How to count bacteria colonies on an Easygel plate.
The Easygel method from Micrology Laboratories makes estimating bacteria in water or other sources very easy. I have no doubt it is the easiest and most useful method I've seen. It is specially good for students and people without special facilities because the method doesn't require any medium preparation or autoclaving etc. When I was a science teacher way back I tried to make some agar plates for my students but remember that the process was tedious and involved heating and storing agar medium.
Now its dead easy because with Easygel all you need is a bottle of medium, a pre treated dish (plate in Australia), a sterile dropper and a sterile sample bottle. All this comes when you purchase Easygel kits. The method is very cost effective because there is no preparation time and no expensive equipment required. I wish I knew about Easygel years ago (it hadn't even been invented then I'm sure).
Now for the real reason I started this page, how to count the bacteria colonies on a plate.
If there are only a few colonies then all of them can be counted. If you used 2.5 ml of the original sample to add to the Easygel bottle then that is a 2.5/100 fraction of 100 mls, or 1/40th of 100 mls. Colonies are usually reported as Colony Forming Units per 100 mls (CFU's/100 ml). Therfore to convert your count to CFU's per 100 ml multiply the count by 40.
But what if there are too many colonies to count? Then you have to resort to a little trick. Find a piece of clear acetate sheet, the type that is used for the clear cover of a booklet such as on a bound report. Scratch out a small square cm x cm grid using a ruler and a sharp point. You only need one square but a 2 x 2 grid is better.
Now place the grid over the culture plate, usually from the underside closest to the colonies. Count the number of colonies in each grid. Only include colonies on the edges of the squares if they are more than half way inside the square. Now move the grid to another position. Just move it without looking and leave it there for the next count. Don't deliberately choose areas where there are more colonies - or you will get an artificially high result. Write down each count then average them to get an average per square cm. There are close to 57.4 square cms in an Easygel dish so then multiply that average by 57.4 This will give an estimate of the number of colonies per dish. Now, how many mls of sample did you use? Again if it was 2.5 ml then the multiplying factor is 40. Here is an example. The dark colonies are E coli and the pink colonies are coliforms.
The more counts you can make the better. Move the grid randomly each time. In the lab if I'm getting fairly consistent results and there are lots of colonies I stop at around 10 counts otherwise go to 15 to 20 counts.
Here is a spreadsheet template (Excel) that you can use to perform all the calculations. Just enter individual squ. cm counts and sample size. It already has the calculations for E coli for the example above so you can see how it works. Bacteria colony calculator.
I'd probably recomend to the owner of this water supply that he doesn't drink the water without treatment!
Notes: You can use this same method for Aerobic plate count, just count the clear to colourless colonies on the Easygel plate after using the AOPC media. Also, if there are large numbers of colonies on a plate then you may have to draw 1/4 squ. cm grids on the clear acetate sheet and count colonies in each smaller grid. Just multiply by 4 to get one squ. cm counts.
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