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GATE BT-2017 Answer Key | Solutions Discussion
(02-20-2017, 01:10 AM)swarnava Wrote:
(02-19-2017, 04:39 PM)sngm Wrote: B option is  correct bcoz lps has rol in endotoxin not in staning....

As you will see in Chapter 4, different kinds of bacteria react
differently to the Gram stain because structural differences in
their cell walls affect the retention or escape of a combination of
crystal violet and iodine, called the crystal violet–iodine (CV–I)
complex. Among other differences, gram-positive bacteria have
a thicker peptidoglycan (disaccharides and amino acids) cell wall
than gram-negative bacteria. In addition, gram-negative bacteria
contain a layer of lipopolysaccharide (lipids and polysaccharides)
as part of their cell wall (see Figure 4.13, page 85). When applied
to both gram-positive and gram-negative cells, crystal violet and
then iodine readily enter the cells. Inside the cells, the crystal violet
and iodine combine to form CV–I. This complex is larger than
the crystal violet molecule that entered the cells, and, because of its
size, it cannot be washed out of the intact peptidoglycan layer of
gram-positive cells by alcohol. Consequently, gram-positive cells
retain the color of the crystal violet dye. In gram-negative cells,
however, the alcohol wash disrupts the outer lipopolysaccharide
layer, and the CV–I complex is washed out through the thin layer
of peptidoglycan. As a result, gram-negative cells are colorless
until counterstained with safranin, after which they are pink. OPTION A IS CORRECT REFER TORTORA MICROBIOLOGY PAGE 69
Gram Stain Mechanism:



Gram Positive Cell Wall:



Gram-positive bacteria have a thick mesh-like cell wall which is made up of peptidoglycan (50-90% of cell wall), which stains purple. Peptidoglycan is mainly a polysaccharide composed of two subunits called N-acetyl glucosamine and N-acetyl muramic acid. As adjacent layers of peptidoglycan are formed, they are cross linked by short chains of peptides by means of a transpeptidase enzyme, resulting in the shape and rigidity of the cell wall. The thick peptidoglycan layer of Gram-positive organisms allows these organisms to retain the crystal violet-iodine complex and stains the cells as purple.

Lipoteichoic acid (LTA) is another major constituent of the cell wall of Gram-positive bacteria which is embedded in the peptidoglycan layer. It consists of teichoic acids which are long chains of ribitol phosphate anchored to the lipid bilayer via a glyceride. It acts as regulator of autolytic wall enzymes (muramidases: Bacterial enzymes located in the cell wall that cause disintegration of the cell following injury or death.)







Medical Relevance of Gram Positive Cell Wall:



LTA also has antigenic properties that stimulate specific immune responses when it is released from the cell wall after cell death. Cell death is mailnly due to lysis induced by lysozymal activities, cationic peptides from leucocytes, or beta-lactam antibiotics.



Gram Negative Cell Wall:



Gram-negative bacteria have a thinner layer of peptidoglycan (10% of the cell wall) and lose the crystal violet-iodine complex during decolorization with the alcohol rinse, but retain the counter stain Safranin, thus appearing reddish or pink. They also have an additional outer membrane which contains lipids, which is separated from the cell wall by means of periplasmic space.







Medical Relevance of Gram Negative Cell Wall:



The cell wall of Gram-negative bacteria is often a virulence factor that enables pathogenic bacteria to cause disease. The virulence of Gram-negative bacteria is often associated with certain components of the cell wall, in particular, the lipopolysaccharide ( otherwise known as LPS or endotoxin). In humans, LPS elicits an innate immune response characterized by cytokine production and activation of immune system. Inflammation occurs as a result of cytokine production, which can also produce host toxicity.



Stain Reaction:



The four basic steps of the Gram Stain are:





1) Application of the primary stain Crystal Violet (CV) to a heat-fixed smear of bacterial culture.

CV dissociates in aqueous solutions into CV+ and Cl – ions. These two ions then penetrate through the cell wall and cell membrane of both Gram-positive and Gram-negative cells. The CV+ ions later interacts with negatively charged bacterial components and stains the bacterial cells purple.




2) Addition of Gram’s Iodine.

