7-4 Typical results for biochemical tests

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Below are pictured typical results for the 12 test species. Because of variation in incubation time, size of inoculum and variable skill in the experimenter, results for each species may be slightly different. With that caution, familiarize yourself with these. Pay special attention to the colony morphology, as that can be very distinctive for each species.

Figure 7-8 Gram stains of 9 isolates

Culture Reaction Description
Escherichia coli a reaction in medium This microbe appears as gram-negative short rods. Older cultures (more than 24 hours) may appear as cocci.
Klebsiella planticola a reaction in medium This microbe may appears as gram-negative short rods. However, the large amount of capsule surrounding these cells often interferes with the destaining process, causing them to stain Gram positive. Older cultures (more than 24 hours) may appear as cocci.
Serratia marcescens a reaction in medium This microbe forms gram-negative short rods
Pseudomonas fluorescens a reaction in medium This microbe forms gram-negative rods
Chromobacterium violaceum a reaction in medium This microbe forms gram-negative rods
Micrococcus luteus a reaction in medium This microbe forms gram-positive cocci
Enterococcus faecalis a reaction in medium This microbe forms gram-positive cocci
Lactobacillus plantarum a reaction in medium This microbe forms gram-positive rods
Bacillus subtilis a reaction in medium This microbe forms gram-positive rods. Endospores may be visible in older cultures as clear areas inside the cells. Older cultures may stain pink, due to the deterioration of cell walls.
Bacillus cereus a reaction in medium This microbe forms gram-positive rods. Endospores may be visible in older cultures as clear areas inside the cells. Older cultures may stain pink, due to the deterioration of cell walls.
Bacillus polymyxa a reaction in medium This microbe may appears as gram-negative short rods. However, the large amount of capsule surrounding these cells often interferes with the destaining process, causing them to stain Gram positive. Older cultures (more than 24 hours) may appear as cocci.
Staphylococcus epidermidis a reaction in medium This microbe appears as gram-positive cocci.

The Gram reactions of the 9 isolates.

Figure 7-9 Colony morphology of test strains

Culture Reaction Description
Escherichia coli a reaction in medium This microbe forms medium sized colonies with a regular margin and convex elevation
Klebsiella pneumoniae a reaction in medium This microbe forms slightly gummy/wet looking colonies that are circular, convex with an entire margin.
Serratia marcescens a reaction in medium Colonies of this microbe are red in appearance, circular and have an entire margin
Pseudomonas fluorescens a reaction in medium Colonies are circular, convex with an entire margin
Chromobacterium violaceum a reaction in medium Colonies are circular, convex with an entire margin and appear purple on most medium.
Micrococcus luteus a reaction in medium Colonies are punctiform, convex with an entire margin and appear yellow on most medium
Enterococcus faecalis a reaction in medium Colonies are punctiform, convex with an entire margin. The smallness of the colonies is due to the inefficient metabolism of these microbes. They can take 3-4 days to reach appreciable size.
Lactobacillus plantarum a reaction in medium Colonies are punctiform, convex with an entire margin. The smallness of the colonies is due to the inefficient metabolism of these microbes. They can take 3-4 days to reach appreciable size.
Bacillus subtilis a reaction in medium Colonies are larger, but not as large as B. cereus. The margin is undulate, with circular form and flat elevation.
Bacillus cereus a reaction in medium Colonies are large, irregular and flat with an undulate margin
Bacillus polymyxa a reaction in medium Colonies are large, irregular and flat with an undulate margin
Staphyloccocus epidermidis a reaction in medium Colonies are circular, convex with an entire margin

Note the margin, shape and elevation of the test strains. Also record the color of each strain.

Figure 7-18 Catalase reactions for test strains

Culture Reaction Description
Escherichia coli a reaction in medium Note the bubble formation. Catalase positive
Klebsiella pneumoniae a reaction in medium Note the bubble formation. Catalase positive
Serratia marcescens a reaction in medium Note the bubble formation. Catalase positive
Pseudomonas fluorescens a reaction in medium Note the bubble formation. Catalase positive
Chromobacterium violaceum a reaction in medium Note the bubble formation. Catalase positive
Micrococcus luteus a reaction in medium Note the bubble formation. Catalase positive
Enterococcus faecalis a reaction in medium Note the lack of bubbles. This microbe uses peroxidase to detoxify H2O2, an enzyme that does not evolve oxygen.
Lactobacillus plantarum a reaction in medium Note the lack of bubbles. This microbe uses peroxidase to detoxify H2O2, an enzyme that does not evolve oxygen.
Bacillus subtilis a reaction in medium Note the bubble formation. Catalase positive
Bacillus cereus a reaction in medium Note the bubble formation. Catalase positive
Bacillus polymxya a reaction in medium Note the bubble formation. Catalase positive
Staphylococcus epidermidis a reaction in medium Note the bubble formation. Catalase positive

Note the catalase reactions of each of the test strains. Look for evolution of bubbles.

