biology

Phenotypic testing of mycobacteria

Phenotypic testing of mycobacteria is a biology topic covered in the lgStudy science library. This page brings together a partial reference excerpt, illustrations, worked examples, real-world applications and a short study plan, so you can understand Phenotypic testing of mycobacteria rather than just read about it. In short: In microbiology, the phenotypic testing of mycobacteria uses a number of methods. The most-commonly used phenotypic tests to identify and distinguish Mycobacterium strains and species from each other are described below.

Key takeaways

Phenotypic testing of mycobacteria belongs to biology; place it in that map before memorising details.
Learn the definition first, then one example that makes the definition concrete.
Connect Phenotypic testing of mycobacteria to a quantity you can measure, compute or draw — that is where exam questions come from.
Reproduce the core statement of Phenotypic testing of mycobacteria from memory before moving on to harder problems.

Reference excerpt

In microbiology, the phenotypic testing of mycobacteria uses a number of methods. The most-commonly used phenotypic tests to identify and distinguish Mycobacterium strains and species from each other are described below.

Tests Acetamide as sole C and N sources

Media: KH2PO4 (0.5 g), MgSO>4*7H20 (0.5 g), purified agar (20 g), distilled water (1000 ml). The medium is supplemented with acetamide to a final concentration of 0.02M, adjusted to a pH of 7.0 and sterilized by autoclaving at 115°C for 30 minutes. After sloping, the medium is inoculated with one loop of the cultures and incubated. Growth is read after incubation for two weeks (rapid growers) or four weeks (slow growers).

Arylsulfatase test

Arylsulfatase enzyme is present in most mycobacteria. The rate by which arylsulfatase enzyme breaks down phenolphthalein disulfate into phenolphthalein (which forms a red color in the presence of sodium bicarbonate) and other salts is used to differentiate certain strains of Mycobacteria. 3 day arylsulfatase test is used to identify potentially pathogenic rapid growers such as M. fortuitum and M. chelonae. Slow growing M. marinum and M. szulgai are positive in the 14-day arylsulfatase test.

Catalase, semiquantitative activity

Most mycobacteria produce the enzyme catalase, but they vary in the quantity produced. Also, some forms of catalase are inactivated by heating at 68°C for 20 minutes (others are stable). Organisms producing the enzyme catalase have the ability to decompose hydrogen peroxide into water and free oxygen. The test differs from that used to detect catalase in other types of bacteria by using 30% hydrogen peroxide in a strong detergent solution (10% polysorbate 80).

Citrate

Sole carbon source

Egg medium

Growth on Löwenstein–Jensen medium (LJ medium)

L-Glutamate

Sole carbon and nitrogen source

Growth rate

The growth rate is the length of time required to form mature colonies visible without magnification on solid media. Mycobacteria forming colonies visible to the naked eye within seven days on subculture are known as rapid growers, while those requiring longer periods are termed slow growers.

Iron uptake

The ability to take up iron from an inorganic iron containing reagent helps differentiate some species of mycobacteria.

Lebek medium

Lebek is a semisolid medium used to test the oxygen preferences of mycobacterial isolates. Aerophilic growth is indicated by growth on (and above) the surface of the glass wall of the tube; microaerophilic growth is indicated by growth below the surface.

MacConkey agar without crystal violet

Niacin accumulation (paper strip method)

Niacin is formed as a metabolic byproduct by all mycobacteria, but some species possess an enzyme that converts free niacin to niacin ribonucleotide. M. tuberculosis (and some other species) lack this enzyme, and accumulate niacin as a water-soluble byproduct in the culture medium.

Nitrate reduction

Mycobacteria containing nitroreductase catalyze the reduction from nitrate to nitrite. The presence of nitrite in the test medium is detected by addition of sulfanilamide and n-naphthylethylendiamine. If nitrate is present, red diazonium dye is formed.

Photoreactivity of mycobacteria; Some mycobacteria produce carotenoid pigments without light; others require photoactivation for pigment production. Photochromogens produce non-pigmented colonies when grown in the dark, and pigmented colonies after exposure to light and re-incubation. Scotochromogens produce deep-yellow-to-orange colonies when grown in either light or darkness. Non-photochromogens are non-pigmented in light and darkness or have a pale-yellow, buff or tan pigment which does not intensify after light exposure.

