Microorganisms found in pharmaceutical and healthcare conditions require identification in order to determine the particular species. This is important so that the origin of contamination can be assessed and the origin of contamination determined. This is typically performed by using a standing technique the Gram stain, which is based is really a type of “phenotypic identification method” also it undertaken so that the microbiologist can understand the general profile for microorganisms.
The first step of most identification schemes is to describe the colony and cellular morphology of the microorganism. Colony morphology is usually described by directly observing growth on agar, where the colony will appear as a particular shape (such because raised, crenated, spherical and so on) and the colony will have a particular color. Some microbiologists will attempt to identify the microorganism based on such visual identification. This is not normally encouraged because considerable experience is required to do this as well as the variety of microflora cannot be characterised with any degree of accuracy. Furthermore, you will of a microorganism are often dependent upon the kind of culture medium used. Nevertheless, the description of the morphology can assist with further stages of identification.
Mobile staining provides important information relating to the composition of the microbial cell wall structure, as well as the shape of the organism. Of these, the most frequently used method is the G stain.
The Gram stain method employed includes the four-step method: Crystal violet (primary stain); iodine (mordant); alcohol (decolorizer); and safranin (counter stain). Done correctly, Gram-positive organisms retain the crystal violet spot and appear blue; Gram negative microorganisms lose the crystal violet spot and contain only the counter-stain safranin and thus appear red. Common stumbling blocks in this method are that temperature fixation may cause Gram-positive cells to stain Gram-negative and older civilizations may give Gram-variable reaction; using a lot of decolorizer could result in a false Gram-negative result and not using enough decolorizer may yield a false Gram-positive result.
The Gram reaction is founded on the differences in the cell wall composition for the two cellular ‘groups’. The particular bacteria that retained the stain (the Gram-positive bacteria) have a higher peptidoglycan and lower lipid articles than those that do not retain the stain (the Gram-negative bacteria). The effect of the solvent is to dissolve the lipid layer in the cell wall of the Gram-negative bacteria, thereby causing the very violet to leach out; whereas for Gram-positive bacteria the solvent dehydrates the thicker cell walls, blocking any diffusion of the violet-iodine complex, which closes the skin pores of the cell and retains the particular stain. There are now several automated Gram stain devices available on the market that can slow up the labour requirement required when performing several multiple Gram stains and, possibly, improve accuracy.
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In addition to the difference based on cell wall, microscopic study of the stains allows the mobile shape to be determined. Bacteria generally fall into the categories of coccus (spherical), rod, vibrio (curved), spirilla (spiral) and plemomorphic (variable).