5.58 Flashcards
For ruling out syphilis (2)
•Rapid plasma reagin (RPR) test
•Venereal Diseases Reference
Laboratory(VDRL) test
•Venereal Diseases Reference
Laboratory(VDRL) test
•These tests have a low
false negative rate (very sensitive)
Follow up a positive (2) test
to confirm syphilis (need a very
— test)
RPR or VDRL
specific
Diagnosing infections by microscopy
• Direct microscopic examination of clinical specimens
• Direct microscopic examination of clinical specimens
Syphilis -
spirochetes observed in scrapings of lesions
Diagnosing infections by microscopy
• Stains
Gram stain and acid-fast stain
Gram stain and acid-fast stain (3)
- Bacteria in normally sterile body fluid (CSF, pleural fluid, urine)
- Staining properties part of larger effort at diagnosis
- Actual diagnosis (sometimes)
e.g. Gram-negative diplococci in leukocytes of CSF
= meningococci (agent of meningitis)
e.g. acid-fast bacteria in smears of sputum
= Mycobacterium tuberculosis
Gram stain
4
•Apply the primary stain (crystal violet) to a
heat-fixed smear of bacteria.
•Add a trapping agent (or mordant) (Gram
s
iodine).
•Decolorize with acetone or alcohol. Thick
peptidoglycan layer of Gram positives keeps
crystal violet-iodine complex trapped within
cells. This step must be done quickly
(seconds); if too long the crystal violet will be
washed from both Gram-negatives and Gram
positives).
•Counterstain with safranin to stain
decolorized cells.
acid-fast stain (Ziehl-Neelsen method)
3
•Apply the primary stain (fuschin) + mordant
(carbolic acid) (combination is called
carbolfuchsin) to a heat-fixed smear of
bacteria. Place piece of absorbant paper
soaked with carbolfuchsin over the smear and
heat the slide to drive the stain+mordant into
cells.
•Decolorize with dilute acid in alcohol. The
carbolfuchsin will wash out of most cells, but
not those with high levels of mycolic acid in
their membranes (e.g. mycobacteria)
•Counterstain with methylene blue to stain
decolorized cells.
Gram-negative diplococci in urethral pus =
gonococci (gonorrhea) (only see counterstain)
• Antibody-based identification -
Direct Fluorescent Antibody (DFA) test
Diagnosing infections by culture
most bacteria:
culturing involves use of numerous kinds of growth media
can provide preliminary information about biochemical nature of bacterium
additional biochemical tests used following isolation
Diagnosing infections by culture
some bacteria are not routinely cultured
rickettsias, chlamydiae, and mycoplasmas
identified with special stains, immunologic tests, or molecular methods
Open-ended sampling versus looking for a particular pathogen
Pus specimen from brain abscess
Throat culture examined for
Streptococcus pyogenes
selective media
prepared by the addition of specific substances to a culture medium that will permit growth of one group of bacteria while inhibiting growth of some other groups
ex. SS