticles and unbound cells in the supernatant onto sorbitol MacConkey agar. Key parameters, including the amount of nanoparticles and immunoreaction time, were optimized with different concentrations of E. coli O157:H7 in phosphate-buffered saline. MNCs presented a minimum capture efficiency of 94% for E. coli O157:H7 ranging from 1.6 x 10(1) to 7.2 x 10(7) CFU/ml with an immunoreaction time of 15 min without any enrichment. Capture of E. coli O157:H7 by MNCs did not interfere with other bacteria, including Salmonella enteritidis, Citrobacter freundii, and Listeria monocytogenes. The capture efficiency values of MNCs increased from 69 to 94.5% as E. coli O157:H7 decreased from 3.4 x 10(7) to 8.0 x 10(0) CFU/ml in the ground beef samples prepared with minimal steps (without filtration and centrifugation). An enrichment of 6 h was done for 8.0 x 10(0) and 8.0 x 10(1) CFU/ml of E. coli O157:H7 in ground beef to increase the number of cells in the sample to a detectable level. The results also indicated that capture efficiencies of MNCs for E. coli O157:H7 with and without mechanical mixing during immunoreaction were not significantly different (P > 0.05). Compared with microbeads based immunomagnetic separation, the magnetic nanoparticles showed their advantages in terms of higher capture efficiency, no need for mechanical mixing, and minimal sample preparation.     

Distribution patterns of Escherichia coli O157:H7 in ground beef produced by a laboratory-scale grinder

This study determined the distribution patterns of Escherichia coli O157:1H7 in ground beef when a contaminated beef trim was introduced into a batch of uncontaminated beef trims prior to grinding in a small-scale laboratory grinder. A beef trim (15.3 +/- 2 g) was inoculated with a rifampicin-resistant strain of E. coli O157:H7 (E. coli O157:H7rif) and introduced into a stream of noncontaminated beef (322 +/- 33 g) prior to grinding. Seven inoculum levels (6, 5, and 4 total log CFU [high]; and 3, 2, 1, and 0 total log CFU [low]) were studied in triplicate. E. coli O157:H7rif was not detected in 3.1 to 43% of the ground beef inoculated with the high levels or in 3.4 to 96.9% of the ground beef inoculated with the low levels. For all inoculum levels studied, the five ground beef fractions (each 7.8 +/- 0.6 g) with the highest pathogen levels accounted for 59 to 100% of the total pathogens detected. For all inoculum levels, there was a linear relationship between the quantity of ground beef containing E. coli O157:H7rif and the inoculum level. The quantity of E. coli O157:H7rif in the beef remaining in the grinder was proportional to the inoculum level and was related to the location in the grinder. Different components of the grinder accumulated E. coli O157:H7rif in different quantities, with the most significant accumulation being in the nut (collar) that attaches the die to the blade. This study determined specific distribution patterns of E. coli O157:H7rif after the grinding of a contaminated beef trim along with uncontaminated trims, and the results indicate that the grinding operation should be regarded as a means of distribution of microbial contamination in risk analyses of ground beef operations.     

Inactivation of Escherichia coli O157:H7 in ground beef by single-cycle and multiple-cycle high-pressure treatments

The effect of single- and multiple-cycle high-pressure treatments on the survival of Escherichia coli CECT 4972, a strain belonging to the O157:H7 serotype, in ground beef was investigated. Beef patties were inoculated with 10(7) CFU/g E. coli O157:H7, and held at 4 degrees C for 20 h before high-pressure treatments. Reduction of the E. coli O157:H7 population by single-cycle treatments at 400 MPa and 12 degrees C ranged from 0.82 log CFU/g for a 1-min cycle to 4.39 log CFU/g for a 20-min cycle. Multiple-cycle treatments were very effective, with four 1-min cycles at 400 MPa and 12 degrees C reducing the E. coli O157:H7 population by 4.38 log CFU/g, and three 5-min cycles by 4.96 log CFU/g. The color parameter L* increased significantly with high-pressure treatments in the interior and the exterior of beef patties, whereas a* decreased in the interior, and b* increased in the exterior-changes that might diminish consumer acceptance of the product. Kramer shear force and energy were generally higher in pressurized than in control ground beef. Maximum values for these texture parameters, which corresponded to tougher patties, were reached after one 10-min cycle in the case of single-cycle treatments or two 5-min cycles in the case of multiple-cycle treatments. High-pressure treatments had no significant effect on Warner-Bratzler shear force.     

