Microbiology Laboratory Unknown Identification

A set of flashcards covering the key laboratory tests, media, and biological concepts used in the identification of unknown bacterial species as described in the lecture notes.

Bacterial Metabolism in Identification

The unique metabolic requirements regarding what and how an organism eats and breathes, which are used to distinguish species that appear morphologically identical.

Exoenzymes

Enzymes secreted by bacteria to break down food externally into absorbable nutrients, a process that alters the external environment of the organism.

Chemolithotrophs

Bacteria and Archaea that use alternative final electron acceptors such as Nitrates ($NO_{3}$) as an alternate oxygen source or Sulfur ($S^{-2}$) for respiration.

Fermentation

A metabolic process used by organisms lacking oxygen alternatives to harvest energy from glycolysis, providing significantly less energy than respiration and producing an acidic solution.

EMB Agar (Eosin-Methylene Blue)

A selective and differential medium used to identify Gram-negative organisms and differentiate coliform bacteria based on lactose fermentation.

Eosin

A pink acidic dye in EMB agar that acts as a color indicator and turns black under acidic conditions.

Methylene Blue

A blue basic dye in EMB agar that is cytotoxic to Gram-positive organisms and helps select for Gram-negative bacteria.

Strong Lactose Fermenters (EMB Agar)

Organisms that precipitate the EMB salt, causing the bacterial colonies to turn green due to the pH change.

Coliform Bacteria

Non-spore forming, Gram-negative, bacillus organisms that survive in both soil and fecal matter.

MacConkey Agar

A selective and differential agar containing Crystal Violet, Bile Salts, Neutral Red, and Sucrose, designed to inhibit Gram-positive growth and identify lactose fermenters.

PEA (Phenylethyl alcohol) Agar

A selective medium that inhibits the DNA synthesis of most Gram-negative organisms, allowing for the growth of Gram-positive organisms.

Thioglycolate Broth

A medium used to determine aerotolerance by employing sodium thioglycolate and thioglycolic acid to reduce oxygen to water, creating layered oxygen concentrations.

Obligate Aerobe

An organism that strictly utilizes $O_{2}$ to create energy and grows only above the oxygenation line in thioglycolate broth.

Obligate Anaerobe

An organism for which $O_{2}$ is toxic and that uses alternatives to oxygen for energy, growing only below the oxygenation line.

Facultative Anaerobe

An organism that utilizes $O_{2}$ if present but can perform fermentation or use alternative acceptors if $O_{2}$ is absent, characterized by growth throughout a thioglycolate tube.

Aerotolerant Anaerobes

Organisms that do not use $O_{2}$ for energy but can tolerate its presence, growing equally well with or without oxygen.

Microaerophile

An organism that uses $O_{2}$ for energy but requires specific low concentrations ($\text{about } 0.2\text{ atm.}$), growing only at the oxygenation line.

Phenol Red Broth

A differential test used to detect the fermentation of a single type of sugar (Glucose, Lactose, or Sucrose) and the production of gas.

Durham Tube

A small upside-down test tube used within broth to detect the production of gas as a byproduct of fermentation.

MSA (Mannitol Salt Agar)

A medium selective for the genus Staphylococcus due to a $7.5\%$ NaCl concentration and differential for Staphylococcus aureus based on mannitol fermentation.

Nitrate A and Nitrate B

Reagents (sulfanilic acid and Dimethyl-$\alpha$-napthylamine) used to detect the presence of nitrous acid, a byproduct of partial nitrate breakdown.

Zinc Powder (Nitrate Test)

A reagent added to a clear nitrate test to determine if nitrates are still present; a color change to red after its addition indicates a negative result.

Indole Test

A differential test identifying an organism's ability to digest the amino acid tryptophan into indole, detected by a red ring after adding Kovac’s reagent.

SIM Media

A semi-solid agar used to test for three characteristics: Sulfide production, Indole production, and Motility.

Cytochrome C Oxidase

The final enzyme in the electron transport chain before ATP synthase, found only in aerobes.

Oxidase Test Reagent

N,N,N,N-Tetramethyl-p-phenylenediamine, which turns blue within $30$ seconds in the presence of Cytochrome C oxidase.

Phenylalanine Deaminase

An enzyme that removes ammonia ($NH_{3}$) from the amino acid phenylalanine, producing phenyl-pyruvic acid.

Ferric Chloride ($FeCl_{3}$)

The reagent used in the Phenylalanine Deaminase test that reacts with phenyl-pyruvic acid to produce a green color.

Permease

The enzyme required by an organism to intake exogenous citrate for use as a sole carbon source.

Simmons Citrate Indicator

Bromothymol blue, which is green at $pH < 7.6$ and turns blue at $pH > 7.6$ when citrate is broken down into alkaline ammonia.

Urease

An enzyme that catalyzes the breakdown of Urea into Ammonia ($NH_{3}$), raising the pH and turning phenol red indicator to magenta.

MRVP (Methyl Red/Voges Proskauer)

Two tests used to differentiate between pure lactic acid fermentation (Methyl Red positive) and the butanediol pathway (Voges Proskauer positive).

TSI (Triple Sugar Iron) Agar

An agar containing Glucose, Lactose, and Sucrose, as well as phenol red and sources of sulfur and iron, used to differentiate enteric bacteria.

Catalase

An enzyme produced by some bacteria to break down the disinfectant hydrogen peroxide ($H_{2}O_{2}$) into water and oxygen, visible as bubbles.

Lipase

An enzyme detected by Spirit Blue agar that breaks down Triglycerides into three fatty acids and glycerol.

Litmus Milk Curd

A solid mass formed from the denaturing of milk proteins (proteolysis) during bacterial growth in milk solution.

Staphylocoagulase

The specific type of coagulase produced by Staphylococcus aureus that converts fibrinogen to insoluble fibrin to form a clot.