I am now working on screening of biofungicide candidate against the growth of plant pathogens. The problem I face is when I grow my fungal candidate in broth and tested for inhibition assay using fungal filtrate obtained, several times my fungal filtrate could inhibit the growth of plant pathogens while sometimes when I try to repeat the experiment it does not provided the same result. Anybody know the reasons for this and how to deal with this problem then? Thank you very much.
Hi Rampai: I prefer you to use a agar medium like PDA, because the liquid medium is very fast to exposed by bacteria, and when the media in contaminated by bacteria you will get confusing results.
I would like to echo what na Wang has said above. Stability is key in performance of any biological pesticide.
It would be important to always use fungal filtrate grown for the same time period. If you wait for even 12h more/less it might exhibit different results. I would first do a time series testing for the best timing. So start growing your fungus at the same time in liquid cultures and then test the filtrates starting after 12h and going up, 24, 36, etc... Also are you using any additives like antibiotics in the broth ? This could alter your results too.
Thank you very much for all answers. I did grow my fungi in broth (no antibiotics included) not in agar medium since I just would like to use the fungal filtrate directly for inhibition assay. However, I am sure that there is no contamination in the media since I all the time done every procedure in laminar. After grow my fungi in broth for three days, I just collected the supernatant and filtered through 0.2 micron filter membrane and used the fungal filtrate for the inhibition assay. However the cultivation period is not exactly 72 hrs for every time where I done on fungal filtrate collection (as mentioned in Ioannis Baltzakis's comment), so this might be the possible reason why my result is not stable. I will try to check with this and let's see for the result then. Thank you very much.
The fungal cultures loose the capacity to synthesis the metabolite for every subculture. Because the fresh culture have maximum production slowly they tend to loose the synthesis. Optimization of media, temp, pH etc may be other reasons. So always you fresh culture for extraction.
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