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2.2 Microbiology > Tests/Agar > Flashcards

Tests/Agar Flashcards

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1
Q

Staphylococcus

A

Blood agar
Mannitol salt
Chromogenic media
- SAIDE ID
- Chromagar MRSA
- BioMerieux
- Oxoid chromogenic
- ChromID MRSA

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2
Q

Mannitol salt

A

7.5% salt = selective
Mannitol + phenol red = differential

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3
Q

Oxoid CHromogenic MRSA Agar

A

Chromogens detect phosphatase activity in S. aureus -> this will produce denim-blue colonies

Cefoxitin (methicillin) will inhibit MSSA colonies

This allows for the accurate detection of MRSA

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4
Q

BioMérieux Chrome ID

A

SAID -> S. aureus ID

Chromogens detect alpha glucuronidase in S. aureus

This forms green colonies i.e. S, aureus = green colonies

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5
Q

ChromID MRSA agar

A

MRSA stains indicated by green colonies

Results from alpha-glucuronidase producing colonies in the presence of cefoxitin

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6
Q

Staphylococcus confirmatory tests

A

Coagulase test
DNase test
Vitek
MALDI

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7
Q

Slide coagulase test

A

Detects bound coagulase only

Mix bacterial saline suspension with rabbit plasma (source of fibrinogen), observe agglutination

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8
Q

Tube coagulase test

A

Detects free and bound coagulase

Mix 9 drops of 3hr incubated test suspension with 1 drop rabbit plasma -> incubate over night -> observe clot

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9
Q

Staphaeurex Plus

A

Rapid latex agglutination test for the identification of S. aureus

In addition to coagulase, protein A is also found on the surface of 95% of human strains of S. aureus

Certain MRSA possess a capsule that masks both protein A and the coagulase

With Staphaurex, rapid agglutination will occur using any three of these components:
- Reaction between fibrinogen and coagulase
- Binding between Protein A and the Fc portion of IgG
- Binding between MRSA capsular polysaccharide and specific IgG

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10
Q

DN’ase test

A

Deoxyribonuclease Agar

Nutrient agar that has 0.2% DNA added

Spot inoculate DNase agar and incubate overnight

Flood with 1 molar HCL

HCL precipitates DNA which turns the medium cloudy

DNase-positive cultures -> DNA is unavailable for precipitation so there will be a clear zone around the inoculum e.g. S. aureus

DNase negative will have no zone of clearing e.g. CNS

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11
Q

Novobiocin test

A

Nutrient agar with novobiocin discs

Lawn inoculum

S. aureus/CNS are novobiocin susceptible

S. saprophyticus are novobiocin resistant

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12
Q

Methicilin suscpetibility for MRSA

A

Cefoxitin disk

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13
Q

N. meningitidis tests

A

Direct specimen microscopy
Latex agglutination (gpB, C, Y and W)
Blood agar
Chocolate agar
Chocolate broth
TAXO sugars

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14
Q

N. gonorrhoea tests

A

Thayer Martin medium or GC/NYC medium
TAXO sugars

Gold standard = commercial PCR deterction on day 1

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15
Q

TAXO sugars

A

Carbohydrate utilisation tests - discriminatory test
Neisseria use carbohydrates oxidatively - not fermentation tests
Test ability to oxidise glucose, maltose, lactose and sucrose
Media e.g. Choc or GC agar minus antibiotics
Lawn inoculum and apply Taxo sugar discs
Incubate overnight in 5-10% CO2
Add phenol red indicator - yellow = positive oxidation reaction

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16
Q

M. catarrhalis

A

Blood or chocolate (but not fastidious will grow on nutrient agar)
TAXO sugars (no fermentation)
DN’ase
Growth (nutrient agar)
Tributyrin hydrolisis

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17
Q

Catarrhalis disc test

A

Organism produces butyrate esterase which hydrolyses tributyrin to release indoxyl which reacts with oxygen to turn blue-green

Rub test colony on disk and observe for a blue/green colour within 2 minutes

M. catarrhalis is positive for tributyrin hydrolysis

Neisser is negative

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18
Q

Haemophilus tests:

A

Latex agglutination for Hib
X and V factors (chocolate agar for isolation)

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19
Q

Pasteurella Multocida
(3)

A

Fermentation of glucose and sucrose
Indole production
Ornithine decarboxylase breakdown

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20
Q

B. pertussis

A

Bordet-Gengou agar
Citrate utilisation
Antigen detection (direct fluorescent antibody test)

