Help on bacterial identification using biochemical tests - (Apr/19/2008 )
Greetings everybody. I am new to this board. I hope I am not asking something that is old.
In my final year project I am tasked to identify 2 unknown bacteria. I am to assume that I don't know what these two samples are (it is m.luteus and e.faecealis) and I am to find out their identity.
Now, people would tell me to preform DNA extraction and followed by PCR and gel extraction and DNA sequencing, but it just happens that I am VERY HORRIBLE at precise handwork. I am born with such clumsy hands that I can only accurately preform easier tasks like streaking microbes on selective agars or inoculating broths (I should have known that being a scientist is mostly about handwork skills. Groan...)
Perhaps I will do the molecular biology tests later, with the help of a fellow colleague, but for now I would like to stick with what I do best, and besides I am dying to know how do microbiologists in the past manage to identify bacteria without the use of DNA sequencing.
The problem is I have no idea how to go with this task. I don't have a good source of which I can base my work on. I have heard from many microbiologists of consulting the Bergey's Manual of Determinative Bacteriology, but the book is unavailable in the library and I just' can't afford to buy one.
Common sense tells me that I should start with the most distintive feature of the unknown samples. For example, the m.luteus sample is gold-pigmented and gram-positive, and thus I can eliminate the rest of the gram-negative and non-pigmented bacterial and proceed with the next distinctive feature which is its spherical shape (correct me if I am wrong?). But I don't have a complete list of all bacteria arranged by gram cell wall type, shape, endospore-forming ability and the like.
Does anyone know of a complete list that I can use in my research?Any help would be greatly appreciated.
In my final year project I am tasked to identify 2 unknown bacteria. I am to assume that I don't know what these two samples are (it is m.luteus and e.faecealis) and I am to find out their identity.
Now, people would tell me to preform DNA extraction and followed by PCR and gel extraction and DNA sequencing, but it just happens that I am VERY HORRIBLE at precise handwork. I am born with such clumsy hands that I can only accurately preform easier tasks like streaking microbes on selective agars or inoculating broths (I should have known that being a scientist is mostly about handwork skills. Groan...)
Perhaps I will do the molecular biology tests later, with the help of a fellow colleague, but for now I would like to stick with what I do best, and besides I am dying to know how do microbiologists in the past manage to identify bacteria without the use of DNA sequencing.
The problem is I have no idea how to go with this task. I don't have a good source of which I can base my work on. I have heard from many microbiologists of consulting the Bergey's Manual of Determinative Bacteriology, but the book is unavailable in the library and I just' can't afford to buy one.
Common sense tells me that I should start with the most distintive feature of the unknown samples. For example, the m.luteus sample is gold-pigmented and gram-positive, and thus I can eliminate the rest of the gram-negative and non-pigmented bacterial and proceed with the next distinctive feature which is its spherical shape (correct me if I am wrong?). But I don't have a complete list of all bacteria arranged by gram cell wall type, shape, endospore-forming ability and the like.
Does anyone know of a complete list that I can use in my research?Any help would be greatly appreciated.
I had a project like that. I used Bergey's Manual of Systematic Bacteriology for that information.
Is there any way I can read the manual online for free (or at least, with a small fee)? The closest thing I have is the Determinative Bacteriology manual at the national library and it's like 30, 40 years old. Certainly NOT what I would like to cite as my reference in my thesis, given the pickiness of my supervisors. 
has your lab no acces to this document online?
Most schools and labs have acces to scientific databases.
There should be some sort of book you can use ? if your lab does research in the field of microbiology, they sure must have such book? or have connections with a school that has such book?
I'll check if I can find an online version in our database, but I am not sure we have this book in pdf.
Not sure what level of education you are pursuing. Your question betrays a very elementary level of micro awareness. My advice is to get one the net and learn something about these microbes. For example, Micrococcus luteus is, as you say a Gram postive coccus. Unlike many other such microbes it is oxidase positive and a strict aerobe.
You sure don't need a complete set of Bergey's volumes to establish a confirm id for such well-known bugs.
Does anyone know of a complete list that I can use in my research?
There is really only one generally accepted answer for that.
Bergey's Manual of Determinative Bacteriology 9th edition is a single paperbound volume for $110.
http://www.bergeys.org/pubinfo.html
You could try asking your microbiology instructor if the school would buy a copy. However, that might take more time than you have.
you could do an API 20E, but that would only ID the GNR (E. faecalis)
What is API 20E?
The API 20E is a biochemical strip test that identifies enterobacteriaceae. Basically, it looks like a 12 inch plastic strip that has tiny little cuvets or tubes attached to it. each tube has a chemical in it and you disperse the mystery bacteria into each cuvet. after incubating it overnight at 37 degrees, you observe the chemical reaction, record the results, and then look up in an index your results and it tells you what bacteria you have.
If you were given an identification project in your final year, then presumably you must have taken a microbiology lab course where you did biochemical tests.
Go take a look in your lab manual. If you don't have it anymore, borrow any microbiology lab manual from your library. I'd say all basic microbiology lab manuals have procedures for these biochemical tests.
The accompanying microbiology textbook (for lectures) also discuss each group of bacteria. You'll see that the primary grouping is based on Gram stain reaction.
Then see how they further subcategorized each group of bacteria.
A lot of microbiology manuals actually have flow charts to help you determine which tests to do. They'd start with Gram stain, usually followed by oxygen requirement tests.
You have two unknown bacteria and you have no idea what their genera are (at least that's what I got from your first post). The Bergey's Manual is most helpful only when you're already at that stage when you already know the genus of your unknown. Bergey's has tons of tables per genus that shows the different biochemical characteristics of each species.