The water testing laboratory is the last line of defense against water-borne infections. Accurate results are critical to consumer safety. A robust Quality Assurance/Quality Control program will give the laboratory confidence in its results. Laboratories need reliable positive and negative microorganism controls to establish a strong quality control (QC) program. The QC strains they choose are typically based on Environmental Protection Agency (EPA), Standard Methods, and ISO 17025 recommendations.

Uses for QC Microorganisms

Water laboratories use QC microorganisms for many purposes including:

• QC of new batches of media and reagents
• Initial and ongoing demonstration of capability
• QC of test batches
• Verification and validation
• Spiking matrices
• Meeting ISO 17025 requirements:
  • ANSI/ISO/IEC 17025:2005, General requirements for the competence of testing and calibration laboratories, 5.9.1 states, “The laboratory shall have quality control procedures for monitoring the validity of tests and calibrations undertaken…This monitoring shall be planned and reviewed and may include, but not be limited to, the following: a) regular use of certified reference materials and/or internal quality control using secondary reference.”
• Meeting Standard Methods for water and wastewater requirements:
  • Section 9020 Quality Assurance/Quality Control states, “Positive and negative control cultures: Use certified reference cultures. For each lot of media received, each lot of media received, each lab prepared batch of media, and each lot of purchased prepared media, verify appropriate response by testing with known positive and negative control cultures for the organism(s) under test.”

EPA Recommendations

The following EPA¹ table lists several bacterial species that are recommended as controls for several groups of microorganisms commonly targeted in water testing methods.

Group Targeted	Positive Controls	Negative Controls
Total Coliform	Escherichia coli Enterobacter aerogenes	Staphylococcus aureus Pseudomonas aeruginosa Proteus vulgaris
Fecal Coliform	Escherichia coli Klebsiella pneumoniae (thermotolerant)	Enterobacter aerogenes

 

Group Targeted	Positive Controls	Negative Controls
Escherichia coli	Escherichia coli (MUG positive strain)	Enterobacter aerogenes Klebsiella pneumoniae (thermotolerant)
Fecal Enterococci/Streptococci	Enterococcus faecalis Enterococcus faecium	Staphylococcus aureus Escherichia coli Serratia marcescens

1 EPA Manual for the Certification of Laboratories Analyzing Drinking Water, Criteria and Procedures Quality Assurance

Recommended QC Strains for Water Testing

Based on EPA, Standard Methods, ISO 17025 and other standards, Microbiologics has developed a list of suggested strains for use in a water microbiology laboratory’s quality control program. If using a kit, consult the manufacturer’s QC recommendations.

Microorganism Strain	Strain Characteristics
Aerococcus viridans derived from ATCC® 10400™*
Enterobacter cloacae subsp. cloacae derived from ATCC® 35030™*
Enterococcus faecalis derived from ATCC® 19433™*
Enterococcus faecalis derived from ATCC® 29212™*
Enterococcus faecalis derived from ATCC® 7080™*
Enterococcus faecium derived from ATCC® 35667™*
Enterococcus faecium derived from ATCC® 6057™*	Listed as an appropriate QC strain by U.S. Environmental Protection Agency, Manual for Certification of Laboratories Analyzing Drinking Water.
Escherichia coli derived from ATCC® 11775™*	Thermotolerant
Escherichia coli derived from ATCC® 25922™*	Listed as an appropriate QC strain by U.S. Environmental Protection Agency, Manual for Certification of laboratories Analyzing Drinking Water; Microbiologics enumerated E. coli, 0335, derived from ATCC® 25922™*, is not recommended for Fecal Coliform testing on mFC agar at 44.5°C. It is best to test a suspension of E. coli, 0335, prepared from fresh colonies grown overnight.
Escherichia coli derived from ATCC® 35218™*
Escherichia coli derived from ATCC® 35421™*
Escherichia coli derived from ATCC® 51813™*
Escherichia coli derived from ATCC® 8739™*	Listed as an appropriate QC strain by U.S. Environmental Protection Agency, Manual for Certification of Laboratories Analyzing Drinking Water.
Klebsiella aerogenes derived from ATCC® 13048™* (formerly Enterobacter aerogenes)	Listed as an appropriate QC strain by U.S. Environmental Protection Agency, Manual for Certification of Laboratories Analyzing Drinking Water.
Klebsiella pneumoniae subsp. pneumoniae derived from ATCC® 13883™*	Listed as an appropriate QC strain by U.S. Environmental Protection Agency, Manual for Certification of Laboratories Analyzing Drinking Water; Not thermotolerant. Does not grow at 44.5°C.
Klebsiella pneumoniae subsp. pneumoniae derived from ATCC® 27736™*	Thermotolerant
Klebsiella variicola derived from ATCC® 31488™*	Thermotolerant
Klebsiella pneumoniae subsp. pneumoniae derived from ATCC® 33495™*	Thermotolerant
Kocuria rhizophila derived from ATCC® 9341™*
Legionella pneumophila subsp. pneumophila derived from ATCC® 33152™*	Used for quality control of buffered charcoal yeast extract (BCYE) agar used in tests for water quality. Fluorescent antibody serogroup 1.
Proteus mirabilis derived from ATCC® 25933™*
Proteus hauseri derived from ATCC® 13315™*	Listed as an appropriate QC strain by U.S. Environmental Protection Agency, Manual for Certification of Laboratories Analyzing Drinking Water.
Pseudomonas aeruginosa derived from ATCC® 10145™*
Pseudomonas aeruginosa derived from ATCC® 27853™*	Listed as an appropriate QC strain by U.S. Environmental Protection Agency, Manual for Certification of Laboratories Analyzing Drinking Water.
Serratia marcescens derived from ATCC® 14756™*	Listed as an appropriate QC strain by U.S. Environmental Protection Agency, Manual for Certification of Laboratories Analyzing Drinking Water.
Serratia marcescens derived from ATCC® 43862™*
Staphylococcus aureus subsp. aureus derived from ATCC® 25923™*
Staphylococcus aureus subsp. aureus derived from ATCC® 6538™*	Listed as an appropriate QC strain by U.S. Environmental Protection Agency, Manual for Certification of Laboratories Analyzing Drinking Water.

Visit our website to find the strains listed above in several easy-to-use formats including certified reference materials (CRM). Contact our Technical Support team at 1.320.229.7045 or techsupport@microbiologics.com for help determining what format is right for your laboratory.

References

International Organization for Standardization. (2004). ISO 11731-2: Water quality-Detection and enumeration of Legionella – Part 2: Direct membrane filtration method for waters with low bacterial counts (1st ed.). Switzerland: ISO.

Root, P., Hunt, M., Fjeld, K., & Kundrat, L. (2014). Microbiological Water Methods: Quality Control Measures for Federal Clean Water Act and Safe Drinking Water Act RegulatoryCompliance. 97(2).

Standard Methods Committee, 2005. Hunt, M., Braun-Howland, E., Dichter, G., Hall, N., & Harris, S. 9020 Quality Assurance/ Quality Control; Standard Methods for the Examination of Water and Wastewater. Washington, DC: American Public Health Association.

United States Environmental Protection Agency. (2005). Chapter V Critical Elements for Microbiology; Manual for the Certification of Laboratories Analyzing Drinking Water Criteria and Procedures, Criteria and Procedures Quality Assurance (5th ed.). Cincinnati, Ohio: US EPA Office of Water.