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Yes, the age of the culture may influence the results of the stain.
With the current theory behind gram staining, it is thought that in gram-positive bacteria, the crystal violet and iodine combine to form a larger molecule that precipitates out within the cell. The alcohol/acetone mixture then causes dehydration of the multilayered peptidoglycan in the gram-positive call wall, thus decreasing the space between the molecules and causing the cell wall to trap the crystal violet-iodine complex within the cell. In the case of gram-negative bacteria, the alcohol/acetone mixture, being a lipid solvent, dissolves the outer membrane of the gram-negative cell wall (and may also damage the cytoplasmic membrane to which the peptidoglycan is attached). The thin layer of peptidoglycan is unable to retain the crystal violet-iodine complex and the cell is decolorized.
It is important to note that gram-positivity (the ability to retain the purple crystal violet-iodine complex) is not an all-or-nothing phenomenon but a matter of degree. There are several factors that could result in a gram-positive organism staining gram-negatively:
1. The method and techniques used. Overheating during heat fixation, over decolorization with alcohol, and even too much washing with water between steps may result in gram-positive bacteria losing the crystal violet-iodine complex.
2. The age of the culture. Cultures more than 24 hours old may lose their ability to retain the crystal violet-iodine complex.
3. The organism itself. Some gram-positive bacteria are more able to retain the crystal violet-iodine complex than others.
What else can I help you with?
What is gram variable bacilli?
Gram-variable bacilli are rod-shaped bacteria that do not consistently take up Gram stain, appearing either Gram-positive or Gram-negative under a microscope. This variability can result from factors such as the age of the culture, the composition of the cell wall, or the presence of certain conditions during staining. Gram-variable bacilli can include various species, some of which are pathogenic and associated with human diseases. Examples include certain strains of Corynebacterium and Mycobacterium.
Why would bacteria stain gram variable?
A gram-negative cell will lose its outer membrane and the peptidoglycan layer is left exposed. or it is best to use younger cells ( 12-24hr) because older gram positive bacteria are subject to break down of the cell wall by enzymes that are produced with age which may result ingram variable staining.
Can Gram-positive organism appear as Gram-negative?
Staphylococcus are Gram positive because, in Gram's test, the decolorizing agent (ETOH) cannot penetrate the thick cell walls, and the stain remains behind: hence Gram positive. I am not aware of a gram negative staph species and, considering the degree of mutation that would be needed to form such a strain, I do not believe that it's possible. I should add that a not-so-brief scan of the net showed me no Gram negative staph mentioned anywhere. So -- for now -- no. That said, let's see what the future holds.
What are the three basic Theories on the principle of gram staining reactions?
The gram stain is a basic differential stain used to determine if a bacterial cell is gram positive or negative. Gram positive cells have a thick peptidoglycan layer that will trap the crystal violet iodine crystalls and apear purple. Gram negative cells only have a thin peptidoglycan layer that allows the crystals to diffuse out of the cell and will only be seen with the application of a counterstain, such as safranin which turns the cells pink.
Can you tell how old a blood stain is?
No, the age of a blood stain cannot be determined solely by looking at it. Factors like environment, fabric type, and blood volume can affect the appearance of a stain. In forensic investigations, chemical testing and analysis are used to estimate the age of a blood stain.
What are 2 conditions in which an organism might stain gram variable?
An organism might stain gram variable due to the presence of a cell wall structure that is intermediate between gram-positive and gram-negative bacteria, often seen in some members of the Actinobacteria or Firmicutes phyla. Additionally, factors such as the age of the culture, where older cells may lose their ability to retain the crystal violet stain, or exposure to certain environmental stresses, such as changes in temperature or pH, can also lead to a gram variable result.
You preformed a gram stain on a sample from a pure culture of bacteria and observed a field of red and purple cocci adjacent cells were not always the same color what do you conclude?
This is a Gram Stain. If the technique was proper, the red rods are Gram-negative and the purple cocci are Gram-positive. This staining technique is used to help identify various bacteria. The Gram-positive bacteria that are purple hold the stain due to it's layered cell membrane. It contains a peptidoglycan layer that acts as a lattice trapping the crystal violet-Iodine dye complex.
