Multiple‐locus variable‐nucleotide tandem repeat subtype analysis implicates European starlings as biological vectors for Escherichia coli O157:H7 in Ohio,USA

Aims: To provide molecular epidemiological evidence of avian transmission of Escherichia coli O157:H7 between dairy farms in Ohio, this study was designed to identify genetic relatedness between isolates originating from bovine faecal samples and intestinal contents of European starlings captured on these farms. Methods and Results: During a three‐year period (2007–2009), cattle (n = 9000) and starlings (n = 430) on 150 different dairy farms in northern Ohio were sampled for the presence of E. coli O157:H7. Isolates were subjected to multiple‐locus variable‐nucleotide tandem repeat analysis (MLVA). Distinct allelic groups were identified on most farms; however, isolates clustering into three MLVA groups originated from both cattle and birds on different farms. Conclusions: Sharing of indistinguishable epidemiologically linked E. coli O157 MLVA subtypes between starlings and cattle on different farms supports the hypothesis that these birds contribute to the transmission of E. coli O157:H7 between dairy farms. Significance and Impact of Study: A continued need exists to identify and to improve preharvest measures for controlling E. coli O157:H7. Controlling wildlife intrusion, particularly European starlings, on livestock operations, may be an important strategy for reducing dissemination of E. coli O157:H7 between farms and thereby potentially decreasing the on‐farm prevalence of E. coli O157:H7 and enhancing the safety of the food supply.     

Shedding of Escherichia coli O157:H7 in Dairy Cattle Housed in a Confined Environment following Waterborne Inoculation

A study of Escherichia coli O157:H7 transmission and shedding was conducted with bull calves housed in individual pens within a confined environment. For comparative purposes, the numbers and duration of E. coli O157:H7 shedding in naturally infected calves were monitored after a single purchased calf (calf 156) tested positive prior to inoculation. During the next 8 days, the calves in adjacent pens and a pen directly across a walkway from calf 156 began to shed this serotype O157:H7 strain. Five of the eight calves in this room shed this O157:H7 strain at some time during the following 8 weeks. The numbers of E. coli O157:H7 isolates shed in these calves varied from 60 to 10⁵ CFU/g of feces, and the duration of shedding ranged from 17 to >31 days. The genomic DNAs from isolates recovered from these calves were indistinguishable when compared by using XbaI digestion and pulsed-field gel electrophoresis. Inoculation of calves with 1 liter of water containing ca. 10³ to 10⁴ CFU of E. coli O157:H7/ml resulted in shedding in 10 of 12 calves (trial 1, 4 of 4 calves; trial 2, 6 of 8 calves). The inoculated calves shed the inoculation strain (FRIK 1275) as early as 24 h after administration. The duration of shedding varied from 18 to >43 days at levels from 10² to 10⁶ CFU/g of feces. The numbers of doses necessary to initiate shedding varied among calves, and two calves in trial 2 never shed FRIK 1275 after four doses (ca. 10⁶ CFU per dose). Results from this study confirm previous reports of animal-to-animal and waterborne dissemination of E. coli O157:H7 and highlight the need for an effective water treatment to reduce the spread of this pathogen in cattle.     

Impact of Dexamethasone-Induced Immunosuppression on the Duration and Level of Shedding of Escherichia coli O157:H7 in Calves

The goal of this study was to determine whether immunosuppression plays a role in the level and duration of fecal shedding of Escherichia coli O157. Immunosuppression was induced in calves by administering dexamethasone. Six 1-week-old Holstein bull calves were injected intramuscularly with dexamethasone and orally inoculated with 10⁹ CFU of a mixture of three nalidixic-acid resistant strains of E. coli O157:H7. Five 1-week-old Holstein bull calves that were given the same oral inoculation of E. coli O157:H7, but not the dexamethasone injections, served as controls. All calves were examined daily and fecal samples were collected three times a week for detection and enumeration of the nalidixic-acid resistant E. coli O157. Four weeks after the last calf stopped shedding, all calves were necropsied and samples from the gastrointestinal tract were taken for the detection of the nalidixic-acid resistant E. coli O157. Dexamethasone-injected calves shed at higher levels (P = 0.04) on days 4 and 7 postinoculation, but not thereafter. None of the samples collected at necropsy were positive for E. coli O157. Data from this study suggest that there may be a time-dependent relationship between dexamethasone immunosuppression and the fecal concentration of E. coli O157 but that transient immunosuppression does not appear to prolong shedding of E. coli O157.     

