Inactivation of Escherichia coli by ozone treatment of apple juice at different pH levels

This research investigated the efficacy of gaseous ozone on the inactivation of Escherichia coli ATCC 25922 and NCTC 12900 strains in apple juice of a range of pH levels, using an ozone bubble column. The pH levels investigated were 3.0, 3.5, 4.0, 4.5 and 5.0. Apple juice inoculated with E. coli strains (10⁶ CFU/mL) was treated with ozone gas at a flow rate of 0.12 L/min and ozone concentration of 0.048 mg/min/mL for up to 18 min. Results show that inactivation kinetics of E. coli by ozone were affected by pH of the juice. The ozone treatment duration required for achieving a 5-log reduction was faster (4 min) at the lowest pH than at the highest pH (18 min) studied. The relationship between time required to achieve 5 log reduction (t_5d) and pH for both strains was described mathematically by two exponential equations. Ozone treatment appears to be an effective process for reducing bacteria in apple juice and the required applied treatment for producing a safe apple juice is dependant on its acidity level.     

Occurrence of Escherichia coli O157, O111 and O26 in raw ewe's milk and performance of two enrichment broths and two plating media used for its assessment

The occurrence of Escherichia coli O157, O111 and O26 in 159 raw ewe's milk samples was examined. Sample-aliquots were incubated simultaneously in TSB added with yeast extract (YETSB) and mTSB with novobiocin (N-mTSB). Serogroup-specific immunomagnetic separation (IMS) was then used and IMS beads were plated in a cefixime tellurite (CT)-containing media (CT-SMAC, CT-SBMAC and CT-RMAC for E. coli O157, O111 and O26, respectively) and E. coli O157:H7 chromogenic ID agar. A sweep of confluent growth from each medium was examined for the presence of E. coli O157 and O111 using PCR, and for E. coli O26 using a latex agglutination test. Enumeration of E. coli O157 and O111 was performed in the samples tested positive for the correspondent serogroup using the most probable number (MPN) method combined with PCR. Percentage occurrences of E. coli O157, O111 and O26 were 18.2, 8.2 and 5.7, respectively. Mean E. coli O157 and O111 levels were 0.22 and < 0.04 MPN/mL, respectively. Enrichment in YETSB resulted in higher detection rates of E. coli O157 and O26 than in N-mTSB. When YETSB was used as enrichment broth and for these last two serogroups, the analysis of the confluent growth from the CT-media gave more positive results than that from E. coli O157:H7-ID medium.     

Concentration-dependent inhibition of Escherichia coli O157:H7 and heterocyclic amines in heated ground beef patties by apple and olive extracts,onion powder and clove bud oil

The effects of plant compounds on Escherichia coli O157:H7 and two major heat-induced heterocyclic amines (HCAs) MeIQx and PhIP in grilled ground beef patties were determined. Ground beef with added apple and olive extracts, onion powder, and clove bud oil was inoculated with E. coli O157:H7 (10⁷ CFU/g) and cooked to reach 45 °C at the geometric center, flipped and then cooked for another 5 min. Cooled samples were taken for microbiological and HCA analyses. Olive extract at 3% reduced E. coli O157:H7 to below detection. Reductions of up to 1 log were achieved with apple extract. Olive and apple extracts reduced MeIQx by up to 49.1 and 50.9% and PhIP by up to 50.6 and 65.2%, respectively. Onion powder reduced MeIQx and PhIP by 47 and 80.7%, respectively. Inactivation of E. coli O157:H7 and suppression of HCAs in grilled meat were achieved by optimized amounts of selected plant compounds.     

