Enumeration of Escherichia coli O157 in cattle faeces using most probable number technique and automated immunomagnetic separation

Fegan, Narelle; Higgs, Glen; Vanderlinde, Paul; Desmarchelier, Patricia

doi:10.1046/j.1472-765x.2003.01439.x

Cited by 60 publications

(37 citation statements)

References 20 publications

(23 reference statements)

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“…A recent attempt to retain sensitivity and at the same time obtain quantitative results employed the most-probable-number analysis of IMS enriched samples (5). However, such methods are extremely labor intensive and consume large amounts of materials.…”

Section: Discussionmentioning

confidence: 99%

Sensitivity of Escherichia coli O157 Detection in Bovine Feces Assessed by Broth Enrichment followed by Immunomagnetic Separation and Direct Plating Methodologies

LeJeune

Hancock²,

Besser

2006

J Clin Microbiol

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In order to more precisely predict food safety risks, the fecal presence of food-borne pathogens among animals at slaughter must be correctly determined. Quantification of Escherichia coli O157 is also desirable. In two separate experiments, detection and enumeration of a nalidixic acid-resistant strain of E. coli O157 in bovine feces was assessed by culture on MacConkey agar supplemented with nalidixic acid (MACnal) and compared to overnight broth enrichment followed by immunomagnetic separation (IMS) and to direct plating of dilutions of bovine feces onto sorbitol MacConkey agar containing cefixime and tellurite (SMACct). The sensitivity of detection of E. coli O157 by both direct plating and IMS was highly dependent upon the initial concentration of the target organism in the sample. Sensitivity of detection by IMS was poor below 100 CFU/g but was better, and not affected by initial E. coli O157 numbers, above this concentration. Sensitivity of detection of E. coli O157 in bovine feces at low initial concentrations is very poor for both direct plating and IMS. Direct plating of dilutions of bovine feces on SMACct can be used to determine the magnitude of fecal E. coli excretion among cattle excreting greater than 100 CFU/g. Among positive samples identified by direct plating on SMACct, the direct counts of E. coli O157:H7 were highly correlated with the estimates obtained with the MACnal plates (r ‫؍‬ 0.88, P < 0.001). Because the majority of cattle excrete less than 10 2 CFU E. coli O157/g feces, most studies, including those using IMS methods, probably grossly underestimate the prevalence of E. coli O157 in cattle.Contamination of foods, water, and other environmental niches with bovine manure has emerged as an important public health concern, especially with regard to the epidemiology of Escherichia coli O157:H7 infections in both humans and animals. Since cattle harbor E. coli O157:H7 in their digestive tracts, albeit transiently, they have been considered the primary reservoir for this important food-borne pathogen. Extensive efforts to understand the epidemiology of E. coli O157:H7 in the bovine host have been made. Theoretical models and empirical data have demonstrated that control of the magnitude and prevalence of E. coli O157:H7 in live animals (preharvest) may have a great impact on reducing the rate of contamination of foods of bovine origin with this pathogen (4, 7). However, to date, limited data on the magnitude of fecal E. coli O157:H7 shedding among animals destined for slaughter are available. In order to quantify the risks associated with the slaughter of E. coli O157:H7-positive animals and the effectiveness of potential preharvest mitigation strategies, accurate estimates of the prevalence and the frequency distribution of E. coli O157:H7 concentrations in bovine feces are required.To increase the diagnostic sensitivity, enrichment of large volumes (10 g or more) of bovine feces, followed by immunomagnetic separation (IMS), has been widely used for epidemiological studies of E. coli O157...

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Section: Discussionmentioning

confidence: 99%

Sensitivity of Escherichia coli O157 Detection in Bovine Feces Assessed by Broth Enrichment followed by Immunomagnetic Separation and Direct Plating Methodologies

LeJeune

Hancock²,

Besser

2006

J Clin Microbiol

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“…The Most Probable Number (MPN) procedure has been used for estimating the numbers of Salmonella on poultry carcases and rinses (Jetton et al 1992;Whittemore 1993;Humbert et al 1997) and a combination of MPN and IMS has been used for the enumeration of E. coli O157 in meats (Chapman et al 2001) and in cattle faeces (Fegan et al 2004). In this study, we used AIMS to determine the prevalence of Salmonella in cattle faeces and a combination of MPN and AIMS to estimate the numbers of Salmonella present in cattle faeces at slaughter.…”

