Samples grow on Broth but not in Agar - (Sep/23/2008 )

Hello everyone!

I'm having some problems in growing my samples on Agar media. They grow well (and fast!) on Broth media but not in Agar.
Well, I trying to grow the following species:
- Vibrio anguillarum
- Vibrio carchariae
- Photobacterium damselae subsp. damselae
- Enterococcus faecium
- Enterococcus faecalis

All were grown on Tryptic Soy Broth enriched with 3% NaCl with no problem, in less than 24 hours the Eppendorf tubes were turbid (compared with a negative tube).

Then I've streaked a loop in the following Agar medias:
- Thiosulfate Citrate Bile Salts Sucrose Agar (TCBS) from Difco for Vibrio and Photobacterium
- Confirmatory Enterococcus Agar (CEA) from Himedia for Enterococcus

The plates were incubated according with the manufacturer's instructions:
- 35ºC for 24-48 hours.

After the incubation time only Photobacterium showed (an excellent!) growth.

By the way, my samples were stored in liquid nitrogen with glicerol as cryoprotector!

Does someone has any idea?
Thanks in advance!

Have you checked (microscopically...) if you have not accidentally grown some contaminating bacteria in your tubes (just to give you an idea: bacillus or some coliforms)?
Maybe you should try a larger volume of liquid culture medium in 10 mL tubes, as the nutrients can be depleted and your bacteria are in a bad physiological state in your eppendorf tubes, so they grow very slow.

And agree with Hanming86, add TSA as "positive control" when plating your bacteria