Gas Production
Positive: Presence of bubble (small or big depending upon the
amount of gas produced) will be seen in the inverted
Durham’s tube.
Negative: Absence of bubble in the inverted Durham’s tube.
3.11 Gelatin
Liquefaction
The test is used to examine the ability of the symbionts to produce
gelatinase. This test is used to confirm the ability of the organism to
generate extracellular proteolytic enzymes, gelatins that hydrolyze
gelatin. The reaction happens in two consecutive steps: in the first
reaction, gelatin hydrolyzes gelatin into polypeptides, and later,
polypeptides are later converted into amino acids.
3.11.1 Materials
Required
l
Media: Gelatin.
l
Organisms: Bacteria Xenorhabdus and Photorhabdus.
3.11.2 Procedure
1. Using a bacterial needle, inoculate culture gelatin deeps. Allow
it to stay for 30 days.
2. Put away the tube in the refrigerator for 30 min to determine
whether liquefaction has occurred.
3. Take the tube from the refrigerator and check the tube for
liquefaction.
4. If negative, continue incubation until liquefaction occurs.
3.11.3 Expected Result
Positive: Robust liquefaction occurs within 3 days.
Weak liquefaction occurs in 4–30 days.
Negative: No liquefaction after 30 days.
3.12 Phenylalanine
Deaminase or
Phenylpyruvic Acid
(PPA)
Phenylalanine deaminase medium determines the ability of an
organism to produce the enzyme deaminase. Microorganisms capable of developing phenylalanine deaminase remove the amine
(NH 2 ) from phenylalanine and release the amine group as free
ammonia. The deamination of phenylalanine by oxidative enzymes
leads to the formation of phenylpyruvic acid.
3.12.1 Materials
Required
l
Media: Phenylalanine deaminase agar.
l
Reagents: Ferric chloride.
l
Organisms: Bacteria Xenorhabdus and Photorhabdus.
3.12.2 Procedure
1. Carefully prepare phenylalanine deaminase agar medium.
2. Inoculate the phenylalanine slant (with a loop on the surface)
with a test organism.
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Laboratory Techniques for Symbiotic Bacteria
Positive: Presence of bubble (small or big depending upon the
amount of gas produced) will be seen in the inverted
Durham’s tube.
Negative: Absence of bubble in the inverted Durham’s tube.
3.11 Gelatin
Liquefaction
The test is used to examine the ability of the symbionts to produce
gelatinase. This test is used to confirm the ability of the organism to
generate extracellular proteolytic enzymes, gelatins that hydrolyze
gelatin. The reaction happens in two consecutive steps: in the first
reaction, gelatin hydrolyzes gelatin into polypeptides, and later,
polypeptides are later converted into amino acids.
3.11.1 Materials
Required
l
Media: Gelatin.
l
Organisms: Bacteria Xenorhabdus and Photorhabdus.
3.11.2 Procedure
1. Using a bacterial needle, inoculate culture gelatin deeps. Allow
it to stay for 30 days.
2. Put away the tube in the refrigerator for 30 min to determine
whether liquefaction has occurred.
3. Take the tube from the refrigerator and check the tube for
liquefaction.
4. If negative, continue incubation until liquefaction occurs.
3.11.3 Expected Result
Positive: Robust liquefaction occurs within 3 days.
Weak liquefaction occurs in 4–30 days.
Negative: No liquefaction after 30 days.
3.12 Phenylalanine
Deaminase or
Phenylpyruvic Acid
(PPA)
Phenylalanine deaminase medium determines the ability of an
organism to produce the enzyme deaminase. Microorganisms capable of developing phenylalanine deaminase remove the amine
(NH 2 ) from phenylalanine and release the amine group as free
ammonia. The deamination of phenylalanine by oxidative enzymes
leads to the formation of phenylpyruvic acid.
3.12.1 Materials
Required
l
Media: Phenylalanine deaminase agar.
l
Reagents: Ferric chloride.
l
Organisms: Bacteria Xenorhabdus and Photorhabdus.
3.12.2 Procedure
1. Carefully prepare phenylalanine deaminase agar medium.
2. Inoculate the phenylalanine slant (with a loop on the surface)
with a test organism.
128
Laboratory Techniques for Symbiotic Bacteria