Regardless the kind of determination method, the most problematic point likely would be to obtain a sample which is really representative for your system. Normally, in agitated liquid culture filamentous fungal organisms form particles (pellets) that may easily reach sizes of some mm. This is quite different from more or less homogeneous bacterial or pure yeast suspensions, where already a small sample amount (e.g. some hundered µl up to 1 ml) would ensure a rerpesentative analyses. With filamentous fungi, you should consider bigger sample volumes (according to my experience, 10 ml could work, taken with pipettes with cutted tips to catch also bigger particles). In any way, dry weight determinations (e.g. separation of biomass by vacuum filtration and drying on pre-weighted filter papers at 80 or 90°C over night) are a reproducible and very simple way to determine your total biomass amount.

You could make a microbial count whit dilutions, in an agar plate, you must use one sugar and metile red to discriminate everi strain , and you can count and discrimitate beteween strains

You can check OD at different time points and u can have rough idea about their growth. U can serial dilute the same culture and correlate with od. Then total count would be no. of colonies*dilution factor. It is the simplest.

Hi, I need to clarify that I have to quantify the growth during fermentation. And the broth not only consists of the fungis but also other biomass such as starch. Checking the OD would not be accurate isn't it, as it would also be influenced by the presence of other biomass?

other type of analisys is to do a real time pcr using a primer for eachstrain of fungi

If you have predominantly yeasts or other small units, the easiest and quickest way to follow growth is to count the cells in a normal microscopic cell-counting chamber. There are several sizes with different volumes available, among which you choose according to your expected cell sizes.

You can employ Millipore membranous filtration procedure using a pressure pump system with a known volume of the broth. Then culture the Microfilter membrane on appropriate fungi culture medium and do a cfu count. This way, you can without discrimination count the total number of the colonies of the different species and calculaute accordinglyper volume used.

Regardless the kind of determination method, the most problematic point likely would be to obtain a sample which is really representative for your system. Normally, in agitated liquid culture filamentous fungal organisms form particles (pellets) that may easily reach sizes of some mm. This is quite different from more or less homogeneous bacterial or pure yeast suspensions, where already a small sample amount (e.g. some hundered µl up to 1 ml) would ensure a rerpesentative analyses. With filamentous fungi, you should consider bigger sample volumes (according to my experience, 10 ml could work, taken with pipettes with cutted tips to catch also bigger particles). In any way, dry weight determinations (e.g. separation of biomass by vacuum filtration and drying on pre-weighted filter papers at 80 or 90°C over night) are a reproducible and very simple way to determine your total biomass amount.

if there arent any other filamentous fungi in the broth, counting cell number is the best way as yeast cells are large enough to to so. If there are other filamentous fungi, the above suggestion is very fine.

u can use total biomass method, u can filterate the growth in millipore filters & the dry in oven at 40 OC for particular time & weight out the mass

I think the best method for counting the mixture of fungi is in solid media like OGYE or PDA, although it takes several days. I agree with Dietmar Schlosser because fungal organisms form large particles that can not be counted in the microscopic counting chamber.

Hi Dietmar, where do I get the cutted pipette tips?

You could take 10-ml-pipettes made of glass, and just cut the tip with a glass cutter. You may also use 5-ml- or 10-ml-tips (e.g. for Eppendorf pipettes, made of plastics), just cut the tip with a a scissors to obtain an opening of the desired size, and then autoclave before use.

cheers, Dietmar

Thanks Dietmar, that's a really fine idea...

Dietmar, have you got any experience measuring the growth of filamentous fungis? What is the initial inoculum dosage and what long did you run your fermentation? I would like to have an idea whether the growth measurements are sufficiently significant.

In 2- or 5-L (lab scale) fermenters (stirred tanks): 5% inoculum, fermentations over 10 to 12 days (maximum)

Also, anybody knows of any paper dealing with growth of mixed culture?

As you dont require growth of individual species and you require overall growth so you should take dry mass.

Look at the work of Nik Money at the Miami University of Ohio. He was my Masters mentor (a lifetime ago) and is now an expert in that field. I bet if you called him he would help you in any way that he could.

Christopher Burlak, could you give me more information?

http://www.cas.muohio.edu/botany/people/profiles/Money.html

Yafetto, L., Davis, D. J., and Money, N. P. 2009. Biomechanics of invasive growth by Armillaria rhizomorphs. Fungal Genetics and Biology 46: 688-694. doi:10.1016/j.fgb.2009.04.005

Money, N. P. 2008. Insights on the mechanics of hyphal growth. Fungal Biology Reviews 22: 71-76.

Money, N. P. 2007. Biomechanics of invasive hyphal growth. In: Howard, R. J., and Gow, N. A. R., eds. The Mycota, Volume 8, Biology of the Fungal Cell, 2nd edition. Springer Verlag, New York, pp. 237-249.

Good Luck.

These links may help you

http://www.massey.ac.nz/~ychisti/BioTech.pdf

http://arrow.dit.ie/cgi/viewcontent.cgi?article=1092&context=sciendoc

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