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The topic of food safety continues to receive increased attention and has ramifications on various human, environmental, policy, and economic levels worldwide. By garnering feedback from 30 food industry experts, this study was undertaken to identify the most critical issues facing the food industry in relation to food safety. According to expert opinion and after three rounds of Delphi inquiry, food contamination detection, outbreaks, and prevention along with governmental oversight, education for and communication with consumers and employees, and globalization were identified as the main areas at the forefront of food safety. Delphi and constant comparative research methods are explained, and suggestions on how to make meaning from the results to progress in this area are discussed. HIGHLIGHTS

Salsa-associated outbreaks, including the large multistate outbreak in the United States in 2008 caused by jalapeño and serrano peppers contaminated with Salmonella Saintpaul, have raised concerns about salsa as a potential vehicle for transmission. Despite these events, there has been relatively limited research on the potential growth of pathogenic bacteria in salsa. The aim of this study was to characterize the survival and growth of Salmonella, including the outbreak strain of Salmonella Saintpaul (E2003001236), in freshly made salsa and its main ingredients. Chopped tomatoes, jalapeño peppers, cilantro, and onions were tested individually or mixed according to different salsa recipes. Samples were inoculated with five Salmonella serotypes at 3 log CFU/g: Saintpaul (various strains), Typhimurium, Montevideo, Newport, or Enteritidis. Samples were then stored at room temperature (23°C) for up to 12 h or 3 days. The Salmonella Saintpaul levels reached approximately 9 log CFU/g after 2 days in tomato, jalapeño pepper, and cilantro. Growth was slower in onions, reaching 6 log CFU/g by day 3. Salsa recipes, with or without lime juice, supported the growth of Salmonella Saintpaul, and final levels were approximately 7 log CFU/g after 3 days at 23°C. In contrast, the counts of Salmonella Typhimurium, Salmonella Montevideo, Salmonella Newport, and Salmonella Enteritidis increased only 2 log CFU/g after 3 days in any of the salsas. Other Salmonella Saintpaul strains were able to grow in salsas containing 10% lime juice, but their final levels were less than 5 log CFU/g. These findings indicate the enhanced ability of the Salmonella Saintpaul outbreak strain to grow in salsa compared with other Salmonella strains. Recipe modifications including but not limited to adding lime juice (at least 10%) and keeping fresh salsa at room temperature for less than 12 h before consumption are strategies that can help mitigate the growth of Salmonella in salsa.

Bacteriophages are considered a potential antimicrobial food intervention due to their ability to inactivate target bacteria in different environments. This study was undertaken to isolate, characterize, and identify a mixture of phages that could be used for control of Listeria monocytogenes. A total of 37 phages were isolated from raw sewage sludge using L. monocytogenes serotypes 4b, 1/2a and 1/2b strains. The host range of these phages was determined by spot-testing. Ten lytic phages were selected for further evaluation with efficiency of plating (EOP) analysis and characterization by pulsed field gel electrophoresis (PFGE). According to the spot tests and EOP results, the isolated phages against L. monocytogenes 4b strains were found to be capable of lysing a wider range of ribotypes. Also, three phages specific for 1/2a and 1/2b serotypes had similar lysis capabilities over a range of target Listeria with high EOP values between 0.5 and 1.0. PFGE analysis identified 5 distinct restriction profiles of the 10 selected phages whose DNA was digested with either EcoRI or HinDIII. Transmission electron microscopy (TEM) pictures revealed that five selected phages belonged to the Myoviridae and one phage to the Podoviridae family. Based on screening results, a phage cocktail (LP1) comprised of six lytic phages was used to inactivate a mixed culture of five L. monocytogenes strains. LP1 was able to lyse bacterial cells within 30–60 min at multiplicity of infection values (MOI, PFU/CFU) of 1000 and maintained the target cells under detection levels for 24 to 48 h at 4, 10, and 30 °C. These results suggested the LP1 phage collection contained effective and strongly lytic phages that may serve as antibacterial preparation to control L. monocytogenes of different serotypes.

