There are two ways to determine motility.
1. Through the use of a medium.
2. Microscopic examination.
1. Using a medium
The medium used is OF (Oxidative-Fermentative) medium.
[Oxidative-Fermentative is another biochemical test.]
OF medium has an agar that is in semisolid form which can also be used to determine if the bacteria are motile or non motile. Semisolid agar (soft agar) allows the bacteria to be diffused throughout the medium by the presence of flagella. Since one tube of OF medium can be used for 2 biochemical tests (Oxidative-Fermentative test and motility test), there is no need to use the motility medium which can only be used for motility test.
Methods:
a. Pick a few colonies and mix with a tube of saline to obtain a McFarland turbidity standard of 0.5.
b. Use a disposable inoculating needle to obtain a drop of the suspension and stab into the medium.
c. Incubate at 35°C in O2 conditions for 18 – 24 hours.
d. Examine the medium.
Results:
Motile bacteria: The inoculation line cannot be clearly seen as the bacteria are able to grow away from the inoculation line and diffuse into the surrounding medium.
Non motile bacteria: The inoculation line [stab] is clearly seen whereas the surrounding medium is clear. (The bacteria remain localized at the inoculation line).
(non motile bacteria)
2. Microscopic examination
This method is known as the hanging drop. It is used when the results of the previous method is not significant (Not sure whether it is motile or non motile).
Methods:
a. Prepare a glass slide with a ring of plasticine.
b. Use a wooden stick to obtain a drop of suspension from the tryptophan broth containing the bacteria and place it on the cover slip. [Tryptophan broth is used for another biochemical test. The OF medium used in the previous method cannot be used as it is not in liquid form.]
c. Invert the glass slide and press it against the cover slip. The drop of suspension should be hanging inside the ring of plasticine.
d. Observe under the microscope at 40X objective.
Results:
Non-motile: The bacteria appear to be stationary.
Motile: The bacteria are moving in forward or in spinning direction.
LeeJin
TG02
5 comments:
cornelyus here. I was just wondering, the only way for the motile bacteria is the flagella is it? Are there any more ways for the bacteria to move around in the media?
Hey leejin
Erm .. I'm curious what the agar contains; like what's the component. And also, there's only one type of agar or different type? like is there different degree of softness or hardness?
Sorry. and thanks. =)
Lyn
0611027D
TG02
Hi Cornelyus
There are three ways for the bacteria to move, depending on the type of bacteria. Flagellum or flagella is the most common mechanism used in bacteria. Besides that, some bacteria such as Spirochaetes have axial filament. It is a specialized internal structure that is between the cell membrane and the outer membrane in helical bacteria which helps the bacteria to rotate and move in a spiral form. Another mechanism is gliding motility. One example is Cyanobacteria. It is the movement of cells over surfaces such as wet glass surface but the exact mechanism is still unknown.
LeeJin
TG02
Hi lyn
OF medium contains pancreatic digest of casein, NaCl, Bromthymol Blue, Dipotassium phosphate, purified water and agar. The first 5 components are for OF test whereas the concentration of the agar is meant for motility test. The agar itself is a component. Normally, the concentration of the agar is between 1% - 2% but for motility test, the ideal concentration of the agar is between 0.05% – 0.5%. This reduced agar concentration allow the bacteria to migrate from the inoculation site. For the medium that I used, the agar concentration is 0.4%.
Hope this answers your question.
LeeJin
TG02
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