Pseudomonas aeruginosa

Pseudomonas aeruginosa &
Related Species
Dr. Suprakash Das
M.D (Microbiology)

Introduction
 Pseudomonas and related nonfermentative rods are opportunistic
pathogens of plants, animals, and humans.
 Despite the many genera, most clinically significant isolates are
members of five genera:
1) Pseudomonas,
2) Burkholderia,
3) Stenotrophomonas,
4) Acinetobacter, and
5) Moraxella.
 The genus Pseudomonas originally consisted of a large
heterogeneous collection of nonfermentative bacteria that were
grouped together because of their morphologic similarity.
 They were referred to as pseudomonads because they are
commonly arranged in pairs of cells that resemble a single cell.
 There are still almost 200 species in Pseudomonas.
 Pseudomonas aeruginosa is the most important species.

Introduction
Habitat-
 Members of the genus are found in
 soil,
 decaying organic matter,
 vegetation, and
 water.
 Unfortunately, they are also found throughout the hospital environment in
 moist reservoirs, such as food, cut flowers,
 sinks,
 toilets,
 floor mops,
 respiratory therapy and
 dialysis equipment, and even
 in disinfectant solutions.
 It is uncommon for carriage to persist in humans as part of the normal microbial flora,
except in
 hospitalized patients and
 ambulatory,
 immunocompromised hosts.

HISTORY-
 In 1850, it was noted by Sédillot that there were sometimes
blue-green discharges on surgical dressings that were
associated with infection, and
 In 1862 Luke noted rod-shaped microscopic entities within
the blue-green pus.
 In 1882, Gessard isolated the organisms and originally
designated them as Bacillus pyocyaneus.
 In 1925, Osler thought the organism to be more of a
secondary or opportunistic invader of damaged tissues as
opposed to a primary cause of infection in healthy tissues.
 P. aeruginosa emerged as a major human pathogen in the
1960s because of its ability to cause infections in
immunocompromised and burned hosts as well as CF
patients,
 all of whom were surviving much longer with modern
medical treatments.

Molecular Taxonomy
 At one time, most of the species belonging to the genera
Brevundimonas, Burkholderia, Ralstonia, and Acidovorax were
members of the genus Pseudomonas.
 Organisms in these genera have many similar phenotypic
characteristics but are genetically distinct.
 These organisms have been categorized into five unrelated rRNA
homology groups through ribosomal ribonucleic acid (rRNA)–
deoxyribonucleic acid (DNA) hybridization studies.
 Pseudomonas spp. belong to rRNA homology Group I;
 Burkholderia and Ralstonia spp. belong to rRNA homology Group
II;
 Acidovorax spp. belong to rRNA homology Group III (along with
genera Comamonas, Delftia, Hydrogenophaga spp.);
 Brevundimonas spp. make up rRNA homology Group IV
 Stenotrophomonas spp. belong to Group V.

Morphology
 Phenotypically, they are all
 aerobic,
 non–spore-forming,
 straight or slightly curved,
 slender, gram-negative bacilli with
 cells that range from 1 to 5 µm long and 0.5 to 1 µm
wide.
 All species except Burkholderia mallei are motile, having
one or several polar flagella.
 At the ultrastructural level, P. aeruginosa produces a single
or monotrichous polar flagellum and many cell surface
fimbriae or pili.
 Almost all strains carry the biosynthetic genes to produce an
extracellular polysaccharide known as alginate because of
its chemical similarity to seaweed alginate.

Morphology
 This material has also been referred to as mucoid
exopolysaccharide and its overproduction is the basis for the
mucoid colony phenotype associated with isolates from CF patients
as well as
 occasional isolates from other patients with chronic P. aeruginosa
infections, such as chronic obstructive pulmonary disease, or
 chronic infections in patients with indwelling urinary catheters.
 In general, strains of P. aeruginosa isolated from the environment
and nosocomial infections are considered nonmucoid, but in
reality most nonmucoid strains can express low levels of the alginate
polysaccharide when grown in vitro.
 Recently identified extracellular polysaccharides involved in the
formation of
 Pellicles - termed Pel and
 Biofilms- termed Psl, also appear to be important factors involved
in pathogenesis.

