..fungal titers?

nmaccallister at webtv.net nmaccallister at webtv.net
Tue Feb 20 04:04:03 EST 2001


Thanks for your reinforcing the safety and cross-contamination issues
involved in any microbial work.  All my platings and transfers are done
in an externally exhausted (double HEPA filtered) validated flow-hood.
I wash that hood with IPA before and after bacterial work, and a dilute
bleach solution after fungal work.  I always wear gloves, Tyvek sleeve
guards, filter mask, and safety glasses.
My purpose is to test the kill rate of some commonly used preservatives.
Most protocols, including the USP, start from cultures grown on solid
media,.. yet many state that liquid cultures are also acceptable.  I
liked the thought of liquid cultures exactly for the reason of reducing
the opportunities of unplanned releases of spores, whether in the hood
or in the incubator.  But while I have significant bacterial growth
experience, I've done very little with molds.  Because of that void, I
thought I would save some time and ask for suggestions, especially
regarding the potential for growing Aspergillus out in a broth culture.
As a test case, I watched some air-catch penicillin-like molds grow
dandelion like structures about one inch under the surface of TAT broth.
They remained a white, floating, "angel-hair" sphere for several
days,... then I shook the bottle vigorously, and spread plate several
drops on SabDex agar. I got nice colonies at 3 days/25*.  Unfortunately,
I didn't have the time to quantify the procedure, so all I learned was
that it works for some molds,.. and that it was fun!
I'm guessing that the fresh growth in liquid culture provided mycelial
stage cells,.. and that the vortexing just fragmented that, and produced
a number of "growing points" (Thanks Mr. Reade!) on the subsequent
agars.
I guess I'll find out if this will work for A. niger this week
(..Kwik-sticks arrive tomorrow!) Until I do, I'll expect to be using a
TAT or Letheen Broth "wash" of a solid media outgrowth,.. (and I'll use
Petri-Seal tape on the pates if I have to protect the incubator!) 
I guess there is no easier way than hard work and careful record
keeping!  I did get some valuable suggestions though, so I thank you
all. And I hope you ALL had a great 3-day weekend!






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