prof . dr. ihsan edan alsaimary department of microbiology - college of medicine - university of basrah - basrah -IRAQ ihsanalsaimary@gmail.com 00964 7801410838

1. Prof dr. Ihsan Edan Alsaimary
department of microbiology – college of medicine – university of basrah – mobile: 07801410838
e.mail : ihsanalsaimary@gmail.com
PSEUDOMONAS

2. Pseudomonas and related organisms
Aerobic gram-negative nonfermentative rods
Pseudomonas aeruginosa: opportunistic infections of
multiple sites
Burkholderia cepacia: RT infection in cystic fibrosis patients,
UTI, opportunistic infections
Burkholderia pseudomallei: opportunistic pulmonary
infections or sepsis
Stenotrophomonas maltophilia: opportunistic infections
Acinetobacter baumannii: opportunistic infections of RT
Moraxella catarrhalis: opportunistic RT infections

3. Pseudomonas
Structure and Physiology
Gram-negative rods.
Motile with polar flagella.
Obligate aerobe.
Oxidase-positive.
Do not ferment
carbohydrates.
Resistant to multiple drugs.

4. MORPHOLOGY
DR.T.V.RAO MD 4
• They are slender gram negative bacillus, 1.5 – 3 microbes x 0.5
microns
• Monoflgellar ?
• Non capsulated but many strains have mucoid slime layer
• Isolates from Cystic fibrosis patients have abundance of
extracellular polysaccharides composed of alginate polymers
• Escape the defence mechanisms by loose capsule in which
micro colonies of bacillus are enmeshed and protected from
host defences.

5. Ps. aeruginosa
Forms round colonies with a fluorescent
greenish color, sweet odor, and b-
hemolysis.
Pyocyanin- nonfluorescent
bluish pigment;
pyoverdin- (fluorescin)
fluorescent greenish pigment;
pyorubin, and pyomelanin
Pyorubin
Reddish brown pigment
pyomelanin
Brown to black pigment
Some strains have a prominent capsule
(alginate).
Identification of P. aeruginosa is usually based on oxidase test
and its colonial morphology: b-hemolysis, the presence of
characteristic pigments and sweet odor, and growth at 42 oC.
PYOCYANIN PYOVERDIN

6. CULTURAL CHARACTERS
DR.T.V.RAO MD 6
• Obligate aerobe, but grow anaerobically if nitrate is
available
• Growth occurs at wide range of temperatures 6-42 c
the optimum being 37 c
• Growth on ordinary media producing large opaque
irregular colonies with distinctive musty mawkish or
earthy smell.
• Iridescent patches with metallic sheen are seen in
cultures on nutrient agar.
• In broth forms dense turbidity with surface pellicle.

7. Motile (by single or multiple polar flagella)
gram-negative rods
Obligate (strict) aerobes (most strains)
Oxidase (usually) and catalase positive
Nonfermentative chemoheterotrophic
respiratory metabolism
Minimal nutritional reqts.; Many organic
compounds used as C and N sources, but
only a few carbohydrates by oxidative
metabolism
• Glucose used oxidatively
• Lactose negative on MacConkey’s agar
Characteristics of Pseudomonas aeruginosa
DR.T.V.RAO MD 7

8. PIGMENT PRODUCTION
DR.T.V.RAO MD 8
Some strains produce diffusible pigments:
• Pyocyanin (blue); fluorescein (yellow);
pyorubin (red)
P. aeruginosa produces characteristic
grape-like odor and blue-green pus &
colonies
Broad antibiotic resistance

9. BIOCHEMICAL REACTIONS
DR.T.V.RAO MD 9
• Oxidative and Non fermentative
• Glucose is utilized oxidatively
• Indole, MR and VP and H2 S tests are
negative
• Catalase, Oxidase, and Arginine tests
are positive

10. TYPING AND IMPORTANCE
DR.T.V.RAO MD 10
• Important cause of Hospital Infections
• Important for epidemiological purpose
• Serotyping
• Bacteriocins typing
• Pyocyanin
• Aeruginosin typing
• Restriction endonuclease typing with pulsed gel
electrophoresis

