NEET MDS Lessons
General Microbiology
Immunoglobulin (Ig)
Immunoglobulins are glycoprotein molecules that are produced by plasma cells in response to an immunogen and which function as antibodies. The immunoglobulins derive their name from the finding that they migrate with globular proteins when antibody-containing serum is placed in an electrical field
FUNCTION
1. Immunoglobulins bind specifically to one or a few closely related antigens. Each immunoglobulin actually binds to a specific antigenic determinant. Antigen binding by antibodies is the primary function of antibodies and can result in protection of the host.
2. The significant biological effects are a consequence of secondary "effector functions" of antibodies.Phagocytic cells, lymphocytes, platelets, mast cells, and basophils have receptors that bind immunoglobulins. This binding can activate the cells to perform some function. Some immunoglobulins also bind to receptors on placental trophoblasts, which results in transfer of the immunoglobulin across the placenta. As a result, the transferred maternal antibodies provide immunity to the fetus and newborn.
STRUCTURE OF IMMUNOGLOBULINS
The basic structure of the immunoglobulins is illustrated in figure 2. Although different immunoglobulins can differ structurally, they all are built from the same basic units.
A. Heavy and Light Chains
All immunoglobulins have a four chain structure as their basic unit. They are composed of two identical light chains (23kD) and two identical heavy chains (50-70kD)
B. Disulfide bonds
1. Inter-chain disulfide bonds - The heavy and light chains and the two heavy chains are held together by inter-chain disulfide bonds and by non-covalent interactions The number of inter-chain disulfide bonds varies among different immunoglobulin molecules.
2. Intra-chain disulfide binds - Within each of the polypeptide chains there are also intra-chain disulfide bonds.
C. Variable (V) and Constant (C) Regions
When the amino acid sequences of many different heavy chains and light chains were compared, it became clear that both the heavy and light chain could be divided into two regions based on variability in the amino acid sequences. These are the:
1. Light Chain - VL (110 amino acids) and CL (110 amino acids)
2. Heavy Chain - VH (110 amino acids) and CH (330-440 amino acids)\(x = {-b \pm \sqrt{b^2-4ac} \over 2a}\)h the arms of the antibody molecule forms a Y. It is called the hinge region because there is some flexibility in the molecule at this point.
E. Domains
Three dimensional images of the immunoglobulin molecule show that it is not straight as depicted in figure 2A. Rather, it is folded into globular regions each of which contains an intra-chain disulfide bond (figure 2B-D). These regions are called domains.
1. Light Chain Domains - VL and CL
2. Heavy Chain Domains - VH, CH1 - CH3 (or CH4)
F. Oligosaccharides
Carbohydrates are attached to the CH2 domain in most immunoglobulins. However, in some cases carbohydrates may also be attached at other locations.
IMMUNOGLOBULIN FRAGMENTS: STRUCTURE/FUNCTION RELATIONSHIPS
Immunoglobulin fragments produced by proteolytic digestion –
A. Fab
Digestion with papain breaks the immunoglobulin molecule in the hinge region before the H-H inter-chain disulfide bond Figure 6. This results in the formation of two identical fragments that contain the light chain and the VH and CH1 domains of the heavy chain.
Antigen binding – These fragments are called the Fab fragments because they contained the antigen binding sites of the antibody. Each Fab fragment is monovalent whereas the original molecule was divalent. The combining site of the antibody is created by both VH and VL.
B. Fc
Digestion with papain also produces a fragment that contains the remainder of the two heavy chains each containing a CH2 and CH3 domain. This fragment was called Fc because it was easily crystallized.
Effector functions – The effector functions of immunoglobulins are mediated by this part of the molecule. Different functions are mediated by the different domains in this fragment .
Treatment of immunoglobulins with pepsin results in cleavage of the heavy chain after the H-H inter-chain disulfide bonds resulting in a fragment that contains both antigen binding sites . This fragment is called F(ab’)2because it is divalent. The Fc region of the molecule is digested into small peptides by pepsin. The F(ab’)2binds antigen but it does not mediate the effector functions of antibodies.
Classification:
Neutrophiles (pH = 7.0)
- P. aeruginosaqo
- Clostridium sporogenes
- Proteus species
Acidophiles (pH < 7.0)
- Thiobacillus thiooxidans
- Sulfollobus acidocaldaarius
- Bacillus acidocaldarius
Alkaliphiles (pH > 7.0)
- Nitrobacter species
- Streptococcus pneumoniae
Enzymes:
Serum lysozyme:
Provides innate & nonspecific immunity
Lysozyme is a hydrolytic enzyme capable of digesting bacterial cell walls containing peptidoglycan
• In the process of cell death, lysosomal NZs fxn mainly to aulolyse necrotic cells (NOT “mediate cell degradation”)
• Attacks bacterial cells by breaking the bond between NAG and NAM.