Figure 7-10 Indole from tryptophan

Culture Reaction Description
Escherichia coli a reaction in medium E. coli is the only positive microbe of the 10 we study here.
Klebsiella pneumoniae a reaction in medium This microbe is negative for indole production.
Serratia marcescens a reaction in medium This microbe is negative for indole production. It will often give a false positive, as shown here, due to the extraction of cell pigment into the upper organic layer.
Pseudomonas fluorescens a reaction in medium This microbe is negative for indole production.
Chromobacterium violaceum a reaction in medium This microbe is negative for indole production.
Micrococcus luteus a reaction in medium This microbe is negative for indole production.
Enterococcus faecalis a reaction in medium This microbe is negative for indole production.
Lactobacillus plantarum a reaction in medium This microbe is negative for indole production.
Bacillus subtilis a reaction in medium This microbe is negative for indole production.
Bacillus cereus a reaction in medium This microbe is negative for indole production.
Bacillus polymyxa a reaction in medium This microbe is negative for indole production.
Staphylococcus epidermidis a reaction in medium This microbe is negative for indole production.

Record the indole tests for each microbe. Note that only E. coli is positive in this test.

Figure 7-11 Reactions in glucose fermenation broth

Culture Reaction Description
Escherichia coli a reaction in medium This microbe not only ferments glucose to acid, it also produces gas. Observe the bubble in the Durham tube.
Klebsiella pneumoniae a reaction in medium This microbe not only ferments glucose to acid, it also produces gas. Observe the bubble in the Durham tube.
Serratia marcescens a reaction in medium This microbe not only ferments glucose to acid, it also produces gas. Observe the bubble in the Durham tube.
Pseudomonas fluorescens a reaction in medium Pseudomonas is an obligate respirer and cannot ferment glucose
Chromobacterium violaceum a reaction in medium This microbe can ferment glucose
Micrococcus luteus a reaction in medium This microbe cannot ferment glucose
Enterococcus faecalis a reaction in medium This microbe can ferment glucose to acid, but does not produce gas.
Lactobacillus plantarum a reaction in medium This microbe can ferment glucose to acid, but does not produce gas.
Bacillus subtilis a reaction in medium This microbe cannot ferment glucose.
Bacillus cereus a reaction in medium This microbes can ferement glucose
Bacillus polymyxa a reaction in medium This microbe can ferment glucose
Staphylococcus epidermidis a reaction in medium Note the interesting pattern for this microbe. Purple on the inside of the tube, yellow on the outside. This is a positive reaction.

Note the presence of growth, color and presence or absence of a gas bubble for each strain.

Figure 7-19 Reactions in lactose fermenation broth

Culture Reaction Description
Escherichia coli a reaction in medium Note the formation of acid (yellow color) and gas.
Klebsiella pneumoniae a reaction in medium Note the formation of acid (yellow color) and gas.
Serratia marcescens a reaction in medium This particular strain does not ferment lactose.
Pseudomonas fluorescens a reaction in medium The tube remains purple with no gas since the microbe is unable to ferment lactose
Chromobacterium violaceum a reaction in medium This microbe is unable to ferment lactose.
Micrococcus luteus a reaction in medium This microbe is unable to ferment lactose.
Enterococcus faecalis a reaction in medium This microbe ferments lactose
Lactobacillus plantarum a reaction in medium L. plantarum is capable of fermentation of lactose, but does not produce gas.
Bacillus subtilis a reaction in medium This microbe is unable to ferment lactose.
Bacillus cereus a reaction in medium This microbe is unable to ferment lactose.
Bacillus polymyxa a reaction in medium Bacillus polymyxa ferments lactose and produces gas.
Staphylococcus epidermidis a reaction in medium Note the interesting pattern for this microbe. Purple on the inside of the tube, yellow on the outside. This is a positive reaction.

Note the presence of growth, color and presence or absence of a gas bubble for each strain.

