Almost all rapid methods are designed to detect a single target, which makes them ideal for use in quality control programs to quickly screen large numbers of food samples for the presence of a particular pathogen or toxin. A positive result by a rapid method however, is only regarded as presumptive and must be confirmed by standard methods. Although confirmation may extend analysis by several days, this may not be an imposing limitation, as negative results are most often encountered in food analysis.
Most rapid methods can be done in a few minutes to a few hours, so they are more rapid than traditional methods. But, in food analysis, rapid methods still lack sufficient sensitivity and specificity for direct testing; hence, foods still need to be culture-enriched before analysis. Although enrichment is a limitation in terms of assay speed, it provides essential benefits, such as diluting the effects of inhibitors, allowing the differentiation of viable from non-viable cells and allowing for repair of cell stress or injury that may have resulted during food processing.
Evaluations of rapid methods show that some perform better in some foods than others. This can be attributed mostly to interference by food components, some of which can be especially troublesome for the technologies used in rapid methods. For example, an ingredient can inhibit DNA hybridization or Taq polymerase, but has no effect on antigen-antibody interactions and the converse situation may also occur. Since method efficiencies may be food dependent, it is advisable to perform comparative studies to ensure that a particular assay will be effective in the analysis of that food type.
The specificity of DNA based assays is dictated by short probes; hence, a positive result, for instance with a probe or primers specific for a toxin gene, only indicates that bacteria with those gene sequences are present and that they have the potential to be toxigenic. But, it does not indicate that the gene is actually expressed and that the toxin is made. Likewise, in clostridial and staphylococcal intoxication, DNA probes and PCR can detect only the presence of cells, but are of limited use in detecting the presence of preformed toxins.
Currently, there are at least 30 assays each for testing for E. coli O157:H7 and for Salmonella. Such a large number of options can be confusing and overwhelming to the user, but, more importantly, has limited the effective evaluation of these methods. As a result, only few methods have been officially validated for use in food testing .
Reference:
Feng, P. 1997. Impact of Molecular Biology on the Detection of Foodborne Pathogens. Mol. Biotech. 7:267-278.
Tuesday, July 17, 2007
Bacillus Cereus Diarrhetic Enterotoxins
Bacillus cereus is an aerobic spore former that is commonly found in soil, on vegetables, and in many raw and processed foods. Consumption of foods that contain large numbers of B. cereus (106 or more/g) may result in food poisoning. Although certain physiological and cultural characteristics are necessary for identifying B. cereus, its enterotoxigenicity indicates whether a suspect strain may be a public health hazard.
Common procedure to detect Diarrhetic Enterotoxins of Bacillus Cereus is by two methods. This is by using a semisolid agar medium and a serological procedure (the microslide gel double diffusion test) to identify the enterotoxin.
Hypertext Source: Bacteriological Analytical Manual, 8th Edition, Revision A, 1998. Chapter 15. Bacillus cereus Diarrheal Enterotoxin
*Author: Reginald W. Bennett
Common procedure to detect Diarrhetic Enterotoxins of Bacillus Cereus is by two methods. This is by using a semisolid agar medium and a serological procedure (the microslide gel double diffusion test) to identify the enterotoxin.
Hypertext Source: Bacteriological Analytical Manual, 8th Edition, Revision A, 1998. Chapter 15. Bacillus cereus Diarrheal Enterotoxin
*Author: Reginald W. Bennett
Sunday, July 15, 2007
Bacteris Toxins
Staphylococcal Enterotoxins Detection and Identification
When large numbers of enterotoxigenic staphylococci grow in foods, they may elaborate enough toxin to cause food poisoning after the foods are ingested. The most common symptoms of staphylococcal food poisoning, which usually begin 2-6 h after contaminated food is consumed, are nausea, vomiting, acute prostration, and abdominal cramps. To date, 8 enterotoxins (types A, B, C1, C2, C3, D, E, and H) have been identified as distinct serological entities. Current methods to detect enterotoxins use specific polyclonal or monoclonal antibodies (33,42,43).
