SEA-PHAGES | Phages suddenly not fully lysing on M. foliorum

Link to this post \| posted 21 Feb, 2020 20:43
Thiel	Hello, I think that we are having a similar problem to the post above that says "D29 plaques barely visible on M. smeg" but I'm not sure. We are doing the phage isolation for the 2nd semester (so I'm not very experienced) and the students were on their 2nd round of purifications and starting to do the 3rd round and then make webbed plates. Our host is M. foliorum. I take new M. foliorum out of the -80C every 2 weeks, streak a plate and then shake it to make new stocks for them to use. The entire class started getting very similar looking plaques, that were difficult to see and smelled weird. (See image). The plates labeled "old" bacteria is what we had been using. The plates labeled "new" bacteria was when we picked a new colony from the same streak plate and it starting giving us the odd results. The difference in the incubation times was because we thought if we gave the 10^-3 plate a little more time- it would fully lyse, but it didn't. We picked a another new colony from the M. foliorum streak plate, grew it up and it seems to have fixed the problem (no pic). Just in case, I made a new M. foliorum streak plate from a different glycerol stock and will be growing that up for future use for the students. So we might have fixed the problem, but what do you think happened? Did we get lucky and pick a mutant colony? Any insight would be awesome. I'll let you know if the problem is not fixed. 1.27Mb

Link to this post | posted 21 Feb, 2020 20:43

Hello,
I think that we are having a similar problem to the post above that says "D29 plaques barely visible on M. smeg" but I'm not sure.
We are doing the phage isolation for the 2nd semester (so I'm not very experienced) and the students were on their 2nd round of purifications and starting to do the 3rd round and then make webbed plates. Our host is M. foliorum. I take new M. foliorum out of the -80C every 2 weeks, streak a plate and then shake it to make new stocks for them to use. The entire class started getting very similar looking plaques, that were difficult to see and smelled weird. (See image). The plates labeled "old" bacteria is what we had been using. The plates labeled "new" bacteria was when we picked a new colony from the same streak plate and it starting giving us the odd results. The difference in the incubation times was because we thought if we gave the 10^-3 plate a little more time- it would fully lyse, but it didn't.

We picked a another new colony from the M. foliorum streak plate, grew it up and it seems to have fixed the problem (no pic). Just in case, I made a new M. foliorum streak plate from a different glycerol stock and will be growing that up for future use for the students.

So we might have fixed the problem, but what do you think happened? Did we get lucky and pick a mutant colony? Any insight would be awesome. I'll let you know if the problem is not fixed.

Link to this post \| posted 21 Feb, 2020 20:51
viknesh	Hi Sarah, It's likely that the liquid culture was contaminated. As such, your lawn is composed of more than one bacterium - M. foliorum and other bacteria. Phages will still be able to infect M. foliorum, but plaques may not be apparent because contaminating bacteria, which cannot be infected by those same phages) are able to continue to grow evenly on the plate and therefore also within the plaque. The weird smell is likely due to the contaminant. Hope this helps. Vic Thiel Hello, I think that we are having a similar problem to the post above that says "D29 plaques barely visible on M. smeg" but I'm not sure. We are doing the phage isolation for the 2nd semester (so I'm not very experienced) and the students were on their 2nd round of purifications and starting to do the 3rd round and then make webbed plates. Our host is M. foliorum. I take new M. foliorum out of the -80C every 2 weeks, streak a plate and then shake it to make new stocks for them to use. The entire class started getting very similar looking plaques, that were difficult to see and smelled weird. (See image). The plates labeled "old" bacteria is what we had been using. The plates labeled "new" bacteria was when we picked a new colony from the same streak plate and it starting giving us the odd results. The difference in the incubation times was because we thought if we gave the 10^-3 plate a little more time- it would fully lyse, but it didn't. We picked a another new colony from the M. foliorum streak plate, grew it up and it seems to have fixed the problem (no pic). Just in case, I made a new M. foliorum streak plate from a different glycerol stock and will be growing that up for future use for the students. So we might have fixed the problem, but what do you think happened? Did we get lucky and pick a mutant colony? Any insight would be awesome. I'll let you know if the problem is not fixed.

Link to this post | posted 21 Feb, 2020 20:51

viknesh

Hi Sarah,

It's likely that the liquid culture was contaminated. As such, your lawn is composed of more than one bacterium - M. foliorum and other bacteria. Phages will still be able to infect M. foliorum, but plaques may not be apparent because contaminating bacteria, which cannot be infected by those same phages) are able to continue to grow evenly on the plate and therefore also within the plaque. The weird smell is likely due to the contaminant.

Hope this helps.

Vic

Thiel
Hello,
I think that we are having a similar problem to the post above that says "D29 plaques barely visible on M. smeg" but I'm not sure.
We are doing the phage isolation for the 2nd semester (so I'm not very experienced) and the students were on their 2nd round of purifications and starting to do the 3rd round and then make webbed plates. Our host is M. foliorum. I take new M. foliorum out of the -80C every 2 weeks, streak a plate and then shake it to make new stocks for them to use. The entire class started getting very similar looking plaques, that were difficult to see and smelled weird. (See image). The plates labeled "old" bacteria is what we had been using. The plates labeled "new" bacteria was when we picked a new colony from the same streak plate and it starting giving us the odd results. The difference in the incubation times was because we thought if we gave the 10^-3 plate a little more time- it would fully lyse, but it didn't.

We picked a another new colony from the M. foliorum streak plate, grew it up and it seems to have fixed the problem (no pic). Just in case, I made a new M. foliorum streak plate from a different glycerol stock and will be growing that up for future use for the students.

So we might have fixed the problem, but what do you think happened? Did we get lucky and pick a mutant colony? Any insight would be awesome. I'll let you know if the problem is not fixed.

Link to this post \| posted 21 Feb, 2020 20:56
Thiel	Great. Thank you!

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Phages suddenly not fully lysing on M. foliorum