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YERSINIA SELECTIVE AGAR BASE

Code: CM0653

A selective medium for Yersinia enterocolitica when used with Yersinia Selective Supplement SR0109 (Schiemann CIN Medium).

YERSINIA SELECTIVE SUPPLEMENT

Code: SR0109

Directions
Suspend 29g in 500ml of distilled water and bring gently to the boil to dissolve completely. Sterilise by autoclaving at 121°C for 15 minutes. Allow to cool to approximately 50°C and aseptically add the contents of one vial of Yersinia Selective Supplement SR0109 reconstituted as directed in the instructions for use that accompany the product. Mix gently and pour into sterile Petri dishes.

Description
Yersinia Selective Medium (CIN Medium) is based on the formulation of Schiemann^1,2 and is recommended for the isolation and enumeration of Yersinia enterocolitica from clinical specimens and food.

Yersinia enterocolitica is becoming increasingly recognised as a cause of diarrhoeal disease of man. Infection by the organisms results in diarrhoea, malaise, nausea and fever, plus constant abdominal pain over a period of 1-2 days. The organism has also been shown as a cause of polyarthritis, mesenteric adenitis and septicaemia. It is likely that human infections are directly or indirectly derived from animal sources and may be contracted through the ingestion of contaminated food. Initially serotypes 0:3 and 0:9 were implicated in human infections but since then other serotypes, mainly 0:5 and 0:8 have also been involved³. It is important to note that incidence of disease caused by the various serotypes of Yersinia enteroccolitica is currently reported to vary considerably with geographical location. It is expected that with provision of a selective medium, a higher isolation rate will result, and Yersinia enterocolitica will be recognised as more common and widespread than previously suspected.

Yersinia Selective Agar Base and the selective supplement SR0109 have been developed specifically for the optimum growth and recovery of Yersinia enterocolitica after 18-24 hours incubation at 32°C. Schiemann² modified his earlier formulation for CIN Medium by replacing bile salts with sodium desoxycholate (0.5g/l) and by reducing the concentration of novobiocin from 15 to 2.5mg/l in order to eliminate the inhibition of some strains of serotype 0:8.

The typical colonies of Yersinia enterocolitica will develop as a red bull’s-eye surrounded by a transparent border and will vary considerably among serotypes in colony size, smoothness and the ratio of the border to centre diameter. Most other organisms that are capable of growing will produce larger colonies (>2mm in diameter) with diffuse pinkish centres and opaque outer zones. Serratia liquefaciens, Citrobacter freundii and Enterobacter agglomerans may give a colonial morphology resembling Yersinia enterocolitica. These organisms can be differentiated from Yersinia enterocolitica by biochemical tests.

Test for growth on Nutrient and MacConkey Agars, test for indole and urease production and for acid reactions from sucrose, cellobiose, amygdalin, melibiose, rhamnose and reffinose. Carry out tests at 30°C rather than 37°C^4,5.

Technique for culture
Direct plate method
1. Pour plates of Yersinia Selective Agar and dry the surface.
2. Inoculate the plates with a suspension of the food, faeces, etc. to produce single colonies.
3. Incubate at 32°C for 24 hours.

Cold Enrichment in Phosphate Buffered Saline⁶
1. Inoculate food, faeces, etc., into M/15 phosphate buffered saline.
2. Hold at 4°C for up to 21 days.
3. Periodically sub-culture samples on to plates of Yersinia Selective Agar.
4. Incubate at 32°C for 24 hours.

CIN Agar had been used for isolation of Leptospira spp⁷. With enhancement of its nutritional properties and addition of 5-fluorouracil to increase selectivity it has also been used to demonstrate the presence of Arcobacter spp. in ground pork⁸.

Colonial morphology
Typical colonies of Yersinia enterocolitica will develop a red bull’s-eye surrounded by a transparent border. The colony size, smoothness and the ratio of the border to centre diameter will vary considerably among serotypes.

Identification of isolates

Storage conditions and Shelf life
Store the dehydrated medium at 10-30°C and use before the expiry date on the label.
Store the prepared medium at 2-8°C for not more than 24 hours.

Appearance
Dehydrated medium: Straw coloured, free-flowing powder
Prepared medium: Red coloured gel

Quality control

* This organism is available as a Culti-Loop®

Precautions
Some strains of Yersinia enterocolitica may grow poorly or not at all. Other species of Yersinia may grow along with some enteric organisms. It is therefore essential that full identification tests are carried out on suspect colonies.

References
1. Schiemann D. A. (1979) Can. J. Microbiol. 25. 1298-1304.
2. Swaminathan B., Harmon M. C. and Mehlman I. J. (1982) J. Appl. Bact. 52. 151-183.
3. Bisset M. L. (1976) J. Clin. Microbiol. 4. 137-144.
4. Swaminathan B., Harmon M. C. and Mehlman I. J. (1982) J. Appl. Bact. 52. 151-183.
5. Mair N. S. and Fox E. (1986) Yersiniosis: Laboratory Diagnosis, Clinical Features and Epidemiology. Pub. Hlth Lab. Ser. London.
6. Pai C. H., Sorger S., Lafleur L., Lackman L. and Marks M. I. (1979) J. Clin. Microbiol. 9. 712-715.
7. Borcyzk A., Rosa S.D. and Lior H. (1991) Abst. Ann. Meet. Am. Soc. Microbiol. C.267. p.386.
8. Collins C.I., Wesley I.V. and Murano E.A. (1996) J. Food Prot. 59. 448-452