This part of the ISO 16649 standard, developed by the International Organization for Standardization (ISO), This document provides for the detection of the enzyme β-glucuronidase by colony count technique after incubation at 44 °C on a solid medium containing a chromogenic component and resuscitation using membranes. indicates a horizontal method for positive Escherichia coli enumeration.
It applies to products for human consumption, products for animal nutrition, environmental samples in food production and food processing, and samples from the primary production stage such as animal excrement, powder and swab.
β-glucuronidase positive Escherichia coli bacteria forming typical, blue or blue-green colonies at 44 °C on tryptone-bile X-glucuronide agar (TBX) under the conditions specified in this document.
Counting of β-glucuronidase-positive Escherichia coli, determination of the number of colony-forming units (cfu) of β-glucuronidase-positive Escherichia coli per gram of sample, per milliliter, per square centimeter, or per sampling device when analysis is performed according to this document.
A specified amount of the test sample, initial suspension or decimal dilutions, is inoculated onto membranes placed on minerals modified glutamate agar (MMGA), followed by incubation at 37 °C for 4 hours.
For isolation, membranes from the resuscitation step on the MMGA are transferred to tryptonebilized X-glucuronide agar(TBX), followed by incubation at 44°C for 20 hours to 24 hours.
The number of colony-forming units (cfu) of β-glucuronidase positive Escherichia coli per gram or milliliter of sample is calculated from the number of typical blue or blue-green colonies per plate.
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