ml

Search


Resource Type
 Video (49)
 Image (454)
 Animation (133)
 Protocol (41)
Curriculum Archive (85)
Tags
 Cell Biology (253)
 Genetics (81)
Humans (372)
 Environment (274)
 Diversity (108)
 Teaching (367)
Banner

Bacterial Identification by the Analytical Profile Index System—Analytical Profile Index E20 for Enterobacteriaceae Send Print

  Votes (0) | Hits (32394) | Comments (0)
Created: Friday, 30 June 2006
Last update: Wednesday, 11 August 2010
Author
View / Comment
Preview
Animation
View / Comment
Preview
Download Flash Archive
Information

Identification of a microbial isolate usually follows from a sequence of morphological, biochemical, immunological, and genetic techniques. Delineation of the biochemical activities of a microbial isolate is the most convenient way to narrow the search path towards the identity of an unknown strain. Biochemical reactions, for the purpose of identification, often utilize a limited combination of metabolic and enzymatic activities prominent and pertinent to a specific group of microorganisms that share these common activities. Variations in biochemical activities exhibited by members of the same group of organisms are then utilized to differentiate between these individuals and ultimately lead to identification. Among the most commonly utilized microbial biochemical activities are fermentation of sugars (carbohydrates), utilization of certain carbon sources, production of certain unique fermentation products, possession of specific enzymes, etc. 

Since the 1970s, routine sugar rack biochemical tests performed in test tubes headed towards miniaturization. In addition to saving materials, miniaturization enabled the expansion of the number of tests that could be included in a single test panel for a specific group of microorganisms. Furthermore, expansion of the number of tests in biochemical identification schemes led eventually to the development of extensive databases associating biochemical parameters with the identity of microbial species. Many commercial miniaturized biochemical test panels are now available in the market; they cover a significant number of clinically-important groups of bacteria, as well as food- and water-associated microorganisms. Among these test panels, and maybe one of the earliest, is the Analytical Profile Index (API) panel.

 

The Analytical Profile Index (API) is a miniaturized panel of biochemical tests compiled for identification of groups of closely related bacteria. Different test panels are prepared in dehydrated forms which are reconstituted upon use by addition of bacterial suspensions. After incubation, positive test results are scored as a seven-digit number (profile). Identity of the bacterium is then easily derived from the database with the relevant cumulative profile code book or software.

 

API 20E presented herein is a biochemical panel for identification and differentiation of members of the family Enterobacteriaceae. Other API panels for other groups of bacteria, such as staphylococci and streptococci, are also available in the same format, but are not included in this presentation. In API 20E for identification of members of the family Enterobacteriaceae, the plastic strip holds twenty mini-test chambers containing dehydrated media having chemically-defined compositions for each test. These include:
  1. ONPG: test for b-galactosidase enzyme by hydrolysis of the substrate o-nitrophenyl-b-D-galactopyranoside
  2. ADH: decarboxylation of the amino acid arginine by arginine dihydrolase
  3. LDC: decarboxylations of the amino acid lysine by lysine decarboxylase
  4. ODC: decarboxylations of the amino acid ornithine by ornithine decarboxylase
  5. CIT: utilization of citrate as sole carbon source
  6. H2S: production of hydrogen sulfide
  7. URE: test for the enzyme urease
  8. TDA: detection of the enzyme tryptophan deaminase
  9. IND: production of indole from tryptophan by the enzyme tryptophanase. Indole is detected by addition of Kovac's reagent.
  10. VP: the Voges-Proskauer test for the detection of acetoin (acetyl methylcarbinol) produced by fermentation of glucose by bacteria utilizing the butylene glycol pathway
  11. GEL: test for the production of the enzyme gelatinase which liquefies gelatin
  12. GLU: fermentation of glucose (hexose sugar)
  13. MAN: fermentation of mannose (hexose sugar)
  14. INO: fermentation of inositol (cyclic polyalcohol)
  15. SOR: fermentation of sorbitol (alcohol sugar)
  16. RHA: fermentation of rhamnose (methyl pentose sugar)
  17. SAC: fermentation of sucrose (disaccharide)
  18. MEL: fermentation of melibiose (disaccharide)
  19. AMY: fermentation of amygdalin (glycoside)
  20. ARA: fermentation of arabinose (pentose sugar)
The OX test is a test for cytochrome oxidase which is performed separately from the above tests. It is done using a portion of a bacterial colony on a paper strip impregnated by the oxidase reagent N,N,N’,N’-tetramethylphenylenediamine which turns blue if cells possess oxidase enzyme.

 

All test chambers are rehydrated by inoculation with a saline suspension of a pure culture of the bacterial strain subjected to identification (or a manufacturer-supplied suspending medium). After incubation in a humidity chamber for 18 to 24 hours at 37°C, the color reactions are read. The results of the test reactions (plus the oxidase reaction which is done separately) are converted to a seven-digit code. The code can then be looked up in the database book or software or fed into the manufacturer's database via touch-tone telephone where the computer voice gives the genus and species identification of the test microorganism.

 

This Flash-animated movie is an original artwork intended for laboratory, classroom, and educational use by microbiology students. In four sections, it graphically illustrates the description of the test panel, inoculation of panels by bacteria suspensions, colors of expected positive results, and reading and interpretation of results using five models of five quality control bacterial strains. This presentation may be helpful as an educational aid for both students who will perform this test and those who are studying background, concepts, and techniques of biochemical methods of identification, viewing the API as a general example of a type of commercial multitest method.  
ShareIcon Share





Tags: Teaching and learning (375)