Attachment and biofilm formation by Escherichia coli O157:H7 at different temperatures, on various food-contact surfaces encountered in beef processing

Escherichia coli O157:H7 attached to beef-contact surfaces found in beef fabrication facilities may serve as a source of cross-contamination. This study evaluated E. coli O157:H7 attachment, survival and growth on food-contact surfaces under simulated beef processing conditions. Stainless steel and high-density polyethylene surfaces (2 × 5 cm) were individually suspended into each of three substrates inoculated (6 log CFU/ml or g) with E. coli O157:H7 (rifampicin-resistant, six-strain composite) and then incubated (168 h) statically at 4 or 15 °C. The three tested soiling substrates included sterile tryptic soy broth (TSB), unsterilized beef fat-lean tissue (1:1 [wt/wt]) homogenate (10% [wt/wt] with sterile distilled water) and unsterilized ground beef. Initial adherence/attachment of E. coli O157:H7 (0.9 to 2.9 log CFU/cm²) on stainless steel and high-density polyethylene was not affected by the type of food-contact surface but was greater (p < 0.05) through ground beef. Adherent and suspended E. coli O157:H7 counts increased during storage at 15 °C (168 h) by 2.2 to 5.4 log CFU/cm² and 1.0 to 2.8 log CFU/ml or g, respectively. At 4 °C (168 h), although pathogen levels decreased slightly in the substrates, numbers of adherent cells remained constant on coupons in ground beef (2.4 to 2.5 log CFU/cm²) and increased on coupons in TSB and fat-lean tissue homogenate by 0.9 to 1.0 and 1.7 to 2.0 log CFU/cm², respectively, suggesting further cell attachment. The results of this study indicate that E. coli O157:H7 attachment to beef-contact surfaces was influenced by the type of soiling substrate and temperature. Notably, attachment occurred not only at a temperature representative of beef fabrication areas during non-production hours (15 °C), but also during cold storage (4 °C) temperatures, thus, rendering the design of more effective sanitation programs necessary.     

Use of low dose e-beam irradiation to reduce E. coli O157:H7, non-O157 (VTEC) E. coli and Salmonella viability on meat surfaces

This study determined the extent that irradiation of fresh beef surfaces with an absorbed dose of 1 kGy electron (e-) beam irradiation might reduce the viability of mixtures of O157 and non-O157 verotoxigenic Escherichia coli (VTEC) and Salmonella. These were grouped together based on similar resistances to irradiation and inoculated on beef surfaces (outside flat and inside round, top and bottom muscle cuts), and then e-beam irradiated. Salmonella serovars were most resistant to 1 kGy treatment, showing a reduction of ≤ 1.9 log CFU/g. This treatment reduced the viability of two groups of non-O157 E. coli mixtures by ≤ 4.5 and ≤ 3.9 log CFU/g. Log reductions of ≤ 4.0 log CFU/g were observed for E. coli O157:H7 cocktails. Since under normal processing conditions the levels of these pathogens on beef carcasses would be lower than the lethality caused by the treatment used, irradiation at 1 kGy would be expected to eliminate the hazard represented by VTEC E. coli.     

Evaluation of a multiplex-PCR detection in combination with an isolation method for STEC O26, O103, O111, O145 and sorbitol fermenting O157 in food

The aim of the current study was to evaluate a multiplex PCR (mPCR) detection test combined with the evaluation of a previously described isolation method.Minced beef, raw-milk cheese and sprouted seed samples were inoculated with low amounts (7-58 cfu 25 g⁻¹) of non-stressed, cold-stressed or freeze-stressed clinical STEC strains, including serogroups O26, O103, O111, O145, sorbitol fermenting (SF) O157 and non-sorbitol fermenting (NSF) O157. The inoculated pathogen was detected using a 24 h-enrichment followed by an mPCR protocol, and in parallel isolated using an enrichment step of 6 and 24 h, followed by selective plating of the enriched broth and selective plating of the immunomagnetic separation (IMS) product. Recovery results were evaluated and compared.Successful mPCR detection and isolation was obtained for non-stressed and cold-stressed STEC cells in minced beef and raw-milk cheese samples, except for serogroups O111 and SF O157. For freeze-stressed cells and sprouted seed samples, false negatives were often found. Isolation was better after 24 h-enrichment compared to 6 h-enrichment. IMS improved in some cases the isolation of non-stressed and cold-stressed cells belonging to serogroups O111 and O157 from minced beef and raw-milk cheese and freeze-stressed cells of all tested serogroups from minced beef.     

