Microbiology 1

CU20624

Lecture 05 K.F. Wannee

Microbial growth measurement Lecturer Life Sciences,
Chemistry Department,
HZ University of Applied
Sciences
Population Growth
• Quantitative Aspects of Microbial Growth
• The Growth Cycle

Measuring Microbial Growth

• Microscopic Counts
• Viable Counts
• Spectrophotometry
Quantitative Aspects of Microbial Growth
• Exponential growth kinetics
Quantitative Aspects of Microbial Growth
• Bacteria have relatively short generation times (g)
• Generation time is dependent on?

• Division rate (v)?

Generation time (g) = 0.5 h Number of generations (n)
Quantitative Aspects of Microbial Growth
• A mathematical relationship exists between the initial
number of cells present in a culture and the number
present after a period of exponential growth
The Growth Cycle
• Batch culture

Growth phases

Lag Exponential Stationary Death

Turbidity
(optical density)

Viable count
Continuous Culture
• Continuous culture
• Chemostat / bioreactior: most common type of
continuous culture device
Continuous Culture
• Increasing concentration of a limiting nutrient
Continuous Culture

optimum

[fraction of total volume vessel/hour]

Measuring Microbial Growth

• Microscopic Counts
• Viable Counts
• Spectrophotometry
Microscopic Counts

• Microbial cells are enumerated

by microscopic observations

Ridges that support coverslip To calculate number

per milliliter of sample:
Coverslip 12 cells X 25 large squares X 50 X 103

Sample added here. Care must be
taken not to allow overflow; space
between coverslip and slide is 0.02 mm
1 mm). Whole grid has 25 large
( 50
squares (a total area of 1 mm2 and
a total volume of 0.02 mm3).
Number/mm2 (3 X 102)
Microscopic observation; all cells are Number/mm3 (1.5 X 104)
counted in large square (16 small squares):
12 cells. (In practice, several large squares
are counted and the numbers averaged.) Number/cm3 (ml) (1.5 × 107)
Microscopic Counts
• Limitations?
Viable Counts

• Viable cell counts (plate counts): measurement of

living, reproducing population
• Two main ways to perform plate counts:
• Spread-plate method
• Pour-plate method
Spread-plate method
Surface
colonies

Incubation

Sample is pipetted onto Sample is spread evenly over Typical spread-plate results
surface of agar plate surface of agar using sterile
(0.1 ml or less) glass spreader

Pour-plate method

Surface
colonies

Solidification Subsurface
and incubation colonies
Sample is pipetted into Sterile medium is added and Typical pour-plate results
sterile plate mixed well with inoculum
 Sample to
1 ml be counted

1 ml 1 ml 1 ml 1 ml 1 ml

9-ml
broth

Total 1/10 1/100 1/103 1/104 1/105 1/106

dilution (10–1) (10–2) (10–3) (10–4) (10–5) (10–6)
Plate 1-ml samples

159 17 2 0
Too many colonies colonies colonies colonies colonies
to count
159 X 103 = 1.59 X 105
Plate Dilution Cells (colony-forming
count factor units) per milliliter of
original sample
Viable Counts
• “The great plate anomaly”: direct microscopic
counts of natural samples reveal far more
organisms than those recoverable on plates
• Why is this?
Spectrophotometry
Light

• Turbidity measurements
Prism

Incident
light, I0

Filter

Sample containing
cells ( )

Unscattered light, I

Photocell (measures
unscattered light, I)

Spectrophotometer

• Pros and cons? Optical density (OD)

I0
= Log
I
Homework assignments
• Assignment 03+04
• Deadline Sun Sep 24th

• Study theory lecture 06

• Pre-class assignment lecture 07
• Deadline Mon Sep 25th