How to Check for E. Coli in Water

Checking for E. coli in water is more than just a scientific procedure; it’s a critical step in safeguarding public health. From a refreshing sip from a well to a dip in a community swimming pool, the invisible threat of bacterial contamination, particularly from Escherichia coli (E. coli), looms large. While often harmless inhabitants of the human gut, certain strains of E. coli are notorious pathogens, capable of causing severe gastrointestinal illness, kidney failure, and even death. This guide delves into the essential methods for detecting E. coli in water, providing a comprehensive, actionable framework for individuals, businesses, and public health officials alike.

The Invisible Threat: Why E. coli Matters in Water

E. coli is a bacterium commonly found in the intestines of warm-blooded animals, including humans. Its presence in water indicates fecal contamination, suggesting that other, more dangerous pathogens, such as Salmonella, Giardia, or norovirus, might also be present. These pathogens can cause a range of waterborne diseases, from mild diarrhea to life-threatening infections.

Understanding the sources of E. coli contamination is crucial for prevention. These can include:

• Agricultural Runoff: Animal waste from farms can be carried by rain into rivers, lakes, and groundwater.

• Malfunctioning Septic Systems: Leaking or improperly maintained septic tanks can release raw sewage into the environment.

• Sewage Overflows: Heavy rainfall can overwhelm municipal sewage systems, leading to untreated sewage discharge into waterways.

• Wildlife: Feces from birds, deer, and other animals can contaminate private wells and recreational waters.

• Recreational Activities: Swimmers carrying E. coli on their bodies can introduce it into pools or natural bodies of water.

The health risks associated with pathogenic E. coli in water are significant. Symptoms can range from abdominal cramps, diarrhea (often bloody), nausea, and vomiting to more severe conditions like hemolytic uremic syndrome (HUS), a serious complication that can lead to kidney failure, particularly in young children and the elderly. Given these risks, timely and accurate detection of E. coli is paramount.

Understanding the Basics of Water Testing

Before diving into specific methods, it’s important to grasp some fundamental concepts in water microbiology:

• Indicator Organisms: We don’t directly test for every possible pathogen in water because it’s impractical and expensive. Instead, we look for “indicator organisms.” E. coli is an excellent indicator because:
  • It’s abundant in the feces of warm-blooded animals.

  • It’s relatively easy and inexpensive to detect.

  • It generally survives in water longer than most waterborne pathogens, providing a conservative measure of risk.

  • Its presence strongly suggests fecal contamination and the potential presence of other harmful microorganisms.

• Coliforms vs. E. coli:

  • Total Coliforms: This is a broad group of bacteria found in soil, vegetation, and the intestines of warm-blooded animals. While their presence in drinking water indicates a potential problem (such as inadequate treatment or a breach in the distribution system), it doesn’t definitively confirm fecal contamination.

  • Fecal Coliforms: A subset of total coliforms, fecal coliforms are more specifically associated with the feces of warm-blooded animals. While a better indicator than total coliforms, they can also be found in environmental sources not directly related to fecal matter.

  • E. coli: The most specific and reliable indicator of recent fecal contamination. If E. coli is detected in a drinking water sample, it indicates that the water has been contaminated with fecal material and may contain disease-causing pathogens.

Sampling: The Crucial First Step

The accuracy of any water test hinges entirely on proper sample collection. A contaminated sample or an improperly collected one can lead to misleading results, potentially putting health at risk or causing unnecessary alarm.

General Principles for Sample Collection:

1. Sterility is Paramount: Use sterile sample bottles provided by the testing laboratory or purchased from a reputable supplier. Do not rinse the bottle or cap with the water to be tested.

2. Avoid Contamination: Do not touch the inside of the bottle or the inside of the cap. Avoid touching the water source directly with your hands or any non-sterile object.

3. Representative Sample: The sample should accurately reflect the water quality at the point of collection.

4. Timeliness: Analyze samples as soon as possible after collection, ideally within 6 hours, and definitely within 24 hours. Bacterial populations can change rapidly after collection, leading to inaccurate results. Transport samples on ice or in a cooler to maintain temperature and minimize bacterial growth.

Specific Sampling Procedures:

• Tap Water (Household/Building):
  • Choose a cold water tap (not a mixer tap) that is regularly used. Avoid taps that leak or have aerators or filter attachments (remove them if present).

  • Before sampling, clean the faucet outlet with an alcohol swab or paper towel to remove any surface contamination.

