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HDM Final F19 > Bacterial Tests > Flashcards

Bacterial Tests Flashcards

1
Q

Gram Stain

A

Crystal violet (primary stain): cells purple
Gram’s iodine-mordant: fixes crystal violet
95% ethanol-decolorized: destains gram (-) bacteria, dehydrates and locks crystal violet w/ gram (+) bacteria
Safranin-counterstain: stains gram (-) bacteria pink/red, gram (+) remain purple

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2
Q

Acid Fast Stain (Ziehl-Neelsen Method)

A

Identify Mycobacterium sp.
Carbol fuschin (red)- primary stain, lipid soluble
Steam heating enhances stain penetration
HCl/Alcohol: destains non acid fast microorganisms
Methylene blue: counterstains non acid fast microorganisms
Heat fix thin film of microorganisms, flood slide w/ carbol fuschsin (steam heat for 5 min)
Rinse/flood slide w/ acid alcohol (30 sec)
Rinse/flood slide counterstain w/ methylene blue (30 sec)

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3
Q

Selective Media

A

favor growth of distinct group of microorganisms while inhibiting others
Agents used: dyes, antimicrobials and salts
Example: PEA agar, contains phenyl ethyl alcohol which inhibits gram (-) microorganisms

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4
Q

Differential Media

A

distinguishes microorganisms having defined component which allows for observable change when specific chemical reaction takes place
Example: Simmons citrate agar

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5
Q

Selective/Differential Media

A

contains key characteristics of both
Example: Mannitol salt agar
Selective: 7.5% salt inhibits all but Staph sp.
Differential: only S. aureus ferments mannitol, turns red to yellow indicating acid production

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6
Q

PEA Agar

A 
phenyl ethyl alcohol agar w/ blood (PEAB)
support gram (+) bacteria, inhibits gram (-)
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7
Q

Mannitol Salt Agar

A

contains 7.5% salt selective for staph sp.

S. aureus ferments mannitol producing acid which drops pH turning area around colonies yellow (differential)

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8
Q

Blood Agar

A

Hemolysis test: contains 5% sheep blood, tests bacteria’s ability to lyse RBCs, lysis (hemolysis) and catalyzed by enzymes (hemolysins)
Different species contain different classes of hemolysis (i.e. beta: complete reaction; yellow; alpha: green, partial reaction; gamma: red, no reaction)

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9
Q

Chocolate Agar (CHOC)

A

Non selective enriched growth medium
variant of blood agar, contains RBCs (lysed by slow heating to 56C)
Used for growing fastidious (fussy) respiratory bacteria, such as Haemophilus influenzas
Need growth factors (NAD and hematin) inside RBCs
Thus need to lyse RBCs before cells can grow

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10
Q

Tinsdale Medium

A

primary isolation medium for Corynebacterium diphtheria
Variations: chocolate tellurite, Loeffler’s medium
Thiosulfate based medium: K+ tellurite inhibitory to gram (+) and many gram (-) microorganisms
Form black colonies surrounded by dark gray halos

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11
Q

Tinsdale (tellurite) Medium

A

Corynebacterum sp. reduces metal tellurite to tellurium (black precipitate), result: gray to black colonies

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12
Q

Thayer-Martin Agar

A

Blood agar base
contaminating microorganisms inhibited by: colistin, nystatin, vancomycin, trimethoprim, lincomycin
Selective for Neisseria gonorrhoeae, meningitidis
N. gonnorrhoeae: small, mucoid, grayish white to colorless colonies
N. meningitidis: medium to large, mucoid, blue gray colonies

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13
Q

Hemoglobin, Dextrose

A

Promotes growth for Neisseria sp.
Trimethoprim added to inhibit proteus sp.
Colistin inhibits most gram (-) microorganisms including (Pseudomonas)
Vancomycin inhibits gram (+) microorganisms (some Neisseria sp. sensitive to vancomycin)
Lincomycin inhibits gram (+) microorganisms
Nystatin inhibits Candida albicans