Current consumer preferences for both clean label food ingredients and convenience-based foods has provided a unique opportunity to explore the application of novel natural food preservatives in sous vide products. The anaerobic environment and relatively low thermal processing of the sous vide process creates a favorable environment for the survival, germination, and outgrowth of spore-forming bacterium Clostridium perfringens. The aim of this study was to identify effective novel natural ingredient formulations against C. perfringens and apply them within a vacuum-sealed sous vide chicken model exposed to abusive storage and chilling conditions. Among six commercial vinegar-based formulations, liquid vinegar with citrus extract (CE; 1.0%) and with lemon juice concentrate (LJC; 1.5%) were identified as the most effective at inhibiting three individual C. perfringens strains. Both reduced viable cell counts by 5 log CFU/mL (P

The presence of antibiotic-resistant bacteria in the food supply chain is a major concern. There is a great need to revalidate the effectiveness of the intervention strategies commonly used in food processing environments for multidrug-resistant strains. The purpose of this study was to determine the survival of antibiotic-resistant O26 and O103 Shiga-toxin producing Escherichia coli (STEC) strains to lactic acid (2.5, 3.5, 5%), sodium hypochlorite (0.2, 0.5, 1 ppm free chlorine), and heat (60, 61, 62.5 °C) treatments. Six strains from STEC serotypes O26 and O103 with three distinctive antibiotic-resistant profiles (susceptible, low and high resistance) were selected for the study. As expected, the rates of inactivation were faster at increasing biocide concentrations or temperature level. The Weibull model provided a better fit than the traditional log-linear model due to the appearance of nonlinear patterns. A 5% lactic acid treatment for 10 min reduced the population of all STEC strains by 5 log CFU/ml. Sodium hypochlorite treatment (1 ppm for 5 min) achieved a similar reduction. Thermal D values for O26 and O103 serotypes ranged from 0.37 to 2.09 and 0.37–1.71 min, respectively. No statistical differences (P > 0.05) in tolerance were found between the two STEC serotypes to the different treatments. The antibiotic profile was not related to the ability to tolerate any of the treatments. Only the susceptible strain showed a relative higher resistance to the lactic acid and lower resistance to sodium hypochlorite treatment. These results would contribute to assess the risk for the presence of STEC strains in food processing environments.

Hispanic style soft non-fermented cheeses, such as queso fresco (QF) have been linked to outbreaks and recalls. Salmonella is one of the main causes of these incidents. Due to lack of ripening or post-processing antimicrobial treatments, incorporating GRAS antimicrobials to production process may be a suitable approach to minimize microbial risk in QF. The aim of this study was to evaluate the efficiency of nisin (N), caprylic acid (CA) and trans-cinnamaldehyde (CN) as single or combined treatments to reduce Salmonella populations in QF during storage. Batches of QF were inoculated after curding with approx. 4 Log CFU/g of 5-strain cocktails of Salmonella and stored at 8 °C for 20 days. The final Salmonella counts in control samples ranged from 6.96 to 7.14 Log CFU/g. Application of CN at 0.6 g/kg inhibited Salmonella growth during storage, resulting in at least 3 Log CFU/g difference with the untreated controls (p

The use of bacteriophages is considered as a viable alternative to chemical antimicrobials against foodborne pathogens. The objective of this study was to develop a collection of lytic bacteriophages which will be able to infect different pathogenic Salmonella enterica serovars. Phages were isolated from animal feces and sewage samples, purified, characterized morphologically and by DNA fingerprinting, and host ranges were determined. Spot test and efficiency of plaquing (EOP) data indicated that two phages, SEA1 and SEA2 had the broadest host range against Salmonella among all isolated phages. SEA2 was highly efficient to infect S. Typhimurium DT104 (0.5–1 EOP value). Only phage SSA1 was able to infect S. Montevideo. Transmission electron microscopy (TEM) revealed the phages in the collection were mostly (4 out of 6) Siphoviridae, while SEA1 and SEA2 were Myoviridae T4-like phages. SEA1 and SEA2 had the largest genome sizes in the collection, 190 and 170 kb, respectively. Pulsed field gel electrophoresis (PFGE) analysis demonstrated distinct digestion profiles with Eco RI for phages SSA1, STD3, STE3 and STF1. However, SEA1 and SEA2 shared a similar restriction enzyme (RE) digestion pattern with same morphotype, but distinct profiles in lysing Salmonella strains. These anti- Salmonella phages were highly host specific with few exceptions of lytic phages that were able to infect a wide variety of Salmonella . These phages have potential for use in applications controlling Salmonella on different matrices.

