Recently in my agar plates I am seeing something very odd and am wondering if it is a contaminant that anyone has seen before?
It starts off looking like tiny stars embedded inside the agar after only a couple of days, and after I let it for a week the lines of the star were much longer. It is not limited to the streak-lines, it is also in other random places of the agar. It seems to only occur on plates that I have inoculated with my Pseudomonas culture, and not on uninoculated plates. Does anyone know what this is, and/or what the source might be so that I can try and prevent this? I sterilised the agar with a normal autoclave cycle and worked aseptically.
It seems the process of autoclaving it not proper! Use a visual (Tape) indicator to put on the media vessel/ flask/ Petri plates which are subjecting to autoclave. The adhering tape becomes brown/ black if autoclaving it proper.
As it is there only in Pseudomonas inoculated plates and not in uninoculated ones, I suspect its due to motility of Pseudomonas. Just check the percentage of agar that you use. Try staining the smear from those streaks. Also try growing those in broth and do some tests to confirm whether its Pseudomonas or not.
Hey Simon A. Dingsdag
long time no see! Thanks for your reply. Wow do you work with Pseudomonas too? Yes that's King's B. Just this morning I had the same thought and re-filtered the MgSO4, so I will see how that goes! I do have my doubts about the autoclaves here sometimes, but I think it may have happened to me from different agar batches made in diff autoclaves. I'll do a process of elimination though.@Shamsher singh kanwar - thanks, but yes I do already put autoclave tape every single time that I autoclave anything :)
@prerana sharma - thank you, my colleague had a similar thought, but I made regular 1.5% agar. I'll try a smear to see whether it looks like Pseudomonas, but I have a feeling it isn't.
Christie Foster Sure. Do let us know what that is as I am quite curious.
Simon A. Dingsdag
I agree that the contaminants are embedded. But I think the bacteria does not move just over the surface. Yes they are dispersed so I agree with you in that matter. But unless it's checked by either staining or some biochemical test we won't know what it is.
Hi again, after this post I tried restreaking colonies from those plates onto freshly made KB agar, AND streaked the -80 stocks onto new agar too. I also incubated uninoculated control plates. None of the plates had the "stars" in the agar this time, HOWEVER I then left them in the fridge for a week and pulled them out today and the stars had formed! Only on the inoculated plates, and not necessarily near the streak lines.
Further, I tried scooping out a chunk of agar containing a "star" and streaked it onto a plate. It grew colonies, which kind of look similar to pseudomonas (at least just to my eye). So perhaps this all tells me that it is actually pseudomonas, and something weird is going on!? I've never seen this ever before in my past year of working with pseudomonas, so I don't know why it's happening now.
Finally you could find the solution and unwanted contamination issue has been resolved.
It's less likely that it's contamination, but I haven't confirmed that it's definitely pseudomonas. And I'm still not sure why it is growing like this when it's never happened ever before. And why it appeared this time after sitting in the fridge!
I didn't check the MgSO4, because the freshly made and uninoculated plates looked perfectly fine after being incubated (and refrigerated) :-)
But yeah, I guess I can do a gram stain and/or sequencing to check if it is actually pseudomonas behaving oddly.
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