Staphylococcus aureus (11 o'clock) Klebsiella pneumoniae (5 o'clock) and Pseudomonas aeruginosa (8 o'clock) growth on CLED agar-Back View
Table of Contents
CLED stands for cysteine, lactose, and electrolyte-deficient. CLED agar or medium is a differential culture medium used for the isolation and enumeration of bacteria in urine specimens from suspected cases of urinary tract infection (UTI). It favors the growth of all potential urinary pathogens as well as a number of contaminants such as diphtheroid, lactobacilli, and micrococci. Candida also enjoys this medium. This is a sole medium alternative for routine urine culture and inoculation of specimens in a combination of blood agar (BAP) and MacConkey agar (MAC) without compromising the quality and reducing the cost.
The composition of CLED Medium is shown below in the table-
| S.No. | Ingredients | Amount |
| 1. | Peptone | 2 gm |
| 2. | “Lab Lemco” Powder (meat extract) | 1.5 gm |
| 3. | Tryptone | 2 gm |
| 4. | Lactose | 5 gm |
| 5. | L-Cysteine | 0.064 |
| 6. | Bromothymol blue | 0.01g |
| 7. | Agar | 7.5 gm |
| 8. | Distilled water | 500 ml |
| Final pH | 7.3 +/- 0.2 at 25°C |
CLED agar is available in dehydrated powder form by various manufacturers like Mast, Oxoid, Difco, Himedia, and so on.
It can be used for several weeks, but should be free from any change in the appearance of the medium, showing contamination, deterioration, or alteration of pH.
Reduced electrolyte prevents the swarming of Proteus, whereas Cystine promotes the formation of cystine-dependent dwarf colonies. Bromothymol blue is the indicator used in the agar; it changes to yellow in case of acid production during fermentation of lactose or changes to deep blue in case of alkalization. Lactose-fermenting bacteria build yellow colonies. Bacteria that decarboxylate L-Cystine cause an alkaline reaction and build deep blue colonies.
Typical Colony Morphology on CLED Agar is as follows:
| S.No. | Organisms | Colony Morphology |
| 1. | Staphylococcus aureus | Deep yellow colonies of uniform color |
| 2. | S. saprophyticus and other Coagulase Negative Staphylococci (CoNS) | Pale yellow to white colonies, more opaque than Enterococcus faecalis |
| 3. | Enterococci | Small yellow colonies, about 0.5mm in diameter |
| 4. | E. coli: Introduction, Identification Features, Keynotes, and Escherichia coli Footages Escherichia coli | Opaque yellow colonies with a slightly deeper yellow center |
| 5. | Klebsiella species | Large mucoid yellow to whitish-blue colonies |
| 6. | Proteus spp. | Translucent blue-grey colonies |
| 7. | Pseudomonas aeruginosa | Green colonies with typical matted surface and rough periphery |
The difference in color due to different pH effects is as follows-
| pH | Color |
| 7.4 | deep blue |
| 7.0 | bluish grey |
| 6.8 | pale grey |
| 6.6 | pinkish grey |
| 6.4 | bright red with a whitish tinge |
| 6.0 | bright red |
For better results, the medium should not be incubated for more than 24 hours because if lactose fermenters predominate, the entire medium may turn pink, masking the presence of non-lactose fermenters. Inoculate the medium immediately after urine collection.
Typical colony morphology on CLED Medium with Andrade’s Indicator after incubation at 35-37°C for 18-24 hours is as follows:
| S.No | Microbes | Colony Characteristics |
| 1. | Klebsiella aerogenes ATCC 13048 | greyish green, mucoid |
| 2. | Escherichia coli ATCC 25922 | bright pink with a pink halo |
| 3. | Enterococcus faecalis ATCC 29212 | orange-yellow or greenish |
| 4. | Proteus mirabilis ATCC 25933 | blue-green |
| 5. | Staphylococcus aureus ATCC 25923 | golden-yellow |
| 6. | Streptococcus pyogenes ATCC 19615 | greyish green |
(CLED Agar for Urine Culture)
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