What is Ribotyping?
Ribotyping is a method that can identify and classify bacteria based upon differences in rRNA. It generates a highly reproducible and precise fingerprint that can be used to classify bacteria from the genus through and beyond the species level. DNA is extracted from a colony of bacteria and then restricted into discrete-sized fragments. The DNA is then transferred to a membrane and probed with a region of the rRNA operon to reveal the pattern of rRNA genes. The pattern is recorded, digitized and stored in a database. It is variations that exist among bacteria in both the position and intensity of rRNA bands that can be used for their classification and identification. Databases for Listeria (80 pattern types), Salmonella (97 pattern types), Escherichia (65 pattern types) and Staphylococcus (252 pattern types) have been established. In addition over 35 different genera have been tested and RiboPrint® patterns obtained. These databases continue to grow as more bacteria are ribotyped.
Examples of RiboPrint® Patterns
Why Ribotype?
Contamination source bacteria in finished products can originate from any one of the ingredients, processing personnel or the environment. Ribotyping allows the establishment of unequivocal relationships between bacterial isolates recovered from any of these sources and the finished product. Molecular epidemiology A number of diseases in animals that are caused by bacteria can be traced to the consumption of contaminated feed. Ribotyping can help to identify the contaminated feed source for elimination. Spread of infection within a group of animals can also be followed by ribotyping and routes of animal-to-animal transmission identified. Identification RiboPrint® patterns can be used to identify a bacterium if that pattern is in the database. Currently well over 600 patterns are in the database representing both Gram positive and Gram negative bacteria and this database grows continuously The largest number of RiboPrint® patterns have been collected for Listeria, Salmonella, Staphylococcus and Escherichia coli. Custom identification databases can be developed for virtually any need.
How is the RiboPrinter® System's typing method superior?
Key to the utility of a typing method is its reproducibility and standardization. The most common technique for typing is using a panel of antisera which selectively react with a certain degree of nonoverlapping specificity. Establishing a serotyping scheme for a new microorganism is tedious and dependent upon the quality of the panel of antisera. Furthermore the dissemination of this technique to other laboratories is usually limited and the longevity of the assay dependent upon antisera stocks. Virtually no typing method has been developed to the stage that allows standards to be established especially when classifying below a species level. Although a number of techniques have been explored and practiced in different laboratories, there is no method whose output is easily standardized. Ribotyping has over the past 10-15 years been established as a robust classification and typing method. It was originally developed as a manual method, but took a quantum leap in reproducibility and standardization with the development of the RiboPrinter® Microbial Characterization System by Qualicon (a subsidiary of DuPont). This instrument automates all of the steps in the process from cell lysis to data capture and database comparisons. Further, reagent cassettes including the enzymes, enzyme conjugated-hybridization probe, electrophoretic gel and membrane have been developed with aim of delivering consistent performance. Data capture is accomplished via a CCD camera and the patterns stored in a digitized format. It is therefore relatively easy to compare results among laboratories and to exchange data electronically.
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