(Solved): Coliform bacteria are the commonly-used bacterial indicator of sanitary quality of foods and water. ...

Coliform bacteria are the commonly-used bacterial indicator of sanitary quality of foods and water. They are defined as rod-shaped Gram-negative organisms which ferment lactose with the production of acid and gas when incubated at $37^{\circ }\mathrm{C}$. Coliforms are abundant in the feces of warm-blooded animals, but can also be found in the aquatic environment, in soil and on vegetation. In most instances, coliforms themselves are not the cause of sickness, but they are easy to culture and their presence is used to indicate that other pathogenic organisms of fecal origin may be present. Fecal pathogens include bacteria, viruses, protozoa or parasites. Water Sample: Water sample A Water sample B Materials: 2 DSPRLB tubes 1 EMB plate (second lab day) Procedure: Presumptive Test 1. Each group will perform one set of presumptive tests with various water samples. You will test the water samples using double-strength phenol red lactose broth (DSPRLB.) The broth should also contain a Durham tube for trapping gas produced during bacterial growth. 3. Obtain 2 DSPRLB tubes. Label them water sample A or B, group name. 2. Mix the water sample thoroughly before adding to the indicator broth. Transfer 1 $\mathrm{ml}$ of each water sample to separate DSPRLB tubes. 3. Incubate tubes at $37^{\circ }\mathrm{C}$ for 48 hours. Second Lab Day: 1. Observe for evidence of gas production and color change. Color change to yellow and presence of gas in the Durham tube is a positive test result. Photographic atlas reference page 71. Confirmed Test A few other organism may produce acid and gas in lactose broth, so to confirm that the causative agent is coliform, select a tube with gas, and use an inoculation loop to streak a plate of EMB agar (EMB- eosin-methylene blue) to obtain isolated colonies. EMB agar is highly selective for the coliforms and distinguishes between the more reliable sewage indicator Escherichia coli and Enterobacter aerogenes, which may come from soil or vegetation. The eosin and methylene blue dyes in the medium will cause lactosefermenting bacteria to produce a purple color, black center, or metallie sheen. Procedure: 1. Use a marking pen to divide the plate into two sectors. Marks should be made on the plate's base. 2. Label the plate with the water sample, and group name. 3. Spot inoculate with the test water sample. 4. Invert the plate and incubate at $37^{\circ }\mathrm{C}$ for 48 hours. Third Lab Day: 1. Observe the EMB plate for typical coliform colonies, Escherichia coli colonies appear very dark bluish-black, and there is often a green metallic sheen around the colony when light is reflected. Enterobacter aerogenes colonies are typically larger, pink colonies with a dark center "fish-eye" colony. Presence of colonies of either organism is a positive confirmed test. Photographic atlas reference page 11. BIO 3: General Microbiology Laboratory Manual Name: Laboratory Report 15: Water quality test for coliforms bacteria Purpose: Background: Data or Results: