Mistake proofing: changing designs to reduce error.

Mistake proofing uses changes in the physical design of processes to reduce human error. It can be used to change designs in ways that prevent errors from occurring, to detect errors after they occur but before harm occurs, to allow processes to fail safely, or to alter the work environment to reduce the chance of errors. Effective mistake proofing design changes should initially be effective in reducing harm, be inexpensive, and easily implemented. Over time these design changes should make life easier and speed up the process. Ideally, the design changes should increase patients' and visitors' understanding of the process. These designs should themselves be mistake proofed and follow the good design practices of other disciplines.

Detection of Escherichia coli serogroup O103 by real-time polymerase chain reaction.

AIMS: The aims of the study were to identify the specific genes of O-antigen gene cluster from Shiga toxin-producing Escherichia coli (STEC) O103 and to provide the basis for a specific real-time PCR test for rapid detection of E. coli O103. METHODS AND RESULTS: The published primers complementary to JUMPstart and gnd gene, the conserved flanking sequences of O-antigen genes clusters in E. coli and related species, were used to amplify the 12-kbp O103 O-antigen biosynthesis locus of STEC O103. A DNA library representative of this cluster allowed two O103-specific probes to be identified in the flippase (wzx) and UDP-galactose-4-epimerase (galE) genes. Two specific O103 serotyping real-time PCR tests based on these two genes were successfully developed. CONCLUSIONS: These results confirm that the O-antigen gene cluster sequences of E. coli allow rapidly a specific O-antigen real-time PCR assay to be designed. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings increase the number of real-time PCR-assays available to replace the classical O-serotyping among E. coli O-antigen.

Development of a 5'-nuclease PCR assay for detecting Shiga toxin-producing Escherichia coli O145 based on the identification of an 'O-island 29' homologue.

AIMS: A DNA sequence, from Escherichia coli STEC O145, homologous to O-island 29 from STEC O157 is described, together with a real-time PCR assay for detecting it. METHODS AND RESULTS: PCR and sequencing were used to identify the 'O-island 29' homologous DNA sequence from STEC O145 (strain VTH34). The sequence divergence between the STEC O145 and O157 'O-island 29' allowed a STEC O145 5'-nuclease PCR assay to be developed. CONCLUSIONS: The characterization of a novel locus in STEC O145 has allowed a specific O145 serogroup 5'-nuclease PCR assay to be designed. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings increase the number of serogroup PCR assays available as alternatives to classical O-serotyping of E. coli.

Identification of the O-antigen biosynthesis genes of Escherichia coli O91 and development of a O91 PCR serotyping test.

AIMS: The aims of the study were to characterize the O91 O-antigen gene cluster from Shiga toxin-producing Escherichia coli (STEC) O91 and to provide the basis for a specific PCR test for rapid detection of E. coli O91. METHODS AND RESULTS: The published primers complementary to JUMPstart and gnd gene, the conserved flanking sequences of O-antigen genes clusters in E. coli and related species were used to amplify the 10-kbp O91 O-antigen biosynthesis locus of STEC O91. A DNA library representative of this cluster allowed two O91 specific probes to be identified, and two specific PCR O91 serotyping tests to be successfully developed. CONCLUSIONS: These results confirm that the O-antigen gene cluster sequences of E. coli allow rapidly a specific O-antigen PCR assay to be designed. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings increase the number of PCR-assays available to replace the classical O-serotyping among E. coli O-antigen.

A PCR test for detecting Escherichia coli O157:H7 based on the identification of the small inserted locus (SILO157).

AIMS: This paper provides identification of a DNA sequence derived from Shiga toxin-producing Escherichia coli (STEC) O157:H7 and information on its utilization for detecting STEC O157 by PCR. METHODS AND RESULTS: Random Amplified Polymorphic DNA and DNA library were used to identify in STEC O157:H7 (strain EDL 933) a 2634-bp Small Inserted Locus, designated SILO157. Analysis of 211 bacterial strains showed that the PCR assays amplifying the SILO157 region could be used to detect STEC O157 with a good specificity. CONCLUSIONS: Characterization of a novel locus in STEC O157 is attractive since the serotype O157:H7 of STEC is still by far the most important serotype associated with more serious diseases. This island encodes putative proteins and especially one that is predicted to be an outer membrane protein designated IHP1. SIGNIFICANCE AND IMPACT OF THE STUDY: Further investigations could now be developed to appreciate the role of the SILO157 in pathogenicity.

Effects of acid mine drainage from an abandoned copper mine, Britannia Mines, Howe Sound, British Columbia, Canada, on transplanted blue mussels (Mytilus edulis).

