Although I know the best way for spore induction is to mix the nutrient medium with manganese ion, I couldn't get any spores from these microorganisms by using Malachite green and Safranin (endospore staining)!
Normally, even under non stress condition, you should see some spores in your culture, since Bacillus sp. produce spores even uder optimal conditions.
I suggest you to use heat stress to induce sporulation. Then after spore production, heat the cell suspension at 80°C for 10 min to kill vegetative cells and take one drop of that into a solid media. If you could observe colonies after incubation, it means that heat can induce spore formation.
You can also limit nutrient source or decrease the pH of growth during growth.
Normally, even under non stress condition, you should see some spores in your culture, since Bacillus sp. produce spores even uder optimal conditions.
I suggest you to use heat stress to induce sporulation. Then after spore production, heat the cell suspension at 80°C for 10 min to kill vegetative cells and take one drop of that into a solid media. If you could observe colonies after incubation, it means that heat can induce spore formation.
You can also limit nutrient source or decrease the pH of growth during growth.
I did research work on Bacillus cereus two years ago. As far as I know, I got spores of Bacillus cereus after two to three days in agar plate. Culture aging will help you to form spore. Hope this answer useful for you.
lynn
Reply to Tin Lynn: How do you detect the spores on agar plate? Endospore staining? What type of agar did you use to culture B. cereus?
Sporulation can be very different according to the species and the strains itself. As said above, sporulation can be induced in normal conditions with aging cultures. But it is variable. If for B subtilis, you can induce sporulation with manganese and decoyinine in 1 or 2 days, the sporulation of some species like B pumilus is inhibited by manganese and needs at least one week in nomal conditions. First, I would try several cultures ages and check the sporulation by heating as described above.
Dear Kokwai Fung, I used MYP agar to isolate Bacillus cereus from fermented rice noodle. Then, I used Nutrient Agar for sub-culturing. I detect the spore of B. cereus by Endospore staining. Though I got spore, the photo of staining is not very good. Here is my paper about Bacillus cereus. You can take a look at that paper. All the best to you!
Article Molecular Characterization of Enterotoxin Genes from Food-bo...
You can obtain 107 UFC/ml G.sterothermophilus spores suspension by using agar sporulation medium. See manuscript attachmented. It is easy to its observe spores by Gram coloration.
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