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14
Q

MacConkey Agar Medium

A

“Mac”
Selects for gram (-) bacteria contains bile salts which inhibit gram (+) bacteria
Gram (-) ferment lactose (E. coli) changes neutral red indicator in medium to dark pink/red causing those colonies to appear red to dark pink (differential)

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15
Q

EMB Agar

A

Eosin methylene blue agar
Both inhibit gram (+) bacteria and favor gram (-) enterics
Contains lactose, allows differentiation between lactose fermenters and non fermenters
E. coli: lactose fermenter, results in metallic green colonies

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16
Q

Hektoen Enteric Agar

A

Isolates bacteria belonging to Enterobacteriaceae family
(particularly Salmonella and Shigella)
High concentration of bile salts inhibit gram (+) and some gram (-) except Shigella and Salmonella
Contains lactose: distinguishes coliform (ferments lactose) from non fermenters (Salmonella, Shigella)
Lactose fermenters produce acid results in a yellow/orange color in medium
Contains thiosulfate which Salmonella reduces H2S (black precipitate)

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17
Q

Xylose lysine Desoxycholate Agar

A

Primary isolation of Salmonella and Shigella
Sodium desoxylate inhibits gram (+) microorganisms
Xylose fermentation, H2S production and lysine carboxylation distinguishes Salmonella from Shigella (sulfide reacts w/ ferric ions (ferric ammonium citrate)) to produce ferric sulfide, a black precipitate

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18
Q

BG Agar

A

Bordet-Gengou Agar, Regan-Lowe Agar (defibrillated horse blood, charcoal)
Potato-glycerol based w/ 15-20% defibrillated blood
Methicillin to inhibit contaminants
Selective for Bordetella pertussis (-) for urease, nitrate reduction, motility and citrate utilization

19
Q

BCYE Agar

A

Buffered charcoal yeast extract agar
Yeast extract, charcoal and salts
Selective for legionella sp. (alpha ketoglutarate: key ingredient in recovery of legionella sp. from clinical and environmental specimens)

20
Q

TCBS Agar

A

Thiosulfate citrate bile salts agar

Selective and differential medium for vibrio sp. (causative agents for cholera, diarrhea, food poisoning)

21
Q

Cefsulodin-Irgason-Novobiocin (CIN) Agar

A

Selective for yersinia sp.
Inhibits gram (+) and (-) microorganisms
Appears as colonies w/ red pigmented “bulls eye” in middle of each colony
After 48hr. incubator at 25-35C

22
Q

Lowenstein-Jensen Agar

A

Egg based medium
Contaminants inhibited by malachite green
Isolation of mycobacterium sp.

23
Q

Catalase Test

A

Enzyme breaks down H2O2 into H2O and O2
(+)= staph sp.
(-)= strep sp.

24
Q

Coagulase Test

A

Converts fibrinogen to fibrin (clot) causing agglutination of microorganisms
(+)= Staphylococcus aureus
(-)= Other staph sp.

25
Q

Gelatin Liquefaction Test

A

Ability to produce hydrolytic exoenzymes (gelatinases) that digests and liquefies gelatin
Tubes inoculated and incubated, warm temps will liquefy gelatin
After incubation complete, tubes placed in fridge to allow gelatin in (-) control to solidify
(+)= P. aeruginosa, liquid after incubation
(-)= E. coli, still gelatin

26
Q

Oxidase Test

A

Detects cytochrome oxidase enzyme
Able to oxidize tetramethyl-p-phenylenediamine forming purple end product (indophenol)
(+)= indophenol, purple spot
(-)= blank strip

27
Q

Indole Test (E. coli)

A

Detects activity of tryptophanase enzyme on tryptophan
End product indole reacts w/ Kovacs reagent (yellow) turning it a red color
(+)= Red, E.coli
(-)= Yellow, no reaction

28
Q

Urease Test

A

Detects activity of urease enzyme which hydrolyzes urea releasing end product ammonia
pH change shown by change of medium color form pale orange to pink
Key characteristic of proteus sp.
(+)= pink, contains urease
(-)= pale orange, no reaction

