We recently obtained several cyanobacterial strains (Synechocystis, Chroococcus, Synechococcus, etc.) from the culture collection in Göttingen. Upon first inoculation in BG-11 (UTEX recipe), everything was fine and they grew well. After that, two attempts to grow them in BG-11 failed:
- subcultures did not grow further in BG-11.
- fresh inoculation from the original samples (stored at 16 °C, illuminated) failed.
What could be the most likely problem here? Whe have the following hypotheses:
- they need a microelement that is missing in BG-11 but can survive from stored reserves for a while (is there evidence that cyanobacteria don't grow over prolonged periods in BG-11?)
- the BG-11 of the subequent cultivations was not prepared properly (unlikely, as two separate batches of BG-11 failed)
- something else that we missed.
I would greatly appreciate if you could share your experience ("try again, it should be a piece of cake", "don't be surprised, this happens often due to to high illumination/to high CO2/too vigorous shaking") to narrow down the problem!
Hello Dr. Dominik Refardt ,
My first guess would be lowering of pH below 5.5 as at this pH the ionic imbalance in cell membrane causes instability in the peptide bonds of the amino acids due to loss of polarity. At lower pH the chief form of carbon is either carbonic acid or free carbon di-oxide, none of which are preferred by a C1 organism such as cyanobacteria (with reservations) which desire bicarbonates more in their ambient environment. Adding extra bicarbonate might do the trick.
Another cause could be the change in the redfield ratio within the media as greater P than N has always shown to trigger cyanobacterial blooms and not so much the other way around in lakes and rivers.
Acidic pH also would ensure a reduced environment where the N would mostly be present as ammonium where the organisms simply won't feel the need to fix atmospheric nitrogen and thus might deviate from their existing metabolic habit resulting in induced stresses.
A change in ambient external pH could also cascade into a shift in the internal pH which would obviously affect the enzyme functionality.
Absence or lack of Molybdenum and Iron might have caused it too as they require both these elements for the functionality of their FeMo center of Nitrogenase enzyme. Shortage of chelating agent in the media could have rendered the iron to precipitate out of the solution phase upon sterilization as insoluble hydroxides resulting in the dearth in iron.
These are all educated guesses based on which you can decide on your course of action.
Thanking you,
Dr. Abhishek Mukherjee
If you did not pH it then it is likely that it is too low. The BG-11 media should work fine if it is what they suggest. Most cyanobacteria do fine on it. It could be that some of your chemicals are not pure enough or have degraded to toxic compounds. You might want to increase the temperature as well.
pH is critical. Also checking the shelf life on compiounds used can make this fail as well. We make our own...
For basic BG11 medium refer to:
http://microbiology.ucdavis.edu/meeks/BG11medium.html
The two main causes may be low pH and temperature. In addition, check for frequent change in water composition (especially pH, alkalinity or chlorine)
After more than two years, it is time to solve the puzzle: we learned that our strain did not grow in BG-11 but was perfectly fine in Z-medium. Why? I don't know. But once we switched media, everything was fine.
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