Tellurite Sorbitol MacConkey Agar Research Articles

Isolation and identification of Escherichia coli O157:H7 from houseflies (Musca domestica L) at cattle barns in Al-Qadisiyah Province, Iraq

Recent studies identified that Escherichia coli O157:H7 (ECO157H7) causes hemorrhagic colitis in humans, a rare spectrum of diarrheal illness. According to the findings of epidemiological investigations, cattle serve as the principal reservoirs for this bacterium. This study was conducted to understand the incidence of ECO157H7 in houseflies (HFs), a key disease vector. HFs (n = 40) were collected from cattle barns. The HFs were cultured on MacConkey (MC) agars, and any suspected colonies were grown on Eosin Methylene Blue (EMB) agar. EMB-based metallic sheen colonies were grown on specific media, such as sorbitol Chromagar (SCA) and cefixime tellurite sorbitol-MacConkey (CT–SMAC) agar. Sorbitol non-fermenting (SNF) isolates were subjected to a commercial latex agglutination kit to identify the existence of O157 group. The results recorded 31/40 (77.5%) E. coli isolates identified from HF samples. The findings revealed that only 18/31 (58%) were from the SNF isolates. The identities of the isolates were further examined by targeting the rfbO157 and fliCH7 genes, as genetic markers in a PCR method. The PCR results reported 10/18 (55.5%) and 12/18 (66.6%) isolates that carried the rfbO157 and fliCH7 genes, respectively. Our research revealed that HFs represent a harbor for the pathogenic E. coli. The ecology and way this bacterium spreads among animals and across the environment may depend heavily on the presence of ECO157H7 in agricultural environments.

Isolation, prevalence of bacteria and fungi from carcasses and different locations within Yola abattoir and antimicrobial resistance profile of Escherichia coli O157:H7

The absence of modern slaughtering facilities and an unhygienic environment in Yola abattoir may result in an outbreak of antibiotic-resistant pathogenic microorganisms, leading to economic loss and human death. Hence, this study was carried out to isolate and assess the prevalence of bacteria and fungi within the abattoir environment as well as to determine the antimicrobial resistance profile of E. coli O157:H7 from different locations and carcasses. A cross-sectional study was conducted where a simple random sampling method was used for sampling. A total of 156 samples were collected. Isolation and identification of bacteria and fungi were done using standard methods. Pseudomonas was the most dominant bacterial species with a total of 151 isolates, followed by E. coli 143 and Klebsiella with 123 isolates. The highest prevalence in fungi was recorded in Aspergillus flavus at 17.6 %, followed by Aspergillus niger at 15.1 % and Trichophyton rubrum and Penicillium that had 14.5 % respectively. The highest mean air-borne coliform count was in the Lairage, it ranged between 1.1 x 106 CFU/m3 and 1.7 x 106 CFU/m3. A hundred and forty-three E. coli were isolated out of which six (4.2 %) were characterized to be E. coli O157:H7 producing characteristic colorless colonies on cefixime tellurite sorbitol MacConkey (CT-SMAC) agar. All the six E. coli O157:H7 were resistant to at least two antibiotics, ampicillin, and amoxicillin-clavulanic acid while ciprofloxacin, trimethoprim-sulfamethoxazole, and tetracycline were effective against the six E. coli O157:H7. The MAR ranged between 0.2 and 0.7. The results obtained in this study showed that there is a need for modern facilities in Yola abattoir, training of workers on hygiene, and a hygienic environment to avoid microbial contamination and spread of antimicrobial-resistant pathogenic strains like E. coli O157:H7 and mycotoxin producer Aspergillus flavus.

