Microbiological Diagnosis of Chest Infection Flashcards
What diagnostic techniques are used for chest infections?
- Microscopy and culture of blood and sputum
- Antigen detection methods
- Nucleic acid amplification (PCR)
- Serology (antibody measurement)
How valuable is gram staining for organisms and pus cells?
Limited value
What are the 3 major respiratory pathogens?
- Strep pneumoniae
- Hameophilius influenzae
- Moraxella Catarrhalis
What may you expect to find in sputum cultures?
Normal upper respiratory flora such as viridans streps
What does previous antibiotic therapy select out?
Organisms that are not respiratory pathogens such as:
- E. coli
- Staph aureus
How must sputum cultures be interpreted?
Using the clinical presentation:
- Community pneumonia
- Aspiration pneumonia
- Ventilated patient pneumonia
How is TB diagnosed?
Using ZN or auramine phenol stain
How is TB described?
Acid and Alcohol Fast Bacilli (AAFB)
How is a broncho-alveolar lavage done?
- Lower airway sample collected at bronchoscopy
- Catheter aspirate in ventilated patient
What is BAL used in the diagnosis of?
Ventilator associated pneumonia
What is an advantaged of BAL?
Less liable to contamination- more accurate diagnosis of LRTI
What does BAL produce?
Quantitative culture of BAL- colony forming units/ml (cfu/ml)
Who are blood cultures taken from?
Any patient with severe sepsis
What do many patients with pneumonia have in their blood?
Bacteraemia (viable bacteria in the blood)
Why is blood culture carried out?
There are too few organisms in blood to see on microscopy
How is blood culture carried out?
Blood inoculated into 2 bottle containing culture media and incubated
What happens on day 1 of blood culture?
- Microscopy positive in 6-48 hours (Gram stain)
- Result is phoed to the clinician and interim recommendations on antibiotics
- Overnight subculture of sensitivity and identification tests
What happens on day 2 of blood culture?
-Full ID and sens- clinical significance is re-assessed
What organisms are not easily cultured and are known as atypical causes?
- Legionella pneumophila
- Mycoplasma pneumoniae
- Chlamydia psittaci
- Coxiella burnetti
What is antigen detection?
- Non-cultural methods of demonstrating the presence of an organism
- Detect specific antigen immunologically
What antigens can be detected in urine?
Legionella and pneumococcal
What antigens can be detected in naso-pharyngeal secretions?
Viruses
What techniques are used in antigen detection?
- Agglutination
- EIA
- Immunofluorescence
How is latex agglutination done?
- Latex particles coated with monoclonal antibodies that are specific for the organism in question are mixed with the clinical specimen
- Agglutination (clumping) visible by naked eye
What does EIA stand for?
Enzyme Immuno-Assay
What does ELISA stand for?
Enzyme Linked Immunosorbent Assay
How is a positive EIA/ELISA result generated?
Linked enzyme on 2nd antibody generates colour change
What is monoclonal antibody the product of?
A single clone of plasma cells
Give an example of how ELISA/EIA works.
- Monoclonal anti-Legionella coated on well of microtitre plate
- Incubated with patients’s urine- antigen binds if present
- Incubated with enzyme linked monoclonal anti-Legionella
- Enzyme substrate added- colour change if enzyme present
What technique is used for DNA detection?
Polymerase chain reaction (PCR)
What specific primers are used in PCR?
Short oglionucleotide sequences
How does PCR work?
- Short oglionucleotide sequences bind to target DNA sequences
- Multiple copies of target sequence are produced
- Amplified copies of DNA easily detected
What is an advantage of PCR?
Very sensitive
What is a disadvantage of PCR?
False positives due to contamination
What does real time PCR allow?
Enables product of the reaction to be measured as the reaction progresses
What is the generic term for the technical variations of PCR?
Nucleic acid amplification test (NAAT)
What samples must be collected for diagnosis by PCR?
Throat swabs or other respiratory sample
What kind of swab is used for PCR?
Flocked swab
What is routinely screened for using PCR?
Mycoplasma pneumoniae
and a wide range od respiratory viruses (multiples assays)
What respiratory viruses are screened for using multiplex assays?
- Influenza A and B
- Adenovirus
- RSV
- Parainfluenza
- Metapneumovirus
- Coronavirus
- Rhinovirus
What does antibody detected by serology measure?
Host antibody response to organism
What is it usually difficult to do with antibody detection by serology?
Culture or detect the organism directly
What was antibody detection by serology previously used for?
Viral infections and so called atypical causes of pneumonia
What is IgM a marker of?
Current infection
What is IgG a marker of?
Previous infection
What must be demonstrated if total antibody is measured?
- A significant increase in levels during the coarse of the illness
- Takes some time for antibody levels to rise
What is antibody measurement now mostly used to determine?
Vaccine response
What organisms cause difficult to diagnose bacterial infections?
- Legionella penumophilia
- Chamydia psittaci
- C penumoniae
- Coxiella burnetti