Direct detection of E. Coli O157:H7 in selected food systems by a surface plasmon resonance biosensor

The Spreeta^TM, surface plasmon resonance (SPR)-based biosensor, was used to detect Escherichia coli O157:H7 spiked in milk, apple juice and ground beef extract using specific antibodies. In the Spreeta^TM biosensor light from an LED is reflected off a gold surface, and the angle and intensity corresponding to the SPR minimum is measured and represented as a refractive index (RI) change corresponding to the antigen-antibody coupling at the sensor surface. Milk, apple juice, and ground beef patties spiked with E. coli O157:H7, at varying concentrations, were injected on the sensor surface immobilized with antibodies against the pathogen at a rate of 1 ml/min for a total of 2 min. The change in RI due to the binding of O157:H7 corresponding to each concentration was computed as an average of three replications over a 2 min interaction period. Assays were conducted at near real-time and results obtained after about 30 min of sample injection. Sensitivity of the E. coli O157:H7 assay was 10²-10³ colony forming unit (CFU)/ml. The biosensor assay was also specific to E. coli O157:H7 as other organisms (E. coli K12 and Shigella) did not produce any appreciable change in the sensogram. Further experiments are needed to establish well-defined methods for detecting other food-borne pathogens in more complex and specific food matrices.     

Multiplex PCR detection of Shiga toxin-producing Escherichia coli strains belonging to serogroups O157, O103, O91, O113, O145, O111, and O26 experimentally inoculated in beef carcass swabs, beef trim, and ground beef

Numerous foodborne outbreaks are attributed to Shiga toxin-producing Escherichia coli (STEC) and have been recognized for causing gastrointestinal disease in humans. Beef products have been considered the principal source of STEC. A multiplex PCR assay enabling simultaneous detection of STEC O103, O91, O113, O145, O111, O157, and O26 was developed and evaluated in artificially contaminated beef carcass swabs, beef trim, and ground beef after overnight enrichment. Individual serogroups were experimentally inoculated at low (1 to 10 CFU/ml) and high (11 to 100 CFU/ml) levels, and with a cocktail of strains belonging to two, four, and six serogroups. There was no significant difference in detecting single STEC strains under the different conditions. Only when strains were combined were there significant differences in detection of all cocktail isolates in some of the beef products. To address this issue, four serogroups were experimentally inoculated together at three different estimated levels (10, 10(2), and 10(3) CFU/ml) in all three beef products. Results yielded no significant difference in detecting STEC at the three inoculation levels (10, 10(2), and 10(3) CFU/ml) in trim and carcass swabs, but there was a significant difference in detecting STEC at the lowest levels (10 and 10(2) CFU/ml) in the 80:20 nonirradiated ground beef, and in the detection of STEC in irradiated ground beef. The findings from this study could provide industry and government agencies with a tool to evaluate the prevalence and incidence of STEC in beef products and their processing environments.     

Survival of Escherichia coli O157:H7 in channel catfish pond and holding tank water

Survival of Escherichia coli O157:H7 in channel catfish (Ictalurus punctatus), pond and holding tank water was investigated. Water from three channel catfish ponds was inoculated with ampicillin/nalidixic acid-resistant E. coli O157:H7 transformed with a plasmid encoding for green fluorescent protein at 10⁵, 10⁶, and 10⁷ CFU/ml. Samples were taken from surface, internal organs, and skin scrape of fish and pond water for E. coli O157:H7 enumeration on brain heart infusion (BHI) agar containing ampicillin and nalidixic acid. To determine the survival of E. coli O157:H7 in catfish holding tank water from two farmers markets, the water was inoculated with 10⁷E. coli O157:H7 CFU/ml. E. coli O157:H7 were detected by direct plating for 33 and 69 d in pond and holding tank water, respectively. A rapid decrease of the pathogen was observed in the first 2 weeks to reach 2 log CFU/ml. When E. coli O157:H7 was not recovered by direct plating, the pathogen was isolated by enrichment in TSB for approximately another 30 d from pond and holding tank water. The populations of E. coli O157:H7 found in the internal organs and skin scrape were 5.5 log and 2.5 log CFU/ml, respectively. E. coli O157:H7 from internal organs and water were recovered for at least 12 d. Results suggest that E. coli O157:H7 can survive in channel catfish pond and holding tank water and channel catfish may become a potential carrier of the pathogen.     

