RE: GMS for fungi

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From:Hewlett Bryan <HEWLETT@HHSC.CA>
To:"''" <>


You wrote;

>>The traditional GMS (Gomori or Grocott methenamine silver) stain for fungi

(including Pneumocystis carinii, which alternates weekly between being a 
fungus and a protozoan) uses chromic acid as an oxidizing agent, to cleave 
vicinal diol linkages and thus produce aldehyde groups which then reduce an 
ammoniacal silver reagent (methenamine silver). 

As does Periodic acid.

>>Chromic acid, a very strong oxidizing agent, is used because it produces 
aldehyde groups in very resistant fungal cell walls.

Again, true. However, So does Periodic acid!  

The PAS reaction gives very strong staining of the majority of fungi when
performed correctly.
The only negative reaction that I have seen, in over 40 years of practice,
was a Mycetoma that was apparently fossilized!! It did, in fact, stain very
very weakly with GMS. I attribute this to the much higher contrast of silver
deposits. However, a problem encountered with PAS staining for fungi is the
surrounding positivity, of other elements, making visualization difficult(
e.g. the proteinaceous matrix surrounding the cysts of P. Carinii, whatever
the current taxonomic group). The problem is one of contrast, not of
	The real reason, that Chromic acid is preferred for fungi, lies in
its tendency to over-oxidize the formed aldehydes to carboxylic acid, which
results in a weak schiff reaction. The fungal cell wall, being resistant,
retards this tendency. In contrast, the surrounding formed aldehyde groups
remain unprotected and thus over-oxidize. Periodic acid does not
progressively over-oxidize(at least in any reasonable time frame), thus
giving more intense reactions.
	This effect was first noticed in 1933 by Bauer, who capitalized on
it to produce the Bauer stain(also known as a Bauer-Feulgen reaction) for
Glycogen. Gridley,in 1953, modified the Bauer stain for demonstration of
fungi in tissues by adding counterstains with Gomori's aldehyde fuchsine and
Metanil yellow. The results of Gridley's stain are often quite pale, but
only fungal elements and glycogen stain. Grocott, in 1955, changed the
aldehyde detection system in Gridley's stain from schiff reagent to Gomori's
methenamine silver, in order to increase the intensity, and hence the
contrast. This strategy was so successful, that we all adopted it and GMS
became the standard for fungal staining.

>>Because of its environmental problems, chromic acid has become difficult
use. Kit manufacturers, with a wave of an MBA's hands I suppose, have 
declared that periodic acid, a much weaker oxidizing agent, can be used in 
its stead. But where is the literature to support this change?

It's out there Samurai! From an era long gone!
Try the venerable R.D.Lillie. "Histopathological technic and practical
histochemistry" 3rd Ed. pp 569-574, 1965.

>>Different fungi differ greatly in their demonstrability with this
and ideally one would have a different control for every fungus one wished
demonstrate. Probably the most difficult of common pathogenic fungi is 
Histoplasma capsulatum in old fibrotic or caseous lesions. 

Yes, I agree but see above. 
Perhaps we could apply a version of Lillie's Allochrome procedure to these

>>If I saw a study that found periodic acid to be as good as chromic acid
this "acid test", then I might believe.

Have a little faith in ol' R.D.
As for the study, here I'm with you 100%.

Good to see you back!


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