Section: Introductionmentioning

confidence: 99%

Quantification and prevalence of Salmonella in beef cattle presenting at slaughter

et al. 2004

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Methods and Results: Automated immunomagnetic separation (AIMS) was used for detection and isolation of Salmonella enriched from cattle faeces. Salmonella were enumerated from positive samples using a combination of the Most Probable Number (MPN) technique and AIMS. A total of 310 faecal samples were tested, 155 were from lot-fed cattle and 155 from grass-fed cattle. Salmonella spp. were isolated from 21 (6AE8%) of the cattle and the prevalence amongst grass-fed cattle (4AE5%) was not significantly different to that found in lot-fed cattle (9%). Counts of Salmonella in positive faeces varied from <3 MPN g)1 of faeces to 2AE8 · 10 3 MPN g )1 and 71% of positive samples had counts <10 MPN g )1 faeces. There was no significant difference in the mean log 10 number of Salmonella in faeces of cattle from each production system. Conclusion: Low numbers of beef cattle were found to shed Salmonella at the time of slaughter and the prevalence and the associated faecal concentrations did not vary significantly with the pre-slaughter production system (grass or lot feeding). The faecal concentration of Salmonella in the majority of faeces was low (<10 MPN g )1 ) with few high concentrations up to 3 · 10 3 MPN g )1 , suggesting there may be a low risk of carcase contamination.Significance and Impact of the Study: Beef cattle do not appear to be a major source of entry of Salmonella into the human food chain and the quantitative information contained in this study can be used in quantitative assessments of the associated risk of human salmonellosis.

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“…The high concentration of E. coli O157:H7 associated with individual HF is an important factor in potential transmission. Cattle manure and other components of the cattle environment usually do not contain a high number of E. coli O157:H7 bacteria (10 5 CFU) per gram of sample (5,13). Interestingly, about 30% of the positive HF (n ϭ 125) were collected from a storage shed containing flaked corn.…”

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confidence: 99%

Association of Escherichia coli O157:H7 with Houseflies on a Cattle Farm

Alam

Zurek

2004

Appl Environ Microbiol

138

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The ecology of Escherichia coli O157:H7 is not well understood. The aims of this study were to determine the prevalence of and characterize E. coli O157:H7 associated with houseflies (HF). Musca domestica L. HF (n ‫؍‬ 3,440) were collected from two sites on a cattle farm over a 4-month period and processed individually for E. coli O157:H7 isolation and quantification. The prevalence of E. coli O157:H7 was 2.9 and 1.4% in HF collected from feed bunks and a cattle feed storage shed, respectively. E. coli O157:H7 counts ranged from 3.0 ؋ 10 1 to 1.5 ؋ 10 5 CFU among the positive HF. PCR analysis of the E. coli O157:H7 isolates revealed that 90.4, 99.2, 99.2, and 100% of them (n ‫؍‬ 125) possessed the stx1, stx2, eaeA, and fliC genes, respectively. Large populations of HF on cattle farms may play a role in the dissemination of E. coli O157:H7 among animals and to the surrounding environment.Enterohemorrhagic Escherichia coli O157:H7 is a wellknown causative agent of hemorrhagic colitis and hemolytic uremic syndrome in humans. Outbreaks of the food-borne illness caused by E. coli O157:H7 have been reported throughout the northern hemisphere, most frequently in the United States, Canada, Japan, and the United Kingdom. In the United States alone, E. coli O157:H7 causes more than 73,000 cases of human infection every year (11). E. coli O157:H7 strains commonly carry verotoxins (encoded by the stx1 and stx2 genes) and factors for the attachment to the host mucosa, including intimin (encoded by the eaeA gene) (11). The low infectious dose and high virulence of E. coli O157:H7 make infections severe and life-threatening, particularly for young children, the elderly, and those with weakened immune systems (11). The main reservoir for E. coli O157:H7 is the intestinal tracts of healthy cattle. Individual cattle are transiently colonized and shed E. coli O157:H7 in their feces (1). The sources of E. coli O157:H7, which colonizes cattle, are not well understood, and little is known about the ecology of E. coli O157:H7 in the environment (1). Additionally, the high variability in the prevalence of E. coli O157:H7 among cattle suggests the possibility of a reservoir of E. coli O157:H7 external to cattle. However, other than the detection of E. coli O157:H7 in nonbovine animals, including sheep, horses, dogs, and wild birds (1), the ecology of this pathogen has not been extensively studied.One of the potential modes of dissemination of this pathogen in the environment is by insects that are associated with animal feces and manure, primarily houseflies (HF; Musca domestica L.). HF larvae develop in animal feces, including cattle manure. Consequently, HF commonly build up very large populations on cattle farms and in other animal facilities. Previously, a laboratory-based study demonstrated that E. coli O157:H7 ingested by HF remained viable in the fly excreta and that the HF were able to carry and disseminate E. coli for several days (9). In Japan, HF were implicated in the transmission of E. coli O157:H7 from reservoir animals to...

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Enumeration of Escherichia coli O157 in cattle faeces using most probable number technique and automated immunomagnetic separation

Cited by 60 publications

References 20 publications

Sensitivity of Escherichia coli O157 Detection in Bovine Feces Assessed by Broth Enrichment followed by Immunomagnetic Separation and Direct Plating Methodologies

Sensitivity of Escherichia coli O157 Detection in Bovine Feces Assessed by Broth Enrichment followed by Immunomagnetic Separation and Direct Plating Methodologies

Quantification and prevalence of Salmonella in beef cattle presenting at slaughter

Association of Escherichia coli O157:H7 with Houseflies on a Cattle Farm

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