The survival of Salmonella enterica serovar Typhimurium DT104 and total viable counts on beef surfaces at different relative humidities and temperatures.

AIMS: The aim of this study was to investigate changes in Salmonella and total viable count (TVC) survival on beef carcass surfaces stored for 72 h under different combinations of relative humidity (i.e. RH 75% or 96%) and temperature (5 degrees C or 10 degrees C). METHODS AND RESULTS: The influence of low water activity (a(w)) and temperature on the survival and growth of Salmonella enterica serovar Typhimurium DT104 and the aerobic mesophilic flora on meat pieces from different sites on beef carcasses was investigated, under controlled conditions (75% or 96% RH; 5 or 10 degrees C) in an environmental cabinet. Salmonella counts declined during storage at low a(w) (75% RH) conditions at 5 degrees C or 10 degrees C. Salmonella counts increased during storage at high a(w) (96% RH) at 10 degrees C only. At 5 degrees C, TVCs increased during storage at high a(w), but not at low a(w). TVCs increased on all samples from carcasses stored at high or low a(w) at 10 degrees C, except those samples taken from areas of surface fat. CONCLUSIONS: This suggests that substrate composition dictates growth rates under low a(w) conditions. The results are discussed in terms of the possible protective effects of substrate osmolyte accumulation in bacterial survival and/or growth. SIGNIFICANCE AND IMPACT OF THE STUDY: The data obtained in this study provides useful insights on the influence of a(w) and temperature on pathogen survival on meat surfaces at chill temperature.

Changes in antibiotic susceptibility in staphylococci habituated to sub-lethal concentrations of tea tree oil (Melaleuca alternifolia).

AIMS: To investigate the effect of sub-lethal challenge with tea tree oil (TTO) on the antibiotic resistance profiles of staphylococci. METHODS AND RESULTS: Isolates of methicillin-resistant/-sensitive Staphylococcus aureus (MRSA and MSSA) and coagulase-negative staphylococci (CONS) were habituated to sub-lethal concentrations of TTO (72 h). Following habituation, the minimum inhibitory concentrations (MIC) of antibiotics and TTO were determined. Habituated MRSA/MSSA cultures had higher (P < 0.05) MIC values than control cultures for the examined antibiotics. Habituated MRSA/MSSA cultures also displayed decreased susceptibility to TTO. Although the MIC of habituated MRSA/MSSA for the examined antibiotics reverted to control values after subsequent culture in the absence of TTO, the increased MIC against TTO were maintained. When compared with control cultures, habituated CoNS cultures had higher (P < 0.05) MIC values against three-fifths of the antibiotics examined; no changes in TTO MIC were observed. CONCLUSIONS: TTO habituation 'stress-hardens' MRSA and MSSA, evidenced by transient decreased antibiotic susceptibility and stable decreased TTO susceptibility. Although TTO habituation did not decrease susceptibility of CoNS to TTO, such cultures showed transient decreased antibiotic susceptibility. SIGNIFICANCE AND IMPACT OF THE STUDY: Application of TTO at sub-lethal concentrations may reduce the efficacy of topical antibiotics used with TTO in combination therapies.

A survey for serotyping, antibiotic resistance profiling and PFGE characterization of and the potential multiplication of restaurant Salmonella isolates.

AIMS: The aims of this research were (1) to determine the occurrence of Salmonella in Irish restaurant kitchens and (2) to investigate the serovar, genotype, antibiotic resistance profile and survival/growth profile of the Salmonella under catering chilled storage and temperature abuse conditions. METHODS: Five sites/tools in each of 200 randomly selected restaurant kitchens were examined for the presence of presumptive Salmonella spp. by enrichment. Serotyping, antibiotic resistance studies and genotyping were performed using the Kauffmann-White, CLSI and PulseNet methods, respectively. Survival/growth was investigated in milk, meat and vegetable products. RESULTS: Presumptive isolates from 15 of the 200 restaurant kitchens were recovered and confirmed as Salmonella positive. Seven different serovars showing a variety of antibiotic resistance profiles were detected. PFGE profiles suggested that isolates from geographically adjacent restaurants were related. Salmonella survived in foods stored at typical catering refrigeration temperatures and increased by approximately 0.8 log(10) CFU ml(-1) per day in food products stored under conditions of thermal abuse (20 degrees C). CONCLUSIONS: Inadequate hygiene has resulted in contamination of restaurant kitchens with Salmonella, which may persist/multiply in cross-contaminated foods. SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlights the need for greater hygiene in restaurant kitchens coupled with rapid chilling of food not for immediate consumption and reheating before subsequent serving.