Epidemiological Investigation, Serotypes and Distribution of Verocytotoxigenic <i>Escherichia coli</i> (VTEC) in Raw Milk and Milk Products in Uyo, Nigeria

Food borne diseases are of great concern globally especially in the developing countries where poor sanitation is applied during collection and processing of milk from animals. The epidemiological investigation, serotypes and distribution of verocytotoxin (VTI and VT2)- producing Escherichia coli in raw milk and milk products were determined using structured questionnaire, Cefixime tellurite-sorbitol MacConkey agar, agglutination kits and VTEC-RPLA Toxin detection Kit. Out of 27 milkers, 7.4 % had primary education, 22.2 % washed the milk utensils with cold water and soap, 11.1 % washed their hands before milking, while 7.4 % milkers washed the udder of the animals before milking. All the yoghurts had the product names; 85.7 % had NAFDAC numbers; 80.0% had Batch Numbers, while 71.4 % had Manufacturer s’ Addresses. The unpasteurized milk samples had E. coli 0157 and non 0157 E. coli counts (CFU.ml-1) ranging from 4.0 x 102 to 1.7 x 103 and 6.0 x 102 to 2.0 x 103 , respectively, while E. coli 0157 and non 0157 E. coli counts of milk products were between 1.0 x 102 and 1.0 x 103 CFU.ml-1. E. coli 0157 had the highest percentage occurrence (38.3%), while E. coli 0145 had the lowest percentage occurrence (2.1%). More than 38.3% of the E. coli serotypes produced VT2, while ≥ 12.8% were VT1 producers. The occurrence of VTEC in the unpasteurized milk shows that the milkers should be enlightened on the necessary sanitary practices to adopt during milking and also post-pasteurization contamination of milk products should be avoided.
 Key Words: Verotoxigenic, Escherichia coli, Milk, Yoghurt, Nono, Serotypes.

Detection of Shiga-Toxin Producing E.coli (STEC), Enteropathogenic E.coli (EPEC) And Enterotoxigenic E.coli (ETEC) From Animals By M-PCR

This research investigated the presence of virulence genes encoding F41, K99, eae, Stx1, Stx2 and STa and the antimicrobial resistance of animal Escherichia coli (E. coli) isolates. Clinical isolates (n:233) were evaluated from faecal samples of cattle, sheep, goats, horses, cats and dogs collected between the years of 2010 to 2015 from Turkey. Enterohaemorrahic E.coli (EHEC) O157:H7 was detected by using cefixime tellurite sorbitol MacConkey agar (CT-SMAC) and Wellcollex E. coli (Remel®). The Kirby-Bauer disc diffusion test was performed to detect the resistance pattern of the isolates to ampicillin, Amoxycillin/clavulanic acid, enrofloxacin, ceftiofur, trimethoprim/sulfamethoxazole and tetracycline. The results showed that 40% of the ruminant isolates were identified as Shiga-toxin producing Escherichia coli (STEC). Enterotoxigenic E. coli (ETEC) was detected in samples from cattle (0.9%) and sheep (12%). Enteropathogenic E.coli (EPEC) was detected in samples from cattle (0.9%) and dogs (11.4%). EHEC O157:H7 was not detected any of the isolates. Among all E. coli isolates that carried at least one virulence gene, 8 (19%) were resistant to more than three antimicrobials, 7 (16.7%) were resistant to at least one antimicrobial and 27 (64.3%) were susceptible to all antimicrobials.

Simultaneous Immunomagnetic Separation Method for the Detection of Escherichia coli O26, O111, and O157 from Food Samples

