Measuring Bacteria

[Red Herring]

Hello! I'm a student in high school doing a science fair project. Because of that, I should probably preface this and say that this probably isn't too scientifically rigorous.... I'm doing the best I can, but I'm sure that best falls short of what's normally posted around here.

 

Anyway, the idea is to make pizza boxes recyclable. The oil from the pizza makes the fibers of cardboard stick together. This hampers the whole recycling process. I had the idea to try to use oil eating bacteria (famous mostly for their help cleaning up oil spills in the ocean) to get this oil off. My science teacher and I discussed for a long time how to measure this... It may be pretty easy to throw the bacteria on the cardboard, but it's another thing entirely to see if they're actually eating the oil. I suppose that there probably exist some device that could take a sample before and after of the cardboard and tell you how much oil is or isn't on it... Unfortunately, I'm a high school student, and I don't have access to that sort of technology.

 

What I do have access to is CO₂ Gas Sensors... And that's what we decided to use. Obviously bacteria do cellular respiration and produce carbon dioxide. We thought if you compared oily cardboard and clean cardboard, and both were exposed to the bacteria, you would be able to tell if the bacteria were eating the oil. If CO₂ levels were higher for longer in the oily cardboard, it would be because the bacteria found food and were able to survive longer, thus producing more CO₂ for longer. If the clean cardboard's carbon level was higher or roughly the same as the oily cardboard, then the microbes didn't eat the oil. I also threw in a control: oily cardboard with no microbes at all. In theory, this was supposed to just show what carbon levels are normally like, so the other two have something to compare to.

 

I'm having some problems, though. It seems that the probes (Vernier CO₂ Gas Sensors, for use with "LabQuests". Link can be found at the bottom of this post.) are too sensitive. I tested in the back of my teacher's room for 72 hours, and found there was a huge jump (something like 1800 ppm -> 5000 ppm, but I don't have my data on hand... I can try to post it later if it'll help) while students were in the room breathing out. I didn't think this was going to be a problem... The sensors are oblong tubes that fit down into a specially made plastic bottle. There are no other openings and the sensor fits perfectly in the bottle. I called Vernier. They suggested to really shorten the amount of time, and mentioned how sensitive the sensors were and that I might be getting a whole lot of outside pollution.

 

So I tried again. This time, I did the test for only twelve hours, overnight while no one was in the room. I wrapped plastic tarps around the area (a fume hood) where I'm testing so that outside carbon wouldn't be able to get in. I also put disinfected plastic trashcans over the top of each plastic bottle so that the bottles wouldn't be able to infect each other.

 

The results of that test don't make the slightest bit of sense to me. These say that the control group (cardboard without microbes) actually had the most carbon. Oil and microbes was second after this. Clean cardboard and microbes had the least carbon over the twelve hours. I can't make sense of what this means... The only thing I could think of was that my breathing messed it up. I was in the testing area for almost a minute longer setting things up before I placed the trashcan over the control probe than I was before I set the trashcans over the other two probes. Could my carbon have gotten caught under the bucket and thrown results off?

 

There are probably a couple of other things I should mention:

-I originally wanted to test one particular strain of oil-eating bacteria, Alcanivorax Borkumensis. Unfortunately, it (and other similar strains) are quite expensive, and once again, I don't have an abundance of resources. We ended up calling a company called "MicroSorb" that makes a product out of the bacteria, mainly to see if they could point us to somewhere that sells bacteria more cheaply. They didn't, but they did offer to send us some of their product for free. I took them up on the offer. The information on what bacteria actually comprise the stuff is proprietary, however, so I have no idea what microbes I'm dealing with... Though evidently, they are completely safe.

-MicroSorb requires water to work, so there's a bit of water in the bottom of the platic bottle (50 mL). According to Vernier, water can affect the probes. However, since I've shortened the testing time by so much, there shouldn't be nearly as much condensation on the probes.

-This stuff has a concentration of 20 billion microbes per gram, and I'm using a half gram. I figured that at this concentration, the MicroSorb would far outweigh any bacteria that might be found in the water.

 

Obviously I ought to test more, but I thought I'd get some advice here first. I'm very quickly running out of time to test. I've also omitted a lot of information, and tried to only include things that seem vital.

 

Any suggestions you could provide would be immensely appreciated. Anything would be helpful... Ideas for alternate ways to test, suggestions for how to get less outside carbon, etc.... Thank you so much!

 

Links

Vernier CO₂ Sensor http://www.vernier.com/products/sensors/co2-bta/

MicroSorb http://www.microsorb.org/MicroSorb_Environmental_Solutions,_LLC/Microsorb.org_Home.html

[CharonY]

I have two concerns. First is that the amount of CO2 actually produced under these conditions is quite low, so probably you are more likely to measure random fluctuations rather than true bacterial activity.

In fact, I do not expect too much of bacterial activity under the described conditions as they need quite a bit more to grow actively enough to emit significant amount of CO2.

With regards to oil, you should be aware that what you put on is most likely fat (as e.g. olive oil) and not crude oil or petroleum. Unless you enjoy toxic pizzas, of course. Alcanivorax is famous for being able to degrade petroleum (also slowly). Lipids, on the other hand, (i.e. fats) can be utilized by many bacteria with various efficiency.

 

However, if the only thing the bacteria have is cellulose (cardboard) and lipids, it will not sustain bacterial growth. Additional macronutrients (such as a nitrogen, phosphorus etc) in a suitable form for the bacterium in question are needed. And even then, respiratory activity is likely not to be very high if other parameters (such as pH, temp. osmolarity, micronutrients etc.) are not matched.

 

It will take way longer than a few days (IMO) to get stable CO2 readings that could be used. Sorry.

 

Even counting the bacteria directly is probably not very efficient (again, the conditions are not terribly suitable for growth). Another issue is that if you actually observe growth, it could be tricky to link it exclusively to the utilization of the fats alone.

Edited February 15, 2013 by CharonY