Dilution Theory / Supplementary Pages:

General Problem Set
No. 2
John L's Bacteriology Pages > 
Selected General Topics  > Dilution Theory:
Dilution Theory per se:
" Page 1 – Dilution Plating
" Page 2 – More Dilution Plating
" Page 3 – The MPN Method		 Supplementary Pages:
" Five-Tube MPN Table
" Practice Set 1 (Plating)
" Practice Set 2 (Plating&MPN)

MPN and Dilution Plating Problems from a Problem Set
Given in Bacteriology 102 in May, 2001. Solutions are here.

1. You added 10 ml of a lake water sample to 90 ml of sterile diluent; this is the "first dilution" indicated in the table below. After mixing thoroughly, one ml of this dilution was added to 99 ml of sterile diluent to make the second dilution. To make the third dilution, one ml of the second dilution was added to 99 ml of sterile diluent. From each of these three dilutions, 1 ml or 0.1 ml amounts were inoculated into pour-plates of Plate Count Agar (PCA) and tubes of Lactose Lauryl Tryptose Broth (LLTB) as shown in the table below.

After appropriate incubation, colonies were counted and the tubes were checked for positive results, and the data are summarized below. Subsequent inoculations into BGLB and EC Broth were appropriately made and incubated; their results are also shown below.

dilution of lake water first dilution second dilution third dilution
Amount inoculated into each of two plates
of PCA and each of three tubes of LLTB:		 1 ml 0.1 ml 1 ml 0.1 ml 1 ml 0.1 ml
For convenience, you can indicate the
"plated dilution" or "dilution factor" here:		            
Colony count on PCA plates: TNTC TNTC 205
215		 18
22		 0
2		 0
0
No. of positive LLTB tubes: 3 2 1 0 0 0
No. of positive BGLB tubes: 3 2 0 0 0 0
No. of positive EC Broth tubes: 1 1 0 0 0 0

a. (2 points) What was the "total aerobic plate count" (in CFUs per ml) of the original, undiluted sample of water?



b. (2 points) What was the confirmed, most probable number of fecal coliforms per ml of the original sample of water.





2. (2 points) A sample of yogurt prepared with the usual organisms (i.e., Streptococcus thermophilus and Lactobacillus bulgaricus) was bacteriologically tested as follows: One tablespoon of yogurt was mixed in a sterile flask with four tablespoons of sterile milk. One ml of this mixture was added to 99 ml of saline. One-tenth ml of this last dilution was plated onto a medium known to support the growth of yogurt organisms.

After appropriate incubation, 250 colonies were counted on the plate. Fifty colonies were then picked at random, and half of these colonies were found to be composed of cocci in chains.

What was the number of Streptococcus thermophilus CFUs per gram of the yogurt?





3. (2 points) One-tenth ml of drinking water was added to a petri dish to which 19.9 ml of melted Plate Count Agar were added. After incubation, 240 colonies arose on the plate. What was the count of CFUs per one ml of the water?





4. (1 point) One should expect the same number of CFUs in 10 ml of an undiluted sample as in      ml of a 1/10 dilution of the same sample.




5. (1 point) The same dilution can be obtained in each of the following situations:

a. The addition of 1 ml of a sample to 9 ml of sterile diluent.

b. The addition of      ml of the same sample to 27 ml of diluent.

c. The addition of 10 ml of the same sample to      ml of diluent.


GO 
TO:

" Solutions to these problems.
" MPN Page.

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Department of Bacteriology, U.W.-Madison