These are very good questions. In response to your first question, this is the one you can answer yourself.

If you get a collection with plenty of material then try all the different techniques and make a careful note of the measurements of fresh and dried material. This will give you a good picture of the changes (if any) caused by your methods and readings using your personal microscope equipment/camera.

Do bear in mind that the use of different stains and reagents can have different effects on different types of fungi. With regard to ascomycetes I would recommend Lugol's iodine rather than Melzer's. This is well worth reading:
http://www.gbif-mycology.de/HostedSites/Baral/IodineReaction.htm

best wishes
Chris

To question 1:

yes, toxic media (like Melzer, KOH, ...) lead to a shrinking of cells (some more, some less). But this is only one important point. Killing of th cells also leads to a change - or better loss - of many important vital characters such as content of spores and paraphyses (e.g.).

Best regards, Lothar

 

In fact, also in Basidiomycetes the spores may shrink considerably and irreversibly when they die. This is mainly in those with hyaline thin-walled spores. These effects intuitively led many workers to the use of lactophenol, chloral hydrate, KOH or the like as a standard medium in order to achieve compatible measurements. But, as Lothar wrote, you lose important characters. In Basidiospores you lose the characteristic guttule pattern of the living spores which is often multiguttulate in one group, one big drop in another, and more or less empty spores in a third group. If you mount in CB or MLZ then you see a homogenepous content in all of them, but no difference anymore.

In Ascomycetes it is the same. But especially for asci the shrinking effect is enormous, unlike for basidia, because they need an elastic wall for spore discharge.

Dried specimens should always first be studied in water, because (1) the spores sometimes survive several years and even the asci and paraphyses may do so, and (2) KOH dissolves important exudates in some groups, also lactophenol or KOH often change the colour of pigments.

If you cannot see the croziers at the ascus base, you need to add KOH+ Congo Red, but not from the beginning....

For more details please see here:

https://invivoveritas.de/articles/vital-taxonomy/

Zotto

Thanks very much for the answers!

Excellent discussion, but has anyone published a paper on intraspecific variation vs. variation caused by different reagents?  If the former is greater, then it shouldn't matter what reagent you use to observe microscopic features.  If the latter is greater, then species concepts based on morphology just became more difficult.

Cheers,
Andy

Dear Andy
no doubt that intraspecific variation in e.g. ascus and spore measurements may sometimes be greater than the difference between living and dead cells. But if you neglect the shrinking effect and compare, let's say dead spores of coll. A with living spores of coll. B, then you may erroneously conclude that the spores of B are larger than those of A, although they would have quite the same size when you compare them in the same state.

Consider that reduction in volume may often be around 50%!
Zotto

Please also see the following article:

"The dependence of ascospore measurements on different preparation methods" by Kullman, Jakobson & Rahi
(Agarica 15, 245-254)

which can be found here:
http://www.soppognyttevekster.no/media/1193/agarica-1998-nr-24-25_komp.pdf

Cheers,
Marcel

Hello Otto,

Chris gave the link to your most informative article on iodine
http://www.gbif-mycology.de/HostedSites/Baral/IodineReaction.htm
and I am asking if we may post this onto our North West Fungus Group website, being so insightful ?

Best wishes,

John Watt

You can put a link to it, of course, thanks!
Zotto