Differential Staining of Bacteria: Endospore Stain

Jackie Reynolds1, Rita Moyes2, Donald P. Breakwell3

1 Richland College, Dallas, Texas, 2 Texas A&M University, College Station, Texas, 3 Brigham Young University, Provo, Utah
Publication Name:  Current Protocols in Microbiology
Unit Number:  Appendix 3J
DOI:  10.1002/9780471729259.mca03js15
Online Posting Date:  November, 2009
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Abstract

Endospore production is a very important characteristic of some bacteria, allowing them to resist adverse environmental conditions such as desiccation, chemical exposure, extreme heat, radiation, etc. The identification of endospores is also very important for the clinical microbiologist who is analyzing a patient's body fluid or tissue—there are not that many spore‐forming genera. In fact, there are two major pathogenic spore‐forming genera, Bacillus and Clostridium, together causing a number of lethal diseases—botulism, gangrene, tetanus, and anthrax, to name a few. Curr. Protoc. Microbiol. 15:A.3J.1‐A.3J.5. © 2009 by John Wiley & Sons, Inc.

Keywords: differential stain; endospore stain; malachite green; Bacillus sp.; Clostridium sp.

     
 
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Table of Contents

  • Introduction
  • Basic Protocol 1: Schaeffer‐Fulton Endospore Stain
  • Alternate Protocol 1: Cold Method Endospore Stain
  • Reagents and Solutions
  • Commentary
  • Literature Cited
  • Figures
     
 
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Materials

Basic Protocol 1: Schaeffer‐Fulton Endospore Stain

  Materials
  • Beaker (500 ml or 1 liter) of water
  • Distilled or deionized water
  • Specimens to be stained (Bacillus subtilus is good as a + control)
  • Malachite green solution (see recipe)
  • Safranin solution (see recipe)
  • Hot plate
  • Small wire overlay for the top of the beaker (or another heat‐resistant structure that will hold the slide over the beaker rim while steaming)
  • Inoculating loops
  • Slides
  • Bunsen burner
  • Stain rack
  • Paper towel (cut the size of the slide)
  • Bibulous paper
  • Light microscope
  • Additional reagents and equipment for examining the slides using a light microscope (unit 2.1)
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Figures

Videos

Literature Cited

   Hussey, M. and Zayaitz, A. September 29, 2007. MicrobeLibrary: Endospore Stain Protocol, Retrieved from http://www.microbelibrary.org/Edzine/details.asp?id=2565&Lang=English.
   Giorno, R., Bozue, J., Cote, C., Wenzel, T., Moody, K.S., Mallozzi, M., Ryan, M., Wang, R., Zielke, R., Maddock, J.R., Friedlander, A., Welkos, S., and Driks, A. 2007. Morphogenesis of the Bacillus anthracis spore. J. Bacteriol. 189:691‐705.
Internet Resources
  http://delrio.dcccd.edu/MFarinha/lab_manual/TOC
  Online laboratory manual of this unit's author (J. Reynolds), with information on other stains and tests.
  https://www.afresearch.org/skins/rims/q_mod_be0e99f3‐fc56‐4ccb‐8dfe‐670c0822a153/q_act_downloadpaper/q_obj_a654b234‐ea95‐404f‐b6b4‐009c12dee2a8/display.aspx?rs=enginespage
  Air Force Institute of Technology, thermal inactivation of Bacillus anthracis using laser irradiation of micro‐etched platforms.
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