Infection Sciences Flashcards

There are three specimen categories:

1. Specimens from sterile sites e.g. cerebrospinal
   fluid, blood.
2. Samples from non-sterile sites. e.g. faeces,
   throat swabs
3. Specimens from sites where pathogen is
   localised at a certain site but need to pass
   through a site that contains its microbiota, e.g. voided urine, expectorated sputum.
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   Specimen
   Clinical specimen
   • Should represent the diseased area and other appropriate sites
   • Must be large enough for carrying out a variety of diagnostic tests
   • Must be collected in a manner that avoids contamination
   • Must be forwarded promptly to clinical laboratory
   • Must be obtained prior to administration of antimicrobial agents, if
   possible
   • The clinical microbiologist function is to isolate and identify microbes
   from clinical specimens rapidly
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   Collection methods
   Skin and mucous membranes
4. Sterile swab
   Limited volume can be collected
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   Collection methods
   Skin and mucous membranes 2. Needle aspiration - Blood - Cerebrospinal fluid (CSF) A physician will insert a needle
   into the space between two
   vertebrae in the lower spine
   .
   When the needle is in place,
   fluid can drip out into
   a
   collection vial
   . More than one
   vial may be needed, and the
   procedure can take several
   minutes
   .
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   Collection methods
5. Intubation: insertion of tube into
   body canal or hollow organ
   e.g. stomach specimens
6. Catheter: tubular instrument
   used to withdraw or introduce
   fluids into body cavity e.g. urine
7. Clean-catch method: collection
   of midstream portion of urine
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   Transport of specimen
    Should be timely
    May involve use of special media that preserve microbes in specimen
    Temperature control may be needed
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   Transport of specimen
   Swabs and transport media
   Some swabs may require supplementation to
   support microbial survival or inhibit the
   microbiota, e.g., use of antibacterial compounds
   such as penicillin to ensure recovery of fungi
   • Sustain the viability of microorganisms
   • Prevent the outgrowth of contaminants
   • Usually thioglycolate and cysteine are used in
   anaerobic transport media to reduce the level
   of oxygen. A built-in color indicator system
   (resazurin) is clear and turns lavender in the
   presence of oxygen.
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   What may go wrong when taking specimens
    Incorrect identification of patient
   e.g. patients with same or similar names.
   Date of birth and NHS number help.
    Mislabelling/ no labelling.
    Patient preparation e.g. fasting.
    Sample poorly/ incorrectly taken (blood stained CSF).
    Incorrect container(s) e.g. air in a sample for anaerobic culture.
    Inadequate amount.
    Incorrect storage/ transport (depends on the specimen ice, warm, delay).e.g.
   ◦ Urine 4oC
   ◦ Swabs (4oC for out-of-hours)
   ◦ Stool (4o)
    Loss, breakage etc.
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   Microscopy
   Choice of smear (how the specimen is prepared on the microscope slides):
   • Wet-mount (unfixed and unstained. E.g. urine and CSF)
   • Heat-fixed
   • Chemically fixed (Alcohol/acetic acid solution)
   The choice of microscopy depends on possible pathogen:
   • Bright field microscopy
   • Dark-field microscopy
   • Fluorescence microscopy
   All are compound microscopes
   with an image given by the added (multiplied!) magnification power of ≥2 lenses
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   Compound light microscope A compound microscope is an
   optical microscope that uses light and different lenses to magnify an
   object
   .
   Light travels through specimen and into a single objective lens
   and then trough the eyepiece (ocular).
   Specimen must be thin enough so
   that light must be able to pass through.
   Highest magnification is
   x1000
   (x10
   x100
   )
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   Magnification power
   To calculate the power of magnification,
   multiply the power of the ocular lens by
   the power of the objective
   .
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   Bright
   -field microscope
   • The name "bright
   -field"
   is derived from the fact
   that the specimen is
   dark and contrasted by
   the surrounding bright
   viewing field
   .
   • It
   uses visible light as
   source of illumination
   • It produces an image of
   the object against
   a
   bright background
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   Dark-field microscope
   • It produces a bright image of the object
   against a dark background
   • It is useful to examine transparent or semitransparent specimens which cannot be
   distinguished
   • It shuts out background light and allows
   only scattered light to reach the specimen
   in order to heighten textural detail.
   Borrelia burgdorferi
   Dark-field Bright-field
   Miklossy et al., 2008 Journal of Neuroinflammation CC BY 2.0
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   Fluorescence microscope
   The specimen is stained with a fluorochrome
   and exposed to a UV or VIS light laser.
   A fluorescence microscope uses a much higher
   intensity light source (laser) which excites the
   fluorescent species in the sample of interest.
   This fluorescent species in turn emits a lower
   energy light of a longer wavelength that
   produces the magnified image instead of the
   original light source.
   The resulting image is a bright image of the
   sample resulting from the fluorescent light
   emitted by the specimen with very high
   contrast and visibility.
   M. tuberculosis
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   Preparation of the microscope slide
8. Drying
   The smear is air-dried before fixing
9. Fixing
   It is the process by which internal and external
   structures are preserved and fixed in position.
   Microorganisms are killed and firmly attached to the
   microscope slide.
   Types of fixing:
   Heat fixing

• It preserves the overall morphology but not
  the internal structures
  Chemical fixing
• It protects fine cellular substructures and
  morphology of larger, more delicate
  organisms. E.g. alcohol/acetic acid solution