Inactivation of different strains of Escherichia coli O157:H7 in various apple ciders treated with dimethyl dicarbonate (DMDC) and sulfur dioxide (SO2) as an alternative method

Escherichia coli has been identified as the causative agent in numerous foodborne illness outbreaks associated with the consumption of fresh apple cider. Apple cider has a pH which is normally below 4.0 and would not be considered a medium capable of supporting the growth of foodborne pathogens. The association of unpasteurized apple cider with foodborne illness due to E. coli O157:H7 has however, led to increased interest in potential alternative methods to produce pathogen free cider. Apple cider was prepared from eight different apple cultivars, inoculated with approximately 10⁶–10⁷ CFU of three strains of E. coli O157:H7 per ml (933, ATCC 43889, and ATCC 43895) and tested to determine the effectiveness of sulfur dioxide (SO₂) and dimethyl dicarbonate (DMDC). Bacterial populations for treated and untreated samples were then enumerated by using non-selective media. Eight different ciders were treated with DMDC (125 and 250 ppm) and SO₂ (25, 50, 75, 100 ppm). Greater than a 5-log reduction was achieved at room temperature with 250 ppm of DMDC and 50 ppm of SO₂ after the incubation time of 6 h and 24 h, respectively. Addition of DMDC and/or SO₂ may offer an inexpensive alternative to thermal pasteurization for the production of safe apple cider for small apple cider producers.     

Effect of cooking procedures of kiymali pide,a traditional Turkish fast-food,on destruction of Escherichia coli O157:H7

The objective of the present study was to obtain data about cooking time and temperature of kiymali pide in the restaurants and to investigate thermal inactivation of E. coli O157:H7 during experimental kiymali pide making. A field study was conducted in randomly selected 23 of 87 pide restaurants. Processing parameters including oven temperature, cooking period and post-cooking temperature were determined. Kiymali pide samples were prepared using ground beef filling experimentally inoculated with E. coli O157:H7 (7.6 log₁₀ CFU/g). Pide samples were cooked at a conventional oven at 180 °C for 180, 240, 270, 300 and 330 s. Results of the current study suggest that cooking kiymali pide at 180 °C for at least 330 s (5.5 min) may provide sufficient food safety assurance (≥ 6 log₁₀ CFU/g) for E. coli O157:H7.     

Enhancing the thermal destruction of Escherichia coli O157:H7 in ground beef patties by trans-cinnamaldehyde

The effect of trans-cinnamaldehyde (TC) on the inactivation of Escherichia coli O157:H7 in undercooked ground beef patties was investigated. A five-strain mixture of E. coli O157:H7 was inoculated into ground beef (7.0 log CFU/g), followed by addition of TC (0, 0.15, and 0.3%). The meat was formed into patties and stored at 4 °C for 5 days or at −18 °C for 7 days. The patties were cooked to an internal temperature of 60 or 65 °C, and E. coli O157:H7 was enumerated. The numbers of E. coli O157:H7 did not decline during storage of patties. However, cooking of patties containing TC significantly reduced (P < 0.05) E. coli O157:H7 counts, by >5.0 log CFU/g, relative to the reduction in controls cooked to the same temperatures. The D-values at 60 and 65 °C of E. coli O157:H7 in TC-treated patties (1.85 and 0.08 min, respectively) were significantly lower (P < 0.05) than the corresponding D-values for the organism in control patties (2.70 and 0.29 min, respectively). TC-treated patties were more color stable and showed significantly lower lipid oxidation (P < 0.05) than control samples. TC enhanced the heat sensitivity of E. coli O157:H7 and could potentially be used as an antimicrobial for ensuring pathogen inactivation in undercooked patties. However detailed sensory studies will be necessary to determine the acceptability to consumers of TC in ground beef patties.     