  • Turn on the cold water and let it run at full flow for at least 2-3 minutes to flush the line and clear any stagnant water. This ensures you’re testing the water from the main supply, not just what’s sitting in the tap.

  • Reduce the flow to a steady stream.

  • Carefully fill the sterile sample bottle to the indicated fill line without overflowing. Leave some headspace (usually 1 inch) as indicated on the bottle for mixing.

  • Replace the cap securely.

  • Record the date, time, and location of the sample.

• Well Water:

  • Choose a tap as close to the well as possible, ideally before any treatment systems (softeners, filters) if you want to assess the raw well water quality. If you want to check the quality of water after treatment, collect from a tap after the system.

  • Follow the same flushing procedure as for tap water.

  • If sampling from a spigot outside, ensure it’s clean and has not been recently used for purposes that might introduce contaminants (e.g., filling a bucket with dirty water).

• Natural Water Bodies (Lakes, Rivers, Ponds):

  • Choose a sampling point that represents the area of concern.

  • Submerge the sterile bottle just below the water surface (about 6-12 inches) with the opening facing upstream (if in a flowing body of water) to avoid collecting disturbed sediment.

  • Fill the bottle, leaving headspace, and cap securely.

  • Avoid disturbing the bottom sediments while collecting.

  • If sampling from a boat, ensure the boat is stationary and the sample is taken away from the immediate wake or engine exhaust.

• Swimming Pools/Spas:

  • Collect samples away from skimmers or inlets.

  • Submerge the bottle elbow-deep and away from the body.

  • Follow general sterility guidelines.

Key Information to Record for Every Sample:

• Date and time of collection

• Exact location of collection (e.g., “Kitchen Sink, 123 Main St” or “North end of Lake Clear, 5 feet from shore”)

• Water temperature at the time of collection (if possible)

• Any unusual observations (e.g., cloudy water, strong odor)

• Sampler’s name

Laboratory-Based Testing Methods

For the most accurate and reliable E. coli detection, sending samples to a certified laboratory is highly recommended. These laboratories employ sophisticated methods and trained personnel to ensure precise results.

1. Most Probable Number (MPN) Method

The MPN method is a statistical approach used to estimate the concentration of viable bacteria in a sample. It’s based on the idea that if a sample contains bacteria, diluting it progressively will eventually lead to dilutions where some tubes show growth and others do not.

Principle: Serial dilutions of the water sample are inoculated into multiple tubes containing a liquid growth medium (e.g., Lactose Broth or Lauryl Tryptose Broth) that promotes the growth of coliforms. These tubes are then incubated. The presence of gas production and/or turbidity in the tubes indicates presumptive coliform growth. Positive tubes are then subcultured into a confirmatory medium (e.g., Brilliant Green Bile Lactose Broth for fecal coliforms or EC Medium for E. coli) and incubated at a higher temperature (44.5∘C±0.2∘C). Gas production in these confirmatory tubes suggests the presence of fecal coliforms or E. coli. The pattern of positive and negative tubes across the dilutions is compared to an MPN table to estimate the most probable number of bacteria per 100 mL of sample.

Procedure (Simplified):

• Presumptive Test:
  • Prepare multiple sets of tubes (e.g., 5 tubes of 10 mL, 5 tubes of 1 mL, 5 tubes of 0.1 mL) with a specific broth medium.

  • Inoculate each tube with the corresponding volume of the water sample.

  • Incubate at 35∘C±0.5∘C for 24-48 hours.

  • Observe for gas production (indicated by a gas bubble in an inverted Durham tube) and/or turbidity. Record positive tubes.

• Confirmed Test (for Fecal Coliforms):

  • From each positive presumptive tube, transfer a small amount to a tube containing Brilliant Green Bile Lactose Broth.

  • Incubate at 44.5∘C±0.2∘C for 24 hours.

  • Observe for gas production. Positive tubes indicate fecal coliforms.

• Confirmed Test (for E. coli):

  • From each positive confirmed fecal coliform tube, perform an indole test or subculture to a selective E. coli medium. Alternatively, directly from positive presumptive tubes, subculture to EC Medium for incubation at 44.5∘C±0.2∘C.

  • Further tests (e.g., Indole, Methyl Red, Voges-Proskauer, Citrate – IMViC tests) can be performed on isolates to confirm E. coli.