Morphology
 Environmental and nosocomial isolates also produce a smooth
lipopolysaccharide (LPS) substituted with long (5 to 100 kDa)
polysaccharide O side chains,
 Only approximately 10% to 30% of the LPS molecules contain O
side chains.
 The rest of the LPS molecules are in the form of LPS rough
molecules containing lipid A, inner- and outercore regions but
lacking O side chains.
 The outer membrane is typical of those for gram-negative bacteria
and contains a large array of outer membrane proteins (OMPs)
with a variety of functions critical for cellular growth and
metabolism.
 It is estimated that there may be more than 300 potential OMPs,
and
 Their expression varies dramatically depending on growth
conditions or phenotype of strains used for analysis of the OMPs

Electron micrographs of
Pseudomonas aeruginosa
cell showing ultrastructural
features. A, Single cell with
polar flagellum (F) with part of
the flagellum running under the
cell body. B, Two cells showing
thin, hairlike pili (P).

Epidemiology
 P. aeruginosa is primarily encountered as a nosocomial
pathogen- grow in a variety of environments with minimal
nutritional components.
 Outside the hospital, P. aeruginosa is commonly found in soil,
water, and plants and can occasionally be associated with
colonization of otherwise healthy humans and animals.
 Within the hospital, P. aeruginosa can colonize moist surfaces of
patients on the axilla, ear, and perineum and is also isolated
from other moist, inanimate environments including water in
sinks and drains, toilets, and showers.
 Pathogenic strains have also been isolated from the water used for
flowers in patients’ rooms.
 Hospital equipment that comes in contact with water, such as
mops, respiratory ventilators, cleaning solutions, and food and
food processing machines, can be sources of P. aeruginosa.

Epidemiology
 Community- acquired P. aeruginosa infection does
occur in certain settings, and infection is often
associated with exposure to moist environments.
 P. aeruginosa skin infections related to use of
 hot tubs,
 whirlpools,
 swimming pools are well recognized clinical
presentations of community-acquired infection.
 Individuals who use contact lenses, particularly of the
extended-wear variety that can be kept in the eyes for
several weeks, have a greatly increased risk of P.
aeruginosa ulcerative keratitis associated with the
presence of the bacteria in their contact lens solutions.
 Otitis externa is frequently caused by P. aeruginosa.

Epidemiology
 Puncture wounds through tennis shoes can give rise to
serious P. aeruginosa infection.
 Infections of the interdigital webs of the toes are
associated with maceration from topical antibiotic ointment.
 Perinychia is associated with constant exposure of the
extremities to water, detergents, or mechanical stress.
 P. aeruginosa endophthalmitis after surgery or eye trauma
can result in serious compromise of vision, and
 P. aeruginosa endocarditis can occur in injection drug
users.
 Endogenous P. aeruginosa brought into intensive care
units (ICUs) by patients from the community can serve as
sources of serious infection.
 Patients with significant burn wounds are at high risk for
P. aeruginosa infection.

Epidemiology
 Up to 7% of healthy humans carry P. aeruginosa in the
 Throat,
 Nasal mucosa, or
 On the skin, and
 carriage rates as high as 24% in the Stool have been reported.
 Many patients, particularly those receiving mechanical
ventilation, become colonized with P. aeruginosa in the upper
airways, but only a minority go on to develop pneumonia.
 Gastrointestinal (GI) colonization with P. aeruginosa,
usually secondary to antibiotic use that disrupts the normal
microbial populations of the GI tract, can lead to aspiration,
respiratory tract colonization, and sometimes pneumonia.

Molecular Epidemiology
 Determining the source and prevalence of P.
aeruginosa in the hospital environment is essential for
effective epidemiologic control of infection, and many
newer techniques are available to carry out effective
investigations using modern molecular typing
techniques. Typing by
 Colonial morphology or phenotype,
 Antibiogram,
 serology of variant LPS antigens,
 Bacteriophages,
 Bacteriocins known as pyocins, or
 Biochemical profiles using standard clinical
microbiologic analysis
 is fairly limited and generally has insufficient
discriminatory power.

Molecular Epidemiology
 Preferred techniques target genomic sequences and include
 DNA size analysis by pulsed-field gel electrophoresis (PFGE),
 analysis of DNA restriction enzyme-based fragment
polymorphisms (RFLPs),
 use of randomly amplified pieces of DNA showing a high degree
of polymorphism among strains (termed RAPD for randomly
amplified polymorphic DNA or AFLP for amplified fragment
length polymorphism
 use of Multilocus sequence typing (MLST) wherein short DNA
sequences in defined genes known to be variable among strains are
determined and used for strain classification, and
 use of polymerase chain reaction (PCR)-RFLP) to discriminate
strains on the basis of DNA sequence variations found in the small
subunit ribosomal RNA genes (ribotyping or riboprinting).