Effect of Dietary Stress on Fecal Shedding of Escherichia coli O157:H7 in Calves

Two groups of calves were subjected to dietary stress by withholding of food beginning 1 or 14 days after inoculation with 10¹⁰ CFU of Escherichia coli O157:H7. Following treatment, neither group had a significant increase in fecal shedding of E. coli O157:H7. A third group of calves had food withheld for 48 h prior to inoculation with 10⁷ CFU of E. coli O157:H7. These calves were more susceptible to infection and shed significantly more E. coli O157:H7 organisms than calves maintained on a normal diet.     

Variable agronomic practices, cultivar, strain source and initial contamination dose differentially affect survival of Escherichia coli on spinach

Aims: Greenhouse and field trials were conducted under different agronomic practices and inoculum doses of environmental Escherichia coli and attenuated E. coli O157:H7, to comparatively determine whether these factors influence their survival on leaves and within the rhizosphere. Methods and Results: Hydroponic conditions: E. coli spray‐inoculated at log 4 CFU ml^?1 was recovered from leaf surfaces at a mean population of 1·6 log CFU g^?1 at 15 days. E. coli O157:H7 sprayed at log 2 or 4 CFU ml^?1 levelled off on spinach leaf surfaces at a mean average population of 1·4 log CFU g^?1 after 14 days, regardless of initial dose. Quantitative recovery was inconsistent across leaf developmental age. Field conditions: Average populations of E. coli O157:H7 spray‐inoculated at log 1·45 or 3·4 CFU m^?2 levelled off at log 1·2 CFU g^?1 over a 14‐day period. Pathogen recovery from leaves was inconsistent when compared to regularly positive detection on basal shoot tissue. Pathogen recovery from soil was inconsistent among sampling locations. Moisture content varied up to 40% DW and was associated with 50% (P < 0·05) decrease in positive locations for E. coli O157:H7 but not for E. coli. Conclusions: Overall, similar populations of environmental E. coli and E. coli O157:H7 were recovered from plants despite differences in inoculum dose and agronomic conditions. Strain source had a significant impact on the quantitative level and duration of survival on leaves and in soil. Water availability appeared to be the determinant factor in survival of E. coli and E. coli O157:H7; however, E. coli showed greater environmental fitness. Significance and Impact of the Study: Persistence of surrogate, indicator E. coli and E. coli O157:H7, irrespective of variable growing conditions in spinach is predominantly limited by water availability, strain source and localization within the plant. These findings are anticipated to ultimately be adopted into routine and investigative pathogen testing protocols and mechanical harvest practices of spinach.     

Cattle,weather and water: mapping Escherichia coli O157:H7 infections in humans in England and Scotland

Entero‐haemorrhagic Escherichia coli O157:H7 is a zoonotic pathogen, responsible for a relatively small number of food poisoning and illness outbreaks each year, when compared with other food‐borne bacteria capable of causing infections in the population. Nevertheless, E. coli O157:H7 is a bacterial pathogen associated with severe human illnesses including bloody diarrhoea and haemolytic uremic syndrome occurring in both outbreak and sporadic settings. In England and Wales approximately 1% of all laboratory‐confirmed cases of food poisoning are the result of E. coli O157:H7; however, in Scotland this figure increases to 3%. When the size of the population is taken into account and the rate of E. coli O157:H7 confirmed cases per 100 000 population is examined, the rate of E. coli 0157:H7 infections in Scotland is much greater than England and Wales. The routes of transmission have changed over time, with new routes of transmission such as farm visits emerging. The prevalence of E. coli O157:H7 has a seasonal dependency, with greater faecal shedding of the organism in the warmer months; this is directly mirrored in the increased reporting of E. coli O157:H7 infection among hospitalized patients. This review attempts to suggest why this phenomenon occurs, paying particular attention to weather, animal movement and private water supplies.     

