Procedures and Biochemical Identification of Bacteria
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What does Catalase test for?
Catalase, an enzyme that produces water and oxygen from hydrogen peroxide
What organisms does the Catalase test identify?
- POS - Staphlococcus
- NEG - Streptococcus
What organism does the Coagulase test identify?
- POS - Staph aureus (also S. intermedius and S. hyicus)
- More specific agglutination tests are now performed
What does the PYR test for?
Detects the enzyme L-pyrrolidonyl arylamidase
What organism does PYR differentiate?
- NEG - Staph aureus
- POS - S. lugdensis, S. schleiferi, Strep pyogenes (Group A), and Enterococcus species
What organism does the Bile solubility test identify?
- Strep pneumoniae
- In the presence of bile at 37 degrees celcius, colonies will autolyse within 30 min and disappear from the agar surface
What does the Hippurate Hydrolysis test for?
Detects the bacterial enzyme hipuricase - which hydrolyzes hippurate to glycine and benzoic acid
What organism does the Hippurate hydrolysis test differentiate?
- POS - Group B Strep, Campylobacter jejuni
- NEG - Most other beta-hemolytic Streps, Campylobacter species
What does the Oxidase test for?
Detects chytochrome oxidase that is used in the electron transport system
What organism does the Oxidase test differentiate?
POS - Pseudomonas species, Neisseria gonorrheae
What does the Indole test for?
Detects the bacterial enzyme tryptophanase (tryptophan is broken down by tryptophanase into pyruvic acid, ammonia, and indole)
What does the Urease test for?
Urease breaks down urea to form ammonia (NH3)
What does the Triple sugar iron agar (TSI) test for?
TSI will show the pattern of glucose, lactose, and sucrose fermentation in addition to H2S and gas production
What organisms does the TSI differentiate?
- Alkaline slant/alkaline deep (K/K) - Nonfermentor, NOT Enterobacteriaceae
- Alkaline slant/acid deep (K/A) - Nonlactose and nonsucrose fermenter, glucose fermenter
- Acid slant/acid deep (A/A) - Lactose and/or sucrose fermenter, and glucose fermenter
- Black deep, production of H2S gas - production of hydrogen sulfide (H2S) from sulfer-containing molecules in the medium
- Lead acetate - added to filter paper strips and one end is held in place by the cap. If H2S gas is produced, a black color will form
What does the TSI result of K/K mean?
- NOT Enterobacteriaceae
What does the TSI result of K/A mean?
- Nonlactose and/or Nonsucrose fermenter
- Glucose fermenter
What does the TSI result of A/A mean?
- Lactose and/or sucrose fermenter
- Glucose fermenter
What does Methyl Red (MR) test for?
- pH indicator
- Yellow at acid pH (indicating glucose fermentation)
- Red is NEG
What does Voges-Proskauer (VP) test for?
- Detects the metabolism of glucose to acetyl-methyl-carbinol (acetoin)
- Bacteria are usually MR or VP POS
What does Citrate test for?
Determines if citrate is used as a sole carbon source
What does ONPG (o-nitrophenyl-beta-D-galactopyranoside) test for?
- Detects the presence of beta-galactosidase, an enzyme that cleaves ONPG and lactose
- Test is useful in detecting delayed (late) lactose fermenters that lack or are deficient in beta-galactoside permease
What does the Amino acid degradation test for?
Detects bacterial enzymes that break down various amino acids
What does the Deaminase reaction test for?
Detects the ability of an organism to remove the amino group from specific amino acids
What does the Decarboxylation reaction test for?
Detects the ability of bacteria to remove the carboxyl group from a specific amino acid
What are the reactions in an Amino acid degradation test?
- Deaminase reaction
- Decarboxylation reaction
What are some examples of bacterial enzymes that will turn positive for the Amino acid degradation test?
- Tryptophan (tryptophan deaminase)
- Lysine (lysine decarboxylase)
- Ornithine (ornithine dihydrolase)
What does the carbohydrate fermentation test for?
Detects the ability of bacteria to produce organic acids by the fermentation of various carbs
What does the Nitrate reduction test for?
Determines the ability of an organism to reduce nitrate (NO3) to nitrite (NO2) and nitrogen gas (N2)
What does the Sherlock Microbial Identification System identify?
The fatty acid composition of the bacterial cell wall as determined by gas chromatography
How are the fatty acids in mycobacteria identified?
- The fatty acids in mycobacteria have a larger molecular weight
- Identified via high-performance liquid chromatography in the Sherlock Mycobacteria Identification System
Name the Gram Stain Reagents, in order
- Gram Crystal Violet
- Gram Iodine
- Acetone-Alcohol Decolorizer
- Safranin Counterstain
What are key characteristics of Micrococcus?
- Catalase POS
- Glucose fermentation NEG
- Bacitracin SENSITIVE
- Modified Oxidase POS
Describe the differentiation techniques for Staphlococcus species
Describe the differentiation techniques for Streptococcus species
Name the H2S producers
Name the positive urease organisms
- K. pneumo
Name the ONPG positive organism
Name the IMViC patterns for Enterics
- Clostridium perfringens
- (Reverse CAMP test)
- Positive for Group B beta Strep
- (Positive CAMP test)
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