Pros:

• Can quantify bacterial concentration.

• Relatively robust for turbid samples.

Cons:

• Labor-intensive and time-consuming (results in 2-4 days).

• Requires multiple tubes and specific media.

• Results are statistical estimations, not direct counts.

2. Membrane Filtration (MF) Method

The MF method is generally preferred for water samples with low turbidity and provides a direct count of bacterial colonies.

Principle: A measured volume of water is passed through a sterile membrane filter with a pore size small enough to retain bacteria (typically 0.45 microns). The filter, now containing any trapped bacteria, is then placed onto a selective agar medium (e.g., m-Endo agar for total coliforms, m-FC agar for fecal coliforms, or Chromocult Coliform Agar for E. coli) and incubated. Each visible colony that grows on the filter theoretically represents one bacterium from the original sample.

Procedure (Simplified):

• Sample Filtration:
  • Assemble a sterile filtration apparatus (funnel, filter holder, vacuum flask).

  • Place a sterile membrane filter onto the filter support.

  • Carefully pour a measured volume of the water sample (e.g., 100 mL for drinking water) into the funnel.

  • Apply a vacuum to draw the water through the filter.

  • Rinse the funnel walls with sterile buffered water to ensure all bacteria are drawn onto the filter.

• Incubation:

  • Carefully remove the filter using sterile forceps and place it onto a selective agar plate.

  • Invert the plate and incubate at the appropriate temperature and duration (35∘C for total coliforms, 44.5∘C for fecal coliforms, or 37∘C for E. coli on chromogenic media).

• Colony Counting and Confirmation:

  • After incubation, count the characteristic colonies. For E. coli on chromogenic media, colonies will typically appear blue or purple due to specific enzyme reactions (e.g., glucuronidase activity).

  • Confirm presumptive E. coli colonies with further biochemical tests (e.g., Indole test).

Pros:

• Provides a direct count of bacteria (Colony Forming Units – CFU/100 mL).

• Faster than MPN for results (18-24 hours for E. coli on chromogenic media).

• Less labor-intensive for high sample volumes.

Cons:

• Can be affected by high turbidity in the sample, which can clog the filter.

• Requires specific equipment and sterile technique.

3. Presence/Absence (P/A) Tests

These tests are designed for a simple “yes” or “no” answer regarding the presence of coliforms or E. coli in a fixed volume of water, typically 100 mL for drinking water. They are often used for routine monitoring of potable water.

Principle: A specific volume of water (e.g., 100 mL) is added to a sterile bottle or bag containing a selective growth medium (e.g., Colilert, IDEXX Quanti-Tray system). The medium contains indicator chemicals that change color or fluoresce in the presence of specific enzymes produced by coliforms and E. coli.

Common P/A Systems:

• Colilert (IDEXX): This is a widely used and highly reliable method.
  • Principle: The Colilert reagent contains ONPG (o-nitrophenyl-β-D-galactopyranoside) and MUG (4-methylumbelliferyl-β-D-glucuronide). Total coliforms produce β-galactosidase, which hydrolyzes ONPG, causing the sample to turn yellow. E. coli produces β-glucuronidase, which hydrolyzes MUG, causing the sample to fluoresce under UV light.

  • Procedure: Add 100 mL of water to the Colilert reagent and mix. Incubate at 35∘C for 24 hours. Observe for yellow color (total coliforms) and fluorescence under a UV lamp (365 nm) (for E. coli).

  • Quanti-Tray/2000 (for enumeration): If quantitative results are needed, the yellow/fluorescent sample can be poured into a Quanti-Tray/2000, which is a sealed tray with 97 wells of varying sizes. After incubation, the number of yellow and fluorescent wells is counted, and an MPN table is used to determine the bacterial concentration.

Pros:

• Simple to perform and interpret.

• No filtration or separate media preparation needed.

• Rapid results (24 hours).

• Can detect both total coliforms and E. coli simultaneously.

• Less prone to interference from turbidity.

Cons:

• Only provides presence/absence for a fixed volume, unless using a quantitative version like Quanti-Tray.

• Requires a UV lamp for E. coli confirmation.

4. Polymerase Chain Reaction (PCR) and Quantitative PCR (qPCR)

These molecular methods directly detect the DNA of E. coli (or specific pathogenic strains of E. coli), offering high specificity and rapid results.