We performed a simultaneous immunomagnetic separation (IMS) assay on Shiga toxin–producing Escherichia coli (STEC) serogroups O26, O111, and O157 with immunobeads coated with O26, O111, or O157 antibodies that were simultaneously added to an aliquot of food culture. We also compared the usefulness of CHROMagar STEC medium against various selective isolation agars designed to test for the three serogroups. Samples of sliced beef, ground beef, and radish sprout were artificially contaminated with STEC O26, O111, and O157 strains after incubation in enrichment broth and were subjected to conventional and simultaneous IMS assays. Simultaneous IMS did not affect the sensitivity of target cell detection. For STEC O26, O111, and O157 inoculated into the enriched samples of sliced beef and radish sprout, the detection ability of CHROMagar STEC was similar to or exceeded that of other isolation agars. However for STEC O157 inoculated into ground beef cultures, cefixime tellurite sorbitol MacConkey agar (CT-SMAC) was the superior detection medium. The performance of simultaneous IMS combined with CT-SMAC and CHROMagar STEC detection is similar to that of the Japanese standard method for isolating E. coli O26, O111, and O157. However, the proposed approach involves the same time, materials, and labor costs as the standard E. coli O157 reference procedure but allows detection of three E. coli serotypes rather than a single strain.

Membrane Damage and Viability Loss of E. coli O157:H7 and Salmonella spp in Apple Juice Treated with High Hydrostatic Pressure and Thermal Death Time Disks

Differences in membrane damage including leakage of intracellular UV-materials and loss of viability of Salmonella spp. and Escherichia coli O157:H7 bacteria in apple juice following thermal-death-time (TDT) disk and high hydrostatic pressure treatments were investigated. Salmonella spp. and E. coli O157H:H7 bacteria were inoculated in apple juice to a final 7.8 log10 CFU/ml and were thermally treated with TDT disks at 25, 35, 45, 50, 55 and 60°C for 4 min or pressurized at 350, 400 and 450 MPa at 25, 35, 45, 50, 55 and 60°C for 20 min. Sublethal injury, leakage of UV- materials and viability loss as a function of membrane damage of these bacterial pathogens was investigated by plating 0.1 ml of treated and untreated samples on non selective Trypticase Soy Agar (TSA) and selective Xylose Lysine Sodium Tetradecylsufate (XLT4) for Salmonella and Cefixime Potassium Tellurite Sorbitol-MacConkey (CT-SMACK) agar plates for E. coli bacteria with incubation at 36°C for 48 h. Sub-lethal injury occurred in Salmonella spp. and E. coli populations thermally treated with TDT disk at 55°C and above and at a pressure treatments of 25°C and above. Leakage of intracellular UV-materials and ATP of TDT disk injured cells was lower than the values determined from pressurized cells. Similarly, recovery of TDT injured cells occurred faster than pressurized cells during storage of treated samples at 22°C. The results of this study indicate that pressure treatment of 350 MPa at 35°C for 20 min and thermal treatments of 55 and 60°C and immediate storage of treated samples at 5°C will inhibit recovery and complete inactivation of injured bacteria in apple juice and therefore, will enhance the microbial safety of the treated juice.

Sorbitol-Fermenting, β-Glucuronidase–Positive, Shiga Toxin–Negative Escherichia coli O157:H7 in Free-Ranging Red Deer in South-Central Spain

We investigated the prevalence of Escherichia coli O157:H7 in free-ranging red deer in south-central Spain, to assess their potential as reservoir hosts of sorbitol-fermenting (SF) E. coli O157:H7 strains, which are emerging causes of hemolytic uremic syndrome in Europe. Fecal samples from 264 hunter-harvested Iberian red deer (Cervus elaphus) were collected in 25 different game estates and examined for E. coli O157:H7 by culture and PCR. E. coli O157:H7 was detected and isolated in 4 of the 25 game estates sampled (16%) and the isolates obtained (four in total) were further phenogenotypically characterized. One of them was biochemically typical of E. coli O157:H7, that is, neither fermented sorbitol nor exhibited β-glucuronidase (GUD) activity, and carried genes encoding Shiga toxins (Stx) 1 and 2, the intimin subtype γ1, the enterohemorrhagic E. coli (EHEC)-hemolysin, and the ter gene cluster. The rest of the isolates (three of four) fermented sorbitol, exhibited GUD activity after 18-24 h incubation, and carried genes encoding the intimin subtype γ1 and the EHEC-hemolysin, although no Stx-encoding genes were detected. All these atypical isolates carried the sfp gene cluster, lacked the ter gene cluster, and were unable to grow on cefixime tellurite sorbitol MacConkey agar, which are typical features of SF E. coli O157:H7 strains isolated from patients. In total, SF, GUD-positive, Stx-negative E. coli O157:H7 strains were isolated in 3 of the 25 game estates sampled (12%), with an overall sample-level prevalence of 1.1% (3/264). Our findings indicate that free-ranging red deer may be one of the possible reservoir hosts of Stx-negative derivatives of SF E. coli O157:H7.