Picrate tolerance

Grows on Sauton agar containing picric acid (0.2% w/v) after three weeks

Pigmentation

Some mycobacteria produce carotenoid pigments without light; others require photoactivation for pigment production (see photoreactivity, above).

Pyrazinamide sensitivity (PZA)

The deamidation of pyrazinamide to pyrazinoic acid (assumed to be the active component of the drug pyrazinamide) in four days is a useful physiologic characteristic by which M. tuberculosis-complex members can be distinguished.

Sodium chloride tolerance

Growth on LJ medium containing 5% NaCl

Thiophene-2carboxylic acid hydrazide (TCH) sensitivity

The growth of M. bovis and M. africanum subtype II is inhibited by thiophene-2carboxylic acid hydrazide; growth of M. tuberculosis and M. africanum subtype I is uninhibited.

Polysorbate 80 hydrolysis

A test for lipase using polysorbate 80 (polyoxyethylene sorbitan monooleate, a detergent). Certain mycobacteria possess a lipase that splits it into oleic acid and polyoxyethylated sorbitol. The test solution also contains phenol red, which is stabilised by the polysorbate 80; when the latter 80 is hydrolysed, the phenol red changes from yellow to pink.

Urease (adaptation to mycobacteria)

With an inoculation loop, several loopfuls of mycobacteria test colonies are transferred to 0.5 mL of urease substrate, mixed to emulsify and incubated at 35 °C for three days; a colour change (from amber-yellow to pink-red) is sought.

References

Worked examples

Example 1 — a first encounter with Phenotypic testing of mycobacteria

Start with the simplest possible case. Write down what Phenotypic testing of mycobacteria claims or describes in one sentence, then invent the smallest concrete situation in which that sentence is true. In biology, the smallest case is usually a single object, a single equation or a single measurement. Check that every symbol or term in your sentence has a meaning in that case.

Example 2 — changing one variable

Take the situation from Example 1 and change exactly one quantity: double it, halve it, or set it to zero. Predict what should happen to Phenotypic testing of mycobacteria before you calculate. Comparing your prediction with the result is the fastest way to find out whether you understand the idea or only the words.

Example 3 — an exam-style question

Typical questions about Phenotypic testing of mycobacteria ask you to (a) state it precisely, (b) apply it to given data, and (c) explain a limitation. Practise writing all three answers in under five minutes; the third part is what separates a full-mark answer from an average one.

Applications of Phenotypic testing of mycobacteria

In research: Phenotypic testing of mycobacteria appears in biology research whenever the underlying quantities have to be modelled precisely. Papers usually cite it as a starting assumption and then explore where it breaks down.
In technology and industry: Engineering practice reuses Phenotypic testing of mycobacteria in design rules, simulations and safety margins. Knowing the idea lets you read a specification sheet and understand why the numbers look the way they do.
In the classroom: Phenotypic testing of mycobacteria is common in secondary-school and first-year university syllabi. It links to neighbouring topics Bacteriology, Microbiology techniques, so understanding it makes those chapters shorter.
In everyday life: Look for Phenotypic testing of mycobacteria outside the textbook — in sport, cooking, traffic, electronics or the sky above you. An example you found yourself is remembered far longer than one you were given.

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How to study Phenotypic testing of mycobacteria in 20 minutes

Read the reference excerpt below once, without taking notes.
Close the page and write down what Phenotypic testing of mycobacteria means in your own words.
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Frequently asked questions

What is Phenotypic testing of mycobacteria in simple terms?

Why does Phenotypic testing of mycobacteria matter?

Because it connects several biology ideas at once: it gives you a definition you can apply, a quantity you can calculate, and a way to check whether a result is plausible.

How should I study Phenotypic testing of mycobacteria?

Read the excerpt, restate it from memory, then work through the examples and applications listed on this page. The five-step study plan above takes about twenty minutes.

What does this page cover?

It gives you a compact reference excerpt plus original lgStudy explanations, examples, applications and study material on Phenotypic testing of mycobacteria.