| Malachite green monochloride salt |
| Endospore Stainig Schaeffer & Fulton 1933 |
DESCRIPTION
Endospores are non-reproductive structures produced by some Gram-positive bacteria, being a form of resistance in unfavorable
conditions, unlike the eukaryotic spores wich have a reproductive role. Bacterial endospores exists in a dormant state waiting for
reactivation when conditions become favorable.
Malachite green has been used for controlling external fungal, bacterial, and protozoan infections of fish, and successfully used as a
treatment to control fish saprolegniosis and as a prophylactic treatment to protect fish eggs from infection.
Endospores are non-reproductive structures produced by some Gram-positive bacteria, being a form of resistance in unfavorable
conditions, unlike the eukaryotic spores wich have a reproductive role. Bacterial endospores exists in a dormant state waiting for
reactivation when conditions become favorable.
Malachite green has been used for controlling external fungal, bacterial, and protozoan infections of fish, and successfully used as a
treatment to control fish saprolegniosis and as a prophylactic treatment to protect fish eggs from infection.
Staining technique consists in dyeing the smear with malachite green. Only the
endospore retain the stain. Vegetative cell is decolorized when washed with water. The
counterstain safranin is then used to color vegetative cell.
MATERIALS
Malachite green (synonyms: brilliant green, aniline green, benzaldehyde green, or china
green) solution 5%,
Safranin solution 0.5% or diluted fuchsin.
Use a culture about one week old; it will contain both vegetative cells and endospores.
An older than two weeks culture may have few if any vegetative cells. A younger than a
few days culture may have less or no endospores.
PROCEDURE
Make a bacterial smear. Dry and heat-fix the slide then cover it with malachite green
solution. Heat the slide for 3 minutes; avoid boiling. If not using steam method then
apply stain while keeping the slide wet for 10 minutes; add more stain to keep it wet if
necessary.
Wash in running tap water. Cover the slide with Safranin solution and wait 1 minute.
Remove solution, wash then dry with filter paper.
A demonstration of staining procedure is available in the video section.
RESULTS
Light-green inclusions (endospores) in pink-to-red vegetative cells.
NOTES
Excesive heating may deform the cells.
Short time malachite green staining may result in weak coloration of the spores.
endospore retain the stain. Vegetative cell is decolorized when washed with water. The
counterstain safranin is then used to color vegetative cell.
MATERIALS
Malachite green (synonyms: brilliant green, aniline green, benzaldehyde green, or china
green) solution 5%,
Safranin solution 0.5% or diluted fuchsin.
Use a culture about one week old; it will contain both vegetative cells and endospores.
An older than two weeks culture may have few if any vegetative cells. A younger than a
few days culture may have less or no endospores.
PROCEDURE
Make a bacterial smear. Dry and heat-fix the slide then cover it with malachite green
solution. Heat the slide for 3 minutes; avoid boiling. If not using steam method then
apply stain while keeping the slide wet for 10 minutes; add more stain to keep it wet if
necessary.
Wash in running tap water. Cover the slide with Safranin solution and wait 1 minute.
Remove solution, wash then dry with filter paper.
A demonstration of staining procedure is available in the video section.
RESULTS
Light-green inclusions (endospores) in pink-to-red vegetative cells.
NOTES
Excesive heating may deform the cells.
Short time malachite green staining may result in weak coloration of the spores.
| Green-stained, central, non-deforming spores inside red-stained vegetative cells (Bacillus anthracis) |
(c) Costin Stoica
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