The rate of horizontal transmission of antibiotic resistance plasmids is increased in food preservation-stressed bacteria.

AIM: The aim of this study was to investigate the possibility that sublethal food preservation stresses (high/low temperature, osmotic and pH stress) can alter the rate of horizontal transmission of antibiotic resistance (ABR) plasmids between Escherichia coli strains and between E. coli and Salmonella serotype Typhimurium. METHODS AND RESULTS: Escherichia coli donor cultures, carrying F1 plasmid R386 and Inc I1 plasmid TP307 and E. coli and Salm. Typhimurium recipient cultures were prestressed under a range of sublethal environmental conditions (high/low temperature, osmotic and pH stress). The prestressed donor and recipient cultures were then mated and the transmission rate calculated. The study found that the horizontal transmission rate of plasmids R386 and TP307 was significantly increased (P < 0.05) when prestressed donor and recipient cells are mated under conditions of environmental stress. CONCLUSION: The results from this study indicate that, the sublethal stresses that food pathogens encounter in modern food preservation systems increase the inter- and intra-specific horizontal transmission of selected ABR plasmids. SIGNIFICANCE AND IMPACT OF THE STUDY: Increased use of bacteriostatic (sublethal), rather than bacteriocidal (lethal) food preservation systems, may be contributing to the dissemination of ABR among important food borne pathogens.

The pattern of pleiomorphism in stressed Salmonella Virchow populations is nutrient and growth phase dependent.

AIMS: To describe the interactions of imposed osmotic and nutritional stress on the morphology of stationary and exponential phase S. Virchow cells. METHODS AND RESULTS: This study examined the morphology and viability of osmotically stressed exponential and stationary phase cultures of Salmonella Virchow under nutritionally deficient and competent conditions. In addition to normal morphology, salt-stressed cultures exhibited filamentous and spherical morphotypes, which were capable of reversion to normal morphology on stress removal. Proportions of atypical morphotypes were influenced by the phase of growth when the stress was applied. Salt-stressed exponential phase populations contained 54% filamentous, 30% spherical forms, salt-stressed stationary phase populations contained 16% filamentous, 79% spherical forms. Proportions of morphotypes were also influenced by the nutrient status of the medium, but not by metabolic by-products. CONCLUSIONS: Development of a range of morphotypes in response to stress (osmotic/nutritional), may offer population level advantages, increasing the survival potential of the population. SIGNIFICANCE AND IMPACT OF STUDY: The application of sublethal concentrations of salt may stimulate S. Virchow morphotype diversity, improving survival and rates of poststress recovery.

The influence of attachment to beef surfaces on the survival of cells of Salmonella enterica serovar Typhimurium DT104, at different a(w) values and at low storage temperatures.

This study investigated the influence of attachment to beef surfaces on the survival, injury and death of stationary phase cells of Salmonella enterica serovar Typhimurium DT104, compared to cells free in solution. The effects on cells are considered at different a(w) values and low temperatures in relation to osmotic and cold temperature shock effects. Attachment of cells to meat surfaces prevented cell injury and death from hyperosmosis and low temperatures, compared to meat solutions. Storage of cells for 72h resulted in higher levels of cell death on cells attached to meat surfaces. The improved survival of cells in solutions was considered to be related to adaptation to osmotic stress as a result of exposure to a previous hyperosmotic shock and the ability of the cells to produce cold shock proteins. Pathogen cell growth at low temperatures is discussed in relation to the presence of low levels of NaCl. Finally the data is discussed in relation to pathogen survival on beef carcass surfaces during refrigeration.

Effects of storage and the presence of a beef microflora on the thermal resistance of Salmonella Typhimurium DT104 in beef and broth systems.

