Overview
ISO/TS 26844:2006 (IDF/RM 215:2006) specifies a microbiological tube diffusion test for the detection of a broad range of antimicrobial residues in milk and milk products. The method is applicable to raw milk, heat‑treated milk and reconstituted dried milk and is intended for routine screening to indicate the presence of inhibitory substances that affect microbial growth.
Key topics and technical requirements
- Principle: Two agar test tubes inoculated with Geobacillus stearothermophilus ATCC 10149 (approx. 5 000 000 CFU/ml) are used. In the absence of inhibitors the organism grows and a pH indicator (bromocresol purple) changes colour from purple to yellow. Presence of antimicrobials prevents growth and the indicator remains purple.
- Two tube formats:
- Tube A - pH 7.0, supplemented with chloramphenicol; enhanced sensitivity to tetracyclines (example: oxytetracycline).
- Tube B - pH 8.0, supplemented with trimethoprim; improved detection of β‑lactams, macrolides, aminoglycosides, sulfonamides and trimethoprim.
- Culture media and additives: Basic medium components and specific additions (bromocresol purple indicator, chloramphenicol or trimethoprim solutions) are described for reproducible preparation. pH control at dispensing temperature (63 °C ± 1 °C) is specified.
- Incubation/interpretation: Tubes are incubated under defined conditions (media prepared so colour change occurs within ~4 h 15 min ± 30 min when no inhibitor is present). Lack of colour change indicates inhibitory residues.
- Quality control: New batches of test‑organism suspension must be checked using standard spiked solutions (examples: benzylpenicillin 2 µg/kg, sulfadiazine 150 µg/kg, neomycin 30 µg/kg, erythromycin 10 µg/kg, oxytetracycline 100 µg/kg) to confirm sensitivity.
- Storage: Prepared tubes may be stored at 0 °C to +5 °C for limited periods (e.g., up to 3 days) with evaporation prevention.
Applications and who uses it
- Dairy processors and quality control labs: routine screening of raw and processed milk for antimicrobial contamination before processing or sale.
- Food safety and regulatory laboratories: surveillance and compliance monitoring for veterinary drug residues.
- Third‑party testing services: rapid, cost‑effective screening prior to confirmatory analysis.
The test serves as a presumptive screen; positive samples typically require confirmatory testing by specific methods (e.g., chromatographic or receptor assays).
Related standards
- ISO 4833 - enumeration of micro‑organisms
- ISO 7218 - general microbiological requirements and guidance
- ISO 13969 | IDF 183 - standardized description of microbial inhibitor tests
- ISO 18330 | IDF 188 - guidelines for immunoassays/receptor assays for antimicrobial residues
Keywords: ISO/TS 26844:2006, tube diffusion test, antimicrobial residues, milk testing, Geobacillus stearothermophilus, dairy quality control, screening method.