Iodine (I – or I3 –) acts as a mordant and as a trapping agent. A mordant is a substance that increases the affinity of the cell wall for a stain by binding to the primary stain, thus forming an insoluble complex which gets trapped in the cell wall. In the Gram stain reaction, the crystal violet and iodine form an insoluble complex (CV-I) which serves to turn the smear a dark purple color. At this stage, all cells will turn purple.



3) Decolorization with 95% ethyl alcohol.

Alcohol or acetone dissolves the lipid outer membrane of Gram negative bacteria, thus leaving the peptidoglycan layer exposed and increases the porosity of the cell wall. The CV-I complex is then washed away from the thin peptidoglycan layer, leaving Gram negative bacteria colorless.



On the other hand, alcohol has a dehydrating effect on the cell walls of Gram positive bacteria which causes the pores of the cell wall to shrink. The CV-I complex gets tightly bound into the multi-layered, highly cross-linked Gram positive cell wall thus staining the cells purple.



The decolori.........
[+] 1 user Likes sngm's post
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Ab v clear n ho aapko to plz click....

http://vlab.amrita.edu/index.php/images/...=208&cnt=1
[+] 1 user Likes sngm's post
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(02-20-2017, 01:58 AM)sngm Wrote: Ab v clear n ho aapko to plz click....  

http://vlab.amrita.edu/index.php/images/...=208&cnt=1

alcohol or acetone dissolves the lipid outer membrane of Gram negative bacteria, thus leaving the peptidoglycan layer exposed and increases the porosity of the cell wall. The CV-I complex is then washed away from the thin peptidoglycan layer, leaving Gram negative bacteria colorless.
 you only posted this and it suggests that the outer lps layer is dissolved by alcohol tratment so in question the reason is obviously correct. if the outer lps layer was not dissolved the gram negative would nt have been colorless and counter stained with saffranin
[+] 1 user Likes swarnava's post
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(02-20-2017, 05:55 PM)swarnava Wrote:
(02-20-2017, 01:58 AM)sngm Wrote: Ab v clear n ho aapko to plz click....  

http://vlab.amrita.edu/index.php/images/...=208&cnt=1

alcohol or acetone dissolves the lipid outer membrane of Gram negative bacteria, thus leaving the peptidoglycan layer exposed and increases the porosity of the cell wall. The CV-I complex is then washed away from the thin peptidoglycan layer, leaving Gram negative bacteria colorless.
 you only posted this and it suggests that the outer lps layer is dissolved by alcohol tratment so in question the reason is obviously correct. if the outer lps layer was not dissolved the gram negative would nt have been colorless and counter stained with saffranin
Itss ok......thnkxxx
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I guess I'll get the score around 45. Do you think I will clear GATE.
Can CutOff go up to 50?
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(02-22-2017, 02:18 AM)Sona Wrote: I guess I'll get the score around 45. Do you think I will clear GATE.
Can CutOff go up to 50?
May be.....bcoz after 29 questions ....still Wait for confirmation...... My marks is 44.......then ....cutoffs ..wil be high 55~60.
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(02-22-2017, 02:28 AM)sngm Wrote:
(02-22-2017, 02:18 AM)Sona Wrote: I guess I'll get the score around 45. Do you think I will clear GATE.
Can CutOff go up to 50?
May be.....bcoz after 29 questions ....still Wait for confirmation...... My marks is 44.......then ....cutoffs ..wil be high 55~60.

55-60?? Shhit. Then toh I wouldn't clear it.
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(02-22-2017, 02:32 AM)Sona Wrote:
(02-22-2017, 02:28 AM)sngm Wrote: May be.....bcoz after 29 questions ....still Wait for confirmation...... My marks is 44.......then ....cutoffs ..wil be high 55~60.

55-60?? Shhit. Then toh I wouldn't clear it.
Then i also.......
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(02-22-2017, 02:35 AM)sngm Wrote:
(02-22-2017, 02:32 AM)Sona Wrote: 55-60?? Shhit. Then toh I wouldn't clear it.
Then i also.......

what was the previeous 5 years cut off?
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(02-22-2017, 03:41 AM)Akshay shinde Wrote:
(02-22-2017, 02:35 AM)sngm Wrote: Then i also.......

what was the previeous 5 years cut off?
Wahi koi ....38 ...---> 27 .. Marks only
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