AIMS: To investigate the effects of storage and the presence of a beef microflora on the thermal resistance of Salmonella serotype Typhimurium DT104 on beef surfaces and in a broth system during subsequent heat treatments after extended low-temperature storage (4 degrees C for 14 days) or mild temperature abuse (10 degrees C for 7 days). METHODS AND RESULTS: Surviving Salm. Typhimurium DT104 cells were estimated after heating in a water bath (55 degrees C) by plating beef and broth samples on tryptone soya agar and overlaying with xylose-lysine-deoxycholate agar. In beef and broth systems, D(55) values for Salm. Typhimurium DT104 stored at 4 degrees C or 10 degrees C in the presence or absence of a beef microflora were significantly lower (P < 0.01) than the D values for this organism heat-treated immediately after inoculation. In beef systems, the D(55) values were significantly lower (P < 0.05) in the presence of a beef microflora than the D(55) values obtained in 'pure' culture under all temperature/storage combinations. However, in broth systems, there was no significant difference between the D(55) values obtained in 'pure' culture and the D(55) values obtained from systems containing beef microflora. CONCLUSIONS: Storage of Salm. Typhimurium DT104 significantly reduced the thermal resistance of the pathogen in beef and broth systems. In the presence of high numbers of a Gram-negative beef microflora, the heat sensitivity of the pathogen was further increased on beef surfaces but not in broth. SIGNIFICANCE AND IMPACT OF THE STUDY: Studies investigating the survival of Salm. Typhimurium DT104 in different food systems will help define safe food preservation processes and will aid in the elimination this pathogen from the food production environments.

Surface decontamination of beef inoculated with Salmonella Typhimurium DT104 or Escherichia coli O157:H7 using dry air in a novel heat treatment apparatus.

AIMS: To determine the effectiveness of a novel dry air decontamination apparatus in the deactivation of Salmonella serotype Typhimurium DT104 or Escherichia coli O157:H7 on beef surfaces. METHODS AND RESULTS: A laboratory scale dry air decontamination apparatus, capable of producing repeatable and known heating time-temperature cycles on food surfaces was used in decontamination trials. Beef samples were surface inoculated with 7-8 log10CFU cm(-2) of S. Typhimurium DT104 or E. coli O157:H7 and heated at 60, 75, 90 and 100 degrees C using fast and slow heating rates and subsequently held at these temperatures for up to 600 s. A substantial reduction in pathogen numbers was achieved at higher temperatures (90 and 100 degrees C, 4.18-6.06 log10CFU cm(-2)) using both heating rates, but cell survival at these temperatures was also observed. At the lower temperatures, deactivation was small at 60 degrees C in particular it was less than one log unit after 3 min heating. No significant differences were observed when total reductions in pathogen counts were compared for all the temperature/heat up time combinations tested. During slow heating at 90 degrees C, and both heating rates at 100 degrees C, the pattern of deactivation of S. Typhimurium DT104 or E. coli O157:H7 was triphasic. CONCLUSIONS: This study has shown that heating meat surfaces with dry air can achieve substantial reductions in S. Typhimurium DT104 or E. coli O157:H7. As surface decontamination of beef surfaces with dry air had a negative effect on beef colour and appearance, such a decontamination apparatus would be unsuitable for producing meat for retail sale but it could be used to produce safer meat for use in the catering trade. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides researchers and food processors with data on the dynamic changes in S. Typhimurium DT104 and E. coli O157:H7 counts on intact beef surfaces during heating with dry air under realistic (time-varying) temperature conditions.

Prevalence and level of Escherichia coli O157 on beef trimmings, carcasses and boned head meat at a beef slaughter plant.