Bacterial inactivation in fruit juices using a continuous flow Pulsed Light (PL) system

In this work, the susceptibility to pulsed light (PL) treatments of both a Gram-positive (L. innocua 11288) and a Gram-negative (E. coli DH5-??) bacteria inoculated in apple (pH = 3.49, absorption coefficient 13.9 cm^− 1) and orange juices (pH = 3.78, absorption coefficient 52.4 cm^− 1) was investigated in a range of energy dosages from 1.8 to 5.5 J/cm². A laboratory scale continuous flow PL system was set up for the experiments, using a xenon flash-lamp emitting high intensity light in the range of 100-1100 nm. The flashes lasted 360 ??s at a constant frequency of 3 Hz.The results highlighted how the lethal effect of pulsed light depended on the energy dose supplied, the absorption properties of liquid food as well as the bacterial strain examined. The higher the quantity of the energy delivered to the juice stream, the greater the inactivation level. However, the absorbance of the inoculated juice strongly influenced the dose deliver and, therefore, the efficiency of the PL treatment. Among the bacteria tested, E. coli cells showed a greater susceptibility to the PL treatment than L. innocua cells in both apple and orange juices. Following treatment at 4 J/cm², microbial reductions in apple and orange juices were, respectively, 4.00 and 2.90 Log-cycles for E. coli and 2.98 and 0.93 Log-cycles for L. innocua.Sublethally injured cells were also detected for both bacterial strains, thus confirming that membrane damage is an important event in bacterial inactivation by PL.     

Detection of E. coli O157:H7 in raw ground beef by Pathatrix™ immunomagnetic-separation, real-time PCR and cultural methods

Detection of Escherichia coli O157:H7 by conventional cultural methods can be difficult in food matrices with a high background flora such as raw ground beef. Raw ground beef samples, artificially contaminated separately with five strains of E. coli O157:H7 at low (~ 0.2 cfu/g) and high (~ 2 cfu/g) levels, were enriched by two enrichment protocols; buffered peptone water (BPW) at 37 °C for 5 h and 24 h and modified buffered peptone water with pyruvate (mBPWp) for 5 h at 37 °C followed by adding selective agents and incubating at 42 °C to 24 h. Detection of added E. coli O157:H7 by real-time PCR (RTiPCR) and recovery on isolation agars was performed before and after PATHATRIX™ immunomagnetic separation (IMS). RTiPCR detection and cultural recovery of inoculated E. coli O157:H7 after 5 h enrichment were poor at 0.21-0.24 cfu/g. The addition of IMS after 5 h enrichment did not improve RTiPCR detection but markedly improved recovery by culturing. By extending enrichment to 24 h, RTiPCR detection improved to 76% for either enrichment protocol without IMS. When 24 h enrichment was followed by IMS, RTiPCR detection was also further improved. Cultural recovery after 24 h enrichment was 56% and 84% without IMS and 100% and 92% after IMS for BPW and mBPWp respectively. Extended enrichment to 24 h followed by IMS was found to be sensitive and reliable for detection and cultural recovery from raw ground beef using either enrichment method.     

Incidence of Shiga toxin-producing Escherichia coli serogroups in ruminant's meat

To assess the presences of Escherichia coli, its serogroups, virulence factors and antibiotic resistance properties in ruminant's meat, a total of 820 raw meat samples were collected and then evaluated using culture, PCR and disk diffusion methods. Totally, 238 (29.02%) samples were positive for presence of Escherichia coli. All of the isolates had more than one virulence gene including Stx1, Stx2, eaeA and ehly. All investigated serogroups were found in beef and sheep and all except O145, O121 and O128 were found in goat. The O91, O113, O111, O103, O26 and O157 serogroups were found in camel. Totally, aadA1–blaSHV combination was the most predominant antibiotic resistance gene. The highest resistance of STEC strains was seen against penicillin while resistance to nitrofurantoin and ciprofloxacin was minimal. These findings showed that health care and meat inspection should be reconsidered in Iranian slaughterhouses and butchers.     

