Water Quality Field References

• Temperature
• pH
• Secchi Disk Transparency
• Salinity and Conductivitiy
• Dissolved Oxygen
• E. Coli
• Nitrates
• Phosphates
• Physical Characteristics
• Weather Indications
• Tide Stages

E. Coli

Testing Parameters

Current methodology for the determination of drinking water, wastewaters, and surface water quality generally emphasizes the qualification or quantification of general coliforms and other fecal coliforms or E. coli. The US EPA (http://www.epa.gov/owowwtrl/monitoring/volunteer/stream/vms511.html) has determined that E. coli is the best indicator of health risk to humans from fresh water instead of the traditionally held idea that "fecal coliforms" should be the target organisms. E. coli is a species of fecal coliform that is specific to the intestinal tract of humans and other warm-blooded vertebrates, while the term "fecal coliform" refers only to gram negative, lactose fermenting, rod shaped bacteria which grown at a temperature of 44.5°C.

Unfortunately, not all the bacteria which will grow under this condition are of fecal origin, and some bacteria that will not grow under this elevated temperature may be strains of temperature intolerant coliforms of fecal origin. There are even some strains of E. coli which will not grow at 44.5°C due to genetic or injury factors, which will grow at lower temperatures. Some procedures, such as for mTEC, attempt to compensate for this by incubating initially for two hours at 35°C as a "resuscitation" period prior to 22 hours incubation at the 44.5°C. Therefore, depending upon the use of temperature tolerance to identify "fecal coliforms" can result in misidentification and less numbers than are actually present in a sample.

Testing Parameters

This monitoring procedure is for testing Escherichia Coli (E. coli) with "Coliscan Easygel."

At the Monitoring Site:

• Collect water using a whirl-pak bag*. Please make sure to collect your sample from a depth of one foot (if your sample site has less than two feet of water, collect your sample at the midpoint of the water column) and conduct two tests using two 5 mL sample volumes at your site.

  * When collecting your sample with the whirl-pak bag, label with site name and time sampled, flip then twist to close. Place on ice to transport to your testing area. If you are testing multiple sites, include an indication of which site this sample represents.

   

At your home, school, or other testing station:

• Shake whirl-pak bag vigorously, then carefully open without touching the lip of the bag. Use your disposable dropper to draw the appropriate sample size from the whirl-pak container (1 mL, 3mL, or 5mL). Deposit sample into Easygel bottle, seal and swirl gently. NOTE: Water samples kept longer than 1 hour prior to plating should be kept on ice until plated (up to 6 hours). A Coliscan Easygel bottle that has had sample placed into it for transport longer than 10 minutes should be kept on ice or in a refrigerator (up to 2 hours) until plated.
• Label each petri dish with the following information: Site name, date, volume of sample, and time sample is poured into the petri dish. Then pour the prepared sample (the Easygel solution with the stream water mixed in) slowly into the petri dish. Swirl gently until there is a smooth coating of solution across the bottom of the petri dish. Allow 10 minutes for the media to gel, then place in the incubator.
  1. An ideal temperature may be maintained by creating a simple light bulb/cardboard box incubator. Use a thermometer to determine temperature with a 40-watt light bulb until the temperature is approximately 35 C. At this temperature, colonies should be completely developed at 24 hours. No counts should be made after 32 hours when using incubation temperature control measures.
  2. Count all the dark purple and dark blue colonies and record the results on the CRWN data sheet. Sort and count all colonies and record this and other required information on the CRWN E. coli data sheet.

     Incubation Option: Use a Styrofoam type cooler with a lid, put in one end a beaker or a container with ½ to 1 gallon of hot boiled water, place plate or dish at other end. An alternative to the boiling water, heat in a microwave several blue hot/cold packs. This process will not give a set uniform temperature like an incubator, but will provide a warmer than room temperature for quicker growth. This should give results in 28-36 hours. Incubator kits can be purchased direct from GQF Manufacturing Company, P.O. Box 1552, Savannah, Georgia 31498, phone: 912/236-0651.

  
  Photo of 24 hr membrane filter incubated at 35° C. (22 E.coli, 30 general coliform, 1 teal blue-green, 1 Pseudomonas, and about 20 colorless colonies.

  Specific colonies above are designated 1, 2, 3, 4 or 5 and their interpretation is described below with the corresponding number.

   

  • Colonies with (1) are interpreted as E.coli. (glucuronidase +, galactosidase+)
  • Colonies with (2) are interpreted as general coliform. (glucuronidase-, galactosidase+)
  • Colony with (3) could be E.coli, Salmonella, Shigella or other genus. (glucuronidase+, glactosidase-)
  • Colony with (4). The color is from the bacteria, not the chromogens. Identified as Pseudomonas sp. (glucuronidase-, galactosidase-)
  • Colony with (5) has no color. Not E.coli or general coliform. (glucuronidase-, galactosidase-)

   

• Like fecal coliform, E. Coli is recorded in colonies per 100 mL of water. Therefore, multiply the sample size by the appropriate dilution factor** to determine how many colonies per 100mL your results represent. Record the product on the data sheet in colonies per 100mL.

  **The dilution factor = 100/volume of sample water added to Easygel solution. For example, if you collected a sample size of 1mL in the disposable dropper and added this to the solution, your dilution factor is 100/1 or 100. This means you multiply the number of purple colonies counted X 100 to determine colonies per 100mL. Common dilution factors are 3 mL sample - dilution factor 33.3, 5 mL sample - dilution factor 20.

• Dispose of the treated petri dishes by placing about 1 teaspoon of bleach or alcohol onto the surface of the medium of each plate. Allow dish to sit at least 5 minutes. Place in a water tight sealed bag and discard in trash.

Data Sheet Example:

Sample Size 3 mL Reading #1 (colonies counted) 8 x 33.3 (dilution factor*) = 266 coliform colonies/100mL Sample Size 3 mL Reading #2 (colonies counted) 11 x 20 (dilution factor*) = 220 coliform colonies/100mL *dilution factor = 100 divided by volume of sample used in test (e.g. 3 mL sample = dilution factor 33.3; 5 mL sample = dilution factor 20)

 

Designated Beach = 235 colonies/mL      ¤      Lightly Used Full Body Contact = 406 colonies/100mL