Page 251 - The Veterinary Laboratory and Field Manual 3rd Edition

220 Susan C. Cork and Roy Halliwell

acid reagent (0.8% in 5 M acetic acid) and two of only 0.4% which enables the organism to
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drops of α-naphthol reagent (0.5% in 5M acetic move and movement to be traced. Motility can
acid). An orange/red colour reaction indicates a also be checked by the ‘hanging drop’ method
positive test. Bacteria that produce nitrate reduc- (see below). True motility must not be confused
tase enzyme will produce nitrites in the medium with Brownian movement (vibration caused by
which result in a pink colour. molecular bombardment) or convection cur-
rents. A motile organism is one which actively
changes its position relative to other organisms
Oxidase (cytochrome oxidase) test
present and is seen in some bacteria and many
Add several drops of oxidase reagent to the protozoal organisms.
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bacterial colonies to be tested. If the organisms To assess motility using the ‘hanging drop’
produce oxidase the colonies will turn pink, red, method.
then black. This can be done on a culture plate
or, if colonies are picked off the plate, it can be 1 Use a clean slide and a square coverslip.
done on filter paper. Oxidase reduces the dye 2 On the slide place small pieces of modelling
in the reagent to a blue/black colour. Note that clay to fit the corners of the coverslip or use
the reagent is not particularly stable so it must a ‘well slide’ with a depression in the centre.
be prepared fresh. It also destroys the bacteria 3 Transfer a loopful of culture to the centre of
after a few minutes so if subculture is required the coverslip.
it needs to be done quickly. 4 Gently press the Plasticine on to the cover-
slip, ensuring that the ‘drop’ of culture is in
the centre of the circle, and does not come
Urea hydrolysis test
into contact with the slide.
Inoculate a slope of Christensen’s urea medium 5 With a quick movement, invert the slide, so
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with a heavy load of test bacteria. Incubate for that the coverslip is uppermost and the drop
1–2 days at 37°C. A red colour change indicates of culture is underneath, ‘hanging’.
decomposition of urea in the medium by ure- 6 Examine the drop of culture under the micro-
ase producing bacteria. Urease splits urea with scope, focusing first with the 10× objective,
the formation of ammonia which is alkaline. and when in focus swing round to the 40×
Bacteria which inhabit the renal tract are often objective to investigate motility.
urease positive (see Figure 4.11c – biochemical 7 Discard the preparation into disinfectant.
screening tests).
There are a number of other tests which are
commonly used in some laboratories as well as
Motility test
variations on those described. For more details
Inoculate tubes of motility medium by stabbing consult Quinn et al. (2000) or, preferably, attend
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the medium to a depth of about 5 mm. Incubate a bacteriology training course.
at the appropriate temperature (28 or 37°C), note
that motility can be temperature dependant and
some organisms are not motile at 37°C. Motile commercial identification systems for
organisms migrate through the medium which bacteria
becomes turbid; growth of non-motile organ-
isms is confined to the stab. ‘Motility’ medium is Although the conventional method of identifying
‘sloppy’ or semi-solid with an agar concentration bacteria by phenotype is accurate and reliable, it

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