Clostridium PET For the rapid identification of Clostridium perfringens and Clostridium difficile

• SIMPLE Two tubes to inoculate, one reagent addition.
• EASY TO READ Distinct colour changes.
• ECONOMICAL Significantly less expensive than antisera or multi-tube systems.
• RELIABLE Based on established biochemical reactions.
• RAPID Results within 3 hours.

Clostridium PET

Consists of 2 tubes and spot indole reagent, each tube yields 2 biochemical reactions. These reactions can presumptively identify the major pathogens – Clostridium difficile and Clostridium perfringens from suitable cultures.

Principle
Tube 1 contains a TABLET and yields ONPG (beta galactosidase) and PYR (pyrridonyl aminopeptidase) results. Tube 2 contains a DISC and yields PO4 (alkaline phosphotase) and PRO (proline aminopeptidase) results. Spot Indole (PEP reagent) detects metabolism of tryptophane to indole.

Method
Cultures should be fresh and pure.

TABLET ONPG & PYR
Add 5 drops of distilled water and inoculate heavily with a loop-full of organism from a fresh culture. Incubate aerobically uncovered for 2 hours at 37°C. The appearance of a bright yellow colour is a positive ONPG result, a negative result is no change or a very pale yellow. After recording the ONPG result add 1-2 drops of PEP reagent (spot indole). A colour change to pink or red or dark orange is a positive result for PYR. A negative result is yellow or pale orange.

DISC PO4 & PRO
Add 3 drops of distilled water and inoculate heavily with a loop-full of organism from a fresh culture. Incubate aerobically uncovered for 2 hours at 37°C. The appearance of a bright yellow colour is a positive PO4 result, a negative result is no change or a very pale yellow. After recording the PO4 result add 1-2 drops of PEP reagent (spot indole). A colour change to pink or red or dark orange is a positive result for PRO. A negative result is yellow or pale orange. If the organism gives reactions indicating the possibility of C. difficile perform a spot indole with the reagent provided. C. difficile is indole negative. C. bifermentans and C. sordellii give similar biochemical reactions but are indole positive.

SPOT INDOLE
From blood agar or any medium containing tryptophane, smear a portion of colony onto filter paper, add 1 drop of the PEP reagent, almost instant development of a blue/purple colour is a positive reaction.