Bacterial Growth Curve

Dr. Kali Kishore Reddy Tetala

Assistant Professor Sr.
Centre for Bioseparation Technology (CBST)
Vellore Institute of Technology (VIT), Vellore
Email: kishore.tetala@vit.ac.in / kishore.tetala@gmail.com
Ph.: 9965062572
• https://virtualbioreactor.wordpress.nc
su.edu/virtual-bio-reactor/
 Yeast fermentation experiment

http://www.bch.cuhk.edu.hk/vlab2/animatio
n/fermentation/index.html
 Bacterial Growth curve

https://vlab.amrita.edu/?sub=3&brch=73&si
m=1105&cnt=1
 Objectives

• To study the different phases of bacterial growth.

• To plot standard growth curve of Staphylococcus aureus.
• To determine the generation time of given bacteria.
 Principle

• The increase in the cell size and cell mass during the
development of an organism is termed as growth.
• It is the unique characteristics of all organisms.
• The organism must require certain basic parameters for their
energy generation and cellular biosynthesis.
• The growth of the organism is affected by both physical and
Nutritional factors. The physical factors include the pH,
temperature, Osmotic pressure, Hydrostatic pressure, and
Moisture content of the medium in which the organism is
growing.
• The nutritional factors include the amount of Carbon, nitrogen,
Sulphur, phosphorous, and other trace elements provided in
the growth medium.
 Principle

• Bacteria are unicellular (single cell) organisms.

• When the bacteria reach a certain size, they divide by binary
fission, in which the one cell divides into two, two into four and
continue the process in a geometric fashion. The bacterium is
then known to be in an actively growing phase.
• To study the bacterial growth population, the viable cells of the
bacterium should be inoculated on to the sterile broth and
incubated under optimal growth conditions.
• The bacterium starts utilizing the components of the media
and it will increase in its size and cellular mass.
• The dynamics of the bacterial growth can be studied by
plotting the cell growth (absorbance) versus the incubation
time or log of cell number versus time.
• The curve thus obtained is a sigmoid curve and is known as a
standard growth curve.
 Principle

• Increase in the cell mass of the organism is measured by

using the Spectrophotometer.
• Spectrophotometer measures the turbidity or Optical density
which is the measure of the amount of light absorbed by a
bacterial suspension.
• The degree of turbidity in the broth culture is DIRECTLY
related to the number of microorganism present, either viable
or dead cells, and is a convenient and rapid method of
measuring cell growth rate of an organism.
• Thus the increasing the turbidity of the broth medium indicates
increase of the microbial cell mass.
• The amount of transmitted light through turbid broth decreases
with subsequent increase in the absorbance value.
 https://www.youtube.com/watc
Different phases of growth of a bacteria
h?v=K0irrQ7vM6c

https://www.youtube.com/wat
ch?v=Eu_O0WS2jHc
 How to calculate generation time?

• The generation time can be calculated from the growth curve

 Contd…

• Generation Time = (Time in minutes to obtain the absorbance 0.4) – (Time in

minutes to obtain the absorbance 0.2)

= 90-60 = 30 minutes
Let No = the initial population number

Nt = population at time t

N = the number of generations in time t

Therefore,

Therefore,

The growth rate can be expressed in terms of mean growth rate constant (k), the number of
generations per unit time.
 Contd…
Mean generation time or mean doubling time (g), is the time taken to double its size.

Therefore,

Substituting equation 4 in equation 3

(Since the population doubles t= g)

Therefore,

Mean growth rate constant,

Mean generation time,
 Procedure
• An isolated colony of the organism (Staphylococcus aureus) was inoculated into 15 ml nutrient
broth and kept for overnight incubation
• Following day, the OD of this culture was measured and confirmed.
• In order to adjust the OD of the inoculum to the standard value (0.05) the following dilution
formula was used
• OD1V1 = OD2V2
• Where,
OD1 = OD of the broth culture, inoculated the previous day.
V1 = volume of this broth culture to be added to the inoculums
OD2 = OD of the inoculum (as a standard, this value was adjusted to 0.05)
V2 = volume of the inoculums (in this experiment, 50 ml)
• Substitute the values in the equation and V1 was calculated.
• That much amount (V1) of the inoculums was pipetted out before adding an equivalent
amount of the broth to it, so that the net volume remains constant.
• The OD was checked at every 30 minutes interval and recorded.
• Using this OD value, a standardized growth curve of the organism was plotted. (Absorbance
verses time).
• Generation time was calculated.
 Constructing a standard curve
(Spectrophotometer)

• https://www.youtube.com/watch?v=b-mlIWb1lDQ
• Using the simulator from the link, construct your own Bacterial growth curve (Try to get the curve as)

• A bacterium increases its number from 5 X 103 cells/ml to 2 X 108 cells/ml in 10 hours. Calculate
the mean generation time?

• Report these in the lab notes.

 Gentle reminder:
Assignment 1 Submission date is fixed for 21/02/2021
(Sunday)
https://medcraveonline.com/JABB/JABB-01-00016.pdf
ADYASHA NAYAK Group 1: Fig 1, Fig 3 and Fig 5
R JANANI
PADMANABHAN SIDDHARTH Group 2: Fig 2, Fig 4 and Fig 6
SHAMBHAVI SHARMA
NATEKAR RUCHA DEODATTA Group 3: Fig 2, Fig 7 and Fig 9
ROMIT BISHAYI
Group 4: Fig 3, Fig 8 and Fig 10

VARNIKA SINGHAL Group 5: Fig 2, Fig 6 and Fig 9

DIVYA AHUJA
ABHISHEK PRABHU P
SHREYA YADAV
PAVIKA SHARMA
SHIVAM GUPTA
DISHANT VANJANI

M M ISHWARYA KAVIN D MEGHANA OMPRAKASH

ANUSHKA MAHESH WAIKAR KAUL DIKSHITA PRIYA RICHIE PRANAU R
TEREDESAI SHREYA VINIT ANKAN GHOSH APARAJITA BISWAS
SOUVIK GHOSH ABHIRUP GHOSH MARIYA SHABBIR PATANWALA
SWORAJ SAPKOTA PRAGATHEESKUMAR M ANAND SAIRAM
HARISH R KAVIYA ADITI SAKSHI SACHIN MAHAJAN
KHALSA GURMEET KAUR DEEKSHITHA J