B3-003 Laboratory Diagnosis of Infectious Disease Flashcards
nucleic acid hybridization
NAAT
Sequencing
Mass spectrometry
are examples of
molecular testing
advantages: molecular testing
fast
sometimes increased sensitivity
limitations: molecular testing
expensive
targeted
no susceptibility testing
source of false positives for molecular testing
contamination at collection or lab
sources of false negatives for molecular testing
below limit of detection
biological inhibitors
steps of PCR
extract
denature
anneal
elongate
repeats
detection and quantification
examples of NAAT amplification and detection tests
single panel
multipanel: respiratory panel
most FDA-cleared NAAT test are
amplification + detection
all laboratory developed NAAT tests are
amplification and sequencing
examples of NAAT amplification and sequencing tests
broad range PCR
metagenomics
MALDI is used to
identify cultured bacteria
MALDI technique
isolate is spotted onto a metal plate
laser vaporized
separated by mass-to-charge ratio
profile created
comparison
MALDI may be used in the future for
susceptibility testing
strain typing
direct from sample detection
Method: antigen detection
use lab generated antibodies to detect patient antigen in specimen
specimens: antigen detection
urine, serum, sputum, CSF, stool
Galactomannan serum is used to detect
invasive aspergillus
cryptococcal urine or CSF is used to detect
cryptococcus
method: Antibody detection
use lab generated antigens to detect IgG, IgM, etc.
Specimen: antibody detection
serum
what technique would you use to diagnose Lyme disease?
Serology: antibody detection
Borrelia burgdorferi
what technique would you use to diagnose Syphilis?
Serology: antibody detection
Treponema pallidum
what technique would you use to diagnose Q fever?
Serology: antibody detection
Coxiella burnetti
Agglutination
serology method
antibody or antigen is fixed to latex beans, specimen added and observed for clumping
lateral flow immunoassay
serology method
antibodies attach to pad, wicked along, accumulate as visible line
what kind of diagnostic test is a pregnancy test?
lateral flow assay
advantages of lateral flow assay
rapid, cheap, POC
limitations of lateral flow assay
decreased sensitivity, difficult to read
ELISA has a _______ predictive value
high negative
(high sensitivity, low specificity)
all positive ELISA assays must be
confirmed with a screening confirmation test assay
**if the first is positive, and the second is negative, the result is NEGATIVE
method: ELISA
antibody or antigen immobilized in well
add patient sample
add reported antibodies/antigens
add substrate and read colormetric change
Screen confirmation tests are
labor intensive
low sensitivity but very high specificity
screening confirmation tests
Immunoblot, Immunodiffusion
What diagnostic test should you choose if you are trying to determine vaccination status?
serology
What diagnostic test should you concerned with an organism that won’t grow in culture or is eliminated quickly?
serology
what organisms don’t grow in culture
Lyme, Syphilis, Mono
What organisms are cleared quickly?
west nile, zika, dengue
What diagnostic test should be chosen for HIV, HCV, HBV screening?
serology
what would cause a false positive on a serology test?
heterophil antibodies/ RF
cross reaction to closely related antigens
maternal transfer
what would cause a false negative on a serology test?
immunocompromised
too early/late
gram stain technique
heat fix sample
crystal violet: stains all cells purple
iodine: mordant
destain with ethanol-acetone
safranin counterstain
gram positive organisms will be
purple
gram negative organisms will be
pink
acid-fast stain
mycobacteria
wright-giemsa stain
spirochetes, rickettsiae
gram + cocci in clusters
staphylococcus
gram + cocci in chains
streptococcus
sputum should be collected when _______ is suspected
LRTI
urine is collected when ______ is suspected
UTI
feces is collected when ______ is suspected
diarrhea
when getting a tissue culture, you should get the
center and border of tissues
aspirate _______, avoid swabs
pus
how much blood should be collected for a blood culture?
