Overview
ISO 11866-1:2005 (IDF 170-1:2005) specifies a microbiological method for the enumeration of presumptive Escherichia coli and presumptive coliforms in milk and milk products. The method uses a liquid selective enrichment medium containing 4‑methylumbelliferyl‑β‑D‑glucuronide (MUG) combined with the Most Probable Number (MPN) technique after incubation at 30 °C. It is intended chiefly for samples suspected of containing comparatively low numbers of E. coli (less than 100/g or 10/ml).
Key Topics and Technical Requirements
- Principle: Inoculation of double‑ and single‑strength selective MUG broth, incubation, detection of fluorescence (MUG cleavage) and indole production for presumptive E. coli; gas formation in Durham tubes indicates presumptive coliforms.
- Incubation: 24 h ± 2 h at 30 °C, extendable up to 48 h ± 2 h if required.
- Confirmation steps:
- Add sodium hydroxide and inspect for fluorescence under a long‑wave UV lamp (360–366 nm).
- Add Kovács indole reagent to fluorescent tubes; a red colour in the alcoholic phase indicates indole production (presumptive E. coli).
- Enumeration: Count positive tubes across selected three consecutive dilutions and calculate the MPN index using standardized MPN tables (Annex A).
- Media and reagents: Modified lauryl sulfate tryptose broth with MUG (single and double strength), Kovács indole reagent, NaOH solution; strict preparation, storage and sterilization requirements apply.
- Apparatus: Autoclave, incubator (30 °C ±1 °C), long‑wave UV lamp, Durham tubes, pH meter, pipettes and vortex mixer.
- Limitations: Method is susceptible to variability; results must be interpreted in light of guidance (Clause 12). It is more rapid than some alternatives due to reduced enrichment steps.
Applications and Who Uses It
- Primary use: Enumeration of presumptive E. coli and coliforms in milk and a wide range of dairy matrices including liquid milk, dried milk powders, whey products, cheese and processed cheese, butter, frozen dairy products, custards and cream.
- Intended users:
- Dairy microbiology laboratories and quality control teams
- Food safety auditors and regulatory laboratories
- Manufacturers of milk and milk products performing routine testing or low‑level contamination screening
- Practical value: Supports HACCP, shelf‑life studies and compliance testing where low bacterial counts are expected; enables faster presumptive detection using fluorescence-based MUG cleavage combined with MPN enumeration.
Related Standards
- ISO 7251:2005 - horizontal MPN method for presumptive E. coli
- ISO 4831 - coliform enumeration (reference purposes)
- ISO 7218 - general microbiological requirements
- ISO 8261 / IDF 122 - sample preparation and initial suspensions
- ISO 707 / IDF 50 - sampling of milk and milk products
- ISO 11866‑2 / IDF 170‑2 - colony-count technique at 44 °C using membranes
Keywords: ISO 11866-1:2005, MUG, MPN technique, presumptive Escherichia coli, milk and milk products, coliforms, dairy microbiology, Kovács indole reagent.