involves thefingerprintingof genomicDNArestriction fragments that contain all or part of the genes
coding for the 16S and 23SrRNA. Conceptually, ribotyping is similar to probingrestriction fragmentsof chromosomal
DNA withcloned probes(randomly cloned probes or probes derived from a specific coding sequence such as that of avirulence factor ).Ribotyping is a method that can identify and classify bacteria based upon differences in rRNA. It generates a highlyreproducible and precise fingerprint that can be used to classify bacteria from the genus through and beyond thespecies level. DNA is extracted from a colony of bacteria and then restricted into discrete-sized fragments. The DNAis 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 theposition 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 patterntypes) have been established. In addition over 35 different genera have been tested and RiboPrint® patternsobtained. These databases continue to grow as more bacteria are ribotyped.Fig:examples of ribotype pattern.
Contamination source bacteria in finished products can originate from any one of the ingredients, processingpersonnel or the environment. Ribotyping allows the establishment of unequivocal relationships between bacterialisolates recovered from any of these sources and the finished product. Molecular epidemiology A number of diseasesin animals that are caused by bacteria can be traced to the consumption of contaminated feed. Ribotyping can help toidentify the contaminated feed source for elimination. Spread of infection within a group of animals can also befollowed by ribotyping and routes of animal-to-animal transmission identified. Identification RiboPrint® patterns canbe used to identify a bacterium if that pattern is in the database. Currently well over 600 patterns are in the databaserepresenting both Gram positive and Gram negative bacteria and this database grows continuously The largestnumber 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 typingis using a panel of antisera which selectively react with a certain degree of nonoverlapping specificity. Establishing aserotyping 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 assaydependent upon antisera stocks. Virtually no typing method has been developed to the stage that allows standards tobe established especially when classifying below a species level. Although a number of techniques have beenexplored and practiced in different laboratories, there is no method whose output is easily standardized. Ribotypinghas over the past 10-15 years been established as a robust classification and typing method. It was originallydeveloped as a manual method, but took a quantum leap in reproducibility and standardization with the developmentof theRiboPrinter® Microbial Characterization SystembyQualicon(a subsidiary of DuPont). This instrument