11. RESISTANCE
DR.T.V.RAO MD 11
• Killed at 55oc in on 1 hour
• High resistance to chemical agents
• Resistance to quaternary ammonium
compounds.Chlorxylenol
• Resistant to Hexchlorophenes
• Grows also in antiseptic bottles
• Dettol as cetrimide as selective medium
• Sensitive to acids silver salts, beta glutaraldehyde

12. WHAT ANTIBIOTICS TO USE
DR.T.V.RAO MD 12
• Aminoglycosides
• Gentamycin, Amikacin, Cephalosporins
• Cefotaxime. Ceftazidime. Ofloxacin,
• Piperacillin, ticarcillin
• Local application, colistin, polymyxin

13. PATHOGENICITY
DR.T.V.RAO MD 13
• Blue pus
• Causing the nosocomial infection
• Suppurative otitis
• Localised and generalised infections
• Urinary tract infection after catheterization
• Iatrogenic meningitis
• Post tracheostomy pulmonary infections

14. P. aeruginosa
Pathogenesis and Immunity
This organism is widely distributed in nature
and is commonly present in moist environments
in hospitals. It is pathogenic only when
introduced into areas devoid of normal
defenses, e.g.,
1. Disruption of mucous membrane and skin.
2. Usage of intravenous or urinary catheters.
3. Neutropenia (as in cancer therapy).

15. P. aeruginosa
Antigenic structure,
enzymes, and toxins
Pili and nonpilus adhesins.
Capsule (alginate, glycocalyx):
seen in cultures from patients
with cystic fibrosis.
LPS- endotoxin, multiple
immunotypes.
Pyocyanin: catalyzes
production of toxic forms of
oxygen that cause tissue
damage. It also induces IL-8
production. Pyoverdin: a
siderophore.
Proteases
Serine protease,
metalloprotease and alkaline
protease cause tissue
damage and help bacteria
spread.
Phospholipase C: a hemolysin
Exotoxin A: causes tissue
necrosis and is lethal for animals
(disrupts protein synthesis);
immunosuppressive.
Exoenzyme S and T: cytotoxic to
host cells.
Pathogenesis

16. PATHOGENESIS AND IMMUNITY
DR.T.V.RAO MD 16
• P. aeruginosa can infect almost any
external site or organ.
• P. aeruginosa is invasive and toxigenic. It
attaches to and colonizes the mucous membrane
or skin, invade locally, and produces systemic
diseases and septicemia.
• P. aeruginosa is resistant to many antibiotics. It
becomes dominant when more susceptible
bacteria of the normal flora are suppressed.

17. • Septicaemia
• Endocarditis
• Ecthyma gangrenous
• Infantile diarrhoea
• Shanghai fever
• Disabling eye infections
• Survive with minimal
nutrients
CLINICAL PRESENTATIONS
DR.T.V.RAO MD 17

18. WHO IS MORE SUSCEPTIBLE TO
INFECTION
DR.T.V.RAO MD 18
• This bacterium is of particular concern to
individuals with cystic fibrosis who are highly
susceptible to pseudomonas lung infections.
Pseudomonas aeruginosa is also of grave
concern to cancer and burn patients as well as
those people who are immunocompromised. The
case fatality rate for individuals infected with
Pseudomonas aeruginosa approaches 50
percent.

19. • Pseudomonas aeruginosa
is the most frequently
encountered lung pathogen
in patients with cystic
fibrosis (CF). Following
initial, often intermittent,
episodes of infection, it
becomes a permanently
established component of
the chronically infected
lung in more than 80% of
patients and confers an
adverse prognosis
PSEUDOMONAS AND CYSTIC FIBROSIS
DR.T.V.RAO MD 19

20. • Respirators
• Endotracheal
tubes
• Can be Infected
• All equipment's
to be sterilized
INFECTION OF EQUIPMENT'S
DR.T.V.RAO MD 20

21. PSEUDOMONAS AND URINARY TRACT
INFECTIONS
DR.T.V.RAO MD 21
• Pseudomonal UTIs are usually hospital-acquired
and are associated with catheterization,
instrumentation, and surgery. These infections
can involve the urinary tract through an
ascending infection or through bacteriuic spread.
In addition, these infections are a frequent
source of bacteraemia. No specific
characteristics distinguish this type of infection
from other forms of UTI.

