Step One: set up your heating equipment.
Set the thermostat on your aquarium heater to the best setting. I read that Bacillus subtilis is cultured in natto production between 37 and 42 degrees celsius so this is what I aimed at. My aquarium thermostat only went as high as 35 degrees. So, I positioned it in the Chilly Bin on a slight angle with the heater part of the unit at the top and the thermostat section at the bottom. Due to layering of temperature in still water this caused the water to heat slightly hotter than 35 degrees. But BE CAREFUL. If you slant the unit too steeply then you may over heat your water and may even damage your heater unit. I did this once and the water was so hot it was steaming. I found that the culture worked fine at 35 or 36 degrees so if you are concerned, just leave your heater and thermostat in the horizontal position.
Your chilly bin will need to be big enough to take the aquarium heater-thermostat unit horizontally in it. These units are attached to the side of the chilly bin using suction cups which is just the same as in an aquarium.
Put enough clean water in the chilly bin to cover the heater-thermostat by at least 6-10 cm
Once you have the heater-thermostat unit in position switch on the power and when the water has heated up fully, check the temperature with your thermometer. You don't need to wait for the water to be fully heated before you put your culture in. But when starting out it is a good idea to check the water temperature at regular intervals until you are sure you have it right.
If you are going to leave the same water in your esky for many days then I suggest you add a cap of bleach to stop it from growing algae, etc.
Step Two: Sterilise your culture container and spoons.
In this picture, you only need to sterilse the glass container, lid and spoons. I did this by pouring boiling water on them. Once it cools enough, pour the water down the sink. BE CAREFUL with hot water. Remember, If you accidentally scald yourself get the part under running cold water asap and keep it there for several minutes.
The metal container is the one I am going to float the glass one in.
Step Three: Using your sterilised spoon, put a thin layer of coconut cream in the bottom of your culture container.
You only need about half a cm or even less of cream in the bottom of the container if you are culturing B subtilis as it grows really well on the top of the coconut mixture. However, if using L plantarum you should mix your culture in and you can use much more coconut mixture.
Step Four: Get your culture and spinkle or mix a small amount on or in the coconut cream.
If using B subtilis, you don't need to mix the culture in. The Bacillus will grow happily on the top of the cream. If you are using pure L. plantarum then I recommend stirring it in.
For a pure L. plantarum in Australia I recommend using the Metagenics product called 'Probex' (not shown here). For B. subtilis, use either the natto culture starter or a Probiotic with Bacillus subtils as the main ingredient and preferably no fillers. The one in the shot here is called Enzymedica Pro-Bio™ and I got it from iherb.com. It has Bacillus subtilis and some species of Lactobacillus (including plantarum) and that is all.
The natto starter is handy because the bottle it is in is easy to use to sprinkle the spores from in small amounts.
My favourite to use is the pure L. plantarum as it works well and the culture smells much better than B sutilis (it smells like yoghurt).
Step Five: Put the lid on and float your culture in the water in the chilly bin.
You will need to make sure that your float container is the right size and weight so that it will not sink or tip over. I used a metal one because I was worried a plastic one might get damaged by floating against the heater unit.
Step Six: Monitor the water temperature and leave the culture in for 36 to 48 hours. Then retrieve and open for use.
I usually leave it in for 48 hours. For B subtils, if it has not been disturbed then after the 48 hours you should find a frosty like coating on the top of the coconut cream. If it has been mixed up then it will likely look slightly orange, pink or purple. It will also have a distinct smell which is not really pleasant. L plantarum is only sightly discoloured and smells like yoghurt.
If you are going to use the mixture for inoculation in your sinuses like I did then you will need three things: a neti pot, a plastic syringe and a nose spray bottle with a screw-off lid. For the neti pot you put clean water in it at body temperature and add a spoon full of the culture, mix thoroughly and then do your sinus rinse. For the syringe, you syringe up about 2ml of the culture and some clean water, mix and then inject it in the nose. For the spray, you must thin the culture with some water and make sure there are no clumps that could block the spray nozzle. If you keep it in the fridge then you may need to warm at least the spray tube (take the lid off to do this) in warm water before using as the coconut may have solidified in the fridge. Breath in through your nose while spraying twice both sides.
To use as a probiotic food, I normally add the B subtilis culture to tahini to improve the flavour. Some times I add a drop of pure peppermint oil. L plantarum tastes similar to yoghurt on its own and is quite pleasant.