Pathogenesis
HOST FACTORS IN PATHOGENESIS-
 The primary determinant of the pathogenic potential of P. aeruginosa
virulence factors is the health status of the human host.
 Because healthy humans are generally highly resistant to P. aeruginosa
infection, some compromise in host health status underlies most of the
serious problems with P. aeruginosa infection.
 Some of the important predisposing factors are
 Neutropenia
 Disruption of skin and mucosal surfaces
 Burn wound
 Use of I.V/ Urinary catheters
 Complement deficiency (C3a /C5a)
 Defect in collectins- soluble molecules involved in innate resistance to P.
aeruginosa infection and include the surfactant proteins A (SP-A) and D (SP-
D) and mannose (or mannan)-binding lectin (MBL)
 Expression of various cytokines like-IL-1/ TNF-α
 Defect in Lung macrophages/ CD4+ cells
 Defect in CFRT protein

Defense mechanism of Lung
 Airway defenses render the lung an inhospitable environment to
inhaled microorganisms.
 Bacteria become trapped in the viscous mucous layer, which is
swept out of the lung by the rhythmic unidirectional beating of
millions of cilia.
 Flagella, lipopolysaccharide, and type 4 pili of Pseudomonas
aeruginosa are highly inflammatory and can be recognized by host
pattern recognition receptors such as TLRs to initiate an
inflammatory response via the NFκB signaling pathway.
 Activated alveolar macrophages as well as neutrophils recruited
by IL8 phagocytose and kill P. aeruginosa.
 Dendritic cells sample the lumen of the lung from the basal
lamina and activate the adaptive response (B cells and T cells).
 The lumen of the lung is also made inhospitable for
microorganisms by the presence of secreted antimicrobial
peptides such as a-defensins, lactoferrin, and lysozyme.

P. aeruginosa Virulence factors
 A multitude of virulence factors are produced by Pseudomonas
aeruginosa.
 Flagella and type 4 pili are the main adhesins, capable of
binding to host epithelial gangliosides, asialoGM1 and
asialoGM2.
 Along with lipopolysaccharide, these surface appendages are
also highly inflammatory.
 Once contact with host epithelia has occurred, the T3SS can
be activated, which is able to inject cytotoxins directly into
the host cell.
 Several virulence factors are secreted by P. aeruginosa and
have varying effects on the host.

 Several proteases are produced, which can degrade
host complement factors, mucins, and disrupt tight
junctions between epithelial leading to dissemination of
the bacteria.
 Lipases and phospholipases can target lipids in the
surfactant as well as host cell membranes.
 Pyocyanin, a blue-green pigment, can interfere with
host cell electron transport pathways and redox cycling.
 Pyoverdine captures Fe3+ to allow for a competitive
edge in an environment in which free iron is scarce.

P. aeruginosa –Antimicrobial Resistance Mechanisms

Clinical Manifestations
Pulmonary Infections-
 Asymptomatic colonization
 Tracheobronchitis
 Severe necrotizing bronchopneumonia.
 VAP
 Hospital associated pneumonia
 Community acquited pneumonia
Skin and Soft Tissue Infections-
 Folliculitis- Resulting from immersion in contaminated water (e.g.,
hot tubs, whirlpools, swimming pools). Secondary infections with
Pseudomonas also occur in people who have acne or who depilate
their legs.
 Fingernail infections in people whose hands are frequently exposed
to water or frequent “nail salons.”
 P. aeruginosa is also the most common cause of osteochondritis
(inflammation of bone and cartilage) of the foot after a penetrating
injury (e.g., associated with stepping on a nail).

P. aeruginosa Necrotizing Pneumoniua

Urinary Tract Infections
 Infection of the urinary tract is seen primarily in patients
with long-term indwelling urinary catheters.
 Typically, such patients are treated with multiple courses of
antibiotics, which tend to select for the more resistant
strains of bacteria, such as Pseudomonas.
Ear Infections
 External otitis is frequently caused by P. aeruginosa,
with swimming an important risk factor (“swimmer’s
ear”). This localized infection can be managed with topical
antibiotics and drying agents.
 Malignant external otitis is a virulent form of disease seen
primarily in persons with diabetes and elderly patients. It
can invade the underlying tissues, can damage the cranial
nerves and bones, and can be life threatening.
 P. aeruginosa is also associated with chronic otitis media.

