OP50 contamination on NGM plates

Hi research fellows,

I study in a C. elegans lab in my first year. Lately I started to notice a weird phenomenin in our NGM plates where the bacterial lawn seems to grow much thicker/have extra growth than it used to be (even so when it is left under room temperature for a day or two). This is most evident in mating plates where op50-looking bacteria grows wildly outside the orginally seeded area. The single colonies on a recently streaked plate appeared to be (abnormally) larger. The other members in my lab didn’t really care about this as the worms still grow and reproduce. Just out of curiosity and also a bit concern, I was wondering if anyone else ever experienced the same issue and could enlighten me on this. Could it be some kind of contaminations? Mutations? Or is it a normal variaton? Any thoughts are appreciated!! Thanks in advance!

Hi Yingjie,

“Growing wildly” is not a descriptor I normally associate with OP50. How old is your working stock of OP50? Has it been thawed recently, or has it been propagated on plates for a long time? If you have a frozen stock, have you tried thawing a fresh stock?

If you are certain that your bacterial culture is OK, have you recently started using a new bottle of peptone, or a different brand of peptone? We have found that OP50 grew abnormally on our NGM plates when we switched to a different brand of peptone. In our case, the lawn was sort of crusty rather than growing thicker, but I have seen labs use NGM with higher peptone content to promote richer bacterial lawns when growing large populations of worms.

Aric

1 Like

Hi Aric,

Thank you for your comment!! Yes I would agree growing wild should not be somethinge normal for OP50 as I’m under the impression that its growth is actually restricted on NGM plates. The working stock in question is more than a month old and I have tried to streak a new one from the frozen stock which worked perfectly fine. We always stick to the same kind of peptone so that should not be a cause of variation. I guess I was mainly curious about what is fundemantally wrong with the OP50 other than being too old, that causes it to overgrow so much to the degree worms start to get “stuck” within the food (like they are wading through the lawn instead of crawling freely). But anyways thank you again for your insights really appreciate it!!

You may have slime contamination. Are worms required for you to see the effect? That is, is it already in the OP50 or does it does it only happen when there are worms? If so, it could be the worms are carrying the contamination. Consider doing a bleach prep of some gravid adults which may help clean it up. Slime can be very persistent, surviving 20% bleach, and it does affect worm health and behavior. You can also just take some gravid adults and move them with fresh food to new plates every few hours / days to try and flush it away.