Antibacterial effect of lactoferricin B on Escherichia coli O157:H7 in ground beef.

The antibacterial activity of lactoferricin B on enterohemorrhagic Escherichia coli O157:H7 in 1% peptone medium and ground beef was studied at 4 and 10 degrees C. In 1% peptone medium, 50 and 100 microg of lactoferricin B per ml reduced E. coli O157:H7 populations by approximately 0.7 and 2.0 log CFU/ml, respectively. Studies comparing the antibacterial effect of lactoferricin B on E. coli O157:H7 in 1% peptone at pH 5.5 and 7.2 did not reveal any significant difference (P > 0.5) at the two pH values. Lactoferricin B (100 microg/g) reduced E. coli O157:H7 population in ground beef by about 0.8 log CFU/g (P < 0.05). No significant difference (P > 0.5) was observed in the total plate count between treatment and control ground beef samples stored at 4 and 10 degrees C. The antibacterial effect of lactoferricin B on E. coli O157:H7 observed in this study is not of sufficient magnitude to merit its use in ground beef for controlling the pathogen.     

Thermal inactivation of Escherichia coli O157:H7 in ground beef supplemented with sodium lactate

A study was conducted to investigate the antimicrobial effect of sodium lactate (NaL) (0, 1.5, 3.0, and 4.5%) on the survival of Escherichia coli O157:H7 in 93% lean ground beef. Samples inoculated with a mixture of four strains of E. coli O157:H7 (10(7) to 10(8) CFU/g) were subjected to immersion heating in a water bath stabilized at 55, 57.5, 60, 62.5, or 65 degrees C. Results of statistical analysis indicated that the heating temperature was the only factor affecting the decimal reduction times (D-values) of E. coli O157:H7 in 93% lean ground beef. The change in temperature required to change the D-value (the z-value) was determined as 7.6 degrees C. The thermal resistance of this organism was neither affected by the addition of NaL nor by the interactions between NaL and temperature. Adding NaL to ground beef to reduce the thermal resistance of E. coli O157:H7 is therefore not recommended.     

Quantifying the robustness of a broth-based Escherichia coli O157:H7 growth model in ground beef

The robustness of a microbial growth model must be assessed before the model can be applied to new food matrices; therefore, a methodology for quantifying robustness was developed. A robustness index (RI) was computed as the ratio of the standard error of prediction to the standard error of calibration for a given model, where the standard error of calibration was defined as the root mean square error of the growth model against the data (log CFU per gram versus time) used to parameterize the model and the standard error of prediction was defined as the root mean square error of the model against an independent data set. This technique was used to evaluate the robustness of a broth-based model for aerobic growth of Escherichia coli 0157:H7 (in the U.S Department of Agriculture Agricultural Research Service Pathogen Modeling Program) in predicting growth in ground beef under different conditions. Comparison against previously published data (132 data sets with 1,178 total data points) from experiments in ground beef at various experimental conditions (4.8 to 45 degrees C and pH 5.5 to 5.9) yielded RI values ranging from 0.11 to 2.99. The estimated overall RI was 1.13. At temperatures between 15 and 40 degrees C, the RI was close to and smaller than 1, indicating that the growth model is relatively robust in that temperature range. However, the RI also was related (P < 0.05) to temperature. By quantifying the predictive accuracy relative to the expected accuracy, the RI could be a useful tool for comparing various models under different conditions.     

High levels of background flora inhibits growth of Escherichia coli O157:H7 in ground beef

The influence of natural background flora under aerobic and anaerobic incubation on the growth of Escherichia coli O157:H7 in ground beef was investigated. The background flora from eight different commercial ground beef were added to ground beef spiked with E. coli O157:H7 and stored either aerobically or anaerobically at 12 degrees C. The results showed that the presence of a large number of background bacteria in the ground meat inhibited the growth of E. coli O157:H7 both aerobically and anaerobically. Inhibition was more pronounced under anaerobic conditions. The background floras consisted mainly of lactic acid bacteria of which approximately 80% were Lactobacillus sakei. These results show the importance of the natural background flora in meat for inhibition of growth of E. coli O157:H7.     

