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Selenite cystine broth
Culture Media for Food Microbiology, J.E.L. Corry et al. (Eds.)
437
9 1995 Elsevier Science B.V. All rights reserved
Selenite cystine broth Description and history This is a selective enrichment medium used in procedures for the detection and isolation of salmonellae. Selectivity is conferred by the inclusion of sodium selenite in the medium, shown by Leifson (1936) to be much more inhibitory to other members of the Enterobacteriaceae than to many strains of Salmonella, in particular Salmonella typhi and Salmonella paratyphi. Selenite cystine broth is a modification (North and Bartram, 1953) of the original Leifson (1936) formula, differing only in the addition of L-cystine which is considered to enhance Salmonella growth by reduction of toxicity. Selenite cystine broth is an internationally accepted medium (see ICMSF, 1978; Anon., 1981; 1985; Speck, 1984).
Composition (grams) Tryptone Lactose Di-sodium hydrogen o r t h o p h o s p h a t e 9 12H20 Sodium hydrogen selenite * L-Cystine Distilled or deionized water
5.0 4.0 10.0 4.0 0.01 1000.0
Preparation Base Dissolve the tryptone, lactose and di-sodium hydrogen orthophosphate in water by boiling for 5 min. After cooling add the sodium hydrogen selenite. Adjust the pH to 7.0 + 0.1 at 20~ Store at 4~ m
L-Cystine solution Add 0.1 g cystine to 15 ml N/1 NaOH. Dilute to 100 ml with sterile water in a sterile flask. Do not heat.
* W A R N I N G : Extreme care should be taken with the laboratory use of selenite solutions because of their potentially toxic effect.
438 Complete medium Cool base and add L-cystine solution at the rate of 0.1 ml per 10 ml of base. Adjust the pH to 7.0 _+ 0.1 at 20~ Transfer the complete medium in appropriate quantities e.g. 10 ml, 100 ml, etc. to sterile containers, to give a depth of 50-60 mm.
Physical properties Appearance pH
Pale straw coloured; clear or may have a slight precipitate. A brick red precipitate indicates overheating. 7.0 + 0.1 at 20~ m
Shelf life Ready to use medium
Use on day of preparation.
Inoculation method for samples 1. Where damaged cells are sought transfer 10 ml (or 1 ml) of sample-inoculated and incubated pre-enrichment broth into 100 ml (or 10 ml) Selenite cystine broth
or 2. Add comminuted (food) sample to Selenite cystine broth in the ratio 1 part sample to 10 parts medium.
Incubation method At 37~ for 24 h.
Reading of results and interpretation Inoculated and incubated medium is subcultured after 24 h onto selective diagnostic agar media in such a way as to obtain well-isolated colonies. Suspect colonies are subcultured and their identity confirmed by biochemical and serological tests.
Quality assessment (i) Productivity Test strains
Salmonella enteritidis 50073 Salmonella virchow 50077 Salmonella typhimurium (NCTC 12190)
439 Inoculation method
Dilution to extinction.
Criteria
R e c o v e r y in selenite cystine b r o t h should be within o n e titre unit of the recovery in t r y p t o n e soya b r o t h after 24 h at 37~
(ii) Selectivity Test strains
Enterobacter cloacae ( A T C C 23355) Escherichia coli 50034
Inoculation method
Dilution to extinction.
Criteria
R e c o v e r y in selenite cystine b r o t h should be less t h a n 2 titre units of the recovery in t r y p t o n e soya b r o t h after 24 h at 37~
Comments This m e d i u m m a y be m a r k e d l y influenced by the following: 1. 2. 3. 4. 5.
Source of i n g r e d i e n t s or d e h y d r a t e d c o m p l e t e m e d i u m T h e p r e s e n c e of food or o t h e r organic m a t e r i a l T h e physiological state of the organisms sought T h e c o m p e t i n g microflora Possibly by c o m b i n a t i o n s of the above
Successful p e r f o r m a n c e of the m e d i u m t e s t e d in o n e set of s t a n d a r d conditions may not be achieved if any of t h e s e conditions are altered.
References Anon. (1981) International Organisation for Standardisation. Microbiology - General guidance on methods for the detection of Salmonella. ISO, 6579-1981. Anon. (1985) International Organisation for Standardisation. Milk and milk products - Detection of Salmonella. ISO 6785-2985(E). ICMSF. (1978) Microorganisms in foods, 1. Their significance and methods of enumeration. 2nd ed. University of Toronto Press. Leifson, E. (1936) New selenite enrichment media for the isolation of typhoid and paratyphoid (Salmonella) bacilli. Amer. J. Hyg. 24, 423-432. North, W.R. and Bartram, M.T. (1953) The efficiency of selenite broth of different compositions in the isolation of Salmonella. Appl. Microbiol. 1, 130-124. Speck, M.L. (1984) Compendium of methods for the microbiological examination of foods. 2nd ed. American Public Health Association.
