Last Modified: 2011-07-01
Exercise 7. Bacteriological Analysis of Water- purpose
Does all water from environmental sources contain microbes? If so, what kinds? Harmful?
Yes! They can include bacteria, viruses, protozoa, fungi, etc.
i. Most are harmless to humans
How can we tell if there are harmful microbes in a water source?
Test for the presence of indicator organisms
Indicator organisms: fecal coliforms
- Most are not human pathogens
- Indicate that feces / sewage are contaminating water
- if coliforms are present, its not safe
Allows you to determine a relative number of culturable bacteria in a water sample
Determine the number of “all” culturable bacteria (coliforms and others)
i. Use a sterile, pipet to transfer 0.1 ml of sample to a nutrient agar plate.
ii. Dip a sterile swab in the sterile water provided at your bench in the test tube.
iii. Use the moistened swab to smear the water sample over the surface of the agar plate.
iv. Use a sterile, 1-ml pipet to transfer 1.0 ml of the water sample to a nutrient agar plate.
v. Repeat steps ii. and iii. with a fresh swab.
vi. Invert the plates and incubate at 37°C
Presumptive test for fecal coliforms
a. One test in a series of two tests to determine if fecal coliforms
are present in the water sample
i. Use a sterile, 1-ml pipet to transfer 0.1 ml of the water sample to one tube of single-strength lactose fermentation broth.
ii. Use same pipet to transfer 1.0 ml of the water sample to the other tube of single-strength lactose
improve contrast between different
groping and identifying microbes
Explain the difference in the cell wall structure between Gram+ and Gram-
Gram–positive have cell walls containing peptidoglycan that retain the CV-I complex;
Gram-negative bacteria have cell walls containing many lipids that are dissolved by the decolorizer removing the CV-I complex
Halo forms if bacteria produce DNAse
- Selective & differential
• Only Gram neg can grow
• Gram negs that ferment lactose turn bright pink
Skim milk agar
- Some species produce casease- breaks down casein
• Clear zone around colonies producing casease
• Lipids & blue dye in medium
• Species that produce lipase degrade lipids
• Dye travels to clear zones
• Iodine turns black when contacts starch
• Some species break down starch - clear zone (golden) around colonies
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