Transcript Bacterial culture in laboratory

Bacterial
culture in laboratory
Bacterial
culture in laboratory
Introduction : bacterial growth
Bacterial growth

Multiplication by binary fission:
bacteria grows and divides into
identical daughter cells.
the
two

Characterized by a generation time (G)
variable depending on the species
20 minutes for Escherichia coli, Lactobacillus acidophilus for
100 minutes, 1000 minutes to Mycobacterium tuberculosis
Bacterial growth
Bacterial growth
After n cycles : 2n bactéria
Bacterial growth
For example, Escherichia coli
G= 20 minutes
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After
After
After
After
After
…
After
1
2
3
4
5
hour: 3 cycles : n = 23 = 8 bacteria
hours: 6 cycles : n = 26 = 64 bacteria
hours : 9 cycles : n = 29 = 512 bacteria
hours : 12 cycles: n = 212 = 4096 bacteria
hours : 15 cycles: n = 215 = 32768 bacteria
10 hours : 30 cycles : n = 230 = 1073741824 bacteria
More than a billion bacteria
after 10 hours of culture!
Bacterial
culture in laboratory
Culture media
and inoculation technique
Culture media

Solid by adding a
gelling agent (agar) to
a liquid media

Liquid
Culture in liquid media
Bacteria
Increasing the cell density during the
growth
Clear media
After 16h of
culture =
turbid media
Culture in solid media
Bacteria
Accumulation of cells during the
growth
After 16 - 24h :
1 bacteria  1 visible bacterial
colony
Inoculation technique
In liquid media :
Inoculation
with pipette
Inoculation
with loop
In solid media (usually
with Agar Petri dish) :
Spreading
with a rake
Ridges with a
loop
Ridges with a
swab
Bacterial
culture in laboratory
Aseptic conditions in lab
Aseptic conditions in lab
Problem 1 :
Bacteria from
environment can
contaminate the
preparation
Problem 2 :
Bacteria can contaminate
the technician
Bacteria to
study…
Need a sterile zone around the
preparation !!
Problem 3 :
Technician can
contaminate the
preparation
Aseptic conditions in lab
Equipment to create a sterile area to work…
Microbiological Safety Cabinet :
System which uses an HEPA filter
(High Efficiency Particulate Air) to
generate a flow of sterile air
 Protection for the preparation and
the technician
Aseptic conditions in lab
Equipment to create a sterile area to work…
Bunsen Burner :
Gas burner which delivers
a flame at 1300°C
 provides a sterile air
zone near
Aseptic conditions in lab
Working with a Bunsen Burner :
Sterile air area :
 Approximately
40 cm
diameter
around the
burner
Convection
air movement
 This area is
use to open
tube or agar
Petri dish with
bacteria
Aseptic conditions in lab
Organization of the workspace
Bunsen Burner is place in the center of the workspace
and all instruments are distributed around (aerial view)
:
Rack tubes
Agar Petri
dish
Bunsen
Burner
Box with
instruments
(pipettes,
inoculation
loops…)
Trash for
contaminated
instruments
Aseptic conditions in lab
Organization of the workspace
Aseptic conditions in lab
Consumables materials
Most microbiology labs use sterile disposable equipment :
Inoculation
needles and
loops
Volumetric
pipettes
Spreading
rake
Pipettes