Pseudomonas (page 2)
© Kenneth Todar, PhD
Resistance to Antibiotics
Pseudomonas aeruginosa is notorious for its resistance to
antibiotics and is, therefore, a particularly dangerous and dreaded
pathogen. The bacterium is naturally resistant to many antibiotics due
to the permeabiliity barrier afforded by its Gram-negative outer
membrane. Also,
its tendency to colonize surfaces in a biofilm form makes the cells
impervious
to therapeutic concentrations antibiotics. Since its natural habitat is
the soil, living in association with the bacilli, actinomycetes and
molds,
it has developed resistance to a variety of their naturally-occuring
antibiotics.
Moreover, Pseudomonas maintains antibiotic resistance
plasmids,
both R-factors and RTFs, and it is able to transfer these genes by
means of the bacterial mechanisms of horizontal gene transfer (HGT),
mainly transduction and conjugation.
Only a few antibiotics are effective against Pseudomonas aeruginosa,
including
fluoroquinolones, gentamicin and imipenem, and even these antibiotics
are
not effective against all strains. The futility of treating Pseudomonas
infections with antibiotics is most dramatically illustrated in cystic
fibrosis patients, virtually all of whom eventually become infected
with
a strain that is so resistant that it cannot be treated.
Diagnosis
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 odor, and its
ability to grow at 42°C. Fluorescence under ultraviolet light is
helpful in early identification of P. aeruginosa colonies.
Fluorescence
is also used to suggest the presence of P. aeruginosa in wounds.
Pathogenesis
For an opportunistic pathogen such as Pseudomonas aeruginosa,
the disease process begins with some alteration or circumvention of
normal
host defenses. The pathogenesis of Pseudomonas infections is
multifactorial,
as suggested by the number and wide array of virulence determinants
possessed
by the bacterium. Multiple and diverse determinants of virulence are
expected
in the wide range of diseases caused, which include septicemia,
urinary
tract infections, pneumonia, chronic lung infections, endocarditis,
dermatitis,
and osteochondritis.
Most Pseudomonas infections are both invasive and toxinogenic.
The ultimate Pseudomonas infection may be seen as composed of
three
distinct stages: (1) bacterial attachment and colonization; (2) local
invasion;
(3) disseminated systemic disease. However, the disease process may
stop
at any stage. Particular bacterial determinants of virulence mediate
each
of these stages and are ultimately responsible for the characteristic
syndromes
that accompany the disease.
Colonization
Although colonization usually precedes infections by Pseudomonas
aeruginosa, the exact source and mode of transmission of the
pathogen
are often unclear because of its ubiquitous presence in the
environment.
It is sometimes present as part of the normal flora of humans, although
the prevalence of colonization of healthy individuals outside the
hospital
is relatively low (estimates range from 0 to 24 percent depending on
the
anatomical locale).
The pili of Pseudomonas aeruginosa will adhere to the
epithelial cells
of the upper respiratory tract and, by inference, to other epithelial
cells
as well. These adhesins appear to bind to specific galactose or mannose
or sialic acid receptors on epithelial cells. Colonization of the
respiratory
tract by Pseudomonas requires pili adherence and
may
be aided by production of a protease enzyme that degrades fibronectin
in
order to expose the underlying pilus receptors on the epithelial
cell
surface. Tissue injury may also play a role in colonization of the
respiratory
tract, since P. aeruginosa will adhere to tracheal epithelial
cells
of mice infected with influenza virus but not to normal tracheal
epithelium.
This has been called opportunistic adherence, and it may be an
important
step in Pseudomonas keratitis and urinary tract infections, as
well
as infections of the respiratory tract.
The receptor on tracheal epithelial cells for Pseudomonas pili
is probably sialic acid (N-acetylneuraminic acid). Mucoid strains,
which
produce an exopolysaccharide (alginate), have an additional or
alternative
adhesin which attaches to the tracheobronchial mucin
(N-acetylglucosamine).
Besides pili and the mucoid polysaccharide, there are possibly
other
cell surface adhesins utilized by Pseudomonas to colonize the
respiratory
epithelium or mucin. Also, it is possible that surface-bound exoenzyme
S could serve as an adhesin for glycolipids on respiratory cells.
The mucoid exopolysaccharide produced by P. aeruginosa is a
repeating
polymer of mannuronic and glucuronic acid referred to as alginate.
Alginate slime forms the matrix of the Pseudomonas biofilm
which anchors the cells to their environment and in medical situations,
it protects the bacteria from the host defenses such as lymphocytes,
phagocytes,
the ciliary action of the respiratory tract, antibodies and complement.
Biofilm mucoid strains of Pseudomonas are also less
susceptible
to antibiotics than their planktonic counterparts. Mucoid strains of P.
aeruginosa are most often isolated from patients with cystic
fibrosis
and they are usually found in lung tissues from such individuals.
Invasion
The ability of Pseudomonas aeruginosa to invade tissues depends
upon production of extracellular enzymes and toxins that break down
physical
barriers and damage host cells, as well as resistance to phagocytosis
and
the host immune defenses. As mentioned above, the bacterial capsule or
slime layer effectively protects cells from opsonization by antibodies,
complement deposition, and phagocyte engulfment.
Two extracellular proteases have been associated with virulence
that exert their activity at the invasive stage: elastase and alkaline
protease. Elastase has several activities that relate to
virulence.
The enzyme cleaves collagen, IgG, IgA, and complement. It also lyses
fibronectin
to expose receptors for bacterial attachment on the mucosa of the lung.
Elastase disrupts the respiratory epithelium and interferes with
ciliary
function.
Alkaline protease interferes with fibrin formation and
will lyse fibrin. Together, elastase and alkaline protease destroy the
ground substance of the cornea and other supporting structures composed
of fibrin and elastin. Elastase and alkaline protease together are also
reported to cause the inactivation of gamma interferon (IFN) and tumor
necrosis factor (TNF).
Pseudomonas aeruginosa produces three other soluble proteins
involved
in invasion: a cytotoxin (mw 25 kDa) and two hemolysins.
The cytotoxin is a pore-forming protein. It was originally named
leukocidin
because of its effect on neutrophils, but it appears to be cytotoxic
for
most eucaryotic cells. Of the two hemolysins, one is a phospholipase
and the other is a lecithinase. They appear to act
synergistically
to break down lipids and lecithin. The cytotoxin and hemolysins
contribute
to invasion through their cytotoxic effects on neutrophils, lymphocytes
and other eucaryotic cells.
One Pseudomonas pigment is probably a determinant of virulence
for the pathogen. The blue pigment, pyocyanin, impairs the
normal
function of human nasal cilia, disrupts the respiratory epithelium, and
exerts a proinflammatory effect on phagocytes. A derivative of
pyocyanin,
pyochelin,
is a siderophore that is produced under low-iron conditions to
sequester
iron from the environment for growth of the pathogen. It could play a
role in invasion if it extracts iron from the host to permit bacterial
growth in a relatively iron-limited environment. No role in virulence
is known for the fluorescent pigments.
chapter continued
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