Development and validation of a culture-based method suitable for monitoring environmental survival of Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium in developing countries

A non-GMO culture-based method suitable for studying the fate of Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium in an agricultural environment in developing countries was developed and validated. This method is based on the use of spontaneous rifampicin-resistant mutants of non-toxigenic variants of E. coli O157:H7 and S. Typhimurium combined with counting of colony-forming units (CFU) on selective media, i.e., Cefixime–Tellurite–Sorbitol MacConkey agar containing 100 μg/ml rifampicin, 50 μg/ml cycloheximide and 50 μg/ml nystatin for E. coli O157:H7 and Xylose–Lysine–Tergitol-4 agar containing the same antimicrobials for S. Typhimurium. Validation experiments using gfp- and ds-red-labelled derivatives of the rifampicin-resistant mutants in a non-sterile manure-soil matrix demonstrated that the new culture method was effective in the selection and recovery of the test strains without any detectable interference from background contaminants both in the short and long term. The rifampicin-resistant-based culture method designed provides a feasible low-cost option to study environmental survival of E. coli O157:H7 and Salmonella spp. in developing countries.

The occurrence of Escherichia coli O157:H7 in market and abattoir meat in Plateau State, Nigeria

Escherichia coli O157:H7 is a newly emerging pathogen frequently associated with the consumption of foods of bovine origin. The severity of the infections caused by this food borne pathogen in the young and the elderly has had a tremendous impact on human health and food industry. The present study evaluated the occurrence of E. coli O157:H7 in abattoir and market meat in seven selected sites in Plateau State, Nigeria. One hundred and forty samples of each of the various meat parts including muscle, liver, heart, kidney and intestine were collected to give a total of 700 meat samples. The samples were aseptically collected at monthly intervals from December 2008 to November 2009 after which they were transported to the laboratory for bacteriological analysis. Enrichment medium Tryptone Soya Broth (TSB) and selective agar media, namely; sorbitol MacConkey agar (SMAC)and cefixime tellurite-sorbitol MacConkey agar (CT-SMAC) were used for the isolation of E. coli O157:H7 from the various meat samples. The results of this study indicated that the intestinal samples had the highest percentage occurrence of the organism (3.39%), while muscle tissue had the lowest (0.43%). The organ meats had comparable percentage occurrence of the organism, with 1.0% for liver, 0.86% for kidney and 0.71% for heart samples. Significant difference (P 0.05) existed in the occurrence of the organism with respect to the sampling sites and the sources (abattoir and market). The presence of E. coli O157:H7 in beef should be regarded as being highly hazardous to consumers and handlers alike because of the severity of the infections associated with the organism. Thus, hygienic and good management of meats and meat environments are very essential.

P.004 Comparative evaluation of two commercial rapid tests able to detect hepatitis C virus (HCV) antibody: Flavicheck HCV versus ImmunoComb® HCV II