Impact of the Direct Application of Therapeutic Agents to the Terminal Recta of Experimentally Colonized Calves on Escherichia coli O157:H7 Shedding

Enterohemorrhagic Escherichia coli O157:H7 is an important intestinal pathogen of humans with a main reservoir of domesticated ruminants, particularly cattle. It is anticipated that the risk of human infection can be reduced by controlling the organism within its reservoir hosts. Several options for the control of E. coli O157:H7 in cattle have been proposed, but none have been demonstrated to be successful in the field. Here we describe a novel experimental method, based on the terminal-rectum-restricted colonization described previously, to eliminate fecal carriage of E. coli O157:H7. In experimentally challenged calves, direct application to the rectal mucosa of either of two therapeutic agents, polymyxin B or chlorhexidine, greatly reduced bacterial shedding levels in the immediate posttreatment period. The most efficacious therapeutic agent, chlorhexidine, was compared in orally and rectally challenged calves. The treatment eliminated high-level shedding and reduced low-level shedding by killing bacteria at the terminal rectum. A rapid-detection system based on the ability to identify E. coli O157:H7 from swabs of the rectal mucosa was also assessed. This test was sufficiently sensitive to identify high-level bacterial carriage. Thus, a combination of the detection method and treatment regimens could be used in the field to eliminate high-level fecal excretion of E. coli O157:H7, so greatly reducing its prevalence within this host and the risk of human infection.     

Quantification of Survival of Escherichia coli O157:H7 on Plants Affected by Contaminated Irrigation Water

Enterohemorrhagic E. coli O157: H7 (EHEC) is a major foodborne pathogen capable of causing diarrhea and vomiting, with further complications such as hemolytic‐uremic syndrome (HUS). The aim of this study was to use the real‐time PCR method to quantify the survival of Escherichia coli O157:H7/pGFP in phyllosphere (leaf surface), rhizosphere (volume of soil tightly held by plant roots), and non‐rhizosphere soils (sand and clay) irrigated with contaminated water and compare the results obtained between real‐time PCR method and conventional plate counts. The real‐time PCR probe was designed to hybridize with the (eae) gene of E. coli O157:H7. The probe was incorporated into real‐time PCR containing DNA extracted from the phyllosphere, rhizosphere, and non‐rhizosphere soils irrigated with water artificially contaminated with E. coli O157:H7. The detection limit for E. coli O157:H7 quantification by real‐time PCR was 2.3 × 10³ in the rhizosphere and phyllosphere samples. E. coli O157:H7 survived longer in rhizosphere soil than the non‐rhizosphere soil. The concentration of E. coli O157:H7/pGFP in rhizosphere soils was ≥ 10⁴ CFU/g in both soils at day 12 based on both plate count and real time PCR, with the clay soil significantly (P = 0.05) higher than the sandy soil. This data showed that E. coli O157H:7 can persist in the environment for more than 50 d, and this may pose some risk for both animal and human infection and provides a very significant pathway for pathogen recontamination in the environment.     

Prevalence of Escherichia coli O157:H7 on hides and faeces of ruminants at slaughter in two major abattoirs in Nigeria

Aim: To determine the occurrence of Escherichia coli O157: H7 in hides and faeces of slaughtered ruminants in Nigeria. Methods and Results: A total number of 320 animals were sampled from January to December covering the wet and harmattan seasons. Samples were obtained from the hides and faeces of animals at slaughter. The ISO (ISO 16654:2001, Microbiology of food and animal feedingstuffs – horizontal method for the detection of Escherichia coli O157) method for enrichment and isolation of E. coli O157 incorporating selective enrichment using modified tryptone soya broth with novobiocin (mTSBn),immunomagnetic separation and plating on sorbitol‐MacConkey agar with cefixime tellurite (CT‐SMAC) was used. Overall cattle had a prevalence rate of 49·4% followed by sheep and goats with rates of 6·3% and 2·5%, respectively. There was a significant difference in carriage of E. coli O157 among two different cattle breeds. Conclusions: The prevalence of E. coli O157: H7 is substantial from two abattoirs in the country. The carriage and shedding of E. coli O157: H7 did not differ with season but differed among groups of ruminants and among breeds of cattle in a tropical country. Significance and Impact of the Study: This is the first study on E. coli O157: H7 from abattoir operations in Nigeria. The study emphasizes the risk of E. coli O157: H7 along the meat chain and the need for concerted effort to limit it through best hygiene practices.     