Evaluation of consumer-style cooking methods for reduction of Escherichia coli O157:H7 in ground beef

The objective of this study was to evaluate the thermal inactivation of Escherichia coli O157:H7 in ground beef cooked to an internal temperature of 71.1 degrees C (160 degrees F) under conditions simulating consumer-style cooking methods. To compare a double-sided grill (DSG) with a single-sided grill (SSG), two different cooking methods were used for the SSG: for the one-turnover (OT-SSG) method, a patty was turned once when the internal temperature reached 40 degrees C, and for the multiturnover (MT-SSG) method, a patty was turned every 30 s. Patties (100 g, n = 9) inoculated with a five-strain mixture of E. coli O157: H7 at a concentration of 10(7) CFU/g were cooked until all three temperature readings (for two sides and the center) for a patty were 71.1 degrees C. The surviving E. coli O157:H7 cells were enumerated on sorbitol MacConkey (SMAC) agar and on phenol red agar base with 1% sorbitol (SPRAB). The order of the cooking methods with regard to the cooking time required for the patty to reach 71.1 degrees C was as follows: DSG (2.7 min) < MT-SSG (6.6 min) < OT-SSG (10.9 min). The more rapid, higher-temperature cooking method was more effective (P < 0.01) in destroying E. coli O157:H7 in ground beef. E. coli O157:H7 reduction levels were clearly differentiated among treatments as follows: OT-SSG (4.7 log10 CFU/g) < MT-SSG (5.6 log10 CFU/g) < DSG (6.9 log10 CFU/g). Significantly larger numbers of E. coil O157:H7 were observed on SPRAB than on SMAC agar. To confirm the safety of ground beef cooked to 71.1 degrees C, additional patties (100 g, n = 9) inoculated with lower concentrations of E. coli O157:H7 (10(3) to 10(4) CFU/g) were tested. The ground beef cooked by the OT-SSG method resulted in two (22%) of nine samples testing positive after enrichment, whereas no E. coli O157:H7 was found for samples cooked by the MT-SSG and DSG methods. Our findings suggest that consumers should be advised to either cook ground beef patties in a grill that cooks the top and the bottom of the patty at the same time or turn patties frequently (every 30 s) when cooking on a grill that cooks on only one side.     

Viability of acid-adapted Escherichia coli O157:H7 in ground beef treated with acidic calcium sulfate

The effects of lactic acid, acetic acid, and acidic calcium sulfate (ACS) on viability and subsequent acid tolerance of three strains of Escherichia coli O157:H7 were determined. Differences in tolerance to acidic environments were observed among strains, but the level of tolerance was not affected by the acidulant to which cells had been exposed. Cells of E. coli O157:H7 adapted to grow on tryptic soy agar acidified to pH 4.5 with ACS were compared to cells grown at pH 7.2 in the absence of ACS for their ability to survive after inoculation into ground beef treated with ACS, as well as untreated beef. The number of ACS-adapted cells recovered from ACS-treated beef was significantly (alpha = 0.05) higher than the number of control cells recovered from ACS-treated beef during the first 3 days of a 10-day storage period at 4 degrees C, suggesting that ACS-adapted cells might be initially more tolerant than unadapted cells to reduced pH in ACS-treated beef. Regardless of treatment of ground beef with ACS or adaptation of E. coli O157:H7 to ACS before inoculating ground beef, the pathogen survived in high numbers.     

Draft risk assessment of the public health impact of Escherichia coli O157:H7 in ground beef

An assessment of the risk of illness associated with Escherichia coli O157:H7 in ground beef was drafted in 2001. The exposure assessment considers farm, slaughter, and preparation factors that influence the likelihood of humans consuming ground beef servings containing E. coli O157:H7 and the number of cells in a contaminated serving. Apparent seasonal differences in prevalence of cattle infected with E. coli O157:H7 corresponded to seasonal differences in human exposure. The model predicts that on average 0.018% of servings consumed during June through September and 0.007% of servings consumed during the remainder of the year are contaminated with one or more E. coli O157:H7 cells. This exposure risk is combined with the probability of illness given exposure (i.e., dose response) to estimate a U.S. population risk of illness of nearly one illness in each 1 million (9.6 x 10(-7)) servings of ground beef consumed. Uncertainty about this risk ranges from about 0.33 illness in every 1 million ground beef servings at the 5th percentile to about two illnesses in every 1 million ground beef servings at the 95th percentile.     

