Hi,
Could anyone please tell me how to prevent fungal attack in C.elegans or NGM plates…Many thanks…
Arafath
Using sterile technique is the best way to prevent fungal contamination. Bleaching and/or serially transferring the worm strains can help remove contamination once it is present. (See the “Maintenance of C. elegans” chapter of WormBook by Theresa Stiernagle, at http://www.wormbook.org/.) The antifungal drug nystatin added to the NGM media can be helpful. Nystatin has been used to affect worm cells in patch clamp experiments, so I would be careful what sorts of experiments you do on plates containing nystatin. In addition, nystatin in any aqueous suspension is unstable, which will result in decreasing concentration of drug in your plates as they age. Please also read the previous threads on this site (search using “nystatin”).
Hi there,
This may sound very simplistic and I apologize for that, but being kind of obsessional as follows has helped me greatly, in addition to the suggestions above which are excellent.
Check all your plates for ANY fungus before seeding them and remove any with signs of fungus on them without ever lifting their lids–preferably parafilm them then toss them immediately. Fungal spores disperse very effectively from the suction of opening the lid even once, then will happily land on many of the rest of the plates you’re trying to seed that day.
Once you carefully weed out the fungal ones, go ahead and seed the rest of your plates with OP50. Later, when moving worms to these seeded plates I check all the ones I plan to use and again weed out any with fungus, parafilm and discard. If I accidentally open a fungally contaminated one near my worm manipulation area, I take a break and wipe down the area with some 70% Etoh so the spores are reduced in number at least.
Just some simple precautions, but they’ll cut your contamination rates a lot (though never unfortunately to 0%) before the fungus even gets established.
Good luck.
Hello,
Thanks for the reply…
Thanks for the reply…
Many labs routinely pour and spot plates on the benchtop, where the air handling system can be a source of fungal spores. Also, the level of spores can vary with the seasons, so contamination can pop up in a formerly clean locale. Performing all of your plate manipulations in a laminar flow hood will minimize this problem.
-Harold
If not in a laminar hood, I try to pour the plates with at least two burners close by, and away from the lab door. Crowded areas are also not good for pouring the plates, people tend to produce air currents that will bring the spores to the plates.
Hi all. i have a little question to ask. i read about the wormbook protocol,
“6. Cleaning contaminated C. elegans stocks
Occasionally, C. elegans stocks may become contaminated with other bacteria, yeast or mold. It is easy to rid
your worm stocks of contaminants. Most contaminants will not hurt the worms. (In fact, the worms like to climb
some fungal hyphae and sway back and forth!) But it is much easier to score phenotypes and do transfers when your
stocks are clean. Mold can be removed by chunking (Protocol 3) and serially transferring, allowing the worms to
crawl away from the contaminant. Bacterial contaminants and yeast are easily removed by treating with a
hypochlorite solution, which will kill the contaminant and all worms not protected by the egg shell. This can be
done using an entire plate that is contaminated (Protocol 4), or it can be done quickly using a single hermaphrodite
(Protocol 5)…”
well, i thought most of the fungus will kill the worms ?
C. elegans meets a lot of fungi in its natural environment and in facts feeds on them. Certainly in our lab, it doesn’t seem unhappy on many of the typical fungal contaminants we get. It just makes observing things inconvenient.