22. • Toxic extracellular products
in culture filtrates
• Exotoxin A and S
• Exotoxin A acts as NADase
resembling Diphtheria toxin
• Proteases,elastatese
hemolysins and enterotoxin
• Slime layer and Biofilms
TOXINS AND ENZYMES IN
PSEUDOMONAS
DR.T.V.RAO MD 22

23. PSEUDOMONAS PROMINENT HOSPITAL
ACQUIRED INFECTIONS
DR.T.V.RAO MD 23
• It causes urinary tract infections,
respiratory system infections, dermatitis,
soft tissue infections, bacteraemia, bone
and joint infections, gastrointestinal
infections and a variety of systemic
infections, particularly in patients with
severe burns and in cancer and AIDS
patients who are immunosuppressed.

24. DIAGNOSIS OF P.AEROGINOSA
INFECTION
DR.T.V.RAO MD 24
• Diagnosis of P,aeroginosa infection depends upon
isolation and laboratory identification of the bacterium.
It grows well on most laboratory media and commonly
is isolated on blood agar or eosin-methylthionine blue
agar. It is identified on the basis of its Gram
morphology, inability to ferment lactose, a positive
oxidase reaction, its fruity odour, and its ability to grow
at 42°C. Fluorescence under ultraviolet light is helpful
in early identification of P.s aeruginosa colonies.
Fluorescence is also used to suggest the presence of
P. aeruginosa in wounds.

25. Pseudomonas sp. develop as easily
distinguishable blue-green coloured
colonies, clearly visible under normal
lighting conditions. Other bacterial
species are inhibited or give
colourless colonies. Pseudomonas
aeruginosa, Pseudomonas
fluorescens, Pseudomonas putida
and Pseudomonas fragilis all give
typical blue-green colony colouration
and can be studied directly by
serotyping or biochemical methods.
IDENTIFICATION WITH CHROMAGAR
DR.T.V.RAO MD 25

26. GROWTH ON KINGS B MEDIUM
• Pale white, translucent colonies
• Circular, convex, and smooth in appearance

27. SCVs of P. aeruginosa in CF
• Slow growing subpopulations (3% of the P. aeruginosa
positive sputum specimens)
• SCVs exhibit an increased resistance towards a broad
spectrum of antimicrobial agents
• The recovery of SCV correlates
• with parameters revealing poor
• lung function and an inhalative
• antimicrobial therapy
• Fast growing revertants can
• be isolated from the SCV population
Small colony variants“ (SCV)

28. LABORATORY IDENTIFICATION OF
DIAGNOSIS OF P.AEROGINOSA INFECTIONS
DR.T.V.RAO MD 28
• Diagnosis of P. aeruginosa infection depends upon
isolation and laboratory identification of the bacterium.
It grows well on most laboratory media and commonly
is isolated on blood agar or eosin-methylthionine blue
agar. It is identified on the basis of its Gram
morphology, inability to ferment lactose, a positive
oxidase reaction, its fruity odour, and its ability to grow
at 42° C. Fluorescence under ultraviolet light is helpful
in early identification of P. aeruginosa colonies and may
also help identify its presence in wounds.

29. TREATING PSEUDOMONAS INFECTIONS
DR.T.V.RAO MD 29
• Combined antibiotic therapy is generally
required to avoid resistance that develops
rapidly when single drugs are employed.
Avoid using inappropriate broad-spectrum
antibiotics, which can suppress the normal
flora and permit overgrowth of resistant
pseudomonads.

30. PSEUDOMONAS AERUGINOSA A RESISTANT
PATHOGEN
DR.T.V.RAO MD 30
• Pseudomonas aeruginosa is frequently resistant
to many commonly used antibiotics. Although
many strains are susceptible to gentamicin,
tobramycin, colistin, and amikacin, resistant
forms have developed. The combination of
gentamicin and carbenicillin is frequently used
to treat severe Pseudomonas infections. Several
types of vaccines are being tested, but none is
currently available for general use.