Comparison of the Diatheva STEC FLUO with BAX System Kits for Detection of O157:H7 and Non-O157 Shiga Toxin-Producing <Emphasis Type="Italic">Escherichia coli</Emphasis> (STEC) in Ground Beef and Bean Sprout Samples Using Different Enrichment Protocols

The aim of this study was to assess the performance of the Diatheva STEC FLUO and BAX System real-time PCR assays for detection of Shiga toxin-producing Escherichia coli (STEC) (stx1/stx2 and eae target genes) and O-group identification in ground beef and bean sprout samples. Ground beef (325 g or 25 g) and mung bean sprout (25 g) samples were inoculated with ~?10 CFU of the “top five” STEC (O157:H7, O26, O103, O111, and O145 as specified in EU regulation ISO13136:2012), enriched using different broths and incubation temperatures, and tested using the Diatheva and BAX real-time PCR assays. In ground beef, both molecular methods were able to detect the “top five” STEC, and lower Ct values were observed for the Diatheva kits compared to BAX System. The O111-contaminated samples gave negative results with both methods using mTSB?+?novobiocin for enrichment. In bean sprouts, both methods provided positive results, although detection was not possible using mTSB?+?acriflavin/cefsulodin/vancomycin for enrichment. In conclusion, the Diatheva and BAX methods detected the “top five” STEC in ground beef and bean sprouts when inoculated at low levels. Both assays provided equivalent results in terms of performance and reliability. Thus, the Diatheva kits are comparable to reference STEC-detection methods and could be used by the food industry to reliably detect the “top five” STEC.     

The effects of X-ray radiation on Escherichia coli O157:H7, Listeria monocytogenes, Salmonella enterica and Shigella flexneri inoculated on whole Roma tomatoes

In the last two decades several foodborne disease outbreaks associated with produce were reported. Tomatoes, in particular, have been associated with several multi-state Salmonella outbreaks. Inactivation of inoculated Escherichia coli O157:H7, Listeria monocytogenes, Salmonella enterica and Shigella flexneri on whole Roma tomato surfaces by X-ray at 0.1, 0.5, 0.75, 1.0, and 1.5 kGy was studied. The main purpose of this study was to achieve a 5 log reduction in consistent with the recommendations of the National Advisory Committee on Microbiological Criteria for Foods. Moreover, the effect of X-ray on inherent microflora (mesophilic counts, psychrotrophic counts and yeast and mold counts) of untreated and treated Roma tomatoes, during storage at ambient temperature (22 °C) for 20 days was also determined. Mixtures of three or two strains of each tested organism was spot inoculated (100 μl) onto the surface of Roma tomatoes (approximately 7–9 log per tomato), separately, and air-dried, followed by treatment with X-ray doses at 22 °C and 55–60% relative humidity. Surviving bacterial populations on tomato surfaces were evaluated using a nonselective medium (tryptic soy agar) with a selective medium overlay for each bacteria; E. coli O157:H7 (CT-SMAC agar), L. monocytogenes (MOA), and S. enterica and S. flexneri (XLD). Treatment with X-ray significantly reduced the population of the tested pathogens on whole Roma tomato surfaces, compared with the control. Approximately 4.2, 2.3, 3.7 and 3.6 log CFU reduction of E. coli O157:H7, L. monocytogenes, S. enterica and S. flexneri per tomato were achieved by treatment with 0.75 kGy X-ray, respectively. More than a 5 log CFU reduction per tomato was achieved at 1.0 or 1.5 kGy X-ray for all tested pathogens. Furthermore, treatment with X-ray significantly reduced the inherent microflora on Roma tomatoes. Inherent levels were significantly (p < 0.05) lower than the control sample throughout storage for 20 days.     

The DNA Sequence of the Escherichia coli O22 O-Antigen Gene Cluster and Detection of Pathogenic Strains Belonging to E. coli Serogroups O22 and O91 by Multiplex PCR Assays Targeting Virulence Genes and Genes in the Respective O-Antigen Gene Clusters