29
Q

Citrate Utilization Test

A

Ability to use citrate as sole carbon source
Production of CO2 leads to reaction w/ water and Na+ to produce sodium carbonate (alkaline)
pH rises turning bromythol blue from green to bright blue
(+)= Blue color change (Enterobacter cloacae)
(-)=no change (E. coli)

30
Q

Bile Esculin Test

A

Group D strep can hydrolyze esculin (now called Enterococcus)
Byproducts of hydrolysis react w/ iron salts in medium changing color from orange to black
(+)= Black, Group D strep
(-)= Orange

31
Q

DNAse Test

A

DNAse catalyzes depolymerization of DNA
Medium contains methylene green dye: binds polymerized DNA
Exoenzyme breaks down DNA into smaller fragments releasing it from methylene green dye (clear zone)
(+)= yellow (clear zone around red streak)
(-)= no reaction black streak

32
Q

Optochin Test-P disc Test

A

Presumptive test for Streptococcus pneumoniae
Ethyl-hydrocupreine, S. pneumoniae optochin sensitive
(+)= clear zone around disc
(-)= growth around disc

33
Q

Bacitracin Susceptibility Test

A

Differentiates group A streptococcus from other groups of beta-hemolytic streptococcus
Example: S. pyogenes (+) test result, clear zone around bacitracin
S. agalactiae (-), no reaction

34
Q

O2 requirements (growth patterns)

A

Strict aerobe: growth at top of test tube
Strict anaerobe: growth at bottom of test tube
Facultative anaerobe: growth throughout tube
(-) control: no growth

35
Q

Motility Test

A 
Soft agar deeps (allows limited movement of microorganisms)
Stabbed w/ inoculating needles 
Agar (yellow) 
(+)= agar completely red 
(-)= one red puncture
36
Q

Direct Microscopic Examinations

A

For hair, skin and nails
KOH prep: 10-20% solution to dissolve tissue elements, keratin
Skin scrapping: scalpel, edge of glass slide placed on microscope slide, add KOH and sit for 5-15 min.
Stain w/ lactophenol cotton blue or calcofluor-white stain

37
Q

Lactophenol Cotton Blue Wet Mount

A

Vulvovaginal candidiasis
pH 4-4.5 (normal, yeast buds or spores or hyphae)
KOH prep: facilitates recognition of budding yeasts or hyphae (sensitivity 70%)
(-) up to 50% of culture proven candidal infections

38
Q

KOH w/ Calcofluor White

A

Fluorescent dye: added to KOH prep prior to cover slipping
Binds to polysaccharides present in chitin or cellulose
Fungal elements fluoresce apple green or blue white (depends on filter used)
Any element w/ a polysaccharide skeleton will fluoresce

39
Q

India Ink

A

(-) stain, used to examine CSF specimens for presence of encapsulated yeast (Cryptococcus neoformans)
Drop of India ink mixed w/ drop of centrifuged CSF specimen
Budding yeast surrounded by a large, clear zone against a black back ground
WBCs and other artifacts resemble encapsulated C. neoformans, require careful examination
Latex agglutination test used to detect cryptococcal antigens

40
Q

Periodic Acid-Schiff Stain

A

Fungal studies tissue stains-detect fungal elements in tissue specimens
Attaches to polysaccharides in fungal wall and stains pink, periodic acid oxidizes hydroxyl groups in mucopolysaccharides to aldehyde groups
Aldehyde groups react w/ Schiff reagent to form a red-purple product

41
Q

Grocott-Gomori Methamine-Silver Nitrate Stain

A

Fungal studies tissue stains-detect fungal elements in tissue specimens
Stains fungal elements black
Stains: Candida, Histoplasma, Blastomyces
Chromic acid used to oxidize hydroxyl groups (in mucopolysaccharides) to aldehydes
Silver ions in methamine silver metal which is black

42
Q

Giemsa Stain

A

Detects Histoplasma capsulate in blood bone marrow

Stain yeast a purple-blue surrounded clear halo

43
Q

Masson-Fontana Stain

A

Stains melanin in cell wall a brown color

HDM Final F19 (3 decks)

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