Effect of curli expression and hydrophobicity of Escherichia coli O157:H7 on attachment to fresh produce surfaces

Aim: To investigate the effect of curli expression on cell hydrophobicity, biofilm formation and attachment to cut and intact fresh produce surfaces. Methods and Results: Five Escherichia coli O157:H7 strains were evaluated for curli expression, hydrophobicity, biofilm formation and attachment to intact and cut fresh produce (cabbage, iceberg lettuce and Romaine lettuce) leaves. Biofilm formation was stronger when E. coli O157:H7 were grown in diluted tryptic soy broth (1 : 10). In general, strong curli‐expressing E. coli O157:H7 strains 4406 and 4407 were more hydrophobic and attached to cabbage and iceberg lettuce surfaces at significantly higher numbers than other weak curli‐expressing strains. Overall, E. coli O157:H7 populations attached to cabbage and lettuce (iceberg and Romaine) surfaces were similar (P > 0·05), indicating produce surfaces did not affect (P < 0·05) bacterial attachment. All E. coli O157:H7 strains attached rapidly on intact and cut produce surfaces. Escherichia coli O157:H7 attached preferentially to cut surfaces of all produce types; however, the difference between E. coli O157:H7 populations attached to intact and cut surfaces was not significant (P > 0·05) in most cases. Escherichia coli O157:H7 attachment and attachment strength (S_R) to intact and cut produce surfaces increased with time. Conclusions: Curli‐producing E. coli O157:H7 strains attach at higher numbers to produce surfaces. Increased attachment of E. coli O157:H7 on cut surfaces emphasizes the need for an effective produce wash to kill E. coli O157:H7 on produce. Significance and Impact of the Study: Understanding the attachment mechanisms of E. coli O157:H7 to produce surfaces will aid in developing new intervention strategies to prevent produce outbreaks.     

Sensitivity of Direct Plating for Detection of High Levels of <Emphasis Type="Italic">E. coli</Emphasis> O157:H7 in Bovine Fecal Samples

To assess the sensitivity of direct plating of bovine fecal samples for detection of Escherichia coli O157:H7, calves (n = 28) were orally inoculated with 10⁹ colony-forming units (cfu) per calf of a mixture of three strains of nalidixic acid-resistant E. coli O157:H7, and fecal samples were collected for analysis. One-gram samples from inoculated calves were mixed with 9 mL of Gram-negative broth with vancomycin, cefixime, and cefsoludin. From this suspension, serial dilutions were made (10⁻¹ to 10⁻⁴) and spread plated in triplicate on Sorbitol MacConkey agar with nalidixic acid for enumeration of E. coli O157:H7 in fecal samples. Direct plating samples were streaked for isolation on Sorbitol MacConkey agar with cefixime, and tellurite (SMACct). After incubation overnight at 37°C, morphologically typical colonies from direct streak plates were plated onto blood agar and incubated overnight at 37°C; then an indole test was performed on each colony. Indole-positive colonies were confirmed by O157 agglutination and were then plated on SMAC agar with 20 μg/mL nalidixic acid (SMACnal) to confirm nalidixic acid resistance. Overall sensitivity of detection was 32.5% (110/338 samples). Sensitivity to detect fecal samples shedding at above 5 × 10⁴ cfu/g was 83% (71/86 samples). Based on these data, direct plating of fecal samples might be an effective way to identify cattle that are likely to be shedding E. coli O157 at high levels.     