Numbers of Escherichia coli O157 in food may be low and sensitive techniques are therefore needed for its detection. The objectives of this study were to use carcass meat samples artificially inoculated with various strains of E. coli O157 to compare the sensitivity of enrichment in three different media and to compare immunomagnetic separation followed by culture of magnetic beads to cefixime tellurite sorbitol MacConkey agar with three immunoassays for the detection of E. coli O157 in the enrichment cultures. Duplicate 250, 25 and 2–3 CFU of each of 16 strains of E. coli O157 added to 25-g samples of beef carcass meat were used to compare the sensitivity of (1) enrichment in supplemented tryptone soya broth (sTSB), Reveal™ 8-h and Reveal™ 20-h media, and (2) immunomagnetic separation and culture to cefixime tellurite sorbitol MacConkey agar (IMS/CT-SMAC) with Reveal™, VIP™ and STAT™ immunoassays for detecting the organism. An initial inoculum of 250 CFU/25 g meat was detected in all 32 samples by IMS/CT-SMAC performed on all enrichment media and by Reveal™ performed on Reveal™ 8-h and Reveal™ 20-h media, but in only 30, 19 and 9 samples by Reveal™, VIP™ and STAT™, respectively, performed on sTSB medium. An initial inoculum of 25 CFU/25 g meat was detected in 28, 32 and 30 of 32 samples by IMS/CT-SMAC performed on sTSB, Reveal™ 8-h and Reveal™ 20-h media, respectively, and in 32 and 30 samples by Reveal™ performed on Reveal™ 8-h and Reveal™ 20-h media, but in only 22, 11 and 2 samples by Reveal™, VIP™ and STAT™, respectively, performed on sTSB medium. An initial inoculum of 2–3 CFU/25 g meat was detected in 25, 31 and 28 of 32 samples by IMS/CT-SMAC performed on sTSB, Reveal™ 8-h and Reveal™ 20-h media, respectively, and in 25 and 23 samples by Reveal™ performed on Reveal™ 8-h and Reveal™ 20-h media, but in only 14, 1 and 0 samples by Reveal™, VIP™ and STAT™, respectively, performed on sTSB medium.

Presence of Shiga toxin-producing E. coli O157:H7 in a survey of wild artiodactyls

This study was carried out to evaluate the role of wild artiodactyls as reservoirs of Escherichia coli O157:H7 for livestock and humans. Retroanal mucosal swabs samples from 206 red deer ( Cervus elaphus), 20 roe deer ( Capreolus capreolus), 6 fallow deer ( Dama dama) and 11 mouflon ( Ovis musimon), collected during the hunting season (autumn–winter) in South-western Spain, were screened. Samples were pre-enriched in modified buffered peptone water, concentrated by an immunomagnetic separation technique and cultured onto selective cefixime tellurite sorbitol MacConkey agar. Polymerase chain reaction (PCR) was used to detect the presence of genes coding O157 and H7 antigens and the virulence factors verocytotoxin, intimin and enterohaemolysin. Three E. coli O157:H7 isolates were obtained from red deer (1.5%). Two of them showed inability to ferment sorbitol and lack of β- d-glucuronidase (GUD) activity, however, the other strain investigated was an atypical sorbitol-fermenting E. coli O157:H7 with GUD + activity. This is the first report pointing to red deer as a reservoir of E. coli O157:H7 in Spain.

Detection, Occurrence, and Characterization of Escherichia coli O157:H7 from Raw Ewe's Milk in Spain

A study was carried out in the Castilla y León region of Spain to investigate the presence of Escherichia coli O157:H7 in raw ewe's milk samples collected from several cheese factories during 1 year. All specimens were examined for E. coli O157:H7 by selective enrichment at 41.5 ± 1.0°C, after both 6 and 22 h of incubation, and then immunomagnetically separated and plated on cefixime–potassium tellurite–sorbitol MacConkey agar. No growth was obtained in the enrichment broth after a 6-h incubation. Presumptive colonies obtained after 22 h of incubation were screened by a multiplex PCR assay for the presence of rfbO157 and fliCH7 genes. Of all the ewe's milk samples studied, three were positive for E. coli O157:H7. The E. coli O157:H7 strains that were positive for the rfbO157 and fliCH7 genes were then analyzed by multiplex PCR for the presence of virulence genes (stx1, stx2, ehxA, and eaeA). All E. coli O157:H7 isolates were Shiga toxigenic and harbored additional genes related to virulence (ehxA and eaeA). The predominant Stx toxin type was stx2. These results demonstrate that raw ewe's milk used in cheesemaking may be sporadically contaminated with E. coli O157:H7 strains that are potentially pathogenic for humans.