Eye Infections
 Infections of the eye occur after initial trauma to the cornea (e.g., abrasion
from contact lens, scratch on the eye surface) and then exposure to P.
aeruginosa in contaminated water.
 Corneal ulcers develop and can progress to rapidly progressive, eye-
threatening disease unless prompt treatment is instituted.
 P. aeruginosa endophthalmitis is one of the most feared P. aeruginosa
infections. Loss of sight and very much reduced visual acuity are the most
common outcomes of these infections
Bacteremia and Endocarditis
 Bacteremia caused by P. aeruginosa is clinically indistinguishable from
that caused by other gram-negative bacteria.
 However, the mortality rate in affected patients is higher with P. aeruginosa
bacteremia because of
(1) the predilection of the organism for immunocompromised patients,
(2) difficulty in treating antibiotic-resistant strains, and
(3) the inherent virulence of Pseudomonas.

Contact lens–related Pseudomonas aeruginosa keratitis in a 49-year

 Bacteremia occurs most often in patients with
 Neutropenia,
 Diabetes mellitus,
 Extensive burns, and
 Hematologic malignancies.
 Most bacteremias originate from infections of the
 Lower respiratory tract,
 Urinary tract, and
 skin and soft tissue (particularly burn wound infections).
 Although seen in a minority of bacteremic patients,
characteristic skin lesions (ecthyma gangrenosum) may
develop. [Pic below]
 The lesions manifest as erythematous vesicles that become
hemorrhagic, necrotic, and ulcerated.

 Pseudomonas endocarditis is uncommon, primarily seen in intravenous
drug abusers.
 These patients acquire the infection from the use of drug paraphernalia
contaminated with the waterborne organisms.
 The tricuspid valve is often involved, and the infection is associated with a
chronic course but with a more favorable prognosis than that in patients
who have infections of the aortic or mitral valve.
Other Infections
 Sternoclavicular septic arthritis
 Vertebral osteomyelitis
 Meningitis
 Septic Arthritis of the Symphysis Pubis
 Skull Base Osteomyelitis
 Osteomyelitis Related to Nail Puncture Wounds
 Paronychia (Green Nail Syndrome)
 Hot Hand-Foot Syndrome

Septic Sterno-clavicular arthritis

Diagnosis
Microscopy
 Observation of thin, gram-negative rods arranged singly and in pairs is
suggestive of Pseudomonas but not pathognomonic—Burkholderia,
Stenotrophomonas, and other pseudomonads have a similar morphology.
Culture-
 Because Pseudomonas has simple nutritional requirements, the bacteria are
readily recovered on common isolation media such as blood agar and
MacConkey agar.
 They do require aerobic incubation (unless nitrate is available), so their
growth in broth is generally confined to the broth-air interface, where the
oxygen concentration is the highest.
 There are 5 types of colonies of P. aeruginosa-
1. Circular smooth colonies (shows swarming)
2. Irregular Contoured colonies (shows swarming)
3. Dry, flat colonies (shows swarming)
4. Mucoid colonies (CF Patients)
5. Rugose colonies

Dry, Flat, Irregular margin colonies

Smooth , round colonies
P. aeruginosa on Blood Agar

King B agar-
used
For
Pigmentation
Test/
Production

Diagnosis
Identification
 The colonial morphology (e.g., colony size, hemolytic
activity, pigmentation, odor; and
 The results of selected rapid biochemical tests (e.g.,
positive oxidase reaction) are sufficient for the
preliminary identification of these isolates.
 For example, P. aeruginosa grows rapidly and has flat
colonies with a spreading border, β-hemolysis, a green
pigmentation caused by the production of the blue
(pyocyanin) and yellow-green (pyoverdin) pigments,
and
 Characteristic sweet, grapelike odor.

TREATMENT
Bacteremia-
 Intravenously either
 ceftazidime 2g every 8 hours,
 cefepime 2g every 8 hours,
 meropenem 1.0g every 8 hours, or
 imipenem-cilastatin 0.5 g every 6 hours.
 If piperacillin-tazobactam is used, it should be given as 3.375g
every 4 hours or 4.5g every 6 hours.
 Aztreonam 2g every 8 hours has been used for patients with serious
β-lactam allergy.
 The addition of amikacin 15mg/kg every 24 hours may be
considered.
 Addition of an aminoglycoside to the other regimens is perhaps less
critical and probably depends on the level of resistance to β-lactam
antibiotics in any given institution.
 Currently, the highest levels of susceptibilities are to amikacin in
most regions of the world.

