Bacteria Testing Reference Guide

Why Testing for Bacteria in Water is Important

Controlling mineral scales, biological deposits, foulants and corrosion through the appropriate use of filtration equipment and treatment products helps maintain water system cleanliness, hygiene, and efficiency, as well as minimizing microbial growth. Regularly testing for bacteria contamination can either verify a treatment plan is working or be an indication of a problem that needs to be addressed.

Proper monitoring and maintenance can also prevent the spread of water borne pathogens and diseases such as Legionella and minimize health risks to people and the surrounding environment.  

Special Methods for Collecting Samples for Bacteria Analysis

Appropriate sampling is essential to provide representative samples to the laboratory in charge of testing. Sampling requirements depend on the objective of sampling, and on the method requested.

Sample Containers

For routine sampling, use new sterile bottles. The volume of the bottles should be adequate for analysis of all requested parameters. If the water is being treated by an oxidant (e.g. chlorine, chloramine, bromine, or ozone), a reducing agent should be added to stop the action of oxidation as soon as the sample is taken such as sodium thiosulfate.

Chem Inc. stocks sterile 250 mL and 1000 mL bottles for bacteria sampling. All sterile sample bottles provided by Chem Inc. contain sodium thiosulfate.

Transportation and Storage

Keep the time between sampling and analysis in the laboratory as short as possible. For drinking water, analysis should ideally be started within the same working day.

Cool samples — ideally (5 ± 3) °C — during transport (e.g. by using ice packs or melting ice), unless otherwise stated. Take care not to freeze samples and to protect samples from sunlight.

Time Delay

The delay between sampling and analysis includes transport, receiving, and processing in the laboratory.

Time delay between sampling and analysis may reduce the reliability of the results. Consequently, samplers and analysts shall work together to keep time delays as short as possible. Sample collection and receiving dates are noted on laboratory reports.

Please be aware that for Legionella testing, the maximum hold time is 48 hours for ISO method and 72 hours for CDC Method.

If you are unsure of the specific testing requirements for the analysis requested, please contact lab personnel.