Occurrence of Shiga Toxin–Producing Escherichia coli O157 in Selected Dairy and Meat Products Marketed in the City of Rabat, Morocco

Samples of meat and dairy products taken from the city of Rabat, Morocco, were examined for the presence of Escherichia coli O157 by the selective enrichment procedure followed by plating on cefixime–tellurite–sorbitol MacConkey agar and a latex agglutination test. The ability of isolates to produce Shiga toxins (ST1 or ST2) was also tested by an agglutination test using sensitized latex. Dairy samples (n = 44) included different products commonly consumed in the country. Meat samples (n = 36) were taken from traditional butchers because these products are generally marketed in this way. Random samples were taken from each product during the period of January through May. Of the 80 samples tested, 8 (10%) harbored E. coli O157. Four dairy and four meat samples were contaminated (9.1 and 11.1%, respectively). Of 10 E. coli O157 isolates from contaminated samples demonstrating true antigen-antibody agglutination, 5 (50%) produced either ST2 alone or ST2 plus ST1. Four of the five strains (80%) were meat isolates and produced ST2 with or without ST1, and the fifth was a dairy isolate producing ST2.

RAPID PROTOCOL (5.25 H) FOR THE DETECTION OF ESCHERICHIA COLI O157:H7 IN RAW GROUND BEEF BY AN IMMUNO‐CAPTURE SYSTEM (PATHATRIX) IN COMBINATION WITH COLORTRIX AND CT‐SMAC 1

ABSTRACTA same day protocol for the detection of Escherichia coli O157:H7 by Pathatrix (a novel immuno‐capture method) and Colortrix (a rapid ELISA test) was developed. This study indicated that Pathatrix system, which circulated the pre‐enriched (4.5 h) samples over paramagnet beads with antibodies (0.5 h), increased the concentration of E. coli O157:H7 by 1.2–2.6 log CFU/25 g in 0.5 h from enrichment broth (modified EC broth with novobiocin or buffered peptone water). Colortrix, a rapid ELISA system, was used to detect the pathogens in another 0.25 h. Excellent correlation (100%) between positive Pathatrix/Colortrix (5.25 h system) results and positive Pathatrix/plating method with cefixime tellurite sorbitol‐MacConkey agar, with initial concentration of 0.7 to 2.1 log CFU/25 g was obtained. This new protocol offers a significant benefit to the meat industry allowing compliance with a “test” and “hold” program for E. coli O157: H7 in ground beef.

Isolation of Escherichia coli O157 from human and bovine faeces in the Urbino area, Italy

We examined 476 faecal samples from subjects aged from 0 to >60 years, 283 with diarrhoea and 193 with illnesses involving other sites or clinically healthy, and 154 samples of faeces of healthy cattle, in order to define the diffusion of E. coli O157 in the Urbino area. The samples were seeded by both direct streaking onto cefixime tellurite sorbitol Mac Conkey agar (CT-SMAC) and previous enrichment in cefixime tellurite tryptone soya broth for human specimens and in cefixime vancomicin tryptone soya broth for bovine samples. The strains of E. coliO157 were characterized by verocytotoxin and adhesin eae genes detection. We isolated one strain of E. coli O157 (0.2%) from a man 68 year old who had bloody diarrhoea, and one strain (0.64%) from a weaned calf. Both isolates carried the adhesin eae gene, but only the bovine strain was VT2+. The study shows a low diffusion of E. coli O157 in the Urbino area, confirming the epidemiological data on the national territory.