TREATMENT
Hospital-Acquired Pneumonia among Ventilated Patients-
 The American Thoracic Society and the Infectious Diseases Society of
America (ATS/IDSA) recommend empirical antimicrobial therapy for
ventilator-associated pneumonia with an antipseudomonal β-lactam
plus either an antipseudomonal quinolone or an aminoglycoside.
 The choice of specific antimicrobial agents should take into account the
local antimicrobial susceptibility patterns and avoiding agents to which the
patient was recently exposed.
 If aminoglycosides are chosen, consideration should be given toward a
short (5-day) course, to minimize nephrotoxicity, when used in
combination with a β-lactam to treat P. aeruginosa pneumonia.
 Once susceptibility profiles are available, monotherapy also should be
considered because data supporting combination therapy are lacking.
 Aerosolized antimicrobial agents (aminoglycosides, polymyxins) can
be used as adjunctive therapy to systemic antimicrobial agents in patients
with highly resistant P. aeruginosa ventilator-associated pneumonia.

TREATMENT
Community-Acquired Pneumonia-
 ATS/IDSA guidelines for P. aeruginosa community-acquired
pneumonia recommend an antipseudomonal β-lactam
antimicrobial agent plus either ciprofloxacin, levofloxacin,
or an aminoglycoside.
 An alternative recommended regimen is an aminoglycoside
plus either ciprofloxacin or levofloxacin.
 Tapering to monotherapy once antimicrobial susceptibilities
are available should then be considered.
 The ATS/ IDSA guidelines state that longer durations of 3 to 7
days are rarely necessary but
 may be warranted if there are pulmonary cavities or other
evidence of tissue necrosis.

TREATMENT
Antimicrobial Therapy for Arthritis and Osteomyelitis-
 The IDSA 2013 guidelines for prosthetic joint infections caused by P.
aeruginosa recommend treatment with
 cefepime, 2 g IV every 12 hours, or
 meropenem, 1 g IV every 8 hours.
 Alternative antimicrobial agents to consider include
 ciprofloxacin, 750 mg orally twice a day or 400 mg IV every 12 hours,
 ceftazidime, 2 g IV every 8 hours, for 4 to 6 weeks.
 use of aminoglycosides is optional
Simple Otitis Externa (Swimmer’s Ear)
 Topical antibacterial agents, such as otic tobramycin, ofloxacin, or a
combination of ciprofloxacin, dexamethasone otic, or aluminum acetate
drops, are beneficial.
Malignant Otitis Externa (Necrotizing Otitis Externa)
 Antipseudomonal antimicrobial agents are required for at least 6 weeks. Initial
therapy with intravenous antimicrobial agents should be considered in severe
cases; otherwise,
 oral ciprofloxacin at high doses (750 mg twice a day) is sufficient, as long as
the strain is susceptible.
 Aggressive debridement of necrotic tissue is also indicated.

TREATMENT
Keratitis-
 Mild cases can be successfully treated with topical antipseudomonal
antimicrobial agents, such as tobramycin ophthalmic
Endophthalmitis-
 Vitrectomy is necessary for severe cases and those that do not
respond to antimicrobial agents alone in the first 24 to 48 hours.
 The use of intravenous antipseudomonal antimicrobial agents has
not been studied in clinical trials but is recommended by some
experts in severe cases of endophthalmitis.
UTI-
 Treatment includes removal of the urinary catheter and correction of
underlying urologic problems, if present.
 Avoiding catheter insertion is also the predominant approach to
preventing catheter-associated urinary tract infection.
 Oral ciprofloxacin (500 mg twice a day) for 3 to 5 days for an
uncomplicated urinary tract infection is usually sufficient.
 Prolonging therapy to 2 to 3 weeks is indicated for complications,
including urosepsis and pyelonephritis.

Species Related to Pseudomonas

Burkholderia cepacia Complex
Clinical Manifestations-
 Respiratory Tract Infection in Cystic Fibrosis
 Lung Transplantation
 Hemophagocytic Syndrome
 Bacteremia and Pneumonia
 Skin, Skin Structure, and Joint Infection
 Genitourinary Tract Infection
Treatment-
 The newer fluoroquinolones, such as moxifloxacin, show improved activity
against S. maltophilia.
 However, resistance has developed after single-drug therapy, which can be
prevented by drug combinations including ticarcillin-clavulanic acid and/or
TMP/SMX.
 Moxifloxacin is bactericidal against TMP/SMX-resistant S. maltophilia,
although bacterial regrowth in in vitro experiments cautions use of
moxifloxacin as a single agent.
 Carbapenems, TMP/SMX, chloramphenicol, and tetracycline are active against
most BCC isolates.
 Treatment pending susceptibility testing will depend on experience of the
institution. Final antibiotic choices will depend on the bacterial drug
susceptibility profile.

Pseudomonas aeruginosa

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