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COMMONWEALTH OF AUSTRALIA
Copyright regulations 1969
Warning
This material has been reproduced and communicated to you by or on behalf of the University of Melbourne pursuant to part
VB of the Copyright Act 1968 (the Act).
The material in this communication may be subject to copyright under the act. Any further copying or communication
of this material by you may be the subject of copyright protection under the act.
Do not remove this notice
Opportunistic Infections
Objectives
• To appreciate the nature of opportunistic infections by understanding (1) the nature of opportunistic pathogens, (2) the contribution of host factors to infection, and (3) the reasons for the increased susceptibility of hospitalised patients to infection
• To illustrate some principles of bacterial pathogenesis and molecular epidemiology, using the opportunistic pathogen, Pseudomonas aeruginosa, as an example.
Microbial pathogenicity vs . Host resistance
colonisation
penetration
multiplication
tissue damage
physical and chemical barriers
inflammatory response phagocytosis
adaptive immune response
disease or protection
Pathogen-Host Relationships
disease threshold
Microbial Virulence
Host Resistance
How Microbial Virulence and Host Resistance Affect Disease Outcome
• E. coli • Staphylococcus aureus • Klebsiella pneumoniae • Enterococcus spp. • Pseudomonas aeruginosa • Enterobacter spp. • Serratia spp. • Proteus spp. • Clostridium difficile
Frequently Encountered Opportunistic Pathogens: “The Seed”
* may cause nosocomial epidemics
* * * * * *
• Local • anatomical defects
• surgical and other wounds
• burns
• catheterisation (bladder, IV)
• foreign bodies in general
Host Factors: “The Soil”
• Systemic • extremes of age • leucopenia • malignancy • malnutrition • diabetes • liver disease • certain infections • treatment with antimicrobials • primary (congenital) immunodeficiency
Host Factors (cont.)
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• wound infection
• UTI
• intra-abdominal infection
• pneumonia
• septicaemia (sepsis)
• meningitis (esp. neonates)
Types of Infection
• endogenous (own microbiota)
• exogenous
Sources
Culture appropriate specimen: • wound swab
• pus
• urine
• sputum
• blood
• CSF
• etc.
Diagnosis
• Depends on antibiotic susceptibility
• nosocomial strains are often multiresistant
• may need potent bactericidal agents
Treatment
• Aseptic technique (esp. hand hygiene)
Prevention
Education
Pseudomonas
• G− rod
• Motile
• Aerobes or facultative anaerobes
• Non-fermenting, non-sporing
• Some species, incl. Ps. aeruginosa, produce pigments
• Widespread
• Low nutritional requirements
Morphology of Pseudomonas aeruginosa
Colonies on nutrient agar Gram stain of culture
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Classification
• Divided into species according to:
• Biochemical tests
• Ps. aeruginosa subtyped according to:
• Serotype, biotype (seldom used)
For epidemiological studies, generally use
• RFLP or MLST (multilocus sequence typing)
Pulsed-Field Gel Electrophoresis of Genomic DNA of Ps. aeruginosa Isolates from Children with Cystic Fibrosis
1 2 3 4 5 6 7 8 9 10 11 12 kb
500-
400-
300-
200-
100-
50-
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Species of Pseudomonas and Related Genera
• Ps. aeruginosa* : ubiquitous saprophyte, important opportunistic pathogen
• Burkholderia cepacia* : opportunistic pathogen
• Stenotrophomonas maltophilia* : opportunistic pathogen
• Burkholderia pseudomallei : causes melioidosis
* These species often colonise the respiratory tract of patients with cystic fibrosis, and are intrinsically resistant to many commonly-used antibiotics
Pseudomonas aeruginosa : overview
• Epidemiology
• Diseases
• Pathogenesis (incl. biofilms and quorum sensing)
• Principles of management
Ps. aeruginosa : epidemiology
• Widespread in nature, esp. in moist environments
• Transiently colonises skin, mucous membranes, git
• A leading nosocomial pathogen:
• previously assoc. with burns, febrile neutropenia
• now mainly pneumonia, UTI, sepsis
• Major problem in patients with cystic fibrosis
Ps. aeruginosa : epidemiology (cont.)