Three factors that may affect gram stain reaction?
(i) The age of bacterial culture should not be more than 24 h. At older age cell loses Gram positivity and will appear as Gram negative. (ii) Application of heat during the fixation of smears is another important step. Too much heating during this step will lead to loss in, Gram positiveness. (iii) Overcrowding of cells in smear also affects the result, due to improper decolourization. (iv) Staining reagents should be freshly prepared. (v) In Gram staining decolourizing step is very important. To obtain satisfactory differentiation, the nature and the exposure time of decolourizing agent should be standardized with the material to be stained. Acetone alone is more powerful decolourizing agent than ethanol. (vi) It is also important not to allow a .bacterial smear to dry. There are many variations of original Gram staining procedure
What is gram variable bacilli?
Gram-variable bacilli are rod-shaped bacteria that do not consistently take up Gram stain, appearing either Gram-positive or Gram-negative under a microscope. This variability can result from factors such as the age of the culture, the composition of the cell wall, or the presence of certain conditions during staining. Gram-variable bacilli can include various species, some of which are pathogenic and associated with human diseases. Examples include certain strains of Corynebacterium and Mycobacterium.
How does culture age affect results of a spore stain?
The age of the culture used for a spore stain can impact the results by affecting the viability and sporulation of the organism. A young culture with actively growing cells is more likely to produce good spore stain results, while an older culture with decreased viability and sporulation may lead to unreliable staining outcomes. It is generally recommended to use a fresh culture for spore staining to obtain accurate and reliable results.
What are the factors that can result in variable gram reaction?
Genetics, Age of culture, type of growth medium, and technique used could result in a gram-variable reaction
When gram stain performed incubated for a longer period of time examination of the stain B cereus slides revealed a great deal of color variability ranging from an intense blue to shades of pink?
This variability in color could be due to over-decolorization during the gram staining process, causing the retention of crystal violet dye in Gram-positive organisms like B. cereus. It may also suggest differences in cell wall composition or age of the bacterial culture. To achieve more consistent results, ensure adherence to standardized gram staining procedures and proper timing during the decolorization step.
Why would bacteria stain gram variable?
A gram-negative cell will lose its outer membrane and the peptidoglycan layer is left exposed. or it is best to use younger cells ( 12-24hr) because older gram positive bacteria are subject to break down of the cell wall by enzymes that are produced with age which may result ingram variable staining.
When bacteria culture age does it appear gram negative or gram positive?
Typically, a negative result can be doubted, but a positive staining cannot be. This is because the vast majority of bacteria that do revert (not to say that the majority of bacteria DO revert) will go from positive to negative. This is due to a decreased average level of peptidoglycan synthesis in Agar culture, as opposed to natural function in vivo. The decreased level of peptidoglycan allows the ethanol/acetone decolorizer to more quickly flush the Crystal Violet from Gram+ cells, making them appear either weakly positive, or fully negative.
Why do gram negative cells stain pink?
Gram negative bacteria actually stain red because they have a less complex cell wall than Gram positive bacteria and they are easily decolorised by the decoloriser acetone alcohol. Hence they take up the colour of the counter stain safranin which is red during the Gram staining procedure.
Can Gram-positive organism appear as Gram-negative?
Staphylococcus are Gram positive because, in Gram's test, the decolorizing agent (ETOH) cannot penetrate the thick cell walls, and the stain remains behind: hence Gram positive. I am not aware of a gram negative staph species and, considering the degree of mutation that would be needed to form such a strain, I do not believe that it's possible. I should add that a not-so-brief scan of the net showed me no Gram negative staph mentioned anywhere. So -- for now -- no. That said, let's see what the future holds.
What are advantages and disadvantages of staining a sample?
Some cells produce gram-variable results. Some cells produce no results. The age of the culture can influence the results. It has limited usage in environmental biology and is not as good as molecular techniques.