Role of rpoS in Acid Resistance and Fecal Shedding of Escherichia coli O157:H7

Acid resistance (AR) is important to survival of Escherichia coli O157:H7 in acidic foods and may play a role during passage through the bovine host. In this study, we examined the role in AR of the rpoS-encoded global stress response regulator ς^S and its effect on shedding of E. coli O157:H7 in mice and calves. When assayed for each of the three AR systems identified in E. coli, an rpoS mutant (rpoS::pRR10) of E. coli O157:H7 lacked the glucose-repressed system and possessed reduced levels of both the arginine- and glutamate-dependent AR systems. After administration of the rpoS mutant and the wild-type strain (ATCC 43895) to ICR mice at doses ranging from 10¹ to 10⁴ CFU, we found the wild-type strain in feces of mice given lower doses (10² versus 10³ CFU) and at a greater frequency (80% versus 13%) than the mutant strain. The reduction in passage of the rpoS mutant was due to decreased AR, as administration of the mutant in 0.05 M phosphate buffer facilitated passage and increased the frequency of recovery in feces from 27 to 67% at a dose of 10⁴ CFU. Enumeration of E. coli O157:H7 in feces from calves inoculated with an equal mixture of the wild-type strain and the rpoS mutant demonstrated shedding of the mutant to be 10- to 100-fold lower than wild-type numbers. This difference in shedding between the wild-type strain and the rpoS mutant was statistically significant (P ≤ 0.05). Thus, ς^S appears to play a role in E. coli O157:H7 passage in mice and shedding from calves, possibly by inducing expression of the glucose-repressed RpoS-dependent AR determinant and thus increasing resistance to gastrointestinal stress. These findings may provide clues for future efforts aimed at reducing or eliminating this pathogen from cattle herds.     

Recovery of E. coli O157 strains after exposure to acidification at pH 2

Aims: Rapid detection and selective isolation of E. coli O157:H7 strains have been difficult owing to the potential interference from background microflora present in high background food matrices. To help selectively isolate E. coli O157H7 strains, a useful plating technique that involved acidifying the cultures to pH 2 was evaluated with a large number of E. coli O157:H7 strains to ensure response to treatment was consistent across strains. Methods and Results: Escherichia coli O157, 46 strains including ATCC 35150, were acidified to pH 2 following enrichment and plated onto Tryptic Soy Agar + 0·6% Yeast Extract (TSA‐YE) and Sorbitol MacConkey Agar + cefixime and tellurite (CT‐SMAC). Samples were enumerated and modest decreases in plate counts were observed on TSA‐YE media, with a greater reduction observed on CT‐SMAC. Conclusions: The acid‐resistant character of E. coli O157:H7 is a consistent trait and may be used for improved isolation of the organism from mixed cultures. Significance and Impact of the Study: There was little difference observed between the commonly used laboratory strain E. coli O157:H7 35150 and 45 other strains of E. coli O157 when subjected to acidifying conditions prior to plating, demonstrating that an acid rinse procedure was equally effective across a wide variety of E. coli O157 strains and broadly applicable for isolating unknown strains from food samples.     

Detection of viable Escherichia coli O157:H7 by ethidium monoazide real-time PCR

Aims: The aim of this study was to develop and optimize a novel method that combines ethidium bromide monoazide (EMA) staining with real‐time PCR for the detection of viable Escherichia  coli O157:H7 in ground beef. EMA can penetrate dead cells and bind to intracellular DNA, preventing its amplification via PCR. Methods and Results: Samples were stained with EMA for 5 min, iced for 1 min and exposed to bright visible light for 10 min prior to DNA extraction, to allow EMA binding of the DNA from dead cells. DNA was then extracted and amplified by TaqMan^® real‐time PCR to detect only viable E. coli O157:H7 cells. The primers and TaqMan^® probe used in this study target the uidA gene in E. coli O157:H7. An internal amplification control (IAC), consisting of 0·25 pg of plasmid pUC19, was added in each reaction to prevent the occurrence of false‐negative results. Results showed a reproducible application of this technique to detect viable cells in both broth culture and ground beef. EMA, at a final concentration of 10 μg ml^?1, was demonstrated to effectively bind DNA from 10⁸ CFU ml^?1 dead cells, and the optimized method could detect as low as 10⁴ CFU g^?1 of viable E. coli O157:H7 cells in ground beef without interference from 10⁸ CFU g^?1 of dead cells. Conclusions: EMA real‐time PCR with IAC can effectively separate dead cells from viable E. coli O157:H7 and prevent amplification of DNA in the dead cells. Significance and Impact of the Study: The EMA real‐time PCR has the potential to be a highly sensitive quantitative detection technique to assess the contamination of viable E. coli O157:H7 in ground beef and other meat or food products.     

