Or you can always go for the classical biochemical characterization media: EMB agar, MacConkey agar, Kligler iron agar, TSI agar, lysin-iron agar, Motility Indole Ornithine (MIO) medium, SIM medium, urea broth/agar, Simmons-citrate agar,...
El miércoles, 10 de abril de 2013 15:18:53 UTC+10, Xabier Vázquez Campos escribió:
Some pH indicators commonly use in media:
Below is a list of some commonly used pH indicators.
pH Indicator pH Range Acid Color Alkaline Color
m-Cresol Purple 0.5–2.5 Red Yellow
Thymol Blue 1.2–2.8 Red Yellow
Bromphenol Blue 3.0–4.6 Yellow Blue
Bromcresol Green 3.8–5.4 Yellow Blue
Chlorcresol Green 4.0–5.6 Yellow Blue
Methyl Red 4.2–6.3 Red Yellow
Chlorphenol Red 5.0–6.6 Yellow Red
Bromcresol Purple 5.2–6.8 Yellow Purple
Bromthymol Blue 6.0–7.6 Yellow Blue
Phenol Red 6.8–8.4 Yellow Red
Cresol Red 7.2–8.8 Yellow Red
m-Cresol Purple 7.4–9.0 Yellow Purple
Thymol Blue 8.0–9.6 Yellow Blue
Cresolphthalein 8.2–9.8 Colorless Red
Phenolphthalein 8.3–10.0 Colorless Red
Some chromogenic/differential media:
Chromogenic E. coli/Coliform Medium
Composition per liter:
Chromogenic mix ............................................................ ........... 20.3g
Agar ............................................................ .............................. .. 15.0g
Peptone..................................................... .............................. ....... 5.0g
NaCl ............................................................ .............................. .... 5.0g
Na2HPO4 ............................................................ ........................... 3.5g
Yeast extract..................................................... ............................. 3.0g
Lactose ............................................................ .............................. 2.5g
NaH2PO4 ............................................................ ........................... 1.5g
Neutral Red ............................................................ ..................... 0.03g
pH 6.8 ± 0.2 at 25°C
Preparation of Medium: Add components to distilled/deionized water
and bring volume to 1.0L. Mix thoroughly. Gently heat and bring to
boiling. Dispense into tubes or flasks. Autoclave for 15 min at 15 psi
pressure–121°C. Pour into sterile Petri dishes or leave in tubes.
Use: For the differentiation between Escherichia coli and other coliforms
in cultures produced from food samples. Agar base uses two enzyme
substrates to differentiate between E. coli and other coliforms.
One chromogenic substrate is cleaved by the enzyme glucuronidase
which is specific for E. coli and produced by approximately 97% of
strains. The second chromogenic substrate is cleaved by galactosidase,
an enzyme produced by the majority of coliforms. This results in purple
E. coli colonies, as they are able to cleave both chromogenic substrates
and pink coliform colonies as they are only able to cleave the
galactosidase chromogen.
Chromogenic Enterobacter sakazakii Agar,
DFI Formulation
Composition per liter:
Agar ............................................................ .............................. .. 15.0g
Tryptone ............................................................ .......................... 15.0g
Soya peptone..................................................... ............................ 5.0g
NaCl ............................................................ .............................. .... 5.0g
Ferric ammonium citrate..................................................... .......... 1.0g
Sodium deoxycholate................................................ ................... 1.0g
Na2S2O3 ............................................................ ............................ 1.0g
Chromogen................................................... .............................. ... 0.1g
pH 7.3 ± 0.2 at 25°C
Preparation of Medium: Add components to distilled/deionized water
and bring volume to 1.0L. Mix thoroughly. Gently heat and bring to
boiling. Dispense into tubes or flasks. Autoclave for 15 min at 15 psi
pressure–121°C. Pour into sterile Petri dishes or leave in tubes. Allow
tubes to cool in a slanted position.
Use: For the differentiation and enumeration of Enterobacter sakazakii
from infant formula and other food samples. The enzyme α-glucosidase,
present in E. sakazakii, hydrolyzes the substrate 5-bromo-4-
chloro-3-indolyl-α,D-glucopyranoside, thus producing blue-green colonies
on this pale yellow medium. Proteus vulgaris is also weakly α-
glucosidase positive and could grow to give colonies of a similar color
to E. sakazakii. However, on this medium, Proteus spp. grow as grey
colonies: they produce hydrogen sulphide in the presence of ferric ions
forming ferrous sulphide. Deoxycholate inhibits the growth of most
Gram-positive organisms.