437
9 1995 Elsevier Science B.V. All rights reserved
Selenite cystine broth Description and history This is a selective enrichment medium used in procedures for the detection and isolation of salmonellae. Selectivity is conferred by the inclusion of sodium selenite in the medium, shown by Leifson (1936) to be much more inhibitory to other members of the Enterobacteriaceae than to many strains of Salmonella, in particular Salmonella typhi and Salmonella paratyphi. Selenite cystine broth is a modification (North and Bartram, 1953) of the original Leifson (1936) formula, differing only in the addition of L-cystine which is considered to enhance Salmonella growth by reduction of toxicity. Selenite cystine broth is an internationally accepted medium (see ICMSF, 1978; Anon., 1981; 1985; Speck, 1984).
Composition (grams) Tryptone Lactose Di-sodium hydrogen o r t h o p h o s p h a t e 9 12H20 Sodium hydrogen selenite * L-Cystine Distilled or deionized water
5.0 4.0 10.0 4.0 0.01 1000.0
Preparation Base Dissolve the tryptone, lactose and di-sodium hydrogen orthophosphate in water by boiling for 5 min. After cooling add the sodium hydrogen selenite. Adjust the pH to 7.0 + 0.1 at 20~ Store at 4~ m
L-Cystine solution Add 0.1 g cystine to 15 ml N/1 NaOH. Dilute to 100 ml with sterile water in a sterile flask. Do not heat.
* W A R N I N G : Extreme care should be taken with the laboratory use of selenite solutions because of their potentially toxic effect.
438 Complete medium Cool base and add L-cystine solution at the rate of 0.1 ml per 10 ml of base. Adjust the pH to 7.0 _+ 0.1 at 20~ Transfer the complete medium in appropriate quantities e.g. 10 ml, 100 ml, etc. to sterile containers, to give a depth of 50-60 mm.
Physical properties Appearance pH
Pale straw coloured; clear or may have a slight precipitate. A brick red precipitate indicates overheating. 7.0 + 0.1 at 20~ m
Shelf life Ready to use medium
Use on day of preparation.
Inoculation method for samples 1. Where damaged cells are sought transfer 10 ml (or 1 ml) of sample-inoculated and incubated pre-enrichment broth into 100 ml (or 10 ml) Selenite cystine broth
or 2. Add comminuted (food) sample to Selenite cystine broth in the ratio 1 part sample to 10 parts medium.
Incubation method At 37~ for 24 h.
Reading of results and interpretation Inoculated and incubated medium is subcultured after 24 h onto selective diagnostic agar media in such a way as to obtain well-isolated colonies. Suspect colonies are subcultured and their identity confirmed by biochemical and serological tests.
Quality assessment (i) Productivity Test strains
Salmonella enteritidis 50073 Salmonella virchow 50077 Salmonella typhimurium (NCTC 12190)
439 Inoculation method
Dilution to extinction.
Criteria
R e c o v e r y in selenite cystine b r o t h should be within o n e titre unit of the recovery in t r y p t o n e soya b r o t h after 24 h at 37~
(ii) Selectivity Test strains
Enterobacter cloacae ( A T C C 23355) Escherichia coli 50034
Inoculation method
Dilution to extinction.
Criteria
R e c o v e r y in selenite cystine b r o t h should be less t h a n 2 titre units of the recovery in t r y p t o n e soya b r o t h after 24 h at 37~
Comments This m e d i u m m a y be m a r k e d l y influenced by the following: 1. 2. 3. 4. 5.
Source of i n g r e d i e n t s or d e h y d r a t e d c o m p l e t e m e d i u m T h e p r e s e n c e of food or o t h e r organic m a t e r i a l T h e physiological state of the organisms sought T h e c o m p e t i n g microflora Possibly by c o m b i n a t i o n s of the above
Successful p e r f o r m a n c e of the m e d i u m t e s t e d in o n e set of s t a n d a r d conditions may not be achieved if any of t h e s e conditions are altered.
References Anon. (1981) International Organisation for Standardisation. Microbiology - General guidance on methods for the detection of Salmonella. ISO, 6579-1981. Anon. (1985) International Organisation for Standardisation. Milk and milk products - Detection of Salmonella. ISO 6785-2985(E). ICMSF. (1978) Microorganisms in foods, 1. Their significance and methods of enumeration. 2nd ed. University of Toronto Press. Leifson, E. (1936) New selenite enrichment media for the isolation of typhoid and paratyphoid (Salmonella) bacilli. Amer. J. Hyg. 24, 423-432. North, W.R. and Bartram, M.T. (1953) The efficiency of selenite broth of different compositions in the isolation of Salmonella. Appl. Microbiol. 1, 130-124. Speck, M.L. (1984) Compendium of methods for the microbiological examination of foods. 2nd ed. American Public Health Association.