Antimicrobial efficacy of lactoferrin, its amidated and pepsin-digested derivatives, and their combinations, on Escherichia coli O157:H7 and Serratia liquefaciens

Aims: To evaluate the in vitro bactericidal efficacy of lactoferrin (LF), its amidated (AMILF) and pepsin‐digested (PDLF) derivatives, and their combinations, on Escherichia coli O157:H7 and Serratia liquefaciens. Methods and Results: PDLF exhibited the most potent bactericidal efficacy on E. coli O157:H7 (>2·5 log₁₀ CFU ml^?1 reduction at concentrations ≥1 mg ml^?1), and AMILF on Ser. liquefaciens (1 log₁₀ CFU ml^?1 reduction at 0·25–0·50 mg ml^?1). Some combinations of LF with PDLF or AMILF showed a slight synergy on E. coli O157:H7 and Ser. liquefaciens. However, all combinations of AMILF with PDLF were less active than the sum of the individual effects of the two antimicrobials. Production of capsular polysaccharide by bacteria might be involved in antimicrobial resistance. Conclusions: Escherichia coli O157:H7 and Ser. liquefaciens showed marked differences in the sensitivity to LF and its derivatives. E. coli O157:H7 was strongly inhibited by PDLF, whereas the effect of LF and its derivatives on Ser. liquefaciens was weak to negligible. Significance and Impact of the Study: PDLF was the most promising of the tested antimicrobials on E. coli O157:H7. However, the resistance of Ser. liquefaciens to LF and its derivatives hinders their use in the food industry.     

Reduction of Escherichia coli O157:H7 Populations in Cattle by Addition of Colicin E7-Producing E. coli to Feed

A cattle trial using artificially inoculated calves was conducted to determine the effect of the addition of colicinogenic Escherichia coli strains capable of producing colicin E7 (a 61-kDa DNase) to feed on the fecal shedding of serotype O157:H7. The experiment was divided into three periods. In period 1, which lasted 24 days, six calves were used as controls, and eight calves received 10⁷ CFU of E. coli (a mixture of eight colicinogenic E. coli strains) per g of feed. Both groups were orally inoculated with nalidixic acid-resistant E. coli O157:H7 strains 7 days after the treatment started. In periods 2 and 3, the treatment and control groups were switched, and the colicinogenic E. coli dose was increased 10-fold. During period 3, which lasted as long as period 1, both groups were reinoculated with E. coli O157:H7. The numbers of E. coli O157:H7 were consistently greater in the control groups during the three periods, but comparisons within each time period determined a statistically significant (P < 0.05) difference only at day 21 of period 1. However, when the daily average counts were compared between the period 1 control group and the period 3 treatment group that included the same six animals, an overall reduction of 1.1 log₁₀ CFU/g was observed, with a maximum decrease of 1.8 log₁₀ CFU/g at day 21 (overall statistical significance, P = 0.001). Serotype O157:H7 was detected in 44% of the treatment group's intestinal tissue samples and in 64% of those from the control group (P < 0.04). These results indicated that the daily addition of 10⁸ CFU of colicin E7-producing E. coli per gram of feed could reduce the fecal shedding of serotype O157:H7.     

Vaccination with attenuated Salmonella enterica Dublin expressing E coli O157:H7 outer membrane protein Intimin induces transient reduction of fecal shedding of E coli O157:H7 in cattle

Background  

Escherichia coli serogroup O157:H7 has emerged as an important zoonotic bacterial pathogen, causing a range of symptoms from self-limiting bloody diarrhea to severe hemorrhagic colitis and hemolytic-uremic syndrome in humans. Beef and dairy cattle are considered the most important animal reservoirs for this pathogen. One of the important virulence characteristics of E. coli O157:H7 is the eaeA gene encoding the 97 kDa surface protein intimin. Intimin is required for attachment and effacement during the interaction of enterohemorrhagic E. coli with human and bovine neonatal enterocytes. The present study was undertaken to test the hypothesis that an adaptive mucosal immune response directed against intimin will reduce or prevent enteric colonization and fecal shedding of E. coli O157:H7 in cattle.     

