LABORATORY Flashcards
Pseudomonas aeruginosa: Specimens
- Pus
- Sputum
- Urine
- Blood
- Cerebrospinal fluid
- Other material as indicated by the type of infection
Gram-negative bacilli
No specific morphologic characteristics differentiate [?] in specimens from enteric or other Gram-negative rods.
pseudomonads
P. aeruginosa grow well on routine laboratory media such as
5% sheep BAM, and MAC or EMB
-35-37°C; growth at [?] helps differentiate it from other Pseudomonas species that produce fluorescent pigments, i.e., P. fluorescens, and P. putida)
- [?]or ambient air.
- [?] hours
42°C
CO2
24 to 48
Colonies are large with a spreading periphery and are often β-hemolytic.
Pseudomonas aeruginosa: On BAM
Growth appear to have a metallic sheen and a
scaling appearance sometimes described as
alligator skin morphology.
Pseudomonas aeruginosa: On BAM
Colonies typically produce a sweet “grape-like” or
“corn tortilla-like” odor.
Pseudomonas aeruginosa: On BAM
Can produce multiple colony types: may be
mucoid, or rough.
Pseudomonas aeruginosa: On BAM
P. aeruginosa imparts a blue-green color to the medium. This is produced by combination of two water-soluble and diffusible pigments:
pyocyanin (blue) and pyoverdin (yellow/yellow-green)
An uninoculated tube (A) is shown for comparison. Note the blue-green color (B) that diffuses into the medium.
P. aeruginosa on tryptic soy agar
Disk diffusion antimicrobial susceptibility test of P. aeruginosa on [?]. Note the blue-green pigments.
Mueller-Hinton agar
is a fluorescein pigment that fluoresces white to blue-green under long wavelength (400-nm) ultraviolet light source (e.g., Wood’s lamp)
Pyoverdin
a yellow-green pigment which is water soluble and fluoresces white to blue-green under longwavelength (400-nm) ultraviolet light.
pyoverdin
produces pyocyanin
P. aeruginos
is a fluorescein pigment that fluoresces white to blue-green under long wavelength (400-nm) ultraviolet light source (e.g., Wood’s lamp)
pyoverdin
Production of pyoverdin is particularly enhanced in media with a high phosphate concentration such as [?]. [?] are also suitable for demonstrating fluorescence.
Flo medium
King’s medium B, Sellers’ medium, and Mueller-Hinton agar
Fluorescence may be enhanced if cultures are incubated at [?] rather than at 35°C to 37°C.
20°C to 30°C
water-soluble, diffusible, blue pigment
Pyocyanin
water-soluble, diffusible, green (or yellow-green) pigment
Pyoverdin
red pigment
pyorubin
black pigment; brown to black
pyomelanin
- Non-lactose fermenting (NLF) colonies.
- May show green pigmentation, or metallic sheen.
- May be mucoid.
Pseudomonas aeruginosa: On MAC
[?] demonstrating growth of a mucoid variety of P. aeruginosa typical of the strains isolated from the sputum of patients with cystic fibrosis. Note that in the heaviest growth area, there appears to be some pyocyanin pigment developing.
MAC
To distinguish P. aeruginosa from the gram-negative bacilli of the family Enterobacteriaceae
Oxidase (+)
No acid production in indicates the inability of nonfermenting P. aeruginosa to utilize the lactose, glucose, or sucrose in TSI, or the lactose or glucose in KIA.
TSI or KIA: K/K
IMViC:
- +
Saccharolytic microorganisms degrade glucose either fermentatively or oxidatively.
The end products of fermentation are relatively strong mixed acids that can be detected in a conventional fermentation test medium.
However, the acids formed in oxidative degradation of glucose are extremely weak, and the more sensitive oxidation-fermentation medium of Hugh and Leifson (OF medium) is required for their detection.
Oxidation-Fermentation (OF) Test
Oxidation-Fermentation (OF) Test: Medium
Hugh–Leifson OF medium
The OF medium of Hugh and Leifson differs from carbohydrate fermentation media as follows:
• The concentration of peptone is [?]
• The concentration of carbohydrate is [?]
• The concentration of agar is [?]making it semisolid.
decreased from 1% to 0.2%.
increased from 0.5% to 1.0%.
decreased from 1.5% to 0.3%,
The [?] reduces the formation of alkaline amines that can neutralize the small quantities of weak acids that may form from oxidative metabolism.
lower protein/carbohydrate ratio
The relatively [?] serves to increase the amount of acid that can potentially be formed.
larger amount of carbohydrate
The [?] of the agar permits acids that form on the surface of the agar to permeate throughout the medium, making interpretation of the pH shift of the indicator easier to visualize.
semisolid consistency
[?] can also be observed in this medium.
Motility
Note that two tubes of each carbohydrate medium
are required for the test.
i. Inoculate each tube with the unknown organism, using a straight needle, stabbing the medium three to four times halfway to the bottom of the tube.
ii. To one tube of each pair, cover with a 1-cm layer of sterile mineral oil or melted paraffin. (CLOSED TUBE).
iii. Leave the other tube open to the air. (OPEN TUBE).
iv. Incubate both tubes at 35°C and examine daily for several days.
Oxidation-Fermentation (OF) Test
Oxidation-Fermentation (OF) Test: Result
A change in the initial green color of the medium to a yellow color indicates [?] from utilization of glucose.
acidification
produce acid only in the open tube exposed to atmospheric oxygen.
Oxidizers
produce acid in both tubes.
Fermenters
bacteria are inert in this medium, which remains at an alkaline pH after incubation.
Nonsaccharolytic
Acid (yellow) is seen only in the top portion of the open tube, indicating that the organism is capable of oxidizing glucose but incapable of fermenting glucose.
OXIDATIVE UTILIZATION OF GLUCOSE by P. aeruginosa in Hugh-Leifson OF medium.