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21
Q

Bordet-Gengou agar

A

Sheep blood in potato glycerol agar
+/- antibiotics

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22
Q

Indole test

A

Peptone media is a source of the amino acid trytophan

Hydrolysis of trytophan, leads to accumulation of indole

Add Kovacs reagent

Cherry red colour on surface is positive

Red colour seen in pasteurella

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23
Q

Ornithine decarboxylase

A

Decarboxylase broth contains nutrients, dextrose, pH indicator (purple) plus a single amino acid e.g. ornithine, lysine

Carboxylase breaks down ornithine - alkaline by-products-pH drop-indicator- purple colour change

Control tube - only dextrose ferments dextrose - pH decrease = yellow colour

P.multicoda positive for ornithine decarboxylase

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24
Q

Group B strep (S. agalactiae)

A

Lancefield test (group B)
CAMP positive
Hydrolysis of hippurate
Does not hydrolyse aesculin
May grow on MacConkey

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25
Q

Citrate

A

Citrate slope (green) - sole carbon source
Utilise citrate -> pH increased - blue colour

B. pertussis = citrate positive

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26
Q

Hippurate hydrolysis test
(4)

A

Used to differentiate S. agalactiae from other Beta Haemolytic streps

Hippurate is hydrolysed by hippuricase into Benzoic acid and glycine

Ninhydrin reagent is used as an indicator to detect Glycine

Ninhydrin reacts with glycine to form a deep blue or purple colour

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27
Q

CAMP test
(4)

A

Detects the production of extracellular protein CAMP factor

CAMP factor acts synergistically with the beta lysin produced by S. aureus

A zone of enhanced lysis of sheep or bovine blood

CAMP positive = Group B strep
CAMP negative = Not GBS

28
Q

S. pyogenes

A

Lancefield group A
B-haemolysis
MacConkey -> no growth
Bacitracin -> susceptible
PYR positive

29
Q

What is PYR

A

Pyrrolidonyl arylamidase

30
Q

S. pneumoniae

A

Optochin susceptibility -> susceptible
Bile susceptibility -> susceptible
Vitek
Lancefield group A

31
Q

Enterococcus

A

Lancefield grouping - D
MacConkey - growth
Bile Aesculin - growth
Molecular detection of Van A or Van B genes (VREs)
Chromogenic VRE medium

32
Q

Enterococcus on VRE Medium

A

Bluish green = E. faecalis
Violet = E. faecium

33
Q

Shigella tests:
(6)

A

Direct detection on faeces using real time PCR (Shigella Ipa gene)
XLD agar
DCA agar
Chrome GIT agar
Biochemical tests
UHG reference lab - serotyping

34
Q

DCA agar

A

Shigella and Salmonella agar

Selective factors include sodium deoxycholate and sodium citrate

Differential factors are lactose and sodium ferric chloride

Lactose fermenters = commensals = pink
Shigella = NLF = pale
Salmonella = NLF = pale with H2S positive black centre

35
Q

XLD agar

A

Shigella and Salmonella agar

Selective factor is sodium deoxycholate
Differential factors are Xylose, lysine, H2S

Lactose fermenters = commensals = yellow
Shigella = xylose lysine = red
Salmonella = red with black centre

36
Q

Chrom GIT agar (shigella and salmonella)

A

Teal coloured colonies
Non-H2S producing salmonellas will look the same

37
Q

Biochemical tests for Shigella

A

Indole negative
Methyl red positive
Vogues negative
Citrate negative
urease negative
Lysine decarboxylase negative

Only MR positive

38
Q

Salmonella tests:

A

Direct detection of Inv gene (RTPCR)
Selective enrichment on selenite broth
XLD -> red with black centre
DCA -> pale with black centre
Chromagar GIT -> teal coloured
Bismuth Sulfide agar -> black colonies
Biochemical tests

39
Q

Selenite broth

A

Salmonella enrichment broth

Selenite broth
Enhances culture yield by reducing growth of faecal coliforms and faecal streptococci.
Take sub-cultures of broth from the upper third of the broth column should be at least 5 cm in depth.
Better for enrichment of Salmonella Typhi
37 not exceeding 18 hours

40
Q

Bismuth Sulfite agar

A

Bismuth sulfite agar (BS) is a selective as well as a differential medium for isolation and presumptive identification of Salmonella spp, especially Salmonella Typhi

Uses brilliant green indicator so colonies are black (H2S) or green

41
Q

Why does Salmonella turn black on chromagar

A

H2S production

42
Q

Salmonella biochemical results

A

I : negative
M: positive
V: negative
iC: positive
urease negative
lysine decarboxylase positive

Neg, pos, neg, pos …

43
Q

Lysine decarboxylase test

A

Glucose
Lysine
Bromocresol Purple

44
Q

Yersinia:

A

Real time PCR detection
CIN agar
Biochemical tests

45
Q

CIN agar

A

Cefsulodin
Irgasan
Novobiocin agar

For Yersinia

46
Q

CIN agar

A

Cefsulodin
Irgasan
Novobiocin agar

For Yersinia

47
Q

Yersinia biochemical tests

A

Indole positive
Methyl red negative
Vogues positive
Citrate negative
Urease positive
Lysine negative
Ornithine decarboxylase positive

pos, neg, pos, neg, pos, neg …

48
Q

Proteus:

A

Swarming on agar
No swarm on MacConkey (NLF)
Brown on chrom UTI agar
Biochemical tests

49
Q

Proteus biochem tests

A

Indole negative
MR positive
VP negative
Citrate positive
Urease positive
Lysine decarboxylase negative
Dulcitol negative
Lactose negative
Phenylalanine postiive

50
Q

Klebsiella:

A

Mucoid colonies
Biochemical tests

51
Q

Klebsiella biochem tests

A

Indole negative
MR negative
VP positive
Citrate postiive
Urease V positive (K. pneumo)
Lysine decarboxylase positive
Dulcitol positive

52
Q

Enterobacter tests

A

MacConkey: LF
Biochemical tests

53
Q

Biochemical results for enterobacter

A

Indole negative
MR negative
VP positive
Citrate postiive
Urease negative
Lysine decarboxylase negative
Dulcitol negative

54
Q

Biochemical results for citrobacter

A

I : produces indole -
 M: methyl red positive +
 V: Voges-Proskauer negative -
 iC: cannot use citrate as sole carbon source +
 Urease -
 Lysine Decarboxylase -
 Dulcitol –
 Lactose + LF

55
Q

EHEC/STEC

A

Direct detection of shiga toxin genes by RTPCR - EntericBio RTPCR

MacConkey for all E. Coli

STEC
- STEC Chromagar
- CTSMAC agar

56
Q

CTSMAC agar

A

For shiga toxin producing E. COLI -> EHEC
Cefixime tellurite sorbitol Mac Conkey

O157:H7 STEC = non sorbiol fermenter = colourless

Non o157:H7 = sorbitol fermenter = pink

Detects most common Shiga-toxin E. Coli serotypes -> mauve colonies

Other enterobacterales are coulerless, blue or inhibited

57
Q

Biochemical results for E. coli

A

I: produces indole
M: methyl red positive
V: negative
iC: negative

58
Q

Indole test

A

A
 decompose the amino acid tryptophan to indole
 accumulates in the medium
 Inoculate a peptone water
 Incubate 24 hrs @ 37
 When indole is combined with Kovac’s Reagent
 yellow to cherry red
 an oily layer at the top of the broth

59
Q

Methyl red test

A

 Methyl Red Test
 detection of end products from the
metabolism of glucose.
 Inoculate a buffered glucose
 the fermentation of glucose.
 MR Positive: When the culture medium turns red
after addition of methyl red
 pH at or below 4.4

60
Q

Voges-Proskauer test

A

 Voges-Proskauer Test
 detection of end products from
the metabolism of glucose.
 produce acetoin as the chief end
product of glucose metabolism
 Heavy inoculum
 Add 40% KoH and alpha Napthol
 Shake vigorously
 Colour development within 15 mins
 Voges-Proskauer negative

61
Q

Simmons Citrate

A

 Simmons Citrate Test
 use citrate as sole carbon source

Growth usually results in the bromothymol blue
indicator, turning from green to blue.

The alkaline carbonates and bicarbonates produced as by-products of citrate catabolism raise the pH of the medium to above 7.6

62
Q

E. Coli biochem tests

A

 produces indole
 methyl red positive
 Voges-Proskauer negative
 cannot use citrate as sole carbon source
 Lactose Fermenter
 produces lysine decarboxylase
 ferments sorbitol except E. coli O157 H7
 produce β-glucuronidase except E. coli O157 H7

63
Q

Chromagar GIT

A

E. coli - dark pink to red
Enterococcus - turquoise blue
Klebsiella, Enterobacter, Citrobacter - metallic blue
Proteus - brown hali

Pseudomonas - cream/translucent
S. aureus -> golden/opaque
S. saprophyticus -> pink small

64
Q

Pseudomonas

A

Blood agar (metallic sheen and sweet odour)
Chocolate agar
Moderately selective media - MacConkey media (NLF)
Selective agar - Cetramide agar

Growth at 42 and 4 degrees

65
Q

Cetramide agar

A

Pseudomonas
P, aeruginosa = enhanced pigment = bright green colour

It enhances the production of Pseudomonas pigments such as pyocyanin and pyoverdine which show a characteristic blue-green and yellow-green colour

2.2 Microbiology (60 decks)

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