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2.2 Microbiology > Tests/Agar > Flashcards

Tests/Agar Flashcards

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
Citrate
Citrate slope (green) - sole carbon source Utilise citrate -> pH increased - blue colour B. pertussis = citrate positive
26
Hippurate hydrolysis test (4)
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
27
CAMP test (4)
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
S. pyogenes
Lancefield group A B-haemolysis MacConkey -> no growth Bacitracin -> susceptible PYR positive
29
What is PYR
Pyrrolidonyl arylamidase
30
S. pneumoniae
Optochin susceptibility -> susceptible Bile susceptibility -> susceptible Vitek Lancefield group A
31
Enterococcus
Lancefield grouping - D MacConkey - growth Bile Aesculin - growth Molecular detection of Van A or Van B genes (VREs) Chromogenic VRE medium
32
Enterococcus on VRE Medium
Bluish green = E. faecalis Violet = E. faecium
33
Shigella tests: (6)
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
DCA agar
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
XLD agar
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
Chrom GIT agar (shigella and salmonella)
Teal coloured colonies Non-H2S producing salmonellas will look the same
37
Biochemical tests for Shigella
Indole negative Methyl red positive Vogues negative Citrate negative urease negative Lysine decarboxylase negative Only MR positive
38
Salmonella tests:
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
Selenite broth
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
Bismuth Sulfite agar
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
Why does Salmonella turn black on chromagar
H2S production
42
Salmonella biochemical results
I : negative M: positive V: negative iC: positive urease negative lysine decarboxylase positive Neg, pos, neg, pos ...
43
Lysine decarboxylase test
Glucose Lysine Bromocresol Purple
44
Yersinia:
Real time PCR detection CIN agar Biochemical tests
45
CIN agar
Cefsulodin Irgasan Novobiocin agar For Yersinia
46
CIN agar
Cefsulodin Irgasan Novobiocin agar For Yersinia
47
Yersinia biochemical tests
Indole positive Methyl red negative Vogues positive Citrate negative Urease positive Lysine negative Ornithine decarboxylase positive pos, neg, pos, neg, pos, neg ...
48
Proteus:
Swarming on agar No swarm on MacConkey (NLF) Brown on chrom UTI agar Biochemical tests
49
Proteus biochem tests
Indole negative MR positive VP negative Citrate positive Urease positive Lysine decarboxylase negative Dulcitol negative Lactose negative Phenylalanine postiive
50
Klebsiella:
Mucoid colonies Biochemical tests
51
Klebsiella biochem tests
Indole negative MR negative VP positive Citrate postiive Urease V positive (K. pneumo) Lysine decarboxylase positive Dulcitol positive
52
Enterobacter tests
MacConkey: LF Biochemical tests
53
Biochemical results for enterobacter
Indole negative MR negative VP positive Citrate postiive Urease negative Lysine decarboxylase negative Dulcitol negative
54
Biochemical results for citrobacter
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
EHEC/STEC
Direct detection of shiga toxin genes by RTPCR - EntericBio RTPCR MacConkey for all E. Coli STEC - STEC Chromagar - CTSMAC agar
56
CTSMAC agar
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
Biochemical results for E. coli
I: produces indole M: methyl red positive V: negative iC: negative
58
Indole test
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
Methyl red test
 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
Voges-Proskauer test
 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
Simmons Citrate
 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
E. Coli biochem tests
 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
Chromagar GIT
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
Pseudomonas
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
Cetramide agar
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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