An evaluation of rapid methods for detecting Escherichia coli O157 on beef carcasses

Numbers of Escherichia coli O157 in food may be low and sensitive techniques are therefore needed for its detection. The objectives of this study were to use carcass meat samples artificially inoculated with various strains of E. coli O157 to compare the sensitivity of enrichment in three different media and to compare immunomagnetic separation followed by culture of magnetic beads to cefixime tellurite sorbitol MacConkey agar with three immunoassays for the detection of E. coli O157 in the enrichment cultures. Duplicate 250, 25 and 2–3 CFU of each of 16 strains of E. coli O157 added to 25-g samples of beef carcass meat were used to compare the sensitivity of (1) enrichment in supplemented tryptone soya broth (sTSB), Reveal™ 8-h and Reveal™ 20-h media, and (2) immunomagnetic separation and culture to cefixime tellurite sorbitol MacConkey agar (IMS/CT-SMAC) with Reveal™, VIP™ and STAT™ immunoassays for detecting the organism. An initial inoculum of 250 CFU/25 g meat was detected in all 32 samples by IMS/CT-SMAC performed on all enrichment media and by Reveal™ performed on Reveal™ 8-h and Reveal™ 20-h media, but in only 30, 19 and 9 samples by Reveal™, VIP™ and STAT™, respectively, performed on sTSB medium. An initial inoculum of 25 CFU/25 g meat was detected in 28, 32 and 30 of 32 samples by IMS/CT-SMAC performed on sTSB, Reveal™ 8-h and Reveal™ 20-h media, respectively, and in 32 and 30 samples by Reveal™ performed on Reveal™ 8-h and Reveal™ 20-h media, but in only 22, 11 and 2 samples by Reveal™, VIP™ and STAT™, respectively, performed on sTSB medium. An initial inoculum of 2–3 CFU/25 g meat was detected in 25, 31 and 28 of 32 samples by IMS/CT-SMAC performed on sTSB, Reveal™ 8-h and Reveal™ 20-h media, respectively, and in 25 and 23 samples by Reveal™ performed on Reveal™ 8-h and Reveal™ 20-h media, but in only 14, 1 and 0 samples by Reveal™, VIP™ and STAT™, respectively, performed on sTSB medium.

Prevalence of Escherichia coli O157:H7 in industrial minced beef.

The lack of baseline data on the prevalence of Escherichia coli O157:H7 in retail minced beef in France prompted this survey of industrial minced beef production. An automated enzyme-linked fluorescence immunoassay (ELFA), the VIDAS E. coli O157 method, was used to detect E. coli O157 in industrial minced beef samples. Confirmation of samples positive according to the ELFA was performed using an automated immunoconcentration (ICE) system, VIDAS ICE, which allows the selective capture and release of target organisms. The ICE was followed by culture on cefixime tellurite sorbitol MacConkey agar and a chromogenic medium, O157:H7 ID. Of the 3450 minced beef samples tested, 175 samples were positive with the ELFA method and, of these, four were confirmed by the ICE method. They were identified as sorbitol-negative, O157-positive, H7-positive, mobile, verotoxin-producing E. coli. The prevalence of E. coli O157:H7 in industrial French minced beef was 0.12%, consistent with many other reports. The low infective dose of E. coli O157:H7 presents a major threat. The main means of combating this organism are thermal destruction and good food hygiene covering activities on-farm, in the abattoir and in minced beef industries.

Applicability of the draft standard method for the detection of Escherichia coli O157 in dairy products

According to the draft standard method, EN ISO 16654 for the detection of Escherichia coli O157 in foods, samples of milk products inoculated with E. coli O157 were cultured in modified Tryptone Soya Broth supplemented with novobiocin. After immuno-magnetic separation (IMS) of the micro-organisms with magnetic beads coated with an antibody against E. coli O157 (Dynabeads® anti E. coli O157, Dynal), the enrichment broth was subcultured onto both Cefixim Tellurite Sorbitol MacConkey agar and CHROMagar™ O157. IMS alone appeared not to be very specific to E. coli O157; however, IMS combined with CT-SMAC plating enabled a greater exclusivity. The method displayed a low limit of detection (1–2 cfu/25 g) of E. coli O157 strains in milk products after only 6 h of incubation of the enrichment broth. However, the detection was affected by storage of the inoculum at 4 °C, and another 12 to 18 h of incubation was necessary to recover potentially stressed cells.