Detection Methods and Techniques
1. Microslide gel double diffusion technique
2. Radioimmunoassay agglutination (RIA)
3. Enzyme-linked immunosorbent assay (ELISA)
- "double antibody sandwich" ELISA is the method of choice, because reagents are commercially available in polyvalent and monovalent formats for both toxin screening and serotype specific identification
- An automated enzyme-linked fluorescent immunoassay (ELFA) has been developed and is commercially available. This method has undergone specificity and sensitivity evaluations and has proven to be an effective serological system for the identification of staphylococcal enterotoxin in a wide variety of foods
4. Other methods, which have been used in the identification of the staphylococcal enterotoxins and may have application in foods, are the T-cell proliferation assay, and polyacrylamide gel electrophoresis (PAGE) combined with Western blotting
Summary
Microslide gel double diffusion precipitation test, two manual ELISAs [TecraTM, TransiaTM], an automated qualitative "enzyme-linked fluorescent immunoassay" [ELFATM, VidasTM], and sodium dodecyl sulfate-polyacrylamide gel electrophoresis [SDS-PAGE]-immunoblotting) for the identification of staphylococcal enterotoxin from isolates and from foods. - most common and often practiced and used
Recommended for routine analysis of foods for staphylococcal enterotoxin is the use, initially, of two different polyvalent ELISA kits
Known limits that causes food borne illness
When large numbers of enterotoxigenic staphylococci grow in foods, they may elaborate enough toxin to cause food poisoning after the foods are ingested. The most common symptoms of staphylococcal food poisoning, which usually begin 2-6 h after contaminated food is consumed, are nausea, vomiting, acute prostration, and abdominal cramps. To date, 8 enterotoxins (types A, B, C1, C2, C3, D, E, and H) have been identified as distinct serological entities. Current methods to detect enterotoxins use specific polyclonal or monoclonal antibodies (33,42,43).
When large numbers of enterotoxigenic staphylococci grow in foods, they may elaborate enough toxin to cause food poisoning after the foods are ingested. The most common symptoms of staphylococcal food poisoning, which usually begin 2-6 h after contaminated food is consumed, are nausea, vomiting, acute prostration, and abdominal cramps. To date, 8 enterotoxins (types A, B, C1, C2, C3, D, E, and H) have been identified as distinct serological entities. Current methods to detect enterotoxins use specific polyclonal or monoclonal antibodies (33,42,43).
Detection Methods and Techniques
1. Microslide gel double diffusion technique
2. Radioimmunoassay agglutination (RIA)
3. Enzyme-linked immunosorbent assay (ELISA)
- "double antibody sandwich" ELISA is the method of choice, because reagents are commercially available in polyvalent and monovalent formats for both toxin screening and serotype specific identification
- An automated enzyme-linked fluorescent immunoassay (ELFA) has been developed and is commercially available. This method has undergone specificity and sensitivity evaluations and has proven to be an effective serological system for the identification of staphylococcal enterotoxin in a wide variety of foods
4. Other methods, which have been used in the identification of the staphylococcal enterotoxins and may have application in foods, are the T-cell proliferation assay, and polyacrylamide gel electrophoresis (PAGE) combined with Western blotting
Summary
Microslide gel double diffusion precipitation test, two manual ELISAs [TecraTM, TransiaTM], an automated qualitative "enzyme-linked fluorescent immunoassay" [ELFATM, VidasTM], and sodium dodecyl sulfate-polyacrylamide gel electrophoresis [SDS-PAGE]-immunoblotting) for the identification of staphylococcal enterotoxin from isolates and from foods. - most common and often practiced and used
Recommended for routine analysis of foods for staphylococcal enterotoxin is the use, initially, of two different polyvalent ELISA kits
Known limits that causes food borne illness
When large numbers of enterotoxigenic staphylococci grow in foods, they may elaborate enough toxin to cause food poisoning after the foods are ingested. The most common symptoms of staphylococcal food poisoning, which usually begin 2-6 h after contaminated food is consumed, are nausea, vomiting, acute prostration, and abdominal cramps. To date, 8 enterotoxins (types A, B, C1, C2, C3, D, E, and H) have been identified as distinct serological entities. Current methods to detect enterotoxins use specific polyclonal or monoclonal antibodies (33,42,43).
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