This study investigated the prevalence and level of Escherichia coli O157 on samples of beef trimmings (n=1351), beef carcasses (n=132) and bovine head meat (n=132) in a beef slaughter plant in Ireland. The survey also included an assessment of the prevalence of virulence genes in the E. coli O157 isolates obtained. Samples were examined for the presence of E. coli O157 by direct plating on SMAC-CT and by enrichment/immunomagnetic separation (IMS) with plating of recovered immunobeads onto SMAC-CT agar. Presumptive E. coli O157 isolates were confirmed by PCR targeting a range of genes i.e. vt1, vt2, eaeA, hlyA, fliC(h7) and portions of the rfb (O-antigen encoding) region of E. coli O157. Enterobacteriaceae on head meat samples were estimated by direct plating onto Violet Red Bile Glucose agar. E. coli O157 was recovered from 2.4% (32/1351) of beef trimmings samples, at concentrations ranging from<0.70-1.61 log10 cfu g(-1). Of the 32 positive isolates, 31 contained the eaeA and hylA genes while 30/32 contained the fliC(h7) gene and 31/32 contained vt1 or vt2, or both vt genes. E. coli O157 was recovered from 3.0% (4/132) of carcass samples, at concentrations ranging from <0.70-1.41 log10 cfu g(-1). All of the carcass isolates contained the eaeA, hylA and fliC(h7) genes. E. coli O157 was recovered from 3.0% (3/100) of head meat samples, at concentrations of 0.7-1.0 log10 cfu g(-1). All of the head meat isolates contained the eaeA, hylA, fliC(h7) and vt2 genes. No head meat isolates contained the vt1 gene. Head meat samples (n=100) contained Enterobacteriaceae, at concentrations ranging from 0.70-3.0 log10 cfu g(-1). Overall, the qualitative and quantitative data obtained for E. coli O157 on beef trimming samples in this study could be employed as part of a quantitative risk assessment model.

Impact of a novel spray-chilling system on surface microflora, water activity and weight loss during beef carcass chilling.

Commercially slaughtered and dressed beef carcass sides (n=30) were followed through a standard commercial chill unit fitted with a new "Jasca" air humidification system adjusted to provide intermittent water spraying of carcass sides (spray cycle 2 min on, 1 min off) for 15 h. Immediately after dressing, and after 24h in the chill unit, the surface water activity, and the weight of each side was measured, and 5 cm2 samples were recovered from four locations, i.e. rump, flank, brisket and neck on the surface of each side. These samples, and similar samples from control sides (n=30) processed in a standard commercial chill unit, were subjected to microbiological examination by direct and resuscitation counts on plate count agar (PCA), MacConkey agar (MAC) and violet red bile glucose agar (VRBGA). No significant differences were observed between bacterial numbers on test and control samples on each of the above agars, at each sample point/occasion. Comparison of direct and resuscitation counts suggested the presence of substantial numbers of injured cells, at both stages (pre- and post-chill), on test and control sides. After 24 h in chill units, test sides exhibited an average weight loss of 1.36% (+/-0.36%), which is significantly less (P<0.001) than the average weight loss (1.55%+/-0.24%) from control sides. These results suggest that the Jasca spray-chilling system can limit carcass shrinkage (on average by 0.19%) without significantly increasing the surface populations of selected bacterial groups.

Antimicrobial resistance in Irish isolates of verocytotoxigenic Escherichia coli (E. coli)--VTEC.

This study compared the antimicrobial resistance profiles of Escherichia coli O157:H7 isolates (n=257) recovered from bovine hides, minced beef and human clinical samples in Ireland, to those profiles of a range of Irish non-O157 E. coli (O111 and O26) isolates (n=31) from a variety of clinical and veterinary sources. Four multi-drug resistant (MDR) E. coli O157:H7 food isolates were identified, with resistance to 10 (1 isolate), 6 (1 isolate) and 4 (2 isolates) antimicrobial agents, respectively. Two of these isolates (resistant to 7 and 4 antimicrobial classes) were characterised further by molecular methods and found to contain class 1 integrons along with a beta-lactamase-encoding tem-1 gene. Transfer of antimicrobial resistance (ampicillin, streptomycin and sulphonamides), the tem-1 gene and markers (int1, qacEDelta1, sul1) characteristic of class 1 integrons were evident in one MDR isolate (resistant to 4 antimicrobial classes) when conjugation and transformation experiments were performed. A clinical isolate and a veterinary isolate of the O111 serotype were MDR and resistant to 4 and 3 antimicrobial classes, respectively. These data suggest that the prevalence of antimicrobial resistance among the three VTEC serotypes examined in this study is low. However, these organisms may become a public health risk should they enter the food chain.