31. DR.T.V.RAO MD 31
• Antibiotic resistance can be
attributed to P. aeruginosa’s
development of biofilm
o Allow communities’ adherence to a
variety of surfaces
o Difficult to destroy
o Depending on location, can be
beneficial or detrimental to
environment
5 stages of Biofilm Development
http://microbewiki.kenyon.edu/images/b/ba/P.aerugenosa_biofilm.jpg
A biofilm is any group of microorganisms in
which cells stick to each other and often these
cells adhere to a surface. These adherent cells
are frequently embedded within a self-produced
matrix of extracellular polymeric substance
(EPS). a polymeric conglomeration generally
composed of extracellular DNA, proteins, and
polysaccharides

32. McKnight et al, 2000
Quorum Sensing in P. Aeruginosa
Quorum sensing is a system of stimuli and response correlated to population density. Many species of bacteria
use quorum sensing to coordinate gene expression according to the density of their local population. In similar
fashion, some social insects use quorum sensing to determine where to nest. In addition to its function in
biological systems, quorum sensing has several useful applications for computing and robotics.
Quorum sensing can function as a decision-making process in any decentralized system, as long as individual
components have: (a) a means of assessing the number of other components they interact with and (b) a standard
response once a threshold number of components is detected.
The opportunistic pathogen Pseudomonas
aeruginosa uses quorum sensing to coordinate the
formation of biofilms, swarming motility,
exopolysaccharide production, virulence, and cell
aggregation.[9] These bacteria can grow within a
host without harming it, until they reach a
threshold concentration. Then they become
aggressive, developing to the point at which their
numbers are sufficient to overcome the host's
immune system, and form a biofilm, leading to
disease within the host as the biofilm is a
protective layer encasing the bacteria population.
Another form of gene regulation that allows the
bacteria to rapidly adapt to surrounding changes is
through environmental signaling. Recent studies
have discovered that anaerobiosis can significantly
impact the major regulatory circuit of quorum
sensing. This important link between quorum
sensing and anaerobiosis has a significant impact
on production of virulence factors of this
organism.[10] Garlic and ginseng experimentally
block quorum sensing in Pseudomonas
aeruginosa.[11] It is hoped that the therapeutic
enzymatic degradation of the signaling molecules
will prevent the formation of such biofilms and
possibly weaken established biofilms. Disrupting
the signalling process in this way is called quorum
sensing inhibition.

33. P. aeruginosa
Prevention and Control
Pseudomonas spp. normally inhabit soil, water, and vegetation
and can be isolated from the skin, throat, and stool of healthy
persons.
Spread is mainly via contaminated sterile equipment's and
cross-contamination of patients by medical personnel.
High risk population: patients receiving broad-spectrum
antibiotics, with leukemia, burns, cystic fibrosis, and
immunosuppression.
Methods for control of infection are similar to those for other
nosocomial pathogens. Special attention should be paid to sinks,
water baths, showers, hot tubs, and other wet areas.

34. Stenotrophomonas maltophilia
A common nonfermentative, gram-negative isolate.
It infects debilitated or immunocompromised persons,
and causes a wide spectrum of diseases, including
wound infections, UT infections, pneumonia, sepsis,
meningitis, etc.
It is resistant to many commonly used antibiotics, and
patients receiving long-term antibiotic therapy are
particularly at risk for acquiring infections.
Infections may be acquired from iv catheters,
contaminated disinfectants, respiratory therapy and
monitoring equipment, and ice machines.

35. Burkholderia
They colonize the moist environmental surfaces and are
commonly associated with nosocomial infections.
B. cepacia complex (of 9 species), B. gladioli and B. pseudomallei
are important pathogens.
B. cepacia complex causes RT infections particularly in cystic
fibrosis patients, UT infections and septicemia. Usually non-fatal
except for RT infections in CF patients.
B. pseudomallei usually causes opportunistic infections (called
melioidosis), but may sometimes infect previously healthy persons.
Infection by this organism may result in asymptomatic infection,
acute suppurative cutaneous infection, and chronic pulmonary
infection ranging in severity from mild bronchitis to necrotizing
pneumonia. All may progress to sepsis.