Germinated myxomycete spore experiment

[Walter Piorkowski]

Myxamoebic swarm cell

Swarm cell flagellum

scale of above images @ 10 micrometers/division


Nikon type S microscope
phase contrast
40X BM phase objective
Special photo projection system using a microfilm projection lens.

Sometime back in any earlier posting Doug (Beetleman) asked if I had any images of myxamoebea in my archives. Well unfortunatly I do not, but something just as interesting. The myxamoebea will not form if water is present, instead a creature called a flagullated swarm cell is created. I managed to get one of these from several hundred spores. Which myxo species I don"t remember at this time.

When I began the experiment with several hundred spores I had no idea of how to measure success. I have no lab training of any sort. In the end I got this one cell. The experiment was weeks in duration and plaged with bacterial attacks of my spores. The cell did not last long.

Please don't beat me up too bad on the quality of these images as they were done early on in my photomicrographic dealings. Plus the scale is quite large.

These are for you Doug. Thanks for asking.

Walt

[beetleman]

Well Walter, thank you very much for bringing these out to show us. I think they are pretty awsome. The birth of a very strange creature.

[Ken Ramos]

I would say you did quite well in your experiment Walter, something I may need to try out. The images are pretty good and the flagella of the swarm cell is quite visible. Wonderful images indeed, thanks!

[beetleman]

Maybe you could fill us in on your technique Walter, things you did or might do differently.........And if you get a chance, take a look at this post I placed a few days ago and tell me what your thoughts are on what they are. http://www.photomacrography.net/forum/v ... .php?t=800 Thanks and take care.

Doug

[Ken Ramos]

Yes I agree with Doug , it would be interesting to read about what you did a little more exactly and what you learned from the experiment. Were there or are there anythings that you would have done different, now that the experiment is over, to see if you could have had a more successful turn out of ameobo-flagellates or swarm cells? When you conducted this experiment did you sterilize or autoclave the container(s) in which the spores were germinated, including the culture medium? If so for how long did the sterilization process take and at what psi? If not, then it is understandable for you to have had a problem with bacterial attacks upon the spores. On second thought though, sterilization may or may not have been useful unless one has a clean room but then again, being amateurs at all this, anything is better than nothing.

[Walter Piorkowski]

So you guys want to know all my secret methods!!! I guess my arm can be twisted.

Basically, since you are working with subjects that you cannot see but can isolate, this is my technique.

Materials required are a petri dish, a one by three inch microscope slide and cover slip, a lid from a frozen drink product (or alternative), a clear plastic cover for the dish, a needle or pin, eye dropper and distilled water. The glass parts are cleaned and assembled as my photos illustrate . Water is placed around the steel lid, but not over it. A few drops of water are placed on the center of the slide.

I choose a myxomycete that generated a lot of spores. The spores, which under bright illumination to my eyes can be seen as a dust, are liberated onto a sheet of clean glass. The tip of the needle is placed into the spore mass and will attract some spores. The collected spores invisible to you of course are transferred to the water droplet on the slide. A cover slip is gently lowered onto the water. It is good to have water bleed out around the slip because you will be adding some to this zone. This may cost you some of your spores but there are probably more than enough.

Observe your handy work under the microscope. Hopefully you collected what you wanted. Take some record photos and keep the water around the cover slip for the duration of the experiment. When not under observation the slide must be kept in its petri dish and under the cover to prevent evaporation of the water. This can be a real challenge in dry environment.

Scan the slide frequently for changes in the spores. I make certain that the spores get some indirect sunlight. From here on its just a waiting game. No food was provided.

I can’t begin to figure how the myxamoebae would be given a chance at life other than to start the same way and let the setup dry out, as they supposedly don’t want water. But you would think that there should be some moisture. Glass I doubt is a suitable substrate for amoeba development but how else would you be able to monitor them. Spores will definitely geminate on bark for example and I have thought of a possible high magnification EPI study of a freshly spored bark surface, but it would be a challenge

Walt

[Ken Ramos]

That is simple enough to give it a try, thanks for the rundown there Walt. I envisioned much more but I always try to make things more complicated than they should be. I find numerous short articles on how to grow myxomycetes but have never ran across one that goes into great detail as how to cultivate spore germination to observe the amoebo-flagellates. You are right in your thoughts about needing some water for spore germination and I have read numerous times about spores germinating in "free water" to produce the flagellates, however again, no one has addressed the subject on how germination occurs on moist substrates to where a myxoamoeba is produced or how one would go about observing it under the microscope. If they have I have just plain missed it. Thanks again Walt for your time in showing and telling us of your procedure.

[beetleman]

I second that thank you Walter . Can you hear the gears in Kens & my head going full speed