Overview:
Clostridial Agar (also known as Clostridium Selective Agar) is a selective and enriched solid medium formulated for the isolation and cultivation of pathogenic Clostridium species, such as C. perfringens (gas gangrene), C. tetani (tetanus), and C. botulinum (botulism), from mixed flora in clinical, food, and environmental specimens. Developed to mimic anaerobic conditions through reducing agents, it provides essential nutrients like peptides, carbohydrates, and sulfur sources while incorporating selective inhibitors (neomycin sulfate and sodium azide) to suppress Gram-negative enteric bacteria (e.g., Proteus, Pseudomonas) and other contaminants. The low oxidation-reduction potential supports spore germination and vegetative growth under anaerobic incubation at 35-37�C for 18-48 hours, yielding opaque, flat, 2-5 mm colonies that are typically non-hemolytic. This medium is recommended in clinical microbiology for wound, blood, and tissue cultures, and complies with APHA and ISO standards for food safety testing.
Applications:
?? Selective isolation of Clostridium perfringens, C. tetani, and C. sporogenes from clinical samples (e.g., blood, wounds, feces) for diagnosing clostridial infections
?? Detection in food and dairy microbiology (e.g., canned goods, meat) per FDA-BAM and ISO 7937 for foodborne botulism or gas gangrene risks
?? Environmental monitoring of Clostridium in soil, sewage, and animal products
?? Subculture from enrichment broths (e.g., cooked meat medium) for confirmation via biochemical tests or toxin assays
Key Features:
?? Reducing agents (sodium thioglycollate and sodium formaldehyde sulfoxylate) maintain anaerobic conditions, supporting >50% recovery of stressed ATCC strains like C. perfringens 12924
?? Selective inhibitors (neomycin and azide) suppress >90% of competing flora (e.g., E. coli, S. aureus) while permitting luxuriant Clostridium growth
?? Enriched with L-cystine for cysteine-dependent strains; dextrose for energy without excessive acid production
?? Validated in CLSI and APHA protocols; promotes characteristic opaque, irregular colonies under anaerobic jars (e.g., GasPak) at 35-37�C
?? Opaque, light amber medium post-preparation; suitable for direct plating or tube inoculation from specimens
Composition (per liter):
Tryptone � 17.0 g
Soya Peptone � 3.0 g
Dextrose � 6.0 g
Sodium Chloride (NaCl) � 2.5 g
Sodium Thioglycollate � 1.8 g
L-Cystine � 0.25 g
Sodium Formaldehyde Sulphoxylate � 1.0 g
Neomycin Sulphate � 0.15 g
Sodium Azide � 0.2 g
Agar � 14.5 g
Final pH: 7.0 � 0.2 at 25�C
Preparation:
Suspend 46.4 g of the dehydrated medium in 1 liter of purified/distilled water. Heat to boiling with frequent agitation until completely dissolved (do not overheat). Autoclave at 118�C (15 psi) for 15 minutes. Cool to 45-50�C in a water bath. Mix thoroughly to ensure even distribution and pour into sterile Petri plates (20-25 mL per plate) or tubes for deep inoculation. Allow to solidify at room temperature. For use, streak or stab-inoculate specimens under anaerobic conditions; incubate at 35-37�C for 18-48 hours in an anaerobic jar. Prepared medium should appear light amber, slightly opalescent, and homogeneous without precipitates.
Storage:
Dehydrated medium: Store in a tightly closed container at 10�30�C, protected from moisture, light, and humidity; stable until expiry date (typically 3 years). Once opened, recap tightly to prevent hygroscopic clumping.
Prepared plates/tubes: Store at 2�8�C in sealed containers under anaerobic conditions if possible; use within 2-4 weeks. Discard if dehydrated, contaminated, discolored, or overgrown.
?? For laboratory and research use only. Not for diagnostic procedures without proper validation. Not for human or veterinary consumption. Handle under biosafety level 2; use Universal Precautions for potentially toxigenic specimens�wear gloves, lab coat, and eye protection. Autoclave all waste before disposal. Anaerobic incubation essential; aerobic exposure may inhibit growth. Confirm isolates with motility, spore staining, and toxin detection. Not suitable for non-clostridial anaerobes without modifications.4.8s