I am currently working on the biosurfactant production by Streptomyces sp. in batch mode using shake flasks (submerged fermentation). Based on several literature, the fermentation for this strain will be carried out for 168 hours on a rotary shaker, using the mineral salt medium (MSM) as the fermentation medium with refined palm oil as the carbon source.
However, from my observation, after 72 hours of incubation, contamination can be detected, by observing the characteristics of the medium. It turns milky white in color instead of clear and brownish color. I have been extra careful on the aseptic technique and even ensured that the contents of the medium are from a good source (not oxidized or not beyond its expiry date).
So, I am quite stuck at the moment on why this problem keeps on re-occurring. I have been struggling with this problem for few weeks and I still could not find the cause of this problem. I truly appreciate your assistance and views in this matter. Thank you in advance.
Check https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7074673/
https://www.frontiersin.org/articles/10.3389/fsufs.2021.696518/full
Check also on this paper
https://www.mdpi.com/2076-2607/10/3/610/pdf
Syafiqah Nadhirah Mukhtar: I do not know how you sterilized the oil. Usually, for 100 ml of oil add about 1 ml water and sterilize at 121 deg C for 20 min. before using in the fermenter. Put the oil for sterility check for 24 hr and use. Check your MCB (Master Cell Bank) for sterility (both for bacteria and fungi) and make sure that your MCB & WCB (Working Cell Bank) vials are sterile. Once the WBC is used for inoculation, DO NOT STORE FOR NEXT BATCH.
Can you run a negative control alongside to check whether this is in fact the media or the inoculum? Is your inoculum from spore suspension or pre-culture? Is it possible to look at the broth under a microscope to ascertain any contaminants? What happens to the culture after 72h? Some Streptomyces strains that I grew from spore suspension in a 5L bioreactor would look cloudy for some time before flocs were formed.
3/24/22
Dear Syafiqah,
This is probably NOT the cause of your contamination problem, but it is easy to check: Are you sure your autoclave is working properly? Periodically, I check (or re-certify) the autoclave w/ Bacillus stearothermophilus spore strips. It's possible the temperature and/or steam pressure control is faulty. I hope this information helps you.
You have received good suggestions from the other responders. Good luck with your research.
Bill Colonna Iowa State University, Ames, Iowa, USA [email protected]
Turbidity can be due to growth of Bacteria that you are adding in the medium.
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