Pseudomonas spp

Sharq Elneil College

School of Medical Laboratory

Sciences

Department of Microbiology

Medical Bacteriology course

U.Mahadi Hassan Mahmoud mahadi2010sd@yahoo.com

Bsc, Msc, MIBMS Microbiology

Pseudomonas 1882 Carle Gessard, a chemist and

bacteriologist from Paris, France,

Gram Stain

Gram-

Positive

Gram-

Negative

Cocci Bacilli Cocci Bacilli

Classification of Bacteria

Gram-negatitive Bacilli

Oxidase Test

Oxidase positive Oxidase Negative

O/F

O+/F-

Pseudomonadaceae

O+/F+

Vibrionaceae

O/F

O+/F+

Enterobacteriaceae

General characters:

Not member of enterobacteriaceae.

It is similar to them in diseases, saprophytes and commensal in the intestine of human.

Oxidase positive.

Obligate aerobe.

Does not ferment glucose.

Large group of microorganisms, more than 200 spp, most of them are saprophytes. The most important species according to infection is P. aeruginosa.

Morphology & stain:

Aerobic, opportunistic pathogen

Gram-negative bacillus

Flagella

Culture characters:

Non-fastidious.

Aerobic organism.

Blood agar: Large irregular colonies surround by zone of β-haemolysis.

MacConkey agar: Pale yellow colonies (NLF), Large irregular colonies.

CLED: Green-blue colonies (NLF), large and irregular.

Nutrient agar: Produced pigmented colonies.

Culture characters:

Different type of pigment:

Pyocynin: blue-green, water soluble and need

peptone water as substrate.

Pyoverdin: yellow-green, fluorescence (UV) and

need phosphate as substrate.

pyorubin (red)

Pyomelanin: brown, need 1% tyrosine in media.

Musty smell (grape like smell due to

aminoacetophenone).

On MacConkey : NLF

On MacConkey agar

Pseudomonas on Blood agar

On Nutrient agar Name this pigment?

On Nutrient agar Name this pigment?

Viability:

It can grow & life in water with small

nutrient.

Highly resistant to antimicrobial agents, but

sensitive to the group of aminoglycosides

(e.g.: Gentamicin, tobramycin).

Biochemical reaction:

One of the inert microorganism.

Cannot ferment glucose, but attack it by

oxidation.

Oxidase test: +ve.

Citrate utilization test: +ve.

Motility test: +ve.

Indole test: -ve.

Oxidase test Principle:

Certain organism produce oxidase enzyme that oxidize oxidase reagents to give purple colour.

Methods:

Filter paper method: Test requirements:

Freshly prepared 1% Oxidase reagent (tetramethyl-p-phenylene diamine).

Filter paper.

Wood stick or glass rods.

Take a colonies and put it in filter paper, add drop of oxidase reagents and examined for purple colour.

Oxidase +ve like Neisseria and Pseudomonas.

Oxidation Fermentation Principle:

The test depends on fermentation of carbohydrate on anaerobic condition of oxidation of it in aerobic condition.

Media content: 2 media each contain nutrition, sugar and bromothymol

blue. One of them closed from air by paraffin oil.

Results: Oxidative ferment sugar on open tube (Yellow colour).

Fermentative but anaerobically give yellow on closed tube.

Facultative anaerobic ferment CHO on both tube (Yellow).

Non oxidative- Non fermentative– give Blue colour on both tube.

Attack sugers by Oxiation and Not Fermentation

Results

Non-Saccharolytic O-/F

Alcaligenes faecalis

Open & covered remain green

Oxidative O+/F-

Pseudomonas

Open turns yellow

Fermentative O+/F+

Enterobacteriaceae

Both turn yellow

Reaction 1 Reaction 3 Reaction 2

There are three types of reactions possible

Biochemical reaction:

Urease test: -ve.

H2S production test: -ve.

MR / VP test: -ve.

KIA: Butt Slant H2S Gas

Alkaline Alkaline - -

O F test: Open tube Sealed tube

Y G

Virulance Factors

Pathogenicity: Pulmonary Infections

Burn Wound Infections and other skin and soft tissue infections (life threatening)

UTI’s (especially catheterized)

External Otitis (malignant OE, swimmer’s ear)

Eye Infections and corneal ulceration via contaminated contact lens cleaning fluids

Pseudomonal Endocarditis

Pulmonary Infections Can range from asymptomatic colonization to

severe necrotizing bronchopneumonia

Colonization is seen in patients with cystic fibrosis, chronic lung disease, and neutropenia

Mucoid strains are commonly isolated from chronic pulmonary patients and are more difficult to eradicate

Predisposing conditions include previous therapy with broad spectrum abx

Pseudomonas PNA

Ecthyma Gangrenosum

Ecthyma gangrenosum is a well recognized

cutaneous manifestation of severe,

invasive infection by Pseudomonas

aeruginosa that is usually seen in

immunocompromised, burn patients, and

other critically ill patients

Echtyma Gangrenosum

Malignant Otitis Externa

Pseudomonas Keratitis and Corneal Ulceration

Endocarditis

LAB- Diagnosis:

Specimen: according to the side of infection

( swabs, blood, urine, sputum……etc).

Direct Gram stain (same specimen).

Culture & incubation.

Colonial morphology.

Indirect Gram stain.

Biochemical reaction.

Antimicrobial Susceptibility testing

Sensetive to:aminoglycosides (e.g.:

Gentamicin, tobramycin

Inherently resistant to many abx

Can mutate to more resistant strains during therapy

Combination of active abx generally required for successful therapy

Mechanisms of Antibiotic Resistance in Pseudomonas aeruginosa