Biochemical Unknown Help Please - (Apr/25/2011 )

Nelson dear!! I dont want to say it.. but you were doing many of unnecessary tests. Let's me start form the beginning..

So as I have said, because of the long period of time staying in the fridge and lab time, so of course the bacteria will eventually dies and therefore may give false result for the Gram stain. That is why you want to do Gram staining and all the EMB, MSA, PEA, TSA on the first day of Bio- unknown..

So as you said, you have figured out your Gram + is 3 and Gram - is O righ! DO NOT do all the tests for two unknown bacteria at the same time unless you know what the directions you want to go!! if you mess up the two and get them contaminated!! YOU ARE SCREWED!!!!!!!

Let's first take care of the Gram +
1. doing catalase test. IF bubble=> catalase + => ELIMINATE all of the organisms that are catalase- on your list.


2. test for starch. if you are sure that it is STARCH-, kick the B. sub.

3. Check for MSA. all growth with yellowish?!! on right track. so it should be S. au and S. epi?

4. DOING THE BLOOD AGAR FOR PATTERN OF MERMOLYSIS/ OR COAGULATE TEST... these are the only two ways that can differentiate between S. epi and S. au


Let's crack the Gram-

you dont have to do the catalase for the Gram- since all of them on the list are + for catalase.

1. Fermentation tube: glucose with Durham tube, sucrose. You dont have to do lactose, chech your MAC for patterns and color to decide if it is lactose + or not.
=> get the result, write it down, clearly and neatly.

2.Doing the SIM. After incubation: a. check if it forms black ppt
b. check for the motility
c. swab the surface of the sim deep with sterilize swab and drop one-2 drops of Kovacs reagant. If it is indole, then an immediately pinkish color appears ( if you get the indole +, lucky you because there are only two organisms on the list are indole +. In your case, we dont know if it is + or not... grrrr)

3. Doing MRVP. Read the steps and perform the MRVP. Carefully observe your MR- VP because this is important!!

4. doing citrate test!!!

5. nitrate tests is you still at this time have no idea what yours is.

6. Urea tests

Read all the result, write down clearly the results for each tests properly and you will figure out yours.... Also you have to check carefully for your biochemical tests.. In my case, the E.coli didnt grown metallic green on the MAC but all the biochemical tests were like pointing at the E.coli:). Fun huh??

Good luck and you can always buzz me on facebook if need help

Good luck!!

P/s: send you the data sheet for GRam -

One thing you should take into consideration: if you do not have practical experience, chances are some of the results can be false positives/negatives.
As pito pointed out - MSA is a fairly selective media for isolation of staphylococci, differential for mannitol fermentation - compare on the photos:
http://www.midlandstech.edu/science/kelleherk/225/labmaterials/sel_diff_media.html
But staphylococci are non-motile, so consider your SIM results, for example.