Bacteria that stain purple with the gram staining procedure are termed gram positive. Selection of empirical antibiotics based on gram stain finding. None of the classes of major chemical constituents of the walls of bacteria and yeasts is exclusive to organisms which give a gram positive reaction. To stain bacteria, a uniform smear is applied to a glass side and allowed to dry. In gram staining and afb staining we use alcohol or acidalcohol or acid as a decolorizer but in spore staining. Gram staining is an essential procedure used in the identification of bacteria and is frequently the only method required for studying their morphology. Gram staining is still the cornerstone of bacterial identification and taxonomic division. Microbiology lecture 2 gram positive and gram negative bacteria gram staining principle this second microbiology lecture will explain the properties of gram positive and gram negative bacteria.
It is also known as gram staining or gram s method. The relation of the gram stain to the cell wall and the ribonucleic acid content of the cell. Bacterial classification, structure and function introduction the purpose of this lecture is to introduce you to terminology used in microbiology. The filter gram stain was approximately 100fold more sensitive than the slide gram stain. Gram staining is a differential staining technique that differentiates bacteria into two groups. The gram stain and differentialstaining the gram test is. The most widely used staining procedure in microbiology is the gram stain, discovered by the danish scientist and physician hans christian joachim gram in 1884. Gram staining technique is the widely used differential staining technique in bacteriology. The latter are characterized by high lipid contents. A grease free glass slide is taken and a circle is marked one side of the slide using a wax pencil or a glass marker pen.
Cover different classification schemes for grouping bacteria, especially the use of the gram stain 2. Cellular responses of bacillus subtilis and escherichia. Bacterial characteristics gram staining video khan. Staining of bacterial cells objective to learn the techniques of smear preparation, gram staining, nigrosin staining and correlating the results of gram staining with koh test. Rod shaped bacteria are called bacilli singular bacillus. This suggests gramnegative bacteria are more leaky, causing these thinwalled lipidrich cells to. Cellular responses of bacillus subtilis and escherichia coli to the. Gram staining of bacteria from environmental sources. Marking the slide makes it convenient to identify the surface of the slide that.
The acid fast stain was discovered by ehrlich, who found that after staining. Learn to complete the gram stain protocol effectively. Smears or films of bacterial cultures and clinical specimens are usually fixed by heat, the slide being first throughly dried in air and then heated gently in a flame. Cell structure and quantitative gram stain of bacillus. The gram staining technique differentiates the mixed culture cells into two terms as gram positive bacteria and gram negative bacteria. The filter gram stain detected 2 to 100 bacteria, whereas the slide gram stain failed to detect less than. This smi describes the identification of bacillus species.
Learn to recognize bacterial shapes and arrangements. A guide to bacterial culture identification and results. Cellular responses of bacillus subtilis and escherichia coli to the gram stain. The name comes from the danish bacteriologist hans christian gram, who developed the technique.
At the end of gram staining, gram positive bacteria appear violet or purple whereas gram negative bacteria appear pink. It is a technique used to differentiate types of bacteria using certain physical and chemical characteristics of their cell walls. The terms positive and negative have nothing to do with. The staining characteristics of bacteria in the gram stain. The gram staining test for bacteria is one of the most important tests in microbiology and is often one of the first tests performed in the identification of bacteria. Selection of suitable culture media based on gram stain finding. Fluorescent gram stains wheat germ agglutinin wga is a carbohydratebinding lectin that preferentially binds to gram positive bacteria fig. The bacteria concentrated on the surface of the membrane are stained with a cationic dye toluidine blue that can be visualized by the naked eye. Gram negative bacteria do not retain the dye for two reasons they have an outer membrane getting in the way of the crystal violet and they lack peptidoglycan to retain the stain.
How to do a gram s stain test gram s stain is one of the most frequently used processes in identifying bacteria and is used daily in hospitals. Students prepare bacterial smears and preform gram, acidfast ziehlneelsen, and endospore stains. Introduction observation of bacteria with conventional bright field microscopy yields relatively little useful information. This was developed in 1884 by a danish bacteriologist, hans christian gram. For example, gram staining can help clinicians classify bacterial pathogens in a sample into categories associated with specific properties. Khan academy is a nonprofit with the mission of providing a free, worldclass education for anyone, anywhere. Prior to staining bacteria, a bacterial smear must be heat fixed onto a microscope slide. Principle the gram stain is used to classify bacteria on the basis of their forms, sizes, cellular morphologies, and gram reactions. The organisms described in this document are those which may be isolated from clinical material, although not all have been shown to cause human disease. Principle, procedure and results endospore staining.
Get a printable copy pdf file of the complete article 4. Gram staining is a diagnostic test that gives an early indication of potential bacteria through visualization of the bacteria. After heatfixing the smear, crystal violet is applied. Gram negative bacteria tend to be more resistant to certain antibiotics than gram positive bacteria.
