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In the serological identification of a suspected Salmonella isolate, one utilizes antibodies pre-pared against specific antigens found on the cell wall and flagella. Blood serum containing one or more known antibodies is called antiserum. Agglutination (defined as a clumping of particles - in this case, bacterial cells) is observed where there is a match between antibodies in the antiserum and the homologous antigens on the particles.

For the detection of specific cell wall ("O") antigens, cells are obtained from a solid medium such as Triple Sugar Iron Agar or Kligler Iron Agar. Slide agglutination tests are performed with indi-vidual antisera as demonstrated in our procedure below. For the detection of flagellar ("H") antigens, the tube agglutination test is performed: BHI Broth cultures of the isolate are mixed with antisera, and an antibody-antigen match results in an agglutination of cells which settles rather quickly out of suspension.

To identify a Salmonella isolate with specificity - especially in the interests of epidemiology - one desires an identification more precise than a species name. In fact, species names are usually ignored. In the slide agglutination test, a reaction with antibodies unique to a serological group will lead to application of the group designation for the isolate as follows: Organisms which possess O antigen 2 are designated Salmonella Group A, those with O antigen 4 are designated Salmonella Group B, those with O antigens 6 and 7 are designated Salmonella Group C1, and so on.

With specific antisera, several cell wall and flagellar antigens can be detected for a Salmonella isolate, and identification becomes more precise. Based on the results of the slide and tube agglutination tests, the isolate is given an antigenic formula such as the following: 30:i:e,n,z₁₅ With this format, the numbers given before the first colon (:) represent specific O antigens, the next set of numbers represent flagellar antigens found for a subpopulation of cells, and the last set of numbers represent flagellar antigens found for another subpopulation. This formula, when given with the genus name - such as Salmonella 30:i:e,n,z₁₅ - identifies the isolate as belonging to a taxonomic subgroup called a serovar. For ease in communication, most serovars are also given informal designations which are written out like species names (but should not be construed as being official species names). For example, the serovar with the preceding antigenic formula is designated Salmonella mjordan (named after basketball star Michael Jordan).

Materials

Heat-killed Salmonella culture belonging to serological group B or C1

Pasteur pipettes

Dropper bottles of "anti-B" and "anti-C1" antisera (antisera prepared against Salmonella cell wall antigens unique to serological groups B and C1)

Petri plates with clear, scratch-free lids

1. On a clean glass slide, draw two circles (about 1 cm in diameter) heavily with a red wax marker.
2. Place the slide in an empty petri dish.
3. With a Pasteur pipette place a drop of the cell suspension within each circle. Make sure the drops appear as even, cloudy suspensions.
4. Without touching the dropper to the cells, place a drop of the anti-B antiserum on one suspension and a drop of anti-C1 on the other.
5. After several minutes, hold up the petri dish and observe the slide from the bottom. Gently tap the dish to effect some mixing of the cell suspension. See figure 14-11. Where there is a reaction between anti-bodies in the antiserum and their homologous antigens on the cell wall of the bacteria, the cells will agglutinate, and the drop will appear to contain many small particles. Where there is no agglutination, the cell suspension will maintain its original, evenly-cloudy appearance.
6. Discard the slide appropriately (or clean it off for reuse) and discard the petri plate into the proper container. (Do not place glass petri plates into the pails with the disposable plastics!)

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