..fungal titers?

Austin Reade rbsjunk at hfx.andara.com
Tue Feb 20 16:19:44 EST 2001

Bear in mind that the response of  fungal spores (conidia) to
preservatives/disinfectants may be quite different to that of the mycelial
stage.  Conidia  would usually (always?) be more resistant than the mycelial
stage so an MIC determined on mycelium might not be effective for conidia.
The relevance of this depends on your application.

You refer to "kill rate of preservatives".  I suppose terminology may differ
across disciplines but in my book, preservatives don't necessarily kill they
prevent further growth.  If the preservative effect is subsequently
neutralized, growth may occur, or continue.  In contrast, disinfectants
(biocides) kill the organism and even if neutralized, no growth will occur
in the absence of further inoculum.  Again, the relevance of this will
depend on the exact application in question.

Mold growth in liquid culture can take various forms depending upon specific
conditions.   Low inoculum levels and minimal agitation often results in the
spheres you mention.  We used to call this pellet growth.  Increased
inoculum levels and increased agitation tends to produce diffuse growth with
no discrete pellets.  For fungal biomass production we tried to maintain
diffuse growth to allow for optimal mass transfer of oxygen and nutrients.
Some industrial fermentations are deliberately run in the pellet mode as the
physiology of the mold at different depths into the pellet changes and has
benefits for certain metabolic products.

Fragmenting mycelial growth (usually in a Waring blender) to increase the
number of "growing points" was  used in attempts to minimize the early
slower phase of growth when a starter culture was used to inoculate the next
culture in a series of increasingly large fermentation tanks.  I don't know
to what extent it is used industrially.  We tried it a few times out of
interest.  One has to take care not to over-fragment as a point is reached
where the mycelium becomes excessively damaged and subsequent growth is
actually slowed.

No three-day weekend up here of course [  :) ]but I hope you enjoyed yours.
Then again I'm semi-retired so it's all three-day weekends for me!!

Austin Reade
Reade BioSciences Inc
Halifax,NS, Canada

<nmaccallister at webtv.net> wrote in message
news:27416-3A923303-134 at storefull-117.iap.bryant.webtv.net...
> 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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