10 mL per bottle, 2 bottles x2
swabs are used when _______ is suspected
URTI, STD
a direct gram stain is done on what specimens
sputum, positive blood culture, abcess fluid, CSF, vaginal discharge
presence of PMNs in sputum indicates
active, purulent infection
> 10 epithelial cells per field suggests
contamination with oral flora
enriched media
contain nutrients to support growth of a wide variety of organisms
selective media
inhibit growth of certain groups of organisms
differential
distinguish organisms based on growth characteristics
MacConkey Agar is
selective: inhibits growth of gram+
AND
differential: distinguishes between ability to ferment lactose
obligate aerobe
requires oxygen for growth
facultative anaerobe
will grow in aerobic and anaerobic conditions
microaerophillic
grows best with low concentrations of oxygen
obligate anaerobe
oxygen is toxic
when should you draw blood cultures?
before antibiotics
2 bottle sets of blood cultures
1 aerobic
1 anaerobic
how many sets of blood cultures should you do?
two sets
(4 bottles, 40 mL total)
mycobacterial culture is done on
broth and solid media
fungal cultures are done on
specialized media
incubated at a lower temp
green, incomplete lysis around colony
alpha hemolysis
clear zone around colony
beta hemolysis
no hemolysis
gamma hemolysis
what color colonies are S. aureus?
yellow
what color are Serratia colonies?
red/pink
what color are P. aeruginosa colonies?
blue green
what bacteria grows mucoid colonies?
klebsiella
what organism grows projection-like colonies?
candida albicans
what organism demonstrates swarming on colony growth?
Proteus vulgaris
gas formation (bubbles) indicates
catalase positive
precipitate formation indicates
coagulase positive
blue color indicates
oxidase positive
clostridium
actinomyces
propionibacterium
gram positive anaerobic bacilli
listeria
cornyebacterium
bacillus
nocardia
gram positive aerobic bacilli
catalase +, coagulase + cocci grown in clusters
staphylococcus
catalase -, cocci grown in chains
streptococcus
anaerobic streptococcus
peptostreptococcus
S. pneumoniae
S. mutans
Viridians
alpha hemolytic streptococcus (GAS)
S. pyogenes
S. agalactiae
beta hemolytic streptococcus (GBS)
enterococcus
gamma hemolytic streptococcus
gram negative cocci
neisseria
moraxella
escherichia
klebsiella
enterobacter
gram negative
+ growth on MacConkey
+lactose fermentors
salmonella
shigellaa
proteus
serratia
yersinia
gram negative
+growth on MacConkey
-lactose fermentors
+glucose fermentors
pseudomonas
burkholderia
acinetobacter
gram negative
+growth on MacConkey
-lactose fermentor
-glucose fermentor
haemophilus
bordetella
legionella
brucella
franciella
pasteurella
campylobacter
helicobacter
vibrio
gram negative
- growth on MacConkey
+oxygen tolerance
bacteriodes
fusobacterium
prevotella
porphyromonas
gram negative
- growth on MacConkey
-oxygen tolerance
PCR based detection of the genes associated with resistance to a drug
genotypic antimicrobial susceptibility testing
tests ability of organism to grow in presence of drug
phenotypic antimicrobial susceptibility testing
susceptible
isolate is inhibited
intermediate
response rate for isolate is lower than that for susceptible isolates
resistant
isolates are not inhibited
limitations of antimicrobial susceptibility testing
imprecise
does not account for host response, toxin production, or biofilm formation
mecA gene
MRSA
vanA/B gene
Vanc resistant E. faecium
bla kpc gene
carbapenem resistance
beta lactamase resistant KPC
advantages of genotypic susceptibility testing
fast
very sensitive
very specific
limitations of genotypic susceptibility testing
expensive
single target: does not detect resistance by multiple genes
MIC
minimal inhibitory concentration
minimum concentration of a drug that will inhibit growth of an organism
phenotypic susceptibility testing: microbroth dilution
the concentration without bacterial growth is the MIC
phenotypic susceptibility testing: strip testing
read MIC at area of growth of inhibition
phenotypic susceptibility testing: Kirby Bauer (disk)
does not give MIC
measure zones of growth inhibition