Micro210 Biochemical Test

What 3 sugars are present on TSI?

Glucose, Sucrose, Lactose

What does 6.5% NaCl test for? What does it differentiate?Appearance

• If organism can grow in high salt concentration

• Differentiates Enterococcus sp. from non-Enterococcus sp. and other NH streptococci

• Cloudy = Growth

Purpose of Aerotolerance testing? What media is used and hot to interpret?

To determine is suspect colony is aerobe/capnophile, an obligate anaerobe or facultative

Purpose of KVC disks?

To help in identification of anaerobe Gram-negative bacteria

Expected Results in Bacticard

Purpose and Principle of Bile Esculin

• if organism can grow in bile salts and hydrolize esculin to esculetin which reacts with ferric chloride to produce black brown precipitate

• Differentiate Enterococcus sp. from other organism

• Slant agar

Principle and Purpose of Bile solubility

• Streptococcus pneumoniae has a natural autolytic process (lyse in presence of O2), bile salts speed up autolytic process.

• Differentiates Streptococcus pneumoniae from other AH Strep

Principle and Purpose of BA with S. aureus streak?

• Staphylococcus aureus lyse RBC to release X+V factors and releases its own V factor

• If organism grows around BH zone (satellism) then organism depends on X+V factors to grow (Haemophillus sp.)

Principle and Purpose of Butyrate Esteras

• If Butyrate esterase is present it will release the indoxyl butyrate to produce blue-green colour

• Aids in identification of Moraxella catarrhalis (only perform on oxidase pos, GNC)

  

Principle and Purpose of CAMP test

• Group B Strep produces CAMP factor and when combined with BH reaction of S. aureus it creates an arrow head zone of hemolysis

• Confirmatory of Streptococcus agalactiae

• 

Principle and Purpose of Catalase (What enzyme can cause false positive)

• If bacteria posses catalase it converts hydrogen peroxide into water (bubbles)

• Differentiates Micrococcus and Staphylococcus from Streptococcus

Principle and Purpose of Cefinase (Nitrocefin)

• If beta-lactamase is present and can break beta lactam ring, a red colour will be visible indicating resistance to beta lactam drugs (penicillins or cephalosporin)

• Determines presence of beta-lactamase in Neisseria gonorrhoeae and Haemophilus influenzae

Principle and Purpose of Citrate Utilization

• If organism can use sodium citrate (carbon) and ammonium salt (nitrogen) for energy. If so breakdown of ammonium salt creates alkaline pH changing bromthymol blule indicator to green-blue

• Differentiates members of Enterobacterales

• 

Principle and Purpose of Coagulase Test (What can cause false negative?)

• The S. aureus enzyme coagulase exist free and bound form(clumping factor).

Principle and Purpose of Decarboxylase Test (Lysine or Ornithine)

• If organism posses decarboxylase enzyme in anaerobic-acidic environment (fermentation of glucose) it removes a carboxyl group from an amino acid creating alkaline end product.

• ph Indicator is bromcresol purple, which turn yellow in acid (fermentation of glucose), and back to purple in alkaline (if decarboxylase is present)

• A control is present without amino acid

• Differentiate Enterobacterales

Principle and Purpose of DNase

• If organism has deoxyribonuclease (DNase) it hydrolizes DNA which unbind to methyl green resulting in clear halo

• Positive for Moraxella catarrhalis, Serratia marcescens, Staphylococcus aureus

  

Principle and Purpose of Germ Tube

• Candida albicans incubated in rabbit plasma or fetal calf serum it will produce germ tubes

• Differentiates Candida albicans from other Candida

Principle and Purpose of Gram Stain

Gram Positive have thick peptidoglycan and teichoic/lipoteichoic acids

Gram Negative have thinner layer

Principle and Purpose of Indole Production

• If bacteria posses tryptophanase they hyrolyze tryptophan to indole. If Kovacs reagent is added observe for pink color change

• Differentiates Enterobacterales

Principle and Purpose of Spot Indole

If bacteria can grow in medium with trytophan (BA) then they posses tryptophanase. Indole is detected in pre-soaked filter paper if blue-green color is produced

Kirby-Bauer Susceptibility

Media?0.5 McFarland in CFU? CLSI modifacation?

• MH

• 1.5×10^8 CFU

• MH with 5% sheeps blood

• Incubated in CO2

Principle and Purpose of Motility

• If bacteria has flagella

• differentiates Enterobacterales

Expected results of MIL

Principle and Purpose of Nagler Reaction (Lectinase)

• Clostridium perfringens produces lectinase which breaks down lecithin (in egg yolk)

• Egg yolk agar has one side streaked with antitoxin then suspect organism is streaked across

• If precipitate on antitoxin free side then organism produced lectinase

• If nothing on antitoxin side then antitoxin neutralized the lictinase

• This aids in identification of Clostridium perfringes

Principle and Purpose of Nitrate Reduction Test (What alternative tube can be used to detect gas production.

• All Enterobacterales can reduce nitrates to nitrites but some can further reduce nitrite to nitrogenous compounds, some do not utilize nitrates

• If organism has nitrate reductase it can reduce nitrate to nitrite

• Durham tube (traps gas)

Principle and Purpose of Novobiocin

• Staphylococcus saprophyticus is resistant to Novobiocin <16mm

Principle and Purpose of ONPG

• LF have B-galactosidase + permease, LLF only have B-galactosidase

• If they have B-galactosidase ONPG will be hydrolyzed and produce yellow colour

• This test differentiate LF and LLF from NLF

  

Principle and Purpose of Optochin? What other screen can be performed at the same time?

• This differentiate Streptococcus pnuemoniae from other AH Strep because S. pneumoniae is susceptible to Optochin >=14

• An oxacillin screen can be performed to determine susceptibility of organism to penicilin >=20mm

Principle and Purpose of Oxidase Test

Organism with cytochrome oxidase will oxidize Kovacs reagent into indophenol to creat a dark purple color

Principle and Purpose of Oxidation/Fermentation (O/F) Medium

Differentiates organism based on their ability to oxidize, ferment, or not utilize specific carbohydrates

Principle and Purpose of PathoDX

Identifies BH Streptococci based on which reagents agglutinates with the antigens

Principle and Purpose of Porphyrin Test

• Haemophilus parainfluenzae produces porphobilinogen synthase making it capable of making its own X Factor

• Differentiates Haemophilus influenzae and Haemophilus parainfluenzae

• 

Principle and Purpose of PYR

• Detects enzyme PYRase which is present on Enterococcus and Streptococcus pyogenes

Rhamnose Broth

• Aids in identification of yersinia enterocolitica

• Yersinia enterocolitica is incapable of fermenting rhamnose while most Enterobacterales can

• 

Principle and Purpose of Reverse CAMP

• Aids in identification of Clostridium perfringens

• 

Principle and Purpose of Staphaurex

• Latex particle with fibrinogen and IgG can react with clumping factor and Protein A of S. aureus to cause clumping

Principle and Purpose of SPS

Principle and Purpose of TSI

K = red

Growth Result of Haemophilus sp. on X,V, XV, Disk Test