B3.003 Prework 1 Culture Based Diagnostic Tests Flashcards
Koch’s postulates
- microorganism present in every case of the infectious disease
- microorganism can be isolated in pure culture
- inoculation of pure culture into animals produces similar disease
- same species of microorganism recovered from diseased animal
difference between envelopes of gram - and + bacteria
gram +
-thick peptidoglycan layer
-variable capsule
-potential for pili and flagella within envelope
gram -
-thin peptidoglycan layer
-variable capsule
-cytoplasmic and outer membrane
-periplasmic space
-cytoplasmic membrane> periplasm> peptidoglycan > outer membrane > capsule
-potential for pili and flagella within envelope
what component of the gram - cell wall is responsible for shock
Lipopolysaccharide (LPS) endotoxin
gram stain technique
- heat fix
- crystal violet stains all cells purple
- iodine solution: mordant-complexes with CV
- ethanol-acetone: destain, gram + retain CV/iodine complex
- safranin: counterstain; stains all cells pink
mycobacteria gram stain results
gram positive
but lipids interfere with stain
use acid fast
spirochetes gram stain results
gram negative
too thin to resolve
use wright-giemsa stain
sputum specimen collection
lower respiratory tract infection
avoid saliva
urine specimen collection
urinary tract infection
minimize contact with normal flora
feces specimen collection
diarrhea
rapid transport
large number of cultures
no direct stains
tissue specimen collection
center and border of lesion
pus specimen collection
abscess
aspirate
avoid swab
blood culture specimen collection
bacteremia
avoid contamination
ensure enough volume is collected per draw, 2 draws per patient
swab specimen collection
upper respiratory tract infection
swab inflamed areas
avoid saliva STDs
not for anaerobes
where do natural bacterial populations exist
skin
respiratory tract (oral cavity, pharynx)
GI tract (primary large intestine)
urogenital tract
upper respiratory tract normal flora
staphylococcus
streptococcus
haemophilus
anaerobes
skin normal flora
staphylococcus
coryneform (diptheroids)
propionibacterium
GI tract normal flora
anaerobe enterococcus enterobacteriaceae -Escherichia coli -klebsiella streptococcus lactobacillus candida
genital tract normal flora
lactobacillus
streptococcus
what characteristic of a sputum sample would indicate contamination?
> 10 epithelial cells per field
what feature of a sputum sample would indicate purulent infection?
presence of PMNs (polymononuclear cells)
growth media options
solid- agar
liquid - broth
enriched growth media
contains nutrients to support growth of a wide variety of organisms
blood agar, chocolate agar
selective growth media
inhibit growth of certain groups of organisms
differential growth media
distinguish organisms based on growth characteristics
obligate aerobe
requires oxygen for growth
facultative anaerobe
will grow in aerobic and anaerobic conditions
microaerophilic
grows best with low concentrations of oxygen
obligate anaerobe
oxygen is toxic
MacConkey Agar
selective: inhibits growth of gram + organisms
differential: distinguished between ability to ferment lactose
viral culture
growth on mammalian cells
not all viruses grow in viral culture
replaced by molecular methods in most clinical labs
fungal culture
specialized media
incubation at lower temperature
mycobacterial culture
broth and solid media
alpha-hemolysis
green, incomplete lysis around colony
beta-hemolysis
clear zone around colony
gamma-hemolysis
no hemolysis
yellow pigment production
staphylococcus aureus
red/pink pigment production
serratia marcescens
blue green pigment production
pseudomonas aeruginosa
projections in colony
candida albicans
mucoid colony
klebsiella pneumoniae
swarming in colony
proteus vulgaris
catalase reaction
gas formation
coagulase reaction
precipitate formation
oxidase reaction
blue color
identifies pseudomonas, Neisseria, vibrio
methods of antimicrobial susceptibility testing (AST)
genotypic: PCR based detection of the genes associated with drug resistance
phenotypic: tests ability of organisms to grow in the presence of drug
susceptible
isolate is inhibited by achievable concentrations of the antimicrobial agent
intermediate
response rate isolate may be lower than that for susceptible isolates
resistant
isolates are not inhibited by achievable concentrations of antimicrobial agent
limitations of AST
imprecise
does not take into effect host response, toxin production, biofilm formation
mecA gene
associated with methicillin resistance in s. aureus
vanA/B genes
associated with vancomycin resistance in enterococcus faecium
blaKPC gene
associated with carbapenem resistance
advantages of genotypic testing
fast
very specific
very sensitive
limitations of genotypic testing
expensive
single target
MIC
minimum inhibitory concentration
minimum concentration of a drug that will inhibit growth of the organism
methods of phenotypic testing
agar dilution
broth microdilution
disk diffusion
strip
microbroth dilution
broth + increasing concentration of antibiotic in microtube or 96 well plate
inoculate isolate into each well
semi quantitiative
strip testing
paper strip with increasing concentration of drug inoculate plate with lawn of isolate place strip in center of plate read MIC at area of growth inhibition quantitative
disk diffusion test
paper disk w single concentration of antibiotic place on agar inoculated with testing bacteria
measurement of the zones of growth inhibition
does not give MIC
