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Incubation of Soil Fungi

About microscopic forms of life, including Bacteria, Archea, protozoans, algae and fungi. Topics relating to viruses, viroids and prions also belong here.

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Incubation of Soil Fungi

Postby tomtriglone » Mon Jul 11, 2011 6:48 am

Firstly, let me introduce myself by saying I'm no Soil Scientist, just a little interested in understanding a bit more of the processes involved in testing and monitoring of Soil Fungi.

I am involved in a company looking to augment vermi-compost with biological fungicides and pesticides, in the form of plant-friendly bacteria and fungi. And one of our production quality controls involve monitoring the concentrations of these fungi/bacterias that we add to the vermi-compost.

So I've started reading a little more about fungi and finding out the "concentrations" in cfus.

Our "lab guy" is currently producing a soil fungi (Trichoderma sp.) in liquid form, primarily because it's easier for us to apply directly to the vermi-compost. I want to find out what sort of cfu concentration he has in this liquid (We would be expecting somwhere between 1x10^6 and 1x10^8 cfus/ml). The dilution process I think I understand, my question is about the incubation process.

Once we have the liquid diluted by 1/10^6, 1/10^7 and 1/10^8 times, how should the incubation be done? Is it as simple as providing a sterilized food source for the fungi in the liquid? Any ideas on incubation times?
Basically, can anyone give me some basic pointers on how to understand the process of finding out the cfu count?

If you need any more specifics I'll do my best to provide them.
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Postby JackBean » Thu Nov 03, 2011 9:23 am

cfu is calculated so that you plate your medium with bacteria on some agar (in different concentrations) and calculate the amount of grown colonies. To make it easier later, measure also OD600 (the optical density at 600 nm) and make some correlation graph.
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Postby JorgeLobo » Sat Nov 05, 2011 12:52 am

This is completely off base. The cfu and od measurement are not useful for mycelial fungi such as Trichoderma spp. the growth of which takes place by apical extension without cytokinesis (cell division). One or a 1000 distinct fungal cells can give 1 cfu. Similarly serial dilution is invalid procedure to reduce inoculum concentration.
You could try harvesting and resuspending conidiospores but precise measurment even with these is not readily obtained and multicellular mycelial fragments and conidiospores clumps are very common. I imagine this is what your so-called "lab guy" is producing.

If you're working with fungi - please learn the terminology. Fungi are plural fungus is singular. Similarly, bacteria are plural and bacterium is singular.
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Postby DanielSan » Sat Nov 05, 2011 10:44 am

As JorgeLobo has already pointed out, conventional bacteriological methods are useless in this case.
I`ve been working with filamentous fungi for quite a while now, mostly in liquid cultures. To assess the growth of the fungi, we use indirect measures such as pH and amount of extracellular protein. Depending on the C and N sources, many fungi can produce impressive amounts of extracellular enzymes. Enzymatic activities (we are esp interested in redox enzymes) can also give a clue about the progress of growth and the adaption of the fungus to the respective substrate. Also, determination of wet or dry biomass might be possible in some cases. However, we usually use solid, non-soluble substrates which makes this method obsolete. And when using soluble sugars, we found that most fungi tend to form large clumps.

Macroscopic observation (with a microscope) might also help to monitor growth.
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Postby JackBean » Sat Nov 05, 2011 11:48 am

my bad, I didn't realize he's talking about fungi :roll: :(
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