The Identities Of These Unknown Microorganisms

Published Date: 02 Nov 2017

http://books.google.com.my/books?id=NN663e2SM04C&printsec=frontcover&dq=chemical+and+physical+signature+for+microbial+forensic&hl=en&sa=X&ei=CpGAUbvoIM_yrQfqt4G4CQ&ved=0CDAQ6AEwAA

http://books.google.com.my/books?id=ZAZ-zI_PCqcC&printsec=frontcover&dq=manual+for+identification+of+medical&hl=en&sa=X&ei=HJGAUa2WBcqHrAfn54GIBg&sqi=2&ved=0CC0Q6AEwAA

http://books.google.com.my/books?id=f-VpAAAAMAAJ&q=biochemical+test+for+identification+of+medical+bacteria&dq=biochemical+test+for+identification+of+medical+bacteria&hl=en&sa=X&ei=NpGAUd3HFY7orQfo84HwCg&ved=0CC0Q6AEwAA

Identification means matching characteristics of an "unknown" organism to lists of known organism through clinical lab identification. Several tests are carried out to determine the characteristics of unknown microorganism and see how much it matches with the known microorganism. Identification of pathogens is crucial as it makes effective treatment possible.

There are several methods of identifying unknown microorganism. There are determining the morphology of microorganism, differential staining on microorganism, biochemical test, serology, phage typing, flow cytometry, DNA fingerprinting and Ribosomal RNA sequencing.

Determining the morphology of microorganism refers to the basic shape (cocci, rods, long, filamentous branched cells, comma shaped and spiral cells), the size (ranging from 10 E - 9 to 10E-4) , arrangement characteristic, presence or absence of flagella on microorganism, presence of endospores and etc.

Differential staining on microorganism refer to gram staining. Gram staining is an important technique in comparing basic morphological features, to strain the cell wall of the microorganism to carry out the test. The Gram stain is often chosen as the first and prior step in the identification of unknown microorganism. Staining with a water-soluble dye called crystal violet, decolorization, and counterstaining, usually with safranin are the three processes used in Gram staining. The process involves three steps:

First, unknown cell is stained with crystal violet dye. Next, iodine and potassium iodide is added on the cell to form a complex of crystal violet and iodine. This complex is insoluble in water.

Ethanol, which function as decolorizer is added to the sample cell. It will dehydrate, then shrink and tightened the peptidoglycan layer of the sample cell. The crystal violet-iodine complex can’t penetrate tightened peptidoglycan layer of Gram positive cell. This gives blue-violet colour of sample cell. While Gram negative cell, which has a thinner peptidoglycan layer doesn’t retain the colour of crystal violet-blue complex and thus decolourised.

Lastly safranin is added to the sample cell as counter stain, staining sample cell red. Since the colour of safranin is lighter than the colour crystal violet, it does not disrupt the purple color in Gram positive cells. However, the decolorized Gram negative cells are stained red.

Biochemical test is a series of tests carried out to test the ability of a microbe to produce certain enzymes, to test the ability to make use of certain nutrients and to test the waste products produced through the test. It is a good test as it gives a clear mapping of identification of unknown microorganism.

Serology is a technique works when microorganism enters an organism’s body, its body will form antibodies as defencing agent. A solution of known antibodies can be prepared first hand and treat on unknown bacterium.

Phage typing is a technique which uses specific viruses to attack specific bacteria and causes lysis of the bacteria that are responsive to them. They can be mixed with several likely antisera on slide. First the unknown microorganism to be identified is cultured on agar plates. Then drop of virus will be dripped on it. A clear area formed, namely plaque, when the unknown microorganism is responsive to particular phage. Phage typing helps to narrow down the area of identification of unknown microorganism.

Flow cytometry is a biomarker test using differences in electrical conductivity between microorganism species to identify unknown microorganism. A mixture of cells is first treated to label cells that have certain antigens with fluorescent-antibody markers. Then the cell mixture leaves nozzle in droplet. A laser beam is set parallel to it and it strikes each droplet. Fluorescence detector identifies fluorescent cells by fluorescent light by emitted by cell. Electrode gives positive charge to identified cells. As cells drop between electrically charged plates, the cells with a positive charge move closer to the negative plate. Thus the separated cells fall into different collection tubes.

DNA fingerprinting is using techniques such as polymerase chain reaction (i.e. RAPD-PCR) to study the DNA sequences of an unknown microorganism in order to identify them. It may also use complex molecular biology method andrestriction enzymes (i.e. pulsed field gel eletrophoresis) to study the fingerprinting pattern of unknown microorganism and compare it with known microorganism. The closeness of the relationship between two organisms is directly proportional the the similarity of the fingerprint patterns of the organisms.

Ribosomal RNA sequencing is another way of identifying the characteristics of unknown microorganisms. Ribosomal RNA is believed to be stable and alter less than other nucleic acids over the time. Ribosomal RNA of various microbes is determined and then compared. The similarity of rRNA sequencing of different organisms is believed to trace back in the past the two organisms branched off a common ancestor.