Reduction of Escherichia coli O157:H7 on radish seeds by sequential application of aqueous chlorine dioxide and dry‐heat treatment

Aims: To assess the effectiveness of sequential treatments of radish seeds with aqueous chlorine dioxide (ClO₂) and dry heat in reducing the number of Escherichia coli O157:H7. Methods and Results: Radish seeds containing E. coli O157:H7 at 5·5 log CFU g^?1 were treated with 500 μg ml^?1 ClO₂ for 5 min and subsequently heated at 60°C and 23% relative humidity for up to 48 h. Escherichia coli O157:H7 decreased by more than 4·8 log CFU g^?1 after 12 h dry‐heat treatment. The pathogen was inactivated after 48 h dry‐heat treatment, but the germination rate of treated seeds was substantially reduced from 91·2 ± 5·0% to 68·7 ± 12·3%. Conclusions: Escherichia coli O157:H7 on radish seeds can be effectively reduced by sequential treatments with ClO₂ and dry heat. To eliminate E. coli O157:H7 on radish seeds without decreasing the germination rate, partial drying of seeds at ambient temperature before dry‐heat treatment should be investigated, and conditions for drying and dry‐heat treatment should be optimized. Significance and Impact of the study: This study showed that sequential treatment with ClO₂ and dry‐heat was effective in inactivating large numbers of E. coli O157:H7 on radish seeds. These findings will be useful when developing sanitizing strategies for seeds without compromising germination rates.     

Experimental and Field Studies of Escherichia coli O157:H7 in White-Tailed Deer

Studies were conducted to evaluate fecal shedding of Escherichia coli O157:H7 in a small group of inoculated deer, determine the prevalence of the bacterium in free-ranging white-tailed deer, and elucidate relationships between E. coli O157:H7 in wild deer and domestic cattle at the same site. Six young, white-tailed deer were orally administered 10⁸ CFU of E. coli O157:H7. Inoculated deer were shedding E. coli O157:H7 by 1 day postinoculation (DPI) and continued to shed decreasing numbers of the bacteria throughout the 26-day trial. Horizontal transmission to an uninoculated deer was demonstrated. Although E. coli O157:H7 bacteria were recovered from the gastrointestinal tracts of deer necropsied from 4 to 26 DPI, attaching and effacing lesions were not apparent in any deer. Results are similar to those of inoculation studies in calves and sheep. In field studies, E. coli O157 was not detected in 310 fresh deer fecal samples collected from the ground. It was detected in feces, but not in meat, from 3 of 469 free-ranging deer in 1997. In 1998, E. coli O157 was not detected in 140 deer at the single positive site found in 1997; however, it was recovered from 13 of 305 dairy and beef cattle at the same location. Isolates of E. coli O157:H7 from deer and cattle at this site differed with respect to pulsed-field gel electrophoresis patterns and genes encoding Shiga toxins. The low overall prevalence of E. coli O157:H7 and the identification of only one site with positive deer suggest that wild deer are not a major reservoir of E. coli O157:H7 in the southeastern United States. However, there may be individual locations where deer sporadically harbor the bacterium, and venison should be handled with the same precautions recommended for beef, pork, and poultry.     

Differentiation of live, dead and treated cells of Escherichia coli O157:H7 using FT-IR spectroscopy

Aims: To apply specific collection techniques and spectroscopy to differentiate between live and dead Escherichia coli O157:H7 cells, as well as cells subjected to various inactivation treatments, including heat, salt, UV, antibiotics and alcohol. Methods and Results: Fourier transform‐infrared (FT‐IR) spectroscopy was used to analyse E. coli O157:H7 cells, after filtration or immunomagnetic collection. Partial least squares analysis of the spectra quantified live E. coli O157:H7 in the presence of dead cells with an R² > 0·996. Canonical variate analysis (CVA) not only differentiated between spectra of 100% dead and 100% live cells but also between 1% live : 99% dead and 100% dead. CVA using principal components also differentiated between the spectra of the differentially treated cells at a 95% confidence level, and Cooman plots showed clear separation between clusters of spectra of bacteria exposed to the different inactivation treatments. Mahalanobis distances (MD) corroborated the results of CVA. Conclusions: These results demonstrated the effectiveness of rapid cell collection and FT‐IR spectroscopy techniques to differentiate between live and dead E. coli O157:H7 cells. Significance and Impact of the Study: This technique has potential applications for use with foods subjected to various inactivation treatments.