A 100 ml of contaminated drinking water was given to you by

A 100 ml of contaminated drinking water was given to you by your lab, supervisor for analysis. You were instructed by your supervisor to determine the number of bacterial cells present in the water sample. You prepared three test tubes, each tube contained 9 ml of diluent broth. You then pipetted 1 ml of the original sample of water and transferred it to test tube 1. After shaking test tube 1 (to mix broth sample), you pipetted 1 ml from tube 1 and transferred it to tube 2. After mixing the contents of tube 2, you pipetted 1 ml from tube 2 and transferred it to tube 3. You also plated out the following: 1 ml from tube 1 onto agar plate A,0.1 ml from tube 2 onto plate B: 1 ml from tube 2 onto plate C; 0.1 ml from tube 3 onto plate D; and 1 ml from tube 3 onto plate E. After incubating the plates for 4 days in the incubator, 80 colonies of bacteria formed on agar plate D. Pls answer the following questions: What is the sample dilution in Tube 1 Tube 2 Tube 3 Plate A Plate B Plate C Plate D Plate E What is the number of bacteria/ml in the sample What is the total number of bacteria in the 100 ml contaminated drinking water? Calculate the generation time if 50 Escherichia coli cells growing for 6 hours produced 100,000 cells.

Solution

In tube 1 the dilution will be 1ml of sample/ 1ml of sample+ 9ml of broth = 1/1+9 = 1/10 = 10^-1

similarly in tube 2 it will be 10^-2

in tube 3 it will be 10^-3.

On plate A the plated dilution is 10^-1, On plate B it is 10^-2, On plate C it will be 10^-2, On plate D and E it is 10^-3.

Since 80 colonies have been found on plate D, the dilution plated on plate D is 0.1 mL of 10^-3. The no.of bacteria per mL of sample can be calculated as

80 colonies/ 0.1 mL plated x 10^-3 = 800 x 10³ = 8 x 10⁵ colony forming units/ ml.

Since 1ml of sample contained 8x10⁵ cfu, for 100 ml it will be 8x 10⁵ x 100 = 8 x 10⁷ colony forming unit/ 100ml

Generation time can be calculated as follows:

G= t/n where G= generation time , t= time, n= no.of generation

n can be calculated by the formula b= B x 2ⁿ where b= no. of bacteria at the end of time interval

B= no. of bacteria at the beginning of the time interval,

putting the values will calculate the generation time.