Electrochemically activated water (ECAW), also known as electrolyzed water, and ozonized water are typically effective in inactivating bacteria, but their generation typically uses high current and voltage. A few simpler antimicrobial technologies that are also based on the application of a mild electrical current have been recently marketed to food retail and service customers claiming to have sanitizing properties for controlling bacteria. The objective of this study was to determine the sanitizing effect of some of these commercial technologies on Escherichia coli O157:H7, Listeria monocytogenes, and Salmonella enterica and compare them with sterile water, generated ECAW generated with a pilot size electrolyzing unit, and salt solutions sprayed using commercial device sprays. A concentration of 100 mg/L ECAW had sanitizing effects of at least 5 log CFU/mL reductions on liquid culture and more than 4 log CFU/coupon reductions for E. coli O157:H7, L. monocytogenes and Salmonella dried on stainless steel surface, respectively. No bacterial cells were detected by direct plate counting post-ECAW treatment. In contrast, the treatment of liquid cultures with any of the commercial technologies tested resulted in non-significant bacterial cell reductions greater than 0.5 log CFU/mL. Similarly, when cells had been dried on metal surfaces and treated with any of the water generated with those technologies, no reductions were observed. When the manufacturer's instructions were followed, the reduction of cells on surface was largely due to the physical removal by cloth-wiping after water fraction application. These results indicate that treatment with any of these portable technologies had no noticeable antimicrobial activity. These results would be helpful for guiding consumers when choosing a right sanitization to ensure food safety.

Lettuce and spinach are increasingly implicated in foodborne illness outbreaks due to contamination by Escherichia coli O157:H7. While this bacterium has been shown to colonize and survive on lettuce leaf surfaces, little is known about its interaction with the roots of growing lettuce plants. In these studies, a microarray analyses, mutant construction and confocal microscopy were used to gain an understanding of structure and function of bacterial genes involved in the colonization and growth of E. coli O157:H7 on lettuce roots. After three days of interaction with lettuce roots, 94 and 109 E. coli O157:H7 genes were significantly up- and down-regulated at least 1.5 fold, respectively. While genes involved in biofilm modulation (ycfR and ybiM) were significantly up-regulated, 40 of 109 (37%) of genes involved in protein synthesis were significantly repressed. E. coli O157:H7 was 2 logs less efficient in lettuce root colonization than was E. coli K12. We also unambiguously showed that a ΔycfR mutant of E. coli O157:H7 was unable to attach to or colonize lettuce roots. Taken together these results indicate that bacterial genes involved in attachment and biofilm formation are likely important for contamination of lettuce plants with Shiga toxin-producing E. coli strains.

In this study, saxitoxin dihydrochloride in skim milk was reacted with sodium hydroxide and hydrogen peroxide to yield nontoxic 8-amino-6-hydroxymethyl-iminopurine-3(2H)-propionic acid (AHIPA), which was quantified by fluorescence spectroscopy using excitation and emission wavelengths of 330 and 425 nm, respectively. Samples of saxitoxin dihydrochloride (in 20% ethanol, vol/vol) were used as controls. The limits of detection of AHIPA, based on the concentration of saxitoxin prior to inactivation, were 5 and 10 μg/ml for the control and skim milk, respectively. These values are considerably below the concentration of saxitoxin that corresponds to the lethal dosage of 1 mg for an adult of average weight (70 kg). The inactivation of saxitoxin proceeded at a lower rate in skim milk than in the control, as its reaction rate constant was only 0.004 min(-1) compared with 0.011 min(-1) for the control. We were unable to detect AHIPA in 2% milk contaminated with saxitoxin because of possible interference from what we believed were products of secondary reactions involving milk fat and sodium hydroxide. Our results also indicated that the conversion of saxitoxin to AHIPA increased initially with temperature up to 40°C but decreased thereafter. We observed a decrease in the formation of AHIPA when the concentration of hydrogen peroxide was increased except at 22°C, where there was an initial increase in AHIPA formation between 1.2 and 2.4 mg/ml hydrogen peroxide but its formation decreased thereafter.