Multiplex polymerase chain reaction (PCR) assays were developed for detection of pathogenic strains belonging to Escherichia coli serogroups O22 and O91. The O-antigen gene cluster of E. coli O22 was sequenced to identify genes that could be employed as targets for serogroup-specific PCR assays. The wzx and wzy genes in the O-antigen gene clusters of E. coli O22 and E. coli O91 were selected as target genes. The assays were serogroup-specific when tested against 72 E. coli O22 strains and 57 E. coli O91 strains isolated from food, humans, and animals, representative strains belonging to 168 E. coli O serogroups and non-E. coli bacteria. Furthermore, 72 E. coli O22 strains and 57 E. coli O91 strains isolated from food, water, animals, and humans were tested by the PCR for the presence of six and 19 virulence genes, respectively, associated with pathogenic E. coli strains. Based on the PCR screening results, multiplex PCR assays targeting the O22 wzy gene and the cnf-1 and sfa genes in E. coli O22 and the O91 wzy gene, conserved sequences of stx ₁ and stx ₂ genes, and the astA and cdt-III genes in E. coli O91 were developed to detect and identify pathogenic strains belonging to serogroups O22 and O91. Furthermore, E. coli O22 and O91 were detected by multiplex PCR assays targeting the wzx or wzy genes and conserved sequences of the stx ₁ and stx ₂ genes in ground beef samples inoculated with approximately two colony-forming units (CFU)/25 g after 18-h enrichment. The results demonstrate that the E. coli O22 and O91 wzx and wzy gene sequences were specific for the respective serogroups and can be used as diagnostic markers for rapid identification of these serogroups as an alternative to serotyping. The multiplex PCR assays targeting the O22 and O91 wzx and wzy genes and virulence genes can be used to identify and to detect pathogenic strains of these serogroups in food and fecal samples. Mention of trade names or commercial products is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the US Department of Agriculture.     

Application of caffeine, 1,3,7-trimethylxanthine,to control Escherichia coli O157:H7

The objective of this research was to determine the effectiveness of caffeine on inactivation of Escherichia coli O157:H7 in brain heart infusion (BHI) broth. Overnight samples of five E. coli O157:H7 strains of (E0019, F4546, H1730, 944 and Cider) were used in this study. These strains were individually inoculated at an initial inoculum level of 2 log CFU/ml into BHI broth containing caffeine at different concentrations (0.00%, 0.25%, 0.50%, 0.75%, 1.00%, 1.25%, 1.50%, 1.75%, and 2.00%). Samples were then incubated at 37 °C for 24 h. Bacterial growth was monitored at different time intervals by measuring turbidity at 610 nm using a spectrophotometer. Results revealed that the addition of caffeine inhibited the growth of E. coli O157:H7. Significant growth inhibition was observed with concentration levels of 0.50% and higher. These results indicate that caffeine has potential as an antimicrobial agent for the treatment of E. coli O157:H7 infection and should be investigated further as a food additive to increase biosafety of consumable food products.     

Defining treatment conditions for pulsed electric field pasteurization of apple juice

The influence of temperature and the presence of N^α-lauroyl ethylester (ethyl lauroyl arginate, LAE) on the inactivation caused by continuous pulsed electric field treatments (PEF) in Escherichia coli O157:H7 suspended in apple juice have been investigated to define treatment conditions applicable at industrial scale that promote an equivalent safety level when compared with thermal processing. In the range of experimental conditions investigated (outlet temperature: 20-40 °C, electric field strength: 20-30 kV, treatment time: 5-125 μs) at outlet temperatures equal or lower than 55 ± 1 °C, the inactivation of E. coli O157:H7 treated in apple juice ranged from 0.4 to 3.6 Log₁₀ cycles reduction and treated in apple juice supplemented with LAE (50 ppm) ranged from 0.9 to 6.7 Log₁₀ cycles reduction.An empirical mathematical model was developed to estimate the treatment time and total specific energy input to obtain 5 Log₁₀ cycles reduction in the population of E. coli O157:H7 suspended in apple juice supplemented with 50 ppm of LAE at different electric field strengths and inlet temperatures. Treatment conditions established for E. coli O157:H7 were validated with other PEF resistant Gram-positive (Listeria monocytogenes, and Staphylococcus aureus) and Gram-negative (Salmonella enterica serovar Typhimurium) strains. When the treatment was applied to the apple juice, a treatment of 25 kV/cm for 63 μs corresponding with an outlet temperature of 65 °C and input energy of 125 kJ/kg was required to achieve more than 5 Log₁₀ cycles in the four strains investigated. The addition of LAE reduced the treatment time required to obtain an equivalent inactivation (> 5 Log₁₀ cycles) in the four microorganisms to 38.4 μs, the outlet temperature to 55 °C, and the input energy to 83.2 kJ/kg.