A comparison of methods for recovery of chlorine dioxide-injured Escherichia coli O157:H7 and Listeria monocytogenes

Direct surface-plating, overlay surface-plating and membrane-transferring surface-plating methods were evaluated for their suitability to recover uninjured and injured Escherichia coli O157:H7 and Listeria monocytogenes on green peppers after 0·1–0·5 mg l−1 chlorine dioxide (ClO2) gas for 15–30 min at 20°C under 75–80% relative humidity ClO2 gas treatments. Direct surface-plating on selective media showed the poorest recovery of injured bacteria. No significant difference (P>0·05) in recovery of injured bacteria was observed between overlay surface-plating and membrane-transferring methods. Overall, membrane-transferring surface-plating methods using tryptic soy agar (TSA) and cefixime–tellurite sorbitol–MacConkey agar (CTSMAC) (with a 2-h repair period and a 24-h total incubation at 37°C), and using TSA and modified Oxford agar (MOA) (with a 4-h repair period and a 48-h total incubation at 37°C) were the preferred methods for resuscitation of ClO2-injured E. coli O157:H7 and L. monocytogenes, respectively. Up to 5 log reductions of E. coli O157:H7 and L. monocytogenes determined by the membrane-transferring enumeration method were observed under a 0·5 mg l−1 ClO2 gas treatment for 30 min at 20°C under 80% relative humidity.

Escherichia coli O157 in cattle and sheep at slaughter, on beef and lamb carcasses and in raw beef and lamb products in South Yorkshire, UK

A 1 year study of Escherichia coli O157 in cattle and sheep at slaughter, on beef and lamb carcasses and in raw beef and lamb products from retail butchers’ shops was performed in the Sheffield area. Each month, samples of rectal faeces were collected immediately after slaughter from 400 cattle and 600 sheep, and 400–430 samples of raw meat products were purchased from butchers’ shops. Meat samples were also obtained from 1500 beef and 1500 lamb carcasses. All samples were examined for E. coli O157 by enrichment culture, immunomagnetic separation and culture of magnetic particles onto cefixime tellurite sorbitol MacConkey agar. Raw meat products were also examined for numbers of generic E. coli by a standard membrane culture method. E. coli O157 was isolated from 620 (12.9%) of 4800 cattle, 100 (7.4%) of 7200 sheep, 21 (1.4%) of 1500 beef carcasses, 10 (0.7%) of 1500 lamb carcasses and from 22 (0.44%) of 4983 raw meat products. E. coli O157 was isolated more frequently from lamb products (0.8%) than from beef products (0.4%). Numbers of generic E. coli in meat products reached seasonal peaks in July and August with counts of >10 4/g occurring more frequently in lamb products (50.8 and 42.4%, respectively) than in beef products (19.3 and 23.8%, respectively). The majority of E. coli O157 strains, from animals, carcasses and meat samples, were isolated during the summer. Most were verocytotoxigenic as determined by Vero cell assay and DNA hybridisation, eaeA gene positive and contained a 92 kb plasmid. The isolates were compared with 66 isolates from human cases over the same period. A combination of phage type, toxin genotype and plasmid analysis allowed subdivision of all the E. coli O157 isolates into 96 subtypes. Of these subtypes, 53 (55%) were isolated only from bovine faecal samples. However, 61 (92%) of the 66 isolates from humans belonged to 13 subtypes which were also found in the animal population.