Principle: PCR amplifies specific DNA sequences present in the sample. If E. coli DNA is present, millions of copies of a target gene are created, which can then be detected. qPCR allows for real-time monitoring of this amplification, providing quantitative data on the initial amount of E. coli DNA.

Procedure (Simplified):

• DNA Extraction: DNA is extracted from the water sample. This often involves concentrating the bacteria from a larger volume of water (e.g., by filtration or centrifugation) before extraction.

• PCR Amplification: The extracted DNA is mixed with primers (short DNA sequences that bind to specific E. coli genes), DNA polymerase, and nucleotides. The mixture undergoes cycles of heating and cooling to amplify the target DNA.

• Detection (for qPCR): During amplification, fluorescent dyes or probes bind to the newly synthesized DNA. A specialized instrument monitors the fluorescence intensity in real-time. The more E. coli DNA initially present, the faster the fluorescence signal rises.

• Analysis: The data from qPCR can be used to quantify the amount of E. coli DNA, which correlates to the number of E. coli cells in the original sample.

Pros:

• Highly specific; can differentiate between E. coli and other bacteria, and even identify specific pathogenic strains (e.g., O157:H7).

• Very rapid results (hours, compared to days for culture methods).

• Can detect non-viable but still potentially harmful cells (DNA from dead cells can still be amplified).

• Less affected by sample turbidity or other contaminants.

Cons:

• Requires expensive specialized equipment and highly trained personnel.

• Doesn’t distinguish between live and dead cells (which may be a pro or con depending on the application – dead cells don’t pose a health risk, but their DNA indicates recent contamination).

• More complex data interpretation.

In-Home Testing Kits: Convenience vs. Accuracy

A variety of over-the-counter E. coli test kits are available for homeowners. While convenient, it’s crucial to understand their limitations.

Types of Kits:

• Dip Slides/Paddles: These contain specific growth media on a paddle that you dip into the water sample. After incubation at room temperature, color changes or colony growth indicate the presence of bacteria. Some may offer presumptive E. coli indication.

• Liquid Reagent Kits: Similar to the professional P/A tests, these involve adding a water sample to a bottle with a dry reagent. Color changes or fluorescence indicate the presence of coliforms and/or E. coli.

General Procedure (varies by kit):

1. Collect Sample: Follow the kit’s instructions carefully for collecting a sterile water sample.

2. Add Reagent/Dip Paddle: Introduce the water sample to the test kit according to the instructions (e.g., pour into a bottle with reagent, dip a paddle).

3. Incubate: Place the kit in a warm area, typically room temperature (20−30∘C), for the specified incubation period (often 24-72 hours).

4. Interpret Results: Compare the results (color change, colony appearance, fluorescence) to the provided color chart or guide.

Pros of In-Home Kits:

• Convenient and readily available.

• Relatively inexpensive.

• Provide immediate peace of mind (or alarm).

Cons of In-Home Kits:

• Lower Accuracy and Sensitivity: Less reliable than laboratory tests. They may produce false positives or false negatives due to variations in environmental conditions, user error, or limited detection capabilities.

• Qualitative Only: Most kits only provide a “presence/absence” result, not a quantification of bacteria. This means you don’t know the level of contamination.

• Limited Specificity: Many kits only test for total coliforms or fecal coliforms, which are less specific indicators than E. coli. Even if E. coli is indicated, it might not be confirmed.

• No Confirmation: A positive result from an in-home kit always warrants immediate follow-up with a certified laboratory for confirmation. Do not rely solely on an in-home kit for critical health decisions.

• Improper Incubation: Room temperature incubation may not be optimal for E. coli growth, leading to underestimation or false negatives.

Recommendation: While in-home kits can be a good preliminary screening tool, especially for well water users, they should never replace professional laboratory testing, particularly if there are concerns about water safety or if a positive result is obtained.

Interpreting Your Results

Understanding what your E. coli test results mean is crucial for taking appropriate action.

• Drinking Water Standards: For potable water (drinking water), the presence of any E. coli is unacceptable. The standard is “zero E. coli per 100 mL sample.” A positive E. coli result in drinking water indicates a potential health risk and requires immediate action.

• Recreational Water Standards: Standards for recreational waters (swimming, boating) are typically higher than for drinking water, as exposure is different. These standards often vary by region and agency but generally allow for a certain number of E. coli per 100 mL before issuing advisories or closures. For example, some guidelines might set a single sample maximum of 235 E. coli per 100 mL for primary contact recreation.

