Laboratory 13: Bacterial Population Counts
Why are population counts important?
To determine how many bacteria in a sample Grading milk Bladder infection threshold Food and water purity
How are cells counted using the counting chamber?
You take a slide of a suspension of bacteria and view it under a microscope using a counting chamber. When counting, you ignore the cell touching the upper and left borders of a box, but count the cells found on the lower or right borders To determine the number of cells per ml of solution, use the formula: # of cells/ml = (total # of cells counted in six medium squares X dilution factor X 1000 (adjustment to ml))/(# of medium squares counted (6) X volume of one medium square (0.004 microliters))
turbidity
a spectrophotometer is used to measure the amount of cloudiness as a measure of growth
standard plate counts (SPC) or viable count
bacterial colonies are viewed through magnifying glass against a colony-counting grid; Quebec colony counter
microscopic counts
special slides are used to count cells using a microscope
How many cells were found per ml? Were the numbers counted here higher or lower than the standard plate count method?
1.0125 X 10^9 yeast per ml higher
Understand the math necessary to create a dilution in a standard plate count
1: 100 1 = mls added 100 = total number of mls in solution To determine the amount of bacteria in the original sample, multiply the dilution factor by the # of bacteria counted
Why do you want to count microbes only from a plate with 30-300 colonies?
> 300 most likely the colonies are too crowded < 30 not enough to get an accurate estimate
What is a serial dilution?
A serial dilution is a process of reducing the number of microorganisms in a solution
What advantages does this method have over the standard plate count and vice versa?
Can't distinguish between dead and living cells in the population count Bacteria are in neutral conditions so there isn't bias like there is in the standard plate count method
What are the advantages and disadvantages of the standard plate count method?
The SPC method determines only viable cells, whereas a microscopic count determines both living and dead cells. The SPC method is biased because specific conditions and media are used, and these factors may exclude certain bacteria in the counts Provides an accurate method to determine the number of bacteria in a sample
How are population counts conducted?
Microscopic counts: special slides are used to count cells using a microscope Standard plate counts (SPC) or viable count: bacterial colonies are viewed through magnifying glass against a colony-counting grid; Quebec colony counter Turbidity: a spectrophotometer is used to measure the amount of cloudiness as a measure of growth
Why are serial dilution necessary when conducting population counts?
Serial dilutions are necessary to create a sample of organisms that are capable of being counted