How much E.

coli can a flask

carry?
We grow E. coli bacteria in flasks containing various dilutions and different pH values. We perform
colorimetric assays over a day to observe its growth and we fit the growth data to the Logistic Growth
Curve to find the Intrinsic Rate of Growth ‘r’ and the Carrying Capacity ‘k’ of the population and the
habitat.

1. Preliminary insights on Growth models

It is a fair assumption to start with that the growth of a population directly depends on only its population,
and this could be mathematically put as
dN/dt = rN
where N- population at given time
r- intrinsic rate of growth determined as (b-d+i-e) {birth, death, immigration and emigration rates
respectively}

This gives us an equation exponential in t. Hence, if an E. coli, whose division time is about 20 minutes, is
left to divide, it would reach a volume of 9.304596e+17 cubic metres in 40 hours, which is approximately
the volume of the earth. Fortunately (or unfortunately), we don’t see E. coli earths coming up every 40
hours.
This is because such an uncontrolled growth is possible only given an unlimited amount of resources and
space, which is not the actual case.

2. The Logistic Growth model

It is known that the population of the species is limited by the resources in the habitat. Leibig’s Law of the
Minimum dictates that under steady state, population size is limited by the resource in shortest supply.
Population growth ceases when population reaches a carrying capacity. This is central to the Logistic
Growth model.
This carrying capacity arises due to the fact that in a condition with limited resources, individuals face
intraspecific and interspecific competition. Hence, the Logistic model is a density-dependent growth model.
The equation governing the Logistic model is therefore:

N= Population density at time t , r= intrinsic rate of natural increase, K= carrying capacity

 The growth of the population is a sigmoid curve as
shown in the figure. There is an initial phase of slow
growth called the lag phase, followed by a phase of
rapid growth of acceleration and then a deceleration
phase when the population reaches the carrying
capacity. The population growth onwards is
negligible and is asymptotic to the carrying capacity.
The expression can be reduced to
ra = rm(K-N)/K
rm is the Malthusian rate, max possible for a
population.

This explains when N=K, ra= 0. As the population

Logistic Growth Curve © study.com increases and approaches the carrying capacity, the
rate of growth tends to zero. When N is very low, the
rate is very close to maximum. When N slightly reaches above K, the rate becomes negative and the
population falls down to the carrying capacity.
However, we can clearly observe many species exhibiting cyclical or unpredictable growth, and as any
modelling framework, the Logistic model comes with assumptions.

The Logistic Growth model assumes the following conditions:

1. The carrying capacity of the habitat being constant
2. It does not consider that the population growth is affected by age distribution.
3. Linear change of b and d
4. The population and K interact instantaneously with each other, i.e. there is no lag between the
growth of the population and its dependence on K.
5. Abiotic, density-independent factors do not affect b and d and there is no environmental
stochasticity.
6. Crowding affects every individual equally.

3. Growth of E. coli in the flask

3.1 Materials and methods
To study the growth of bacteria in given nutrients and conditions in the limited space of the flask, we set
up flasks of different pH conditions and varying concentrations (dilution) of nutrients. Nine flasks were set
with combinations of one of
i) three values of pH- 5.5 (acidic), 7 (neutral) and 8.5 (basic) and,
ii) three concentrations of nutrient medium- 1:5 dilution,1:10 dilution and original concentration.
A few ways to measure the number of bacteria in the tube are streaking, serial dilution and measuring the
turbidity. Streaking works only for solid cultures, and serial dilution is tedious and could contaminate the
solution. Hence to measure the number of bacteria at a given point, we perform colorimetric/turbidimetric
assays to measure the optical density/turbidity of the contents of each flask at 30-minute intervals for 12
hours. As the bacteria grows, the solution becomes turbid. The hence measured turbidity or cloudiness of
the solution is a proxy for the number of bacteria present in the flask. The flask was structured with an
attached cuvette/test tube to measure the optical density of the culture just by tilting the flask into the
colorimeter compartment.
3.2 Observations
The turbidity values at half-hour intervals for each of the flasks were measured and are presented in the
table as follows:

pH pH pH pH 7, pH pH 5.5, pH 7, pH 8.5,
Time in 5.5, pH 7, 8.5, 5.5, 1:5 8.5, 1:10 1:10 1:10
Time minutes Ori Ori Ori 1:5 dil dil 1:5 dil dil dil dil

