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Endospore Staining

This document describes a procedure for endospore staining to detect endospores produced by certain bacterial species. Materials needed include cultures of Bacillus subtilis, Bacillus cereus, Staphylococcus aureus and staining reagents. The procedure involves preparing smears, heat fixing, staining with malachite green and safranin, then examining under a microscope. Results show B. subtilis contains green endospores while S. aureus does not, confirming their different abilities to form endospores. The conclusion is that endospore staining is an important technique for visualizing bacterial endospores.

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Gladys Tabane

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0% found this document useful (0 votes)

50 views4 pages

Endospore Staining

Uploaded by

Gladys Tabane

We take content rights seriously. If you suspect this is your content, claim it here.

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Download as DOCX, PDF, TXT or read online on Scribd

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ENDOSPORE STAINING

Endospores are very resistant structures produced by some bacterial species as a

survival mechanism in adverse environmental conditions. (Barthelousens and Mittuer
1950). With modification it is also useful in detecting spores produced by certain
eukaryotes. (Evans et al..,2012). Endospores ae formed by a process called
sporulation, which involves a complex series of transformation of bacteria cell.
Endospores are not metabolically active and can remain viable for a long period of
time.

MATERIALS

3-day cultures of Bacillus subtilis, Bacillus Cereus, Staphylococcus aureus.

Microscope slides
Schaeffer-Fulton endospore staining reagents
Malachite Green, 5%aqueous (aq.)
Safranin 5% aq
Bunsen burner
Forceps
Tripod stands gauzes
Beaker of boiling water

PROCEDURE

A smear of B.cereus, B. subtilis, S. aureus was prepared, air dried and heat fixed. The
slide was placed over a beaker of boiling water, then the smear on the slide was
covered with malchite green and steamed for 5 minutes. The smear was not allowed
to dry before 5 minutes elapsed by adding more dye as required. The slide was placed
on the staining rack using forceps, allowed to cool then rinsed thoroughly with tap
water until the water ran clear with no green dye flowing. The slide was drained, and
counter stained with 0.5% safranin for 1 minute. The slide was rinsed thoroughly with
water. The slide was blot dried and examined through the microscope at 100x (oil
immersion )

RESULTS

Bacteria Characteristi Gram Cell Cell Endospor DRAWIN

cs type arrangeme morpholo es G
nt gy Present or
absent
Bacillus Facultatively Gram Chains Rod Present
cereus anaerobic, positiv shaped
motile and e cells
spore
forming
Staphylococc Facultatively Gram Clustered Cocci absent
us aureus anaerobic, positiv cells
motile and e
spore
forming
Pseudomanas Heterotrophi Gram Single Rod Present
aeruginosa c and motile negativ cells shaped
e cells

Escherichia Non spore Gram Single Rod Present

coli forming negativ cells shaped
e cells

DISCUSSION

The results of the endospore staining of Bacillus subtilis and Staphylococcus aureus
confirm the differential ability of these bacterial species to form endospores. Bacillus
subtilis is known to be strong endospore former ,while staphylococcus aureus does
not produce endospores. The green ovoid structures observed in the Bacillus subtillis
smear are characteristics of endospores staining. The heat used in the staining process
helped to soften the endospore coat ,allowing the stains to penetrate and visualize the
endospores.

CONCLUSION