Survival of antibiotic resistant and antibiotic sensitive strains of E. coli O157 and E. coli O26 in food matrices.

Escherichia coli O157:H7 or E. coli O26, which were AS (antibiotic sensitive), AR (laboratory created antibiotic resistant mutants), or naturally MAR (multi-antibiotic resistant), were inoculated into laboratory media, yoghurt or orange juice and their growth/survival monitored during enrichment at 37 degrees C or storage at 4 degrees C. The strains were also inoculated into minced beef and their thermal inactivation (D-values) examined at 55 degrees C, with and without a prior heat shock at 48 degrees C. The growth kinetics (lag phases, growth rates) of the VTEC (verocytotoxigenic E. coli), incubated over 24 h at 37 degrees C in laboratory media, were similar regardless of the presence or absence of antibiotic resistance. In yoghurt and orange juice, E. coli O157:H7 MAR died off significantly faster (P<0.05) than any of other VTEC strains examined. E. coli O157:H7 MAR was also found to be significantly more heat sensitive (P<0.05) than the other VTEC strains tested. The reasons for the observed differences in survival of the different VTEC strains and the link between antibiotic resistance and survival in VTEC organisms are discussed.

An impedimetric sensor for monitoring the growth of Staphylococcus epidermidis.

There is a need for accurate, reliable methods of detecting bacteria for a range of applications. One organism that is commonly found in urinary catheter infections is Staphylococcus epidermidis. Current methods to determine the presence of an infection require the removal of catheters. An alternative approach may be the use of in vivo sensing for bacterial/biofilm detection. This work investigates electrical impedance spectroscopy to detect the growth of Staphylococcus epidermidis RP62A on gold electrodes fabricated on a flexible substrate. Impedance spectra measured during biofilm formation on the electrode surface showed an increase in charge transfer resistance (RCT) with time.

An investigation of the thermal inactivation of Staphylococcus aureus and the potential for increased thermotolerance as a result of chilled storage.

AIMS: The aims of this study were; (i) to provide thermal inactivation data for Staphylococcus aureus; (ii) to examine the kinetics, including decimal reduction times (D-value) and rate constants (k), that describe the thermal inactivation of Staph. aureus and to compare two different methods of calculating D-values and (iii) to determine whether or not chilled storage would toughen these microorganisms resulting in increased thermotolerance. METHODS AND RESULTS: Isolates of Staph. aureus recovered from domestic refrigerators were grown in shaken culture for 8 h at 37 degrees C, recovered and washed by centrifugation and combined to form a cocktail of five strains. Samples from this cocktail were (a) heat treated at 50, 55 and 60 degrees C or (b) held under simulated domestic refrigeration conditions for 72 h and then heat treated as above. The numbers of Staph. aureus in heat treated and chill held, heat treated samples were enumerated by direct selective plating onto Baird Parker Agar (BPA) and recovery plating on Tryptone Soya Agar (TSA) subsequently overlaid with BPA. D-values were obtained using two different methods both of which may be used when the thermal inactivation follows first order kinetics. In the first method D-values are obtained by plotting the Log(10) of the surviving cells against time and using the equation D = -1/slope. The second method uses the rate constant (k) which is obtained from the slope of a plot of ln N/N(0)vs time and D is obtained using the equation D = 2.303 k(-1). D(50), D(55) and D(60) values ranged from 94.3 to 127.9 min, 13 to 21.7 min and 4.8 to 6.5 min. Prechilling did not enhance thermal resistance. The method of calculation did not affect the D-values obtained because the thermal inactivation of Staph. aureus in this study followed first order kinetics with r(2) values of 0.91-0.99. CONCLUSIONS: The thermal inactivation of Staph. aureus in tryptone soya broth (TSB) follows first order kinetics and in general chilling of these bacteria does not increase the resistance to thermal destruction during subsequent thermal processes such as cooking. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides much needed data on the thermal resistance of Staph. aureus and validates chilling as a food storage activity which does not cause toughening of the microorganisms to subsequent cooking. However, the data generated strongly suggests that Staph. aureus is more thermotolerant than Listeria monocytogenes and should be used as the target microorganism in designing mild thermal treatments for food, in which case the current recommendations for pasteurization (70 degrees C for 2 min, minimum) should be revised.