Chromogenic Candida Agar
Composition per liter:
Chromogenic mix ............................................................ ........... 13.6g
Agar ............................................................ .............................. .. 13.6g
Peptone ............................................................ ............................. 4.0g
Selective supplement solution .................................................10.0mL
pH 6.0 ± 0.2 at 25°C
Source: This medium is available from Oxoid Unipath.
Selective Supplement Solution:
Composition per 10.0mL:
Chloramphenicol............................................. ....................... 500.0mg
Preparation of Selective Supplement Solution: Add chloramphenicol
to distilled/deionized water and bring volume to 10.0mL. Mix
thoroughly. Filter sterilize.
Preparation of Medium: Add components to distilled/deionized
water and bring volume to 1.0mL. Mix thoroughly. Gently heat while
stirring and bring to boiling. Do not autoclave. Cool to 45°C. Pour into
sterile Petri dishes.
Use: For the rapid isolation and identification of clinically important
Candida species. The medium incorporates two chromogens that indicate
the presence of the target enzymes: X-NAG (5-bromo-4-chloro-3-
indolyl N acetyl ß-D-glucosaminide) detects the activity of hexosaminidase.
BCIP (5-bromo-6-chloro-3-indolyl phosphate p-toluidine salt)
detects alkaline phosphatase activity. An opaque agent has been incorporated
into the formulation to improve the color definition on the agar.
The broad-spectrum antibacterial agent chloramphenicol is added to
the agar to inhibit bacterial growth on the plates.
Chromogenic Listeria Agar
Composition per liter:
Peptone ............................................................ ........................... 18.5g
LiCl........................................................ .............................. ....... 15.0g
Agar ............................................................ .............................. .. 14.0g
NaCl........................................................ .............................. ........ 9.5g
Yeast extract..................................................... ............................. 4.0g
Maltose ............................................................ ............................ 4.0g
Sodium pyruvate ............................................................ .............. 2.0g
X-glucoside chromogenic mix ..................................................... 0.2g
Differential lecithin solution.................................................... 40.0mL
Selective supplement solution .................................................20.0mL
pH 7.2 ± 0.2 at 25°C
Source: This medium is available from Oxoid Unipath.
Differential Lecithin Solution:
Composition per 40.0mL:
Lecithin ............................................................ .................. Proprietary
Preparation of Differential Lecithin Solution: Available as premixed
solution.
Selective Supplement Solution:
Composition per 20.0mL:
Nalidixic acid........................................................ ................... 26.0mg
Polymyxin B ............................................................ ................ 10.0mg
Amphotericin ............................................................ ............... 10.0mg
Ceftazidime................................................. .............................. . 6.0mg
Preparation of Selective Supplement Solution: Add components
to distilled/deionized water and bring volume to 20.0mL. Mix thoroughly.
Filter sterilize.
Preparation of Medium: Add components, except differential lecithin
solution and selective supplement solution, to distilled/deionized
water and bring volume to 940.0mL. Mix thoroughly. Gently heat
while stirring and bring to boiling. Autoclave for 15 min at 15 psi pressure–
121°C. Cool to 46°C. Aseptially add differential lecithin solution
and selective supplement solution. Mix thoroughly. Pour into sterile
Petri dishes.
Use: For the isolation, enumeration, and presumptive identification of
Listeria spp. and Listeria monocytogenes. This selective medium contains
the substrate lecithin, which permits differentiation of L. monocytogenes
and L. Ivanovii from other Listeria species. Differential activity
for all Listeria species is due to the addition of a chromogenic
substrate.
Chromogenic Listeria Agar (ISO)
Composition per liter:
Enzymatic digest of animal tissue .............................................. 18.0g
Agar ............................................................ .............................. .. 12.0g
LiCl ............................................................ .............................. ... 10.0g
Yeast extract..................................................... ........................... 10.0g
Enzymatic digest of casein ........................................................... 6.0g
NaCl ............................................................ .............................. .... 5.0g
Na2HPO4, anhydrous................................................... ................. 2.5g
Glucose ............................................................ ............................. 2.0g
Sodium pyruvate ............................................................ .............. 2.0g
Magnesium glycerophosphage ..................................................... 1.0g
MgSO4, anhydrous................................................... ..................... 0.5g
X-glucoside chromogenic mix ................................................... 0.05g
L-α-phosphotidylinositol .........................................................40. 0mL
Selective supplement solution .................................................20.0mL
pH 7.2 ± 0.2 at 25°C
Source: This medium is available from Oxoid Unipath.
Selective Supplement Solution:
Composition per 20.0mL:
Nalidixic acid........................................................ ................... 20.0mg
Ceftazidime................................................. ............................. 20.0mg
Amphotericin ............................................................ ...............10.0mg
Polymyxin B ............................................................ ............. 76,700 U
Preparation of Selective Supplement Solution: Add components
to distilled/deionized water and bring volume to 20.0mL. Mix thoroughly.