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 has been recognized as a major foodborne pathogen responsible for frequent gastroenteritis outbreaks. Phages and essential oils can be used as a natural antimicrobial method to reduce bacterial pathogens from the food supply. The objective of this study was to determine the effect of a bacteriophage cocktail, BEC8, alone and in combination with the essential oil trans-cinnameldehyde (TC) on the viability of a mixture of EHEC O157:H7 strains applied on whole baby romaine lettuce and baby spinach leaves. The EHEC O157:H7 strains used were Nal(R) mutants of EK27, ATCC 43895, and 472. Exponentially growing cells from tryptic soy (TS) broth cultures were spot inoculated on leaves and dried. EHEC cells were placed at low, medium, and high inoculum levels (10(4), 10(5), and 10(6) CFU/mL, respectively). Appropriate controls, BEC8 (approx. 10(6) PFU/leaf), and TC (0.5% v/v) were applied on treated leaves. The leaves were incubated at 4, 8, 23, and 37 °C in Petri dishes with moistened filter papers. EHEC survival was determined using standard plate count on nalidixic acid (50 μg/mL) Sorbitol MacConkey agar. No survivors were detected when both leaves were treated with BEC8 or TC individually at low inoculum levels after 24 h at 23 and 37 °C. When the EHEC inoculum size increased and/or incubation temperature decreased, the efficacy of BEC8 and TC decreased. However, when the two treatments were combined, no survivors were detected after 10 min at all temperatures and inoculum levels on both leafy greens. These results indicated that the BEC8/TC combination was highly effective against EHEC on both leafy greens. This combination could potentially be used as an antimicrobial to inactivate EHEC O157:H7 and reduce their incidence in the food chain.

Foods have been identified as a potential target for bioterrorism due to their essential nature and global distribution. Foods produced in bulk have the potential to have large batches of product intentionally contaminated, which could affect hundreds or thousands of individuals. Bacillus anthracis spores are one potential bioterrorism agent that may survive pasteurization and remain viable throughout the shelf life of fruit juices and cause disease if consumed. This project examined B. anthracis spore survival in orange, apple, and grape juices, as well as wine. Samples of beverages were inoculated with spores of two nonpathogenic B. anthracis strains at approximately 10(6) CFU/ml, and the spore count was determined periodically during storage for 30 days at 4°C. After this time, the counts of survival spores never declined more than 1 log CFU/ml in any of the beverage types. These results indicate that spores can survive, with little to no loss in viability, for at least a month in fruit juices and wine.

This research was conducted to study the growth of Listeria monocytogenes inoculated on frankfurters stored at different conditions as a basis for a safety-based consume by shelf life date label. Three L. monocytogenes strains were separately inoculated at 10 to 20 CFU/cm2 onto frankfurters that were previously formulated with or without high pressure and with or without added 2% potassium lactate (PL) and 0.2% sodium diacetate (SD). Inoculated frankfurters were air or vacuum packaged; stored at 4, 8, or 12 degrees C; and L. monocytogenes and psychrotrophic plate counts were determined for 90, 60, and 45 days, respectively, or until the stationary phase was reached. The data (log CFU per square centimeter versus time) were fitted using the Baranyi-Roberts model to determine maximum growth rates and lag-phase time. The maximum growth rates and the lag time under each growth condition were used to calculate the time to reach 100-fold the initial Listeria population. In frankfurters lacking PL and SD, the count of all strains increased by 2 log after 18 to 50 days at 4 degrees C and 4 to 13 days at 8 degrees C. The growth was inhibited at 4 and 8 degrees C in frankfurters containing PL and SD, but one ribotype was capable of growing, with the time to reach 100-fold the initial Listeria population ranging from 19 to 35 days at 12 degrees C. In most cases, the time to reach 100-fold the initial Listeria population of L. monocytogenes was significantly longer in vacuum-packaged frankfurters as compared with air-packaged samples. Inclusion of PL and SD also inhibited the growth of psychrotrophs, but at all temperatures the psychrotrophic plate counts were greater than 4 log CFU/cm2 at the end of the experiments. These results indicated that despite the use of antimicrobials, certain L. monocytogenes strains could be capable of growing under storage-abuse conditions. Growth kinetics data could be useful for establishing a shelf life date label protocol under different handling scenarios.

