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Problems with Clostridium - (Apr/22/2012 )

Hi,

I´m trying to grow Clostridium sphenoides in a specific liquid medium (Reinforced Clostridial medium) and also in Petri dishes that I have put inside an anaerobic jar. It seems that the bacteria grows in the liquid medium as it becomes muddy or cloudy. However, there is no growth in the agar plate. The medium I have used is adecuate,so I don´t know which the problem is. I would like to extract DNA from this bacteria so that´s why I prefer to inoculate it in an agar plate. Could anyone give me any idea or any advice about it? I would really appreciate it..


Thanks

-vanu-

Assuming the obvious, but I have to ask: you did pre-reduce the agar plates before use, yes? O2 indicator in the canister?

-Paulgs3-

Is this the same medium only with additonal agar for plate culture? Assuming you're confident anaerobiosis was established in the jars, do you think your observation could be due to aerobic storage of plates before inoculation and subsequent anaerobic incubation?
The liquid medium is usually a little hazy - are you sure you saw actual growth?

-Phil Geis-

Yes, I have used the same medium adding the 15g/l of agar, and you´re right maybe the storage of plates before inoculation could be a possible cause (I have used O2 indicator), what can I do in that case? I have the same problem with an aerobic bacteria and a facultative anaerobic one, they grow in liquid medium but not in the agar plates....

-vanu-

vanu on Tue Apr 24 07:48:30 2012 said:


Yes, I have used the same medium adding the 15g/l of agar, and you´re right maybe the storage of plates before inoculation could be a possible cause (I have used O2 indicator), what can I do in that case? I have the same problem with an aerobic bacteria and a facultative anaerobic one, they grow in liquid medium but not in the agar plates....

The aerobic one not growing in the plates does not sound normal...

I am starting to think you are doing something "weird" (wrong) with your plates then...

Are you sure that the only difference between the plates and the liquid is the agar? And what about growht conditions? Temp? etc?

-pito-

I have put the agar plates and the liquid medium together at the same time inside the anaerobic jar, with the same conditions ...I have no idea which is/are the steps where I´ve done something wrong...
Anyway thank you very much

-vanu-

What catalyst are you using? How are you controlling the environment in the jar?

-Paulgs3-