The prevalence and characterisation of Cryptosporidium spp. in beef abattoir water supplies.

The prevalence of Cryptosporidium spp. in 50 l samples of water used to wash beef carcasses at (a) an abattoir with a borehole water (BH) supply (n = 46) and (b) an abattoir with a river water (RW) supply (n = 48) was determined. In addition, a 100 l water sample and post-wash carcass samples (n = 24) were collected from the RW supply on a single day in July. Cryptosporidium spp. was detected in 0% and 26.1% of samples from the BH and RW supply abattoirs, respectively, with oocyst concentrations ranging from 0.02 to 8.6/l. Cryptosporidium spp. was not isolated from post-wash beef carcasses, while it was detected in water samples from that day at a concentration of 0.06 oocysts/l. The species of 3/5 isolates were identified as C. parvum, and the remaining were C. andersoni. This study has demonstrated that water used to wash beef carcasses can be contaminated with Cryptosporidium of human health importance and is a potential source of carcass contamination.

Food safety knowledge of consumers and the microbiological and temperature status of their refrigerators.

The objectives of this study were to examine domestic food safety knowledge levels of consumers, establish the levels and incidence of bacterial contamination and operational temperatures in domestic refrigerators, and identify areas in which consumer food safety education is necessary in Ireland. A food safety knowledge questionnaire applied to a representative sample of households (n = 1,020) throughout the island of Ireland found the gaps in consumer food safety knowledge. Analysis of swab samples (n = 900) recovered from the domestic refrigerators in these households showed average total viable counts of 7.1 log CFU/cm2 and average total coliform counts of 4.0 log CFU/cm2. Analysis of swab samples also detected the incidence of Staphylococcus aureus (41%), Escherichia coli (6%), Salmonella enterica (7%), Listeria monocytogenes (6%), and Yersinia enterocolitica (2%). Campylobacter jejuni and E. coli O157:H7 were not detected in domestic refrigerators. The temperature profiles of a subset of the sampled refrigerators (100) were monitored for 72 h, and 59% were found to operate, on average, at temperatures above the recommended 5 degrees C. Knowledge and temperature survey results varied considerably, but consumers who scored better in terms of basic food safety knowledge had reduced levels of bacterial contamination in their refrigerators and reported a reduced incidence of food-associated illnesses. This study confirms the effect of basic food hygiene knowledge on hygienic practice and identifies specific areasfor emphasis in the development and delivery of effective food safety risk communication messages to consumers.

Detection and recovery rates achieved using direct plate and enrichment/immunomagnetic separation methods for Escherichia coli O157:H7 in minced beef and on bovine hide.

AIMS: To assess the detection and recovery rates achieved with commonly used cultural methods for the enumeration and recovery of Escherichia coli O157:H7 from minced beef and bovine hide. METHODS AND RESULTS: Minced beef and bovine hide were inoculated with varying concentrations (log(10) 1.58-2.58 CFU g(-1) and log(10) 2.42-4.49 CFU 100 cm(2) respectively) of E. coli O157:H7 and recovered using a direct plate method or an enrichment/immunomagnetic separation (IMS) method and then plated onto SMAC or SMAC-CT in both cases. The direct plate method detected the pathogen consistently from minced beef samples with an average recovery of 69.2-91.2%. From faecal material on the bovine hide the recovery of the pathogen ranged from 1.80 to 64.5% with fresh faeces depending on the inocula while from dried faeces on hide the results ranged from no recovery at all to 25.1%. Enrichment/IMS recovered E. coli O157:H7 at all inocula levels tested in minced beef while the pathogen was only detected consistently at an average inocula level of log(10) 2.73 CFU 100 cm(2) from fresh faeces and log(10) 4.49 CFU 100 cm(2) from dried faeces on bovine hide. CONCLUSIONS: The direct count enumeration method for E. coli O157:H7 underestimated the numbers of pathogens present. The enrichment/IMS procedure consistently detected the pathogen from minced beef but did not always detect E. coli O157:H7 from faeces on bovine hide. SIGNIFICANCE AND IMPACT OF THE STUDY: Overall this study highlights that any microbial data, used in either predictive microbiology or risk assessment, must take account of the sensitivity and associated performance of the methods employed, in order to make an accurate reflection of the true microbiology of the examined sample.