8:00
AM 0 0.01 0.03 0.03 0.02 0.01 0.02 0.01 0.01 0.01

8:30
AM 30 0 0.03 0.03 0.04 0.03 0.03 0.01 0.02 0.02

9:00
AM 60 0.02 0.05 0.03 0.03 0.05 0.07 0.02 0.04 0.04

9:30
AM 90 0.04 0.1 0.02 0.07 0.09 0.08 0.04 0.07 0.07
10:00
AM 120 0.07 0.23 0.02 0.12 0.18 0.16 0.03 0.11 0.12

10:30
AM 150 0.18 0.45 0.03 0.19 0.29 0.3 0.06 0.16 0.18

11:00
AM 180 0.3 0.74 0.04 0.29 0.39 0.4 0.09 0.22 0.26

11:30
AM 210 0.5 0.95 0.04 0.39 0.51 0.53 0.12 0.29 0.35

12:00
PM 240 0.64 1.11 0.04 0.53 0.56 0.71 0.18 0.39 0.44

12:30
PM 270 0.79 1.23 0.04 0.65 0.77 0.83 0.18 0.4 0.46

1:00
PM 300 0.91 1.31 0.05 0.81 0.87 0.95 0.34 0.53 0.57

1:30
PM 330 1.06 1.4 0.06 0.95 0.98 1 0.45 0.6 0.58

2:00
PM 360 1.11 1.44 0.08 1.09 1.04 1.06 0.57 0.59 0.58

2:30
PM 390 1.2 1.5 0.1 1.11 1.04 1.07 0.59 0.59 0.57

3:00
PM 420 1.25 1.52 0.14 1.1 1.03 1.05 0.58 0.59 0.57
 3:30
PM 450 1.31 1.58 0.2 1.11 1.05 1.07 0.58 0.59 0.56

4:00
PM 480 1.37 1.61 0.28 1.08 1.03 0.99 0.58 0.58 0.55

4:30
PM 510 1.41 1.63 0.39 1.01 0.97 0.99 0.57 0.58 0.55

5:00
PM 540 1.48 1.69 0.57 1.07 1.02 1.03 0.56 0.57 0.54

5:30
PM 570 1.52 1.75 0.78 1.08 1.02 1.02 0.56 0.57 0.54

6:00
PM 600 1.55 1.78 0.9 1.07 1 1.02 0.55 0.57 0.54

6:30
PM 630 1.59 1.8 1.01 1.07 1.02 1.02 0.55 0.57 0.53

7:00
PM 660 1.62 1.84 1.14 1.05 1.01 1.02 0.54 0.56 0.54

7:30
PM 690 1.65 1.88 1.2 1.06 1.03 1.03 0.53 0.57 0.53
8:00
PM 720 1.68 1.91 1.26 1.06 1.02 1.01 0.54 0.58 0.53

3.3 Graphing and Results

Logistic fit for pH 5.5, Ori Logistic fit for pH 7, Ori Logistic fit for pH 8.5, Ori
Logistic fit for pH 5.5, 1:5 dil Logistic fit for pH 7, 1:5 dil Logistic fit for pH 8.5, 1:5 dil

Logistic fit for pH 5.5, 1:10 dil Logistic fit for pH 7, 1:10 dil Logistic fit for pH 8.5, 1:10 dil

The (proxy) values for k and r based on turbidity were found to be:

pH/Dilution Original dilution 1:5 dilution 1:10 dilution

pH 5.5 k=1.5861 k=1.0849 k=0.5678

r=0.012152 r=0.019822 r=0.025049

pH 7 k=1.7403 k=1.0317 k=0.5842

r=0.014471 r=0.018789 r=0.0196559

pH 8.5 k=1.3676 k=1.0353 k=0.55376

r=0.015644 r=0.02152 r=0.023225
3.4 Analysis and Inferences
i) Effect of concentration of nutritients.
There is a close to linear decrease in k with decrease in concentration/ increase in dilution of nutrients in
all different pH values. Such linearity is very clear in the case of pH=5.5, and in other cases there is a
decrease as the concentration is reduced. This is a obvious result of the nutrient limiting the growth of the
bacteria and hence it shows up as the carrying capacity.
A very peculiar result is the increase in growth rate ‘r’ as we dilute the medium. We steady a steady
increase of r on dilution across any pH value. This is also clear from the graphs showing a steep initial
increase in 1:10. Attempting an explanation at this result, it is possible that the dilution meant easier
assimilation of nutrients by the bacteria when they are at lower populations, and this would imply a higher
growth rate. This could also be because of an increase in the birth rate arising due to the availability of
space and water. Since carrying capacity doesn’t depend on the birth or death rate of individuals, it is
solely the effect of reduction in nutrients that decreases the carrying capacity but the growth rate of the
bacteria is an effect of easier assimilation or some process that enables the bacteria to reproduce more
efficiently, possibly due to the increase in space and water.
It could also be a mathematical/ demographic result that the time taken to reach such a small carrying
capacity is much lower than the others and the growth of bacteria is hence rapid compared to a magnified
lag phase in the other nutrient concentrations. This is again the effect of the increase in space, since our
cultures are of equal volume and, at lower concentrations, the carrying capacity is low, effectively the
bacteria there has a higher per capita space than when the carrying capacity is higher, hence facilitating
rapid growth and lower death.
ii) Effect of pH
There is a decrease of k about neutral pH, except in the case of 1:5 dilution. This anomalous result is
unexplained. However, on comparison between basic and acidic pH, we see that the k is higher in acidic
pH and this means that the species is not very limited by an acidic medium, when compared to a basic
medium. This could be due to biochemical reasons such as the efficiency of enzymes etc.
However, the r is higher in most cases as the pH moves from neutrality. This could be due to a similar
mathematical/demographic result as discussed in the case of nutrients. It is evident again from the r values
that the bacteria is better off in an acidic medium than in a basic medium.
The pH of the medium essentially determines how efficient the individual can survive and reproduce and
hence should result in changes in r. However, we see changes in k, which could possibly imply that the
relative efficiency of nutrient assimilation at different pH is lower, and hence k value is decreased.

4. References

1. https://study.com/academy/lesson/logistic-population-growth-equation-definition-graph.html
2. PAST