The growth variability of three Listeria monocytogenes ribotypes in ready-to-eat (RTE) sliced uncured turkey breast and cured ham was studied under storage conditions that RTE foods are likely to encounter. Three product treatments studied were: (1) a control; (2) a formulation subjected to high pressure processing to reduce initial microbial load (HPP); (3) a formulation containing 2.0% potassium lactate and 0.2% sodium diacetate (PL/SD). After separate inoculation with individual L. monocytogenes ribotypes and packaging each treatment under air and vacuum, the packages were stored at 4, 8, or 12 degrees C and the counts of L. monocytogenes and psychrotrophic bacteria (PPC) were determined for several weeks. The Baranyi model was used to estimate lag times and growth rates. Significant effect of strain difference was noted in both sliced products (P

This study compared the performance of four primary mathematical models to study the growth kinetics of Listeria monocytogenes ribotypes grown at low temperature so as to identify the best predictive model. The parameters of the best-fitting model were used to select the fastest growing strains with the shortest lag time and greatest growth rate. Nineteen food, human and animal L. monocytogenes isolates with distinct ribotype were grown at 4, 8, and 12 degrees C in tryptic soy broth and slurries prepared from cooked uncured sliced turkey breasts (with or without potassium lactate and sodium diacetate, PL/SD) and cooked cured frankfurters (with or without PL/SD). Separate regressions were performed on semi-logarithm growth curves to fit linear (based on Monod) and non-linear (Gompertz, Baranyi-Roberts, and Logistic) equations and performance of each model was evaluated using an F-test. No significant differences were found in the performance of linear and non-linear models, but the Baranyi model had the best fit for most growth curves. The maximum growth rate (MGR) of Listeria strains increased with the temperature. Similarly MGR was found significantly greater when no antimicrobials were present in the formulation of turkey or frankfurter products. The variability in lag times and MGRs in all media as determined by the Baranyi model was not consistent among strains. No single strain consistently had the fastest growth (shortest lag time, fastest MGR, or shortest time to increase 100-fold), but nine strains were identified as fastest growing strains under most growth conditions. The lack of association between serotype and fastest strain was also observed in the slurry media study. The fastest growing strains resulting from this study can be recommended for future use in L. monocytogenes challenge studies in delicatessen meat and poultry food matrices, so as to develop conservative pathogen growth predictions.

Microbiological analyses of fruits and vegetables produced by farms in Minnesota and Wisconsin were conducted to determine the prevalence of Escherichia coli in pre-harvest fruits and vegetables. During the 2003 and 2004 harvest seasons, 14 organic (certified by accredited organic agencies), 30 semi-organic (used organic practices but not certified) and 19 conventional farms were sampled to analyze 2029 pre-harvest produce samples (473 organic, 911 semi-organic, 645 conventional). Before each harvest season, a farmer survey was conducted to collect relevant information on farm management practices that might affect the risk of E. coli contamination in fresh produce. The use of animal wastes for fertilization of produce plants increased the risk of E. coli contamination in organic (OR=13.2, 95% CI=2.2-61.2, P-value

In the current study, temperature dependent growth of Listeria monocytogenes strain H7776, with an initial inoculum level of approximately 0.1 CFU/mL or g, was modeled based on "time to detect" (TTD) using the Arrhenius equation. The activation energies (E(a)) obtained from specific growth rate and lag phase duration in tryptic soy broth (TSB) and frankfurters were compared. At 4 degrees C, the TTD for L. monocytogenes was 217 h in TSB and 48 h in portions of frankfurters. The TTD decreased as temperature increased from 4 to 36 degrees C in both culture media and frankfurters, and this relationship was modeled using the Arrhenius equation. Based on this model, the E(a) values for the TTD in TSB and frankfurters averaged 22.7 and 18.7 kcal/mol, respectively, and were not significantly different (p0.05). Linear regression was performed on the exponential part of the growth curve to evaluate specific growth rate constants at each temperature using the Monod model. The E(a) values were also calculated based on the specific growth rate and the lag phase of L. monocytogenes in TSB and frankfurters incubated in the same range of temperatures. The average E(a) values for the specific growth rates and the lag phase durations in TSB cultures were 19 and 21 kcal/mol, respectively. In frankfurters, the average E(a) values were slightly greater for both specific growth rate and lag phase duration (29 and 35 kcal/mol, respectively), but these values were not significantly different from the E(a) calculated for the TTD in each medium. These results indicate that the TTD concept can be used to develop and validate safety-based shelf life models.