Juvenile mussels (Mytilus edulis) were transplanted to Howe Sound, British Columbia, Canada, along an apparent pollution gradient of acid mine drainage (AMD) from an abandoned copper (Cu) mine. Cages containing 75 mussels each were placed at a total of 15 stations and were exposed to concentrations of dissolved Cu in surface waters ranging from 5 to 1009 micrograms/l for a period of 41 days. Mussels located at stations closer to the source of AMD at the mouth of Britannia Creek bioaccumulated higher concentrations of Cu and zinc (Zn) in their tissues. Mussel growth was adversely affected by Cu tissue concentrations above 20 micrograms/g dry wt., while declines in survival and condition index occurred in mussels that bioaccumulated greater than 40 micrograms/g dry wt. Cu. Tissue Zn concentrations (117-192 micrograms/g dry wt.) were likely not high enough to have a direct impact on mussel health. Reduced survival of transplanted mussels was supported by an absence of natural mussels in contaminated areas. Phytoplankton was also severely reduced in areas contaminated by mine waters. Based on the weight of evidence, AMD from the Britannia mine had a deleterious impact on mussel survival in a zone extending at least 2.1 km to the north and 1.7 km to the south of Britannia Creek on the east shore of Howe Sound.

Comparison between a PCR-ELISA test and the vero cell assay for detecting Shiga toxin-producing Escherichia coli in dairy products and characterization of virulence traits of the isolated strains.

AIMS: This paper provides information on a PCR-ELISA method for detecting Shiga toxin-producing Escherichia coli (STEC), and on their prevalence in dairy products. METHODS AND RESULTS: The sensitivity and specificity of the test was evaluated using pure cultures, spiked and naturally-contaminated samples. A comparative study with vero cytotoxicity testing was conducted, and STEC isolated from naturally-contaminated samples were characterized. The PCR-ELISA test was highly specific and sensitive, and detected 14% more positive samples than the vero cell assay. The prevalence of STEC in raw milk and unpasteurized cheese was 21.5% and 30.5%, respectively, while samples from the 'dairy environment' and from pasteurized cheese were less contaminated. The 34 strains of STEC isolated from natural samples showed that some of them carried virulence genes. CONCLUSION: No conclusion can be drawn at the moment concerning the potential risk to consumers. SIGNIFICANCE AND IMPACT OF THE STUDY: These data show the necessity of valuable screening methods to appreciate the virulence of STEC.

Evaluation of a polymerase chain reaction-based test for detecting Salmonella spp. in food samples: Probelia Salmonella spp.

A commercially available polymerase chain reaction (PCR) kit was evaluated for the detection of Salmonella spp. in food samples. The test combines PCR amplification and sandwich hybridization of the amplified DNA in microtiter plates. The sensitivity and specificity was evaluated with 52 Salmonella strains and 51 non-Salmonella strains and showed that the test was entirely reliable. The threshold sensitivity was 10(2) CFU/ml. The limit of detection of dead cells that determines the minimum detection level of dead cells in food samples was superior to 10(6) CFU/25 g, a level rarely achieved in naturally contaminated samples. After an 18-h pre-enrichment step, the test could detect viable Salmonella in artificially contaminated food samples, even for the lower contamination level (3 CFU/25 g). There was complete agreement between the PCR test and the ISO 6579 bacteriological reference method with artificially contaminated samples. Regarding the accuracy of the results obtained from 253 naturally or noncontaminated foods and from 32 artificially contaminated foods, the agreement percentage was 99.6%. The fidelity of the technique was evaluated in a collaborative study with eight European laboratories and showed a correlation of 98.4%.

Comparison of five typing methods for the epidemiological study of Listeria monocytogenes.

Five typing methods were compared in a study designed to adapt a strategy for epidemiologically typing large numbers of Listeria monocytogenes strains. The methods studied were serotyping, electrophoretic typing of esterases (zymotyping), restriction fragment length polymorphism of ribosomal DNA (ribotyping), random amplified polymorphic DNA (RAPD) and pulsed-field gel electrophoresis (PFGE). Data were analysed by computer-assisted statistical analysis. Included in the analysis were 35 strains of L. monocytogenes, including 14 epidemic strains isolated during outbreaks in France in 1992 and 1993, and 21 strains isolated from food and the environment. Five serotypes, eight zymotypes, ten ribotypes, 13 RAPD patterns and 12 PFGE patterns were identified among the 35 strains. The most discriminating combination of typing methods was ribotyping and PFGE typing [27 types, discriminatory index (D.I.) = 0.978]. A factorial analysis of correspondence for each method differentiated the epidemic strains from the environmental strains. This study shows that computer-assisted statistical treatment of the data, combined with the use of discriminating typing methods, is a powerful tool for the epidemiological analysis of Listeria monocytogenes.

Molecular epidemiological tools for Salmonella Dublin typing.

A total of 32 strains of Salmonella Dublin recovered from cattle were differentiated by electrophoretic typing of their esterases (zymotyping), restriction fragment length polymorphism of ribosomal DNA (ribotyping), arbitrarily primed PCR (AP-PCR) using five primers, PCR based on repetitive extragenic palindromic sequences (REP-PCR) and PCR based on enterobacterial repetitive intergenic consensus sequences (ERIC-PCR). ERIC-PCR and REP-PCR each gave one type, zymotyping gave three, AP-PCR gave five and ribotyping gave seven types. Combination of ribotyping and AP-PCR produced a total of 11 types, whereas 14 different types were obtained by all five methods. Thus a combination of several methods enhanced the discrimination of cattle-adapted strains among the genotypically homogeneous serovar Salmonella Dublin.