• Infections mostly acquired from environment
• Spread in hospital on hands and fomites
• Some strains spread between patients with CF
• All strains are intrinsically resistant to many antibiotics and weak disinfectants
• Resistance also readily acquired from other bacteria
Ps. aeruginosa : diseases
• Superficial infections: • Skin: wound infection, otitis externa, folliculitis
• Eye: keratitis, corneal ulcer, (also deep infection)
• Deep and systemic infections: • Pulmonary: nosocomial pneumonia, chronic
infection in patients with cystic fibrosis
• Other: UTI, endocarditis, osteomyelitis, septicaemia, etc. in immunocompromised patients
Ps. aeruginosa Pathogenesis (1): Adhesion and Invasion
• Does not invade intact skin unless very large numbers
• Adheres weakly to intact epithelium via flagella, pili, LPS
• As with other bacteria, LPS and flagellin bind to Toll-like receptors: affect cytokine production
• LPS core binds to CFTR (CF transmembr. conductance reg.)
• Once adherent, bacteria tend to produce biofilm
• Capsule assists adherence and biofilm formation
Properties of Ps. aeruginosa in Biofilms
• Non-motile
• More capsule material - mucoid phenotype
• More adherent
• Less invasive
• Shorter LPS (no O-antigen)
• Slowed growth
• Increased resistance to antibiotics
Much of this results from the activation of genes that are regulated by quorum sensing
response to environmental signals
Bacterial “cross-talk” (= quorum sensing)
Stages in Biofilm Formation
primary-plus.com
An Example of How Quorum Sensing Works
Broth culture of light- emitting bacteria at
high cell density
B. Bassler
An Example of How Quorum Sensing Works
low cell
density
autoinducer
LuxR LuxR
LuxI
luxR lux
box luxI luxC D A B E
autoinducer
high cell
density
autoinducer
LuxR LuxR
LuxI
luxR lux box luxI luxC D A B E
autoinducer
LuxR
Light
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Properties of Ps. aeruginosa in Biofilms
• Non-motile
• More capsule material - mucoid phenotype
• More adherent
• Less invasive
• Shorter LPS (no O-antigen)
• Slowed growth
• Increased resistance to antibiotics
Ps. aeruginosa Pathogenesis (2): Spread and Multiplication
Spread through tissues and body facilitated by:
• Reduced PMNs
• Flagella
• Exoezymes: proteases, haemolysins, phospholipases, elastase. Act on pulmonary tissue and surfactant
• Exotoxins: exotoxin A; exoenzymes S and U (EXoS, ExoU), etc. Inhibit phagocytosis
• LPS-CFTR-mediated invasion: aggravates corneal damage
What Happens in Patients with Cystic Fibrosis?
• Defect in CFTR → abnormal ion transport, thickened mucus and impaired mucociliary function
• Pseudomonas and Staph not inhibited by high salt concs.
• Bacteria in biofilms resist mechanical removal and are less visible to the innate immune system
• DNA secreted by bacteria and released from dying cells → thickened mucus
• Type of host antibody response may affect outcome.
Ps. aeruginosa Pathogenesis (3): Tissue Damage
Type III secreted proteins • Exoenzymes S, T, U and Y (=ExoS, ExoT, ExoU, ExoY):
act on various host cell targets to interfere with phagocytosis and enhance cytokine production
Other Exotoxins and Enzymes • Exotoxin A: blocks protein synthesis (≡ diphtheria toxin)
• Elastase: damages connective tissue
• Phospholipase: damages cell membranes, degrades surfactant
Ps. aeruginosa : Control
• Reduce risk of susceptible patients by suitable management of burns, neutropenia, catheters, ventilators
• Be on alert for infection and treat early
• Practice high levels of hand hygiene
• Use contact lenses and solutions appropriately
• Lifelong monitoring and treatment of CF patients
Pseudomonas cannot be eradicated from the environment. Therefore management is directed towards prevention.
Ps. aeruginosa : Summary
• Ps. aeruginosa is widespread in the environment
• Its low nutritional requirements and genetic versatility allow it to occupy many different environments
• It is a prototypical opportunistic pathogen and a leading cause of hospital-acquired infections
• In humans, it shows a preference for situations where innate immunity is reduced
• Its intrinsic and acquired resistance to antimicrobials provide a therapeutic challenge
• Prevention is the key to controlling infection
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