Culturing Oil eating Microbes?? - (Dec/17/2010 )

Hello out there.....

I'm not a Microbiologist so this is probably why the process below is
not working....
Any help or comments would be appreciated.

I am trying to culture oil eating microbes and measure their growth by
looking at the co2 levels they are emitting.

I know the normal thing would be to culture them in an agar dish and
score visually but I got the bright idea that measuring co2 emissions
would provide an earler and more sensitive indication of their growth.

Here is how I'm doing it:

Have a quart Ball fruit jar filled with distilled water about 2" below
full.
PH about 6.0-temp is about 70 degrees.
Purchased some Oil Eating Microbes from a science lab, added 1/2
teaspoon to the jar.
Added Butanol as the Hydrocarbon food. Added enough to create about a
1/4" oil slick on top of the jar.

Placed lid on jar.

Jar lid has an air tight inlet and outlet tube that connects to my co2
meter. Co2 meter will measure down to 50ppm.Co2 meter has an injector
pump and I am pulling air from the headspace of the jar, through the
co2 meter, back into the jar.

The incubation has been running 72 hours and I'm not seeing any real
elevation in co2 and no real visible indication of growth.Room c02 is
about 400ppm and the jar co2 is about 1600 ppm however it read that
almost immediately after I started the incubation and that reading has
actually dropped some and is not increasing.

I am using Butanol instead of a light grade oil for food. Light grade
oil is usually supplied with the microbes I bought but I am looking
for bugs that eat Butanol type (light) Hydrocarbons.

The jar is sealed where the normal science kit indicated leaving the
jar open , however I needed it to be sealed in order to trap the c02.
Also I figured there was enough O2 in the headspace of the jar for
some growth.
I have also considered the cO2 could actually choke the bugs but I
don't think co2 levels have ever got to that point yet.

Many have said add sugar or other nutrients to kick start growth but I
don't want to get a Heinz 57 colony growing. I only want to stimulate
Hydrocarbon specific bacteria.

Anybody have any thoughts, ideas, or suggestions?

If someone can help me with this I'll gladly pay a couple hundred
bucks for your time.

Thanks

Hola, only two things oil, lipids are hidrocarbon compounds, and if you don´t add sugars, bacteria use the hidrocarbon skelet of lipids,in fact in biotechnology the main growth medium Luria bertany (mor or less) LB only has aminoacidic substrate and natrium chloride. In fact I think that your medium has to be water, oil and any proteic extract as yeast extract or meat extract (as Bovril), If the strain hasn´t any auxotrophy for any aminoacid or vitamin the culture will grow at 37 ºC, but first you have to sterilize the medium, inoculate in sterility and sake, or stirr the culture during the process. Buena suerte.

Just a quick note:

you can start growing your culture on an agar plate and then select some colonies from that plate and add them to your fluid...
(just to check they are "alive" and also: the preserve some of them.. if you put all your sample in a fluid and it fails.. you will need to buy new one..)


And butanol is that a hydrocarbon? Isnt it an alcohol...?

And what bacteria did you get anyway?
70°C.. seems hot ...

I'm guessing that butanol is a pretty good disinfectant. It's an alcohol, not a hydrocarbon (alkane). I suspect you are killing your cells with butanol. It's not that dissimilar to isopropanol, which is commonly used as a topical disinfectant.

phage434 on Sat Dec 18 17:51:11 2010 said:

what I forgot to mention: it might be needed to let them adept to the butanol (if that is what you really want to break down/test==> I am thinking at breakdown of alcohols, biofuels? rahter then oil..).
So you might want to test different concentrations and start with adding it to some agar plates to see if they grow.
Another thing: you can also try to find the concentration that kills your microbes.. by checking it with a 96well, incubated with different cocentrations etc..

I offcourse do not know what material you have etc.. but you are rushing a bit with your current approach.