Figure 7-12 Starch hydrolysis of test strains

Culture Reaction Description
Escherichia coli a reaction in medium Note the lack of a zone of clearing. This microbe is unable to hydrolyze starch and does not produce amylase.
Klebsiella pneumoniae a reaction in medium Note the lack of a zone of clearing. This microbe is unable to hydrolyze starch and does not produce amylase.
Serratia marcescens a reaction in medium Note the lack of a zone of clearing. This microbe is unable to hydrolyze starch and does not produce amylase.
Pseudomonas fluorescens a reaction in medium Note the lack of a zone of clearing. This microbe is unable to hydrolyze starch and does not produce amylase.
Chromobacterium violaceum a reaction in medium Note the lack of a zone of clearing. This microbe is unable to hydrolyze starch and does not produce amylase.
Micrococcus luteus a reaction in medium Note the lack of a zone of clearing. This microbe is unable to hydrolyze starch and does not produce amylase.
Enterococcus faecalis a reaction in medium Note the lack of a zone of clearing. This microbe is unable to hydrolyze starch and does not produce amylase.
Lactobacillus plantarum a reaction in medium Lactobacillus plantarum does not grow well on starch medium and is difficult to see in the photograph. Note the lack of a zone of clearing. This microbe is unable to hydrolyze starch and does not produce amylase.
Bacillus subtilis a reaction in medium Note the zone of clearing. All the starch in the medium near the microbe has been hydrolyzed by extracellular amylases.
Bacillus cereus a reaction in medium Note the zone of clearing. All the starch in the medium near the microbe has been hydrolyzed by extracellular amylases.
Bacillus polymyxa a reaction in medium Note the zone of clearing. All the starch in the medium near the microbe has been hydrolyzed by extracellular amylases.
Staphylococcus epidermidis a reaction in medium Note the lack of a zone of clearing. This microbe is unable to hydrolyze starch and does not produce amylase.

Observe for zones of clearing around starch plates that have been flooded with iodine. Note that the zone of clearing should be larger than 3 mm.

Figure 7-20 Nitrate Broth reactions

Culture Reaction Description Culture Reaction Description
Escherichia coli a reaction in medium E. coli can reduce nitrate to nitrite.
Klebsiella pneumoniae a reaction in medium This microbe can reduce nitrate to nitrite.
Serratia marcescens a reaction in medium This microbe can reduce nitrate to nitrite.
Pseudomonas fluorescens a reaction in medium Pseudomonas fluorescens reduces nitrate all the way to nitrogen gas N2. This will cause the nitrate broth to not turn red when reagents are added. Subsequent addition of zinc, which will chemically reduce nitrate to nitrite, will not occur, since there is no nitrate to act upon. Thus the tube will not turn red.
Chromobacterium violaceum a reaction in medium This microbe can reduce nitrate to nitrite.
Micrococcus luteus a reaction in medium This microbe is negative for the nitrate test. Addition of zinc to the test tube after adding reagents results in a weak red color.
Enterococcus faecalis a reaction in medium This microbe can reduce nitrate to nitrite
Lactobacillus plantarum a reaction in medium This microbe is negative for the nitrate test. Addition of zinc to the test tube after adding reagents results in a weak red color.
Bacillus subtilis a reaction in medium This microbe can reduce nitrate to nitrite
Bacillus cereus a reaction in medium This microbe can reduce nitrate to nitrite
Bacillus polymyxa a reaction in medium This microbe can reduce nitrate to nitrite.
Staphylococcus epidermidis a reaction in medium This microbe can reduce nitrate to nitrite.

Reaction in nitrate broth for the various test species.

Figure 7-21 Motility of selected strains

Culture Reaction Description
Escherichia coli a reaction in medium This microbe is highly motile and will show turbidity throughout the tube.
Klebsiella pneumoniae a reaction in medium This microbe is nonmotile.
Serratia marcescens a reaction in medium This microbe is motile. Note how the microbe swims to the top of the tube where there is more oxygen.
Pseudomonas fluorescens a reaction in medium This microbe is motile. Note how the microbe swims to the top of the tube where there is more oxygen.
Chromobacterium violaceum a reaction in medium This microbe is motile. Note how the microbe swims to the top of the tube where there is more oxygen.
Micrococcus luteus a reaction in medium This microbe is nonmotile.
Enterococcus faecalis a reaction in medium This microbe is nonmotile.
Lactobacillus plantarum a reaction in medium This microbe is nonmotile. It is often difficult to see in the tube, due to low growth.
Bacillus subtilis a reaction in medium This microbe is motile. Note how the microbe swims to the top of the tube where there is more oxygen.
Bacillus cereus a reaction in medium This microbe is motile. Note how the microbe swims to the top of the tube where there is more oxygen.
Bacillus polymyxa a reaction in medium This microbe is motile. Note how the microbe swims to the top of the tube where there is more oxygen.
Staphylococcus epidermidis a reaction in medium This microbe is nonmotile.

The motility test using semi-solid medium.

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