• Peptidoglycan – the rigid component of cell walls in most bacteria – not found in archaebacteria or eukaryotic cells
• Lysozyme is found in serum, tears, saliva, egg whites & phagocytic cells protecting the host nonspecifically from microorganisms
Superoxide dismutase: catalyzes the destruction of O2 free radicals protecting O2-metabolizing cells against harmful effects
Catalase:
- catalyzes the decomposition of H2O2 into H2O & O2
- Aerobic bacteria and facultative anaerobic w/ catalase are able to resist the effects of H2O2
- Anaerobic bacteria w/o catalase are sensitive to H2O2 (Peroxide), like Strep
- Anaerobic bacteria (obligate anaerobes) lack superoxide dismutase or catalase
- Staph makes catalase, where Strep does not have enough staff to make it
Coagulase
- Converts Fibronogen to fibrin
• Coagulase test is the prime criterion for classifying a bug as Staph aureus – from other Staph species
• Coagulase is important to the pathogenicity of S. aureus because it helps to establish the typical abscess lesion
• Coagulase also coats the surface w/ fibrin upon contact w/ blood, making it harder to phagocytize
The cell cycle
1) Labile cells (GI tract, blood cells)
- Described as parenchymal cells that are normally found in the G0 phase that can be stimulated to enter the G1
- Undergo continuous replication, and the interval between two consecutive mitoses is designated as the cell cycle
- After division, the cells enter a gap phase (G1), in which they pursue their own specialized activities
• If they continue in the cycle, after passing the restriction point (R), they are committed to a new round of division
• The G1 phase is followed by a period of nuclear DNA synthesis (S) in which all chromosomes are replicated
• The S phase is followed by a short gap phase (G2) and then by mitosis
• After each cycle, one daughter cell will become committed to differentiation, and the other will continue cycling
2) Stable cells (Hepatocytes, Kidney)
- After mitosis, the cells take up their specialized functions (G0).
- They do not re-enter the cycle unless stimulated by the loss of other cells
3) Permanent cells (neurons)
- Become terminally differentiated after mitosis and cannot re-enter the cell cycle
- Which cells do not have the ability to differentiate -> Cardiac myocytes
Measurement of Bacterial of Growth
A convenient method is to determine turbidity by photoelectric colorimeter or spectrophotometer.
The cell number can be counted as total cell number as well as viable count. Viable Count Viable number of bacteria can be counted by inoculating the suspension onto solid growth medium and counting the number of colonies. Since each colony is the end product of one viable bacterium, their count gives the number of viable bacteria in the suspension.
Total number of bacteria can be ascertained in specially designed chambers such as Coulter counter.
THE PLASMIDS
The extrachromosomal genetic elements, called as plasmids are autonomously replicating , cyclic ,double stranded DNA molecules which are distinct from the cellular chromosome
Classification
Plasmids can be broadly classified as conjugative and nonconjugative.
Conjugative plasmids are large and self-transmissible i.e. they have an apparatus through which they can mediate their own transfer to another cell after coming in contact with the same. Example: RF and certain bacteriocinogen plasmids.
Nonconjugative plasmids are small in size and can be mobilised for transfer into another cell only through the help of a conjugative plasmid. To this group belong some ‘r’ determinants and few bacteriocinogenic plasmids. Plasmids can also be transferred without cell contact by the process of transfection.
Properties of plasmids
Double stranded DNA , Autonomously replicate in host cell, Plasmd specific, Free DNA is transferred b transfection
Significance of Plasmids :The spread of resistance to antibiotics is one such well known example. These also play an important role in the geochemical cycle by spreading genes for the degradation of complex organic compounds.
DISINFECTION AND STERILIZATION
• Sterilization is the best destruction or com removal_of all forms of micro organisms.
• Disinfection is the destruction of many microorganisms but usually the b spores.
• Antisepsis is the destruction or inhibition of microorganisms in living tissues thereby limiting or preventing the harmful effect of infection.
• Astatic Agent would only inhibit the growth of microorganisms (bacteriostatic, fungistatic, sporostatic).
• Acidal agent would kill the microorganism (bactericidal. virucidal, fungicidal)
• Sterilants are the chemicals which under controlled conditions