Filter sterilize.
Preparation of Medium: Add components, except L-α-phosphotidylinositol
and selective supplement solution, to distilled/deionized
water and bring volume to 940.0mL. Mix thoroughly. Gently heat
while stirring and bring to boiling. Autoclave for 15 min at 15 psi pressure–
121°C. Cool to 46°C. Aseptially add L-α-phosphotidylinositol
and selective supplement solution. Mix thoroughly. Pour into sterile
Petri dishes.
Use: For the isolation, enumeration, and presumptive identification of
Listeria spp. and Listeria monocytogenes. This selective medium contains
the substrate lecithin, which permits differentiation of L. monocytogenes
and L. Ivanovii from other Listeria species.
Chromogenic Listeria Agar (ISO) Modified
Composition per liter:
Enzymatic digest of animal tissue .............................................. 18.0g
Agar ............................................................ .............................. .. 12.0g
LiCl........................................................ .............................. ....... 10.0g
Yeast extract..................................................... ........................... 10.0g
Enzymatic digest of casein ........................................................... 6.0g
NaCl........................................................ .............................. ........ 5.0g
Na2HPO4, anhydrous................................................... ................. 2.5g
Glucose ............................................................ ............................. 2.0g
Sodium pyruvate.................................................... ....................... 2.0g
Magnesium glycerophosphage ..................................................... 1.0g
MgSO4, anhydrous ............................................................ ........... 0.5g
X-glucoside chromogenic mix ................................................... 0.05g
Differential lecithin solution.................................................... 40.0mL
Selective supplement solution .................................................20.0mL
pH 7.2 ± 0.2 at 25°C
Source: This medium is available from Oxoid Unipath.
Differential Lecithin Solution:
Composition per 40.0mL:
Lecithin ............................................................ .................. Proprietary
Preparation of Differential Lecithin Solution: Available as premixed
solution.
Selective Supplement Solution:
Composition per 20.0mL:
Nalidixic acid........................................................ ................... 20.0mg
Ceftazidime................................................. ............................. 20.0mg
Amphotericin ............................................................ ............... 10.0mg
Polymyxin B ............................................................ ............. 76,700 U
Preparation of Selective Supplement Solution: Add components
to distilled/deionized water and bring volume to 20.0mL. Mix thoroughly.
Filter sterilize.
Preparation of Medium: Add components, except differential lecithin
solution and selective supplement solution, to distilled/deionized
water and bring volume to 940.0mL. Mix thoroughly. Gently heat
while stirring and bring to boiling. Autoclave for 15 min at 15 psi pressure–
121°C. Cool to 46°C. Aseptially add differential lecithin solution
and selective supplement solution. Mix thoroughly. Pour into sterile
Petri dishes.
Use: For the isolation, enumeration, and presumptive identification of
Listeria spp. and Listeria monocytogenes. This selective medium contains
the substrate lecithin, which permits differentiation of L. monocytogenes
and L. Ivanovii from other Listeria species.
Chromogenic Salmonella Esterase Agar
(CSE Agar)
Composition per liter:
Agar ............................................................ .............................. .. 12.0g
Lactose..................................................... .............................. ..... 14.65
Peptone ............................................................ ............................. 4.0g
Tryptone.................................................... .............................. ...... 4.0g
Tween™ 20.......................................................... ......................... 3.0g
Lab Lemco....................................................... ............................. 3.0g
Na3-citrate dihydrate................................................... .................. 0.5g
L-cysteine.................................................. .............................. .. 0.128g
Tris ............................................................ .............................. .... 0.06g
SLA-octonoate solution ........................................................... 50.0mL
Novobiocin solution.................................................... .............10.0mL
Ethyl 4-dimethylaminobenzoate solution ................................10.0mL
pH 7.0 ± 0.2 at 25°C
Novobiocin Solution:
Composition per 10.0mL:
Novobiocin.................................................. ............................. 70.0mg
Preparation of Novobiocin Solution: Add novobiocin to distilled/
deionized water and bring volume to 10.0mL. Mix thoroughly.
Filter sterilize.
Ethyl 4-dimethylaminobenzoate Solution:
Composition per 10.0mL:
Ethyl 4-dimethylaminobenzoate ................................................. 0.35g
Methanol ............................................................ ........................8.0mL
Preparation of Ethyl 4-dimethylaminobenzoate Solution:
Add ethyl 4-dimethylaminobenzoate to 8.0mL methanol. Mix thoroughly.
Bring volume to 10.0mL with distilled/deionized water. Mix
thoroughly. Filter sterilize.