Microbiological analyses of fruits and vegetables produced by farms in Minnesota and Wisconsin were conducted to determine coliform and Escherichia coli counts and the prevalence of E. coli, Salmonella, and E. coli O157:H7. During the 2003 and 2004 harvest seasons, 14 organic farms (certified by accredited organic agencies), 30 semiorganic farms (used organic practices but not certified), and 19 conventional farms were sampled to analyze 2,029 preharvest produce samples (473 organic, 911 semiorganic, and 645 conventional). Produce varieties included mainly lettuces, leafy greens, cabbages, broccoli, peppers, tomatoes, zucchini, summer squash, cucumber, and berries. Semiorganic and organic farms provided the majority of leafy greens and lettuces. Produce samples from the three farm types had average coliform counts of 1.5 to 2.4 log most probable number per g. Conventional produce had either significantly lower or similar coliform populations compared with the semiorganic and organic produce. None of the produce samples collected during the 2 years of this study were contaminated with Salmonella or E. coli O157:H7. E. coli contamination was detected in 8% of the samples, and leafy greens, lettuces, and cabbages had significantly higher E. coli prevalence than did all the other produce types in both years for the three farm types. The prevalence of E. coli contamination by produce type was not significantly different between the three farm types during these 2 years, with the exception of organic leafy greens, in which E. coli prevalence was one-third that of semiorganic leafy greens in 2003. These results indicate that the preharvest microbiological quality of produce from the three types of farms was very similar during these two seasons and that produce type appears to be more likely than farm type to influence E. coli contamination.

Samples were collected from 26 organic and conventional farms and 12 county fairs in Minnesota during 2001 and 2002 to identify the presence of Escherichia coli O157. Immunomagnetic separation was used for isolation of E. coli O157. Isolates were further characterized by the presence of virulence marker genes (stx1, stx2, eaeA, E-hly, katP, etpD, and espP), antimicrobial susceptibility profiles, and genotypes. During 2001, E. coli O157 was isolated from 16 (5.2%) of 305 fecal samples and from 7 (36.8%) of 19 farms. During 2002, E. coli O157 was isolated from 6 (4.5%) of 132 fecal samples from weaned calves at 4 (23.5%) of 17 farms. During 2001 and 2002, cattle manure samples were collected from 12 county fairs, and E. coli O157 was isolated from 19 (11%) of 178 samples and 9 (75%) of 12 county fairs. Among 40 E. coli O157 isolates, 17 isolates (43%) had both the stx1 and stx2 genes, and 21 strains (53%) had the stx2 gene only. Thirteen percent of O157 isolates were resistant to tetracycline, and 25% were resistant to sulfadimethoxine. Heterogeneity of E. coli O157 strains was demonstrated by the presence of 22 different pulsed-field gel electrophoresis (PFGE) patterns. Four PFGE patterns matched those of isolates previously found in humans. The presence of E. coli O157 at county fairs suggests the potential for transmission to the public, who may have contact with cattle or their environment.

The effect of lactose at the concentration typically found in milk (134 mM) on the ability of ricin to inhibit protein synthesis in HeLa cells was studied. Ricin (0.001 to 300 mg/ml) that was either not treated or treated with 134 mM lactose was added to test tubes containing 1 ml of HeLa cells (approximately 3 | 10 5 cells in a low-leucine medium). After 2 h of incubation at 37uC, 0.5 mCi of L-[U- 14 C]-leucine was added to each tube and incubated for another 60 min. The cells were harvested by centrifugation and lysed, and cellular proteins were separated. The amount of radioactivity incorporated into the proteins was determined by liquid scintillation. The biological activity of ricin, i.e., the amount of radioactivity in a sample relative to that of the control (cells not treated with ricin), was calculated for each treatment. The inhibitory effect of 134 mM lactose on the biological activity of ricin was only significant at concentrations of ricin below 1 mg/ml. At higher ricin concentrations, the effect of 134 mM lactose decreased as the concentration of ricin increased, resulting in an increase in the inhibition of proteins synthesis. Our results also indicated that bovine milk, when used in place of 134 mM lactose, was more effective for reducing the activity of ricin at concentrations below 1 mg/ml but was ineffective against ricin concentrations greater than 1 mg/ml. These results suggest that milk may not protect against ricin intoxication at the concentration (0.89 mg/ml) equivalent to the lowest limit of its 50% lethal dose for a 20-kg child consuming 225 ml (8 oz) of milk. Ricin is a globular protein that can be used as a warfare

