EPA-OW: 1106.1:  Enterococci in water by MF using mE-EI Agar

  • Summary
  • Analytes
  • Revision
  • Data and Sites
Official Method Name
 Enterococci in water by membrane filtration using membrane-Enterococcus-Esculin Iron Agar (mE-EIA)
Current Revision
July 2006
Media
 WATER
Instrumentation
 Membrane Filtration
Method Subcategory
 Microbiological
Method Source
  EPA-OW
Citation
 USEPA, 2006, Method 1106.1: Enterococci in water by membrane filtration using membrane-Enterococcus-Esculin Iron Agar (mE-EIA): U.S. Environmental Protection Agency Report 821-R-06-008, 42 p.
Brief Method Summary
 This method provides a direct count of bacteria in water based on the development of colonies on the surface of the membrane filter. A water sample is filtered through the membrane which retains the bacteria. Following filtration, the membrane is placed on a selective medium, mE agar, and incubated at 41oC + 0.5oC for 48 + 3 hours. The filter is then transferred to a differential medium, EIA, and incubated at 41oC + 0.5oC for 20-30 minutes. Pink to red enterococci colonies on mE will develop a black or reddish-brown precipitate on the underside of the filter after transfer to EIA. A fluorescent lamp with a magnifying lens is used for counting to give maximum visibility of colonies.
Scope and Application
 This method describes a membrane filter procedure for the detection and enumeration of the enterococci bacteria in ambient water. The enterococci test is recommended as a measure of ambient fresh and marine recreation water quality.
Applicable Concentration Range
 20-80 colonies per membrane
Interferences
 Water samples containing colloidal or suspended particulate materials can clog the membrane filter and prevent filtration, or cause spreading of bacterial colonies which could interfere with enumeration and identification of target colonies.
Quality Control Requirements
 The minimum analytical QC requirements for the analysis of samples using this method include routine analysis of positive and negative controls, filter sterility checks, method blanks, and media sterility checks.
Additional analytical QC for the analysis of samples using this method include an initial demonstration of laboratory capability through performance of the initial precision and recovery (IPR) analyses, and ongoing demonstration of laboratory capability through performance of the ongoing precision and recovery (OPR) analysis. For the IPR and OPR analyses, it is necessary to spike PBS samples with either laboratory-prepared spiking suspensions or BioBalls.
Sample Handling
 Samples are collected by hand or with a sampling device if the sampling site has difficult access such as a dock, bridge, or bank adjacent to surface water. Composite samples should not be collected, since such samples do not display the range of values found in individual samples. The sampling depth for surface water samples should be 6-12 inches below the water surface. Sample containers should be positioned such that the mouth of the container is pointed away from the sampler or sample point. After removal of the container from the water, a small portion of the sample should be discarded to allow for proper mixing before analyses.
Ice or refrigerate bacteriological samples at a temperature of <10oC during transit to the laboratory. Do not freeze the samples. Use insulated containers to assure proper maintenance of storage temperature. Ensure that sample bottles are not totally immersed in water during transit or storage.
Maximum Holding Time
 Analysis preferably within 2 hours of collection; max. transport time to lab is 6 hours, and samples should be processed within 2 hours of receipt at lab.
Relative Cost
 Less than $50
Sample Preparation Methods
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