SLA-Octonoate Solution:
Composition per 50.0mL:
4-[2-(4-octanoyloxy-3,5-dimethoxyphenyl)-
vinyl]-quinolinium-1-(propan-3-yl
carboxylic acid) bromide
(SLPA-octanoate; bromide form) ..................................... 0.3223g
Preparation of SLA-Octonoate Solution: Add SLA-octonoate
to distilled/deionized water and bring volume to 50.0mL. Mix thoroughly.
Filter sterilize.
Preparation of Medium: Add components, except novobiocin solution,
SLA-octonoate solution, and ethyl 4-dimethylaminobenzoate
solution, to distilled/deionized water and bring volume to 920.0mL.
Mix thoroughly. Gently heat and bring to boiling. Autoclave for 15 min
at 15 psi pressure–121°C. Cool to 50°C. Aseptically add 10.0mL novobiocin
solution, 50.0mL SLA-octonoate solution, and 10.0mL ethyl
4-dimethylaminobenzoate solution. Mix thoroughly. Pour into sterile
Petri dishes.
Use: For the detection of Salmonella spp. in clinical specimens. For
the differentiation of Salmonella spp.
Chromogenic Substrate Broth
Composition per liter:
NaCl ............................................................ .............................. .. 10.0g
HEPES (N-[2-hydroxyethyl]
piperazine-N´-[2-ethanesulfonic
acid]) buffer ............................................................ .. 6.9g
(NH4)2SO4................................................... .............................. .... 5.0g
o-Nitrophenyl-β-D-galactopyranoside............. .............................. 0.5g
Solanium.................................................... .............................. ..... 0.5g
MgSO4 ............................................................ .............................. 0.1g
4-Methylumbelliferyl-β-D-glucuronide................... .................. 0.075g
CaCl2....................................................... .............................. ...... 0.05g
Na2SO3 ............................................................ ............................ 0.04g
Amphotericin B........................................................... ...............1.0mg
MnSO4 ............................................................ ........................... 0.5mg
ZnSO4 ............................................................ ............................ 0.5mg
Preparation of Medium: Add components to distilled/deionized
water and bring volume to 1.0L. Mix thoroughly. Distribute into tubes
or flasks. Autoclave for 15 min at 15 psi pressure–121°C.
Use: For the detection of coliform bacteria based on their hydrolysis of
chromogenic substrates by production of β-D-galactopyranosidase. Bacteria
that produce β-D-galactopyranosidase turn the medium yellow.
Chromogenic Urinary Tract Infection (UTI) Medium
Composition per liter:
Chromogenic mix ............................................................ ........... 26.3g
Peptone ............................................................ ........................... 15.0g
Agar ............................................................ .............................. .. 15.0g
pH 6.8 ± 0.2 at 25°C
Source: This medium is available as a premixed powder from Oxoid
Unipath.
Preparation of Medium: Add components to distilled/deionized
water and bring volume to 1.0L. Mix thoroughly. Gently heat while
stirring and bring to boiling. Autoclave for 15 min at 15 psi pressure–
121°C. Cool to 50°C. Pour into sterile Petri dishes.
Use: For the presumptive identification and differentiation of all the
main microorganisms that cause urinary tract infections (UTIs). The
medium contains two specific chromogenic substrates which are
cleaved by enzymes produced by Enterococcus spp., Escherichia coli,
and coliforms. In addition, it contains phenylalanine and tryptophan
which provide an indication of tryptophan deaminase activity, indicating
the presence of Proteus spp., Morganella spp., and Providencia
spp. It is based on electrolyte deficient CLED Medium which provides
a valuable non-inhibitory diagnostic agar for plate culture of other urinary
organisms, while preventing the swarming of Proteus spp. One
chromogen, X-Gluc, is targeted towards β-glucosidase, and allows the
specific detection of enterococci through the formation of blue colonies.
The other chromogen, Red-Gal, is cleaved by the enzyme β-
galactosidase which is produced by Escherichia coli, resulting in pink
colonies. Cleavage of both chromogens occurs in the presence of coliforms,
resulting in purple colonies. The medium also contains tryptophan
which acts as an indicator of tryptophan deaminase activity,
resulting in colonies of Proteus, Morganella, and Providencia spp.
appearing brown.
Lactic Bacteria Differential Agar
Composition per liter:
Agar ............................................................ .............................. .. 15.0g
Casein enzymic hydrolysate ....................................................... 10.0g
Casein acid hydrolysate ............................................................ .... 3.0g
Fructose.................................................... .............................. ....... 2.5g
KH2PO4...................................................... .............................. ..... 2.5g
Papaic digest of soybean meal...................................................... 1.5g