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Being able to differentiate bacterial species is important for a host of reasons, from diagnosing infection or checking food safety, to identifying which species it is that gives a cheese it’s fantastic character. Bacterial species, and even specific strains can be differentiated using a number of molecular techniques such as PCR, quantitative PCR, genome sequencing and mass spectrometry. But even without getting into the molecular nitty gritty, there are phenotypic differences between groups of bacteria that can be used to differentiate them. This includes characteristics like their shape (bacilli vs cocci for example), growth in particular nutrients and preference for high or low oxygen environments. Depending on the characteristic being studied, bacterial species may be broken down into broad groups, but taken together this information can narrow the possible identities greatly. One such useful classification – if a bacterium is Gram positive or Gram negative - is based on the structure of bacterial cell walls.
Difference in structure of Gram positive vs Gram negative bacteria
Gram positive bacteria have a thick peptidoglycan layer and no outer lipid membrane whilst Gram negative bacteria have a thin peptidoglycan layer and have an outer lipid membrane.
Gram positive vs Gram negative stainGram stain procedure - Preparing a sample
If preparing your slide from a liquid bacterial culture: Dab a small drop culture onto the slide using a sterile loop. Gently smear the droplet in a circular motion into an area of approximately 1 cm diameter. For very dense cultures it may be necessary to pre-dilute your culture to ensure individual bacterial cells can be seen under a microscope following staining.If the source material is from a bacterial plate:Resuspend a loop of colony material in sterile phosphate buffered saline (PBS) and then proceed as for a liquid culture.
Gram stain procedure - Gram staining a sample
Gram positive vs Gram negative colorGram positive bacteriaGram positive bacteria have a distinctive purple appearance when observed under a light microscope following Gram staining. This is due to retention of the purple crystal violet stain in the thick peptidoglycan layer of the cell wall. Examples of Gram positive bacteria include all staphylococci, all streptococci and some listeria species. Gram negative bacteria
Reference 1. Gram, H.C. (1884). "Über die isolierte Färbung der Schizomyceten in Schnitt- und Trockenpräparaten". Fortschritte der Medizin (in German). 2: 185–189. Find this article helpful? Download a copy here. Correction: The article erroneously stated that enterococci were examples of Gram negative bacteria, this was updated on June 6, 2022 to correctly identify enterobacter species as examples of Gram negative bacteria.
Gram Staining is the common, important, and most used differential staining techniques in microbiology, which was introduced by Danish Bacteriologist Hans Christian Gram in 1884. This test differentiate the bacteria into Gram Positive and Gram Negative Bacteria, which helps in the classification and differentiations of microorganisms. Principle of Gram StainingWhen the bacteria is stained with primary stain Crystal Violet and fixed by the mordant, some of the bacteria are able to retain the primary stain and some are decolorized by alcohol. The cell walls of gram positive bacteria have a thick layer of protein-sugar complexes called peptidoglycan and lipid content is low. Decolorizing the cell causes this thick cell wall to dehydrate and shrink, which closes the pores in the cell wall and prevents the stain from exiting the cell. So the ethanol cannot remove the Crystal Violet-Iodine complex that is bound to the thick layer of peptidoglycan of gram positive bacteria and appears blue or purple in color. In case of gram negative bacteria, cell wall also takes up the CV-Iodine complex but due to the thin layer of peptidoglycan and thick outer layer which is formed of lipids, CV-Iodine complex gets washed off. When they are exposed to alcohol, decolorizer dissolves the lipids in the cell walls, which allows the crystal violet-iodine complex to leach out of the cells. Then when again stained with safranin, they take the stain and appears red in color. Reagents Used in Gram Staining
Procedure of Gram Staining
InterpretationGram Positive: Blue/Purple Color ExamplesGram Positive Bacteria: Actinomyces, Bacillus, Clostridium, Corynebacterium, Enterococcus, Gardnerella, Lactobacillus, Listeria, Mycoplasma, Nocardia, Staphylococcus, Streptococcus, Streptomyces ,etc. AnimationDownload Animation from Below: Similar Posts:
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