Several outbreaks of Escherichia coli O157:H7 infections have been associated with contaminated alfalfa seeds. A recently isolated E. coli strain Hu194 was capable of inhibiting 22 strains of E. coli O157:H7 and this inhibition was mediated by the production of a colicin named Hu194. The objectives of this study were to test the efficacy of treating alfalfa seeds with colicin Hu194 against E. coli O157:H7 strains, and to characterize this antimicrobial protein. Significant reductions (approximately 5 log CFU ml-1) in the viable cell counts of strains 43890 and 43895 were observed after 1-day incubation with semi-crude colicin, and after 2 days for strain 3081. Strain 43890 was successfully eliminated (5 log CFU g-1) from inoculated alfalfa seeds after soaking in a colicin suspension at a concentration of 10,000 AU/g. Treatment of alfalfa seeds inoculated with strains 43895 and 3081 required 20-fold higher concentrations of colicin Hu194 to achieve as much as 3 log CFU g-1 reductions. The genes encoding the colicin Hu194 operon were located on a 6 kb plasmid, and the sequence analysis revealed that this colicin was an E-type DNAse. From the sequence data, the estimated molecular masses of colicin Hu194, its immunity protein and lysis protein were 61.3, 10.0 and 4.8 kDa, respectively. Based on DNA and protein sequence comparisons with other E-type colicin, colicin Hu194 belonged to the type E2-colicin cluster. However, cross-immunity tests between E-group colicins suggested that Hu194 colicin was divergent from the previously characterized E2 colicins.

A previously identified set of anti-Escherichia coli O157:H7 colicinogenic E. coli were characterized to assess the suitability of these isolates as a preharvest food safety intervention in cattle. This collection of 23 E. coli strains were screened for virulence factors, antibiotic resistance, type of colicin(s) present, and their ability to inhibit other pathogenic E. coli. With the use of PCR, pathogen genes were detected in six of the 23 colicinogenic E. coli. When the nonpathogenic strains were assessed for antibiotic resistance, four strains showed resistance to at least one antibiotic. The remaining set of 14 strains were evaluated for the presence of previously identified colicins. Seven colicins (B, El, E2/E7, E7, Ia/Ib, K, and M) were detected. One half of the strains possessed multiple types of colicins. The most commonly detected colicins were B, E2/E7, and M, which were found in six strains each. DNA sequencing was also performed in order to classify the E2/E7 colicins separately from E7 colicins. The 14 colicinogenic E. coli also were evaluated for their ability to inhibit 10 different non-O157 pathogenic E. coli. Six of the colicinogenic E. coli were capable of inhibiting all 10 pathogens, and the remaining eight strains could each inhibit between six to eight of the pathogenic E. coli. This strain collection has great potential for inhibiting E. coli O157:H7 in cattle.

Grain feeding seems to promote the growth and acid resistance of Escherichia coli in fattening beef cattle, and acid-resistant E. coli are more likely to survive the human gastric stomach. When cattle were fed hay for only five days, the number and acid resistance of E. coli decreased dramatically.

When stationary phase Escherichia coli O157:H7 cells were subjected to extreme acid shock (pH 2·0, 6 h, 37°C) cell survival was as great as 10%, but culture conditions greatly affected the acid resistance. Anaerobic cultures were more resistant to extreme acid shock if the glucose concentration of the growth medium was high, acids accumulated, and pH declined. By varying pH and acetate concentration, it was possible to demonstrate a high correlation (R2=0·86) between undissociated acetate and extreme acid resistance. Because dissociated acetate and extreme acid resistance were poorly correlated (R2