Prevalence and numbers of Escherichia coli O157 on bovine hides at a beef slaughter plant.

In this study, we investigated the prevalence and numbers of Escherichia coli O157 on bovine hides. Samples (n = 1,500) were collected over a 17-month period (30 samples per week) by sponge swabbing approximately 122-cm2 areas of the bovine rump of slaughtered cattle at an early stage of carcass processing (first legging). Sponge samples (n = 1,500) were stomached in buffered peptone water supplemented with novobiocin, directly plated on sorbitol MacConkey with Cefixime tellurite (SMAC-CT), enriched for 24 h, extracted by immunomagnetic separation, and plated onto SMAC-CT agar. Presumptive E. coli O157 colonies from SMAC-CT plates were confirmed by PCR for the presence of eaeA, hlyA, fliCh7, vt1, vt2, and portions of the rfb (O-antigen encoding) region of E. coli O157. Overall, E. coli O157 was recovered from 109 samples (7.3%) at concentrations ranging from less than 0.13 to 4.24 log CFU/100 cm2. PCR analysis revealed a wide diversity of genetic profiles among recovered isolates of verocytotoxigenic E. coli. Of the isolates recovered, 99 of 109 contained the attaching and effacing gene (eaeA) and the hemolysin gene (hlyA), and 78 of 109 had the flagellar H7 antigen-encoding gene (fliCh7). Only 6 of 109 isolates contained both verotoxin-producing genes (vt1 and vt2); 91 of 109 contained the vt2 gene only, whereas 1 of 109 contained the vt1 gene only. The remaining 11 of 109 contained neither vt1 nor vt2.

Prevalence and characterisation of Cryptosporidium species in cattle faeces and on beef carcases at slaughter.

Cattle are known reservoirs and asymptomatic excretors of Cryptosporidium, a protozoan parasite that causes severe and protracted diarrhoea in people. The incidence of Cryptosporidium was investigated in 288 matched samples taken from beef carcases of 1 g samples of faeces retrieved immediately after de-legging, 25 cm2 samples of beef excised from the rump of uneviscerated carcases, and 25 cm2 samples of beef excised from the brisket area of eviscerated carcases. Cryptosporidium species were detected in 21 of the faecal samples after salt flotation and immunofluorescent microscopy. The species isolated from the positive samples were identified by restriction fragment length polymorphism and PCR as Cryptosporidium andersoni (54.5 per cent) and Cryptosporidium parvum genotype 2 (45.5 per cent). In the faecal samples, there was a significantly higher prevalence of the parasite in samples taken in summer (May to July) and winter (November to January) than in spring or autumn. No Cryptosporidium species were recovered from any of the beef samples.

The effect of thermal treatments on the viability and infectivity of Cryptosporidium parvum on beef surfaces.

AIMS: The aim of this research was to examine the effect of thermal treatments on the viability and infectivity of Cryptosporidium parvum oocysts attached to a beef surface. METHODS AND RESULTS: This study examined the effects of heat treatment (60 or 75 degrees C) on the viability of C. parvum oocysts inoculated onto the surface of beef muscle estimated by vital dye assay. The infectivity of the oocysts was assessed against monolayers of HCT-8 cells. At 60 degrees C viability of the oocysts decreased from 100% at T0 to 64.2% at T60. At 75 degrees C the viability of the oocysts decreased from 100% at T0 to 53.7% at T15 and finally to 11.2% at T60. Oocysts were rendered noninfective against monolayers of HCT-8 cells following treatments of 60 degrees C/45 s and 75 degrees C/20 s. CONCLUSION: The washing of carcasses with hot water and standard thermal treatments is sufficient to kill C. parvum on beef. SIGNIFICANCE AND IMPACT OF THE STUDY: This study found that relatively mild heat, currently used to decontaminate and heat treat beef carcasses and to cook meat products, is capable of inactivating C. parvum.