• “Below Detection Limit” or “Not Detected”: This is the ideal result, indicating that E. coli was not found in the tested sample volume using the specific method employed.

• “Present” or “Positive”: Indicates that E. coli was detected.

Actions to Take Based on Results

If E. coli is Detected in Your Drinking Water (e.g., from a well or private supply):

1. Stop Drinking and Cooking with the Water: Immediately switch to bottled water or a safe alternative.

2. Boil Water: If you must use the water, bring it to a rolling boil for at least one minute to kill bacteria before consuming or using for food preparation, brushing teeth, or washing dishes.

3. Identify the Source: Work with a qualified professional (well driller, plumber, environmental health specialist) to investigate the source of contamination. This could involve:

   • Inspecting your well for structural damage, cracks, or loose well caps.

   • Checking the proximity of your well to septic systems, livestock, or agricultural runoff.

   • Assessing the integrity of your plumbing system.

4. Disinfect Your Well (Shock Chlorination): For private wells, a common remediation step is “shock chlorination,” which involves introducing a strong chlorine solution into the well and plumbing system to kill bacteria. This should be done by a professional or with careful adherence to detailed instructions.

5. Re-test: After disinfection and addressing the source of contamination, re-test your water multiple times over a period to ensure the E. coli has been eliminated and the water is safe. Follow local health department guidelines for re-testing frequency.

6. Consider Long-Term Solutions: Depending on the recurring nature of the problem, consider permanent water treatment systems such as:

   • UV Sterilization: A UV light system can effectively kill bacteria and viruses without adding chemicals.

   • Chlorination System: Automatic chlorination systems can provide continuous disinfection.

   • Point-of-Use Filters: Filters certified to remove bacteria can be installed at individual taps, but these are not a whole-house solution for E. coli.

If E. coli is Detected in a Recreational Water Body:

• Public Health Advisories: Local health departments or environmental agencies will typically issue public advisories, warnings, or close beaches/swimming areas if E. coli levels exceed safety thresholds.

• Avoid Contact: Do not swim, wade, or engage in other primary contact recreational activities in water bodies with elevated E. coli levels.

• Shower After Contact: If accidental contact occurs, shower thoroughly with soap and clean water immediately afterward.

• Pets: Keep pets out of contaminated water, as they can also become ill or spread bacteria.

Prevention: The Best Defense

The best way to deal with E. coli in water is to prevent its presence in the first place.

• For Private Wells:
  • Regular Testing: Test your well water at least once a year for total coliforms and E. coli, and more frequently if there are changes in taste, odor, clarity, or if someone in the household experiences unexplained gastrointestinal illness.

  • Proper Well Construction and Maintenance: Ensure your well is properly constructed, sealed, and maintained. The well cap should be secure and free of cracks. The area around the well should slope away to prevent standing water.

  • Septic System Maintenance: Regularly inspect and pump your septic system (typically every 3-5 years) to prevent leaks and overflows.

  • Livestock Management: Keep animal waste away from wellheads and natural waterways.

  • Hazardous Material Storage: Store fertilizers, pesticides, and other chemicals away from your well.

• For Public Water Systems:

  • Support Infrastructure Investment: Advocate for investment in robust municipal water treatment and distribution infrastructure.

  • Report Issues: Report any suspicious leaks, discolored water, or strange odors to your local water utility.

• For Recreational Water Bodies:

  • Don’t Swim When Ill: Avoid swimming if you have diarrhea.

  • Practice Good Hygiene: Shower before entering pools or natural bodies of water.

  • Avoid Fecal Accidents: Take young children for frequent bathroom breaks and check diapers regularly.

  • Don’t Feed Wildlife: Feeding ducks, geese, or other animals near swimming areas can increase fecal contamination.

  • Pick Up Pet Waste: Properly dispose of pet waste, especially near waterways.

• General Hygiene: Always wash your hands thoroughly with soap and water after using the restroom, changing diapers, and before preparing food.

The detection of E. coli in water is a serious matter that demands attention and informed action. While professional laboratory testing remains the gold standard for accuracy and reliability, understanding the principles of sampling, various testing methods, and the interpretation of results empowers individuals and communities to better protect themselves. By prioritizing regular testing, practicing diligent prevention strategies, and responding promptly to any detected contamination, we can collectively ensure the safety and potability of our vital water resources, safeguarding health for all.