microbiologic staining method 顏 慕 庸 高 雄 榮 總. microbiological staining gram stain acid...
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Microbiologic Staining Method
顏 慕 庸
高 雄 榮 總
Microbiological Staining
Gram stain
Acid fast stain
KOH stain
Wet mount
Indian ink
Gram Stain
• Christian Gram, 1884
• crystal violet
• iodine-KI
• alcohol wash
• safranin
Significance
• Culture
• Buy time for golden period
• Diagnosis
Interpretation
• Morphological
• Associate environment
• Site-specific
• Immunity-oriented
Morphologic interpretation
• cocci vs bacilli, cocco-bacilli
• Gram positive vs Gram negative
Morphologic interpretation
• cocci vs bacilli, cocco-bacilli
• Gram positive vs Gram negative
Gram-Positive Cocci
• GPC in group• GPC in chain• GPC w/c capsule
• GPC in pair• lancet shape• kidney shape• sarcinae
Gram-Positive Cocci
• GPC in group• GPC in chain• GPC w/c capsule
• GPC in pair• lancet shape• kidney shape• sarcinae
Gram Negative Cocci
Cocco-bacilli, Kidney-shape
G (+) vs G (-)
Bacilli
• Gram positive bacilli– spore– spindle– box car– palisade– beaded– branching
• Gram negative bacilli– pleomorphic– cocco-bacilli– capsule– curved
Bacilli
• Gram positive bacilli– spore– spindle– box car– palisade– beaded– branching
• Gram negative bacilli– pleomorphic– cocco-bacilli– capsule– curved
Presumptive Diagnosis
• Staphylococcus aureus
• Streptococcus pneumoniae
• Hemophilus influenzae
• Klebsiella pneumoniae
• Nocardia spp
• Candida spp
Associate environment
• Neutrophils, epithelial cells
• Aid for judgement of G(+), G(-)
• Intracellular vs extracellular
• Mixed flora
• w/o PMN, w/o bacteria
• Non-bacteria organisms
Associate environment
• Neutrophils, epithelial cells
• Aid for judgement of G(+), G(-)
• Intracellular vs extracellular
• Mixed flora
• w/o PMN, w/o bacteria
• Non-bacteria organisms
Associate environment
• Neutrophils, epithelial cells
• Aid for judgement of G(+), G(-)
• Intracellular vs extracellular
• Mixed flora
• w/o PMN, w/o bacteria
• Non-bacteria organisms
Site specific
• Gram negative cocci– urine:– CSF:– sputum:
• Gram positive bacilli– CSF:– sputum:
• Gram positive cocci– urine:
Site specific
• Gram negative diplococci
– urine: N. gonorrheae– CSF: N. meningitidis– sputum: M. catarrhalis
Site specific
• Gram positive bacilli– CSF: L. monocytogenes– sputum: Corynebaterium straitum
• Gram positive cocci– urine: Enterococcus , Viridans strep.
Immunity - oriented
• Community-acquired
– GNB in CSF– cocco-bacilli in
sputum
• T-cell immunity– GNB in joint
• Nosocomial infection
– GNB in CSF– cocco-bacilli in
sputum
• T-cell immunity– GPB in CSF
Immunity - oriented
• Community-acquired
– GNB in CSF
• K. pneumoniae
– cocco-bacilli sputum
• H. influenzae
• Nosocomial infection
– GNB in CSF• P. aeruginosa• Enterobactericeae
– cocco-bacilli in sputum• A. baumannii
Immunity - oriented
• T-cell immunity– GNB in joint
– Salmonella spp.
• T-cell immunity– GPB in CSF
– L. monocytogenes
Acid Fast Stain
• Ziehl-Neelsen stain
• Kinyoun modification
– Mycobacteria
– Nocardia, partial stain
– NOT for actinomycosis