Thermal inactivation of Escherichia coli O157:H7 inoculated at different depths of non-intact blade-tenderized beef steaks

Studies evaluated thermal inactivation of Escherichia coli O157:H7 inoculated at different depths of simulated blade-tenderized non-intact steaks. Fresh beef slices (0.3 or 0.6 cm thick) were stacked on top of each other to form 2.4 or 1.2 cm thick steaks. Steaks were blade-tenderized and then inoculated with rifampicin-resistant Escherichia coli O157:H7 (8 strain mixture; 4 log CFU/cm(2)) on the surface or between slices, vacuum-packaged, and stored at 4 or -20 °C for 5 d before cooking. Steaks were cooked by pan-broiling or roasting to a geometric center temperature of 60 °C. Frozen samples were either cooked from the frozen state or after thawing to approximately 4 or 25 °C. In steaks inoculated on the external surface and cooked by pan-broiling, pathogen survivors recovered from thinner (1.2 cm) steaks were greater (P < 0.05) than those recovered from thicker (2.4 cm) steaks. Cooking steaks from a frozen state or after thawing (4 or 25 °C) did not (P ≥ 0.05) affect extent of pathogen inactivation. Survivors after pan-broiling of 2.4 cm thick steaks increased (P < 0.05) from 0.3 to 1.3 log CFU/cm(2) for surface-inoculated steaks to 2.5 to 3.2 log CFU/cm(2) for samples inoculated at the center (1.2 cm depth). In comparison, overall thermal destruction of the pathogen in steaks cooked by roasting was less, and survivor counts were generally not different (P ≥ 0.05) at each depth of inoculation. These data should be useful in development of lethality guidelines to ensure safe consumption of non-intact meat products. PRACTICAL APPLICATION: Results of this study should be useful for developing cooking guidelines, for foodservice establishments and consumers, to ensure safe consumption of non-intact meat products.     

Carvacrol and cinnamaldehyde facilitate thermal destruction of Escherichia coli O157:H7 in raw ground beef

The heat resistance of a four-strain mixture of Escherichia coli O157:H7 in raw ground beef in both the absence and presence of the antimicrobials carvacrol and cinnamaldehyde was tested at temperatures ranging from 55 to 62.5 degrees C. Inoculated meat packaged in bags was completely immersed in a circulating water bath, cooked for 1 h to an internal temperature of 55, 58, 60, or 62.5 degrees C, and then held for predetermined lengths of time ranging from 210 min at 55 degrees C to 5 min at 62.5 degrees C. The surviving bacteria were enumerated by spiral plating onto tryptic soy agar overlaid with sorbitol MacConkey agar. Inactivation kinetics of the pathogens deviated from first-order kinetics. D-values (time for the bacteria to decrease by 90%) in the control beef ranged from 63.90 min at 55 degrees C to 1.79 min at 62.5 degrees C. D-values determined by a logistic model ranged from 43.18 min (D1, the D-value of a major population of surviving cells) and 89.84 min (D2, the D-value of a minor subpopulation) at 55 degrees C to 1.77 (D1) and 0.78 min (D2) at 62.5 degrees C. The thermal death times suggested that to achieve a 4-D reduction, contaminated processed ground beef should be heated to an internal temperature of 60 degrees C for at least 30.32 min. Significantly increased sensitivity to heat (P < 0.05) was observed with the addition and/or increasing levels of carvacrol or cinnamaldehyde from 0.5 to 1.0%. The observed thermal death times may facilitate the design of acceptance limits at critical control points for ground beef at lower times and temperatures of heating.     

Evaluation of culture- and PCR-based detection methods for Escherichia coli O157:H7 in inoculated ground beeft

Currently, several beef processors employ test-and-hold systems for increased quality control of ground beef. In such programs, each lot of product must be tested and found negative for Escherichia coli O157:H7 prior to release of the product into commerce. Optimization of three testing attributes (detection time, specificity, and sensitivity) is critical to the success of such strategies. Because ground beef is a highly perishable product, the testing methodology used must be as rapid as possible. The test also must have a low false-positive result rate so product is not needlessly discarded. False-negative results cannot be tolerated because they would allow contaminated product to be released and potentially cause disease. In this study, two culture-based and three PCR-based methods for detecting E. coli O157:H7 in ground beef were compared for their abilities to meet the above criteria. Ground beef samples were individually spiked with five genetically distinct strains of E. coli O157: H7 at concentrations of 17 and 1.7 CFU/65 g and then subjected to the various testing methodologies. There was no difference (P > 0.05) in the abilities of the PCR-based methods to detect E. coli O157:H7 inoculated in ground beef at 1.7 CFU/65 g. The culture-based systems detected more positive samples than did the PCR-based systems, but the detection times (21 to 48 h) were at least 9 h longer than those for the PCR-based methods (7.5 to 12 h). Ground beef samples were also spiked with potentially cross-reactive strains. The PCR-based systems that employed an immunomagnetic separation step prior to detection produced fewer false-positive results.