Bacteria Testing Reference Table

Bacteria Analysis	Turn Around Time	Results Given and/or Interpretation	Special Notes	When/Why this test is important
Total Aerobic Bacteria	2 days	0 – 3,000 CFU/mL	When a problem is detected, you may want to conduct more testing to determine the nature of the microbial problem.	Aerobic bacteria can cause several problems in water, including slime formations, turbidity, taste and odor, corrosion, health risks, and hygiene risks.
ATP (Adenosine triphosphate) Free, Total, and Microbial	1 day	RLU (Relative Light Units) is directly proportional to the amount of ATP collected from the sample. A high RLU indicates a large amount of ATP. Because colony forming units (CFU) and RLU values are determined using different test methods and measure different substances, RLU values do not consistently correlate to CFU when testing samples.	ATP is an indicator for the presence of biological residues due to its universal presence in all living cells.	Fast and easy test to perform. “Free” measures dissolved ATP that is free in solution (non-microbial ATP), while “Total” measures both free ATP in solution and microbial ATP. Therefore, the difference between Total and Free ATP provides a measure of microbial contamination in the sample.
Total Coliform Bacteria / E coli.	2 days	Present or Absent	Not all coliform bacteria are harmful to your health, but E. coli is. E. coli is a type of fecal coliform bacteria	For drinking water quality Indicator of fecal contamination
Denitrifying Bacteria	Up to 6 days	Aggressive: 1,800,000 – 215,000 CFU/mL Moderate: 25,000 – 3,000 CFU/mL Not Aggressive: 350 - <50 CFU/mL	These bacteria reduce nitrate to nitrite, and some continue nitrification to gaseous nitrogen (complete denitrification). If highly aggressive bacteria are detected, the water should be tested for the presence of coliform bacteria.	The presence of denitrifying bacteria can indicate that the water has been polluted by nitrogen-rich organics from sources such as compromised septic tanks, sewage systems, industrial and hazardous waste sites.
Heterotrophic Plate Count	2 days	0 – 300 CFU/mL	Tests for Heterotrophic bacteria (Total Bacteria Count/ Total Plate Count) including Pseudomonas, Salmonella, Escherichia, Rhizobium, etc. Aerobic and anaerobic	For drinking water quality Some heterotrophic bacteria may be pathogenic and/or decompose organic matter in water.
Hydrogen Sulfide Producing Bacteria*	2 days	Present or Absent	Detection of Salmonella, Citrobacter, Proteus, Edwardsiella, and Klebsiella. Mostly found in fecal material. It is a type of Sulfate Reducing bacteria.	Hydrogen sulfide can cause several significant problems in water. Problems range from “rotten egg” odors to the blackening of equipment, slime formations, and the initiation of corrosive processes.
Iron-Related Bacteria	Up to 10 days	Aggressive: 570,000 – 9,000 CFU/mL Moderate: 2,200 – 25 CFU/mL Not Aggressive: 9 - <1 CFU/mL	Iron-related bacteria can use iron in their metabolism. This test can detect both iron-oxidizing and iron-reducing bacteria. Common iron-related bacteria include Gallionella, Crenothrix, Sphaerotilus, Siderocapsa, and Thiobacillus ferroxidans.	Taste and odor problems and “red water” are common symptoms of problems due to iron-related bacteria.
Legionella	7-10 days	Potable: ≥0.1 CFU/mL Non-Potable: ≥10 CFU/mL	L.pneumophila serogroup 1 and serogroup 2-14 are responsible for most hospitalizations. However, there are over 100 species of Legionella which have the potential to cause harm. Therefore, any positive detection of Legionella should be taken seriously.	Legionella bacteria transmits through water aerosols such as mist or spray. Therefore, locations such as showerheads, sink faucets, cooling towers, hot tubs, and fountains are potential sources for human infection.
Myco-bacterium	Up to 21 days	0 – 500 CFU/mL	Test can detect the presence of mycobacterium species including Mycobacterium tuberculosis, Mycobacterium kansasii, Mycobacterium scrofulaceum, Mycobacterium intracellulare, and Mycobacterium fortuitum	Mycobacterium can cause diseases such as tuberculosis in humans if exposed. Problem in waste, surface, recreational (Fountains), and ground Creates biofilms
Nitrifying Bacteria	5 days	Very Aggressive: 100,000 CFU/mL Aggressive: 10,000 CFU/mL Moderate: 1,000 CFU/mL Not Aggressive: None Detected	Nitrifying bacteria recycle organic nitrogenous materials from ammonium (the endpoint for the decomposition of proteins) to nitrates. In water, aggressive nitrifiers can produce high concentrations of nitrates. Nitrates in water can be a potential health risk, particularly to infants who have not yet developed a tolerance to nitrates.	Aggressive nitrifying bacteria can indicate that the water may have been polluted by nitrogen-rich organics from sources such as compromised septic tanks, sewage systems, industrial and hazardous waste sites and is undergoing an aerobic form of degradation.
Sulfate Reducing Bacteria*	Up to 11 days	Aggressive: 2,200,000 – 6,000 CFU/mL Moderate: 1,400 – 75 CFU/mL Not Aggressive: 20 - <1 CFU/mL	Sulfate-Reducing bacteria are a group of anaerobic bacteria that generate hydrogen sulfide (H2S). SRB microorganisms are difficult to detect because they are anaerobic and tend to grow deep down within biofilms (slimes) as a part of a microbial community. SRB may not be present in the free-flowing water over the site of the fouling.	SRB can cause several significant problems in water. Problems range from “rotten egg” odors to the blackening of equipment, slime formations, and the initiation of corrosive processes.
Yeast & Mold	Up to 2.5 days	1-150 CFU/mL	Both require free oxygen for growth and pH range of 2 – 9. Indicates organic contamination, high dissolved oxygen, and possible biofilm formation.	Tested in drinking water. May cause poor odor and taste. Should be done along with HPC

* Hydrogen Sulfide Producing Bacteria is a type of Sulfate Reducing Bacteria (SRB)

Abbreviations and Terms

ATP	Adenosine triphosphate
CFU/mL	Colony forming units per milliliter (unit of concentration)
DN	Denitrifying Bacteria
HPC	Heterotrophic Plate Count / Heterotrophic Bacteria
IRB	Iron Related Bacteria
RLU	Relative Light Units
SRB	Sulfate Reducing Bacteria