After staining, the membrane is treated with ethanolacetic acid ph 2. Walls of gram positive bacteria are richer in mucopeptide than are those of gram negative bacteria. Bacteria are generally studied when fixed and stained. Understand the role of the gram stain as an important differential. Screening of the quality of the clinical specimens such as sputum that should contain many. 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. Microbiology gram stain for bacteria simply re pinning because i never see anything for medical lab techs on here. Laboratory perspective of gram staining and its significance in investigations of infectious diseases yunusa thairu 1, idris abdullahi nasir 1, yahaya usman 2 1 department of medical microbiology, university of abuja teaching hospital, gwagwalada, abuja, nigeria 2 department of medical laboratory science, faculty of medicine, ahmadu bello university, zaria, kaduna, nigeria. Gram stain or gram staining, also called gram s method, is a method of staining used to distinguish and classify bacterial species into two large groups. Learn for free about math, art, computer programming, economics, physics, chemistry, biology, medicine, finance, history, and more. This can result in areas that are under decolorized, thus organisms that are truly gram negative will appear gram positive.
Cell structure and quantitative gram stain of bacillus megaterium. The heat kills the bacteria and attaches the sample to the slide so that it does not easily wash away. The gram stain is a differential method of staining used to assign bacteria to one of two groups gram positive and gram negative based on the properties of their cell walls. Bacteria are very difficult to study microscopically unless stained. The gram stain is the most widely used staining procedure in bacteriology. It is a primary test which quickly and cost effectively divides bacteria into one of two types. A smear is a sample of bacteria suspended in a small amount of water on a slide. Rapid presumptive diagnosis of diseases such as bacterial meningitis. You can use 2 slides, 1 for each bacterium, or you can divide one slide in half and smear each bacterium on the divided slide. It is almost always the first test performed for the identification of bacteria. The gram staining method was first described in 1844 by the danish bacteriologist hans christian gram, after whom the test was named. Learn to recognize gram positive from gram negative bacteria. Gram staining 2 free download as powerpoint presentation. Gram staining is an empirical method of differentiating bacterial species into two large groups gram positive and gram negative based on the chemical and physical properties of their cell walls.
Gram staining is a simple procedure if done with precision and accuracy can guide in diagnosis of several bacterial infections. Objectives to learn the technique of smear preparation. It is called a differential stain since it differentiates between gram positive and gram negative bacteria. Endospore bacterial stain procedure science prof online. The gram stain easily divides bacteria into two groups, grampositive and gram negative, on the basis of their. Gram staining method, the most important procedure in microbiology, was developed by danish physician hans christian gram in 1884. The procedure is named for the person who developed the technique, danish bacteriologist hans christian gram.
Microbiology lecture 2 gram positive and gram negative. When colonies of gramnegative bacteria are treated with 3% koh solution, a slimy suspension is produced, probably due to. Pdf methods for confirming the gram reaction of gramvariable. The gram stain helps to differentiate the organism, whether it is gram positive or gram negative. Two basic methods provide foundations for differential staining and detection of bacteria. Summary in a strain of bacillus megaterium the gram reaction was investigated by comparing its effect on intact bacilli with its effects on bacilli treated with lysozyme in several different ways. The lysozymetreated bacteria varied from bacilli showing only polar separation of the cell wall from the protoplasm to protoplasts free from cell wall.
Rapid method for detection of grampositive and negative. To use and relate the gram stain to the study of bacterial cell morphology, and as an important step in the identification of a bacterial species. The primary stain of the gram s method is crystal violet. Gram staining allows for quick visualization of bacterial morphology and broad cellular distinction from a wide range of environmental samples. In bacteriology, gram positive bacteria are bacteria that give a positive result in the gram stain test, which is traditionally used to quickly classify bacteria into two broad categories according to their cell wall gram positive bacteria take up the crystal violet stain used in the test, and then appear to be purplecoloured when seen through an optical microscope. The relationship between the nature of the cell wall and. Learn to complete the simple stain protocol effectively. Gram staining lab introduction gram staining is a very important technique used in biology labs all over the world. Gram positive bacilli with clumps of free spores and bacteria distended with ovoid subterminal spores. To learn the techniques of gram staining, nigrosin staining and koh test. Other shapes will be considered later in the course. Pdf cellular responses of bacillus subtilis and escherichia coli to.
Gram staining tips page 2 of 2 some specimens on preparation will have areas on the slide that are thicker than others. This differential staining procedure separates most bacteria into two groups on the basis of cell wall composition. Gram stain, a widely used microbiological staining technique that greatly aids in the identification and characterization of bacteria. Laboratory perspective of gram staining and its significance in. Pdf bacillus is a predominant genus of bacteria isolated from tobacco. It was devised by a danish physician, hans christian gram, in 1884. Occurrence of a phosphoric ester in certain bacteria. Flood slide with crystal or gentian violet 60 seconds. This test differentiate the bacteria into gram positive and gram negative bacteria, which helps in the classification and differentiations of microorganisms. The gram reaction reflects fundamental differences in the biochemical and structural properties of bacteria.
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