Characterization of Bacteria Responsible for Background ...

Characterization of Bacteria Responsible for Background ...

Characterization of Bacteria Responsible for Background Anomalies in the Enumeration of Fecal Coliform Bacteria by the Membrane Filter Technique Esther P. Tennant-Clegg Jerald D. Hendrix Apogee Colony Counting System Background interference caused by non-fecal coliforms interfered with both automated and manual fecal coliform enumeration.

Inconsistent Colony Counts Two plates created from identical samples. The plate on the left is overgrown with typical blue colonies formed by fecal coliforms The plate on the right shows a zero fecal coliform count. Bacteria Isolated from Water Samples on mFC agar: Typical and Atypical Group Designation

Identification Typical fecal coliform Escherichia coli A lactosenonfermenting variety Escherichia fergunsonii of Escherichia Nonfermentative Pseudomonas maltophilia gram-negative bacillus Pseudomonas putrefaciens Acinetobacter calcoaceticus Acinetobacter lwoffi Flavobacterium odoratum Bordetella bronchiseptica Growth on mFC agar

Inhibited by Rosolic Acid? Blue colonies Blue background No Yellow colonies Yellow background No

Yellow colonies Yellow background No Atypical: gramGram-positive streptococcus Blue colonies positive streptococcus Blue background No Standard Membrane Filter Technique 0.45

m nitrocellulose filter mFC Agar Rosolic acid to inhibit background interference Incubation at 44.5C 0.5C for 24 hrs in water bath Plates are incubated in a watertight bag or box The Effect of Selected Bacteria on Agar Surface pH 9

8.5 Final Surface pH (Initial pH = 7.5) 8 7.5 7 6.5 6 5.5 5 4.5 Escherichia coli Pseudomonas

maltophila Pseudomonas putrefaciens Acinetobacter lwoffi Common Features of Atypical Bacteria on mFC Produce higher pH (up to 8.5) in the mFC

Agar Agar turns transparent yellow Aerobic Non-fermentative Not inhibited by rosolic acid Methods Used to Identify Bacteria

Gram Stain Wet mount slide to observe motility Catalase and Oxidase Test SIMS Agar to discover Indole reactions, and Oxygen requirements Fermentation Tubes (Sucrose, Lactose, Glucose) Oxiferm and Enterotube rapid biochemical test (Beckton Dickinson) Limited Oxygen Incubation Method Agar

is poured close to the top lip of plate limiting the area between bacterial growth and lid. Each plate is individually wrapped with parafilm prior to incubation. All other aspects of the Standard Method of Membrane Filter Technique remain unchanged. Effect Of Limited 02 on Background and Growth of E. coli Atmospheric O2 Levels Limited O2 Levels

Effect Of Limited 02 on Background and Growth of Atypical Isolates Atmospheric O2 Levels Limited O2 Levels Growth of Bacteria in Aerobic and Limited O2 Conditions: Effect on Agar Surface pH Final Surface pH (Initial pH = 7.5) 9 8.5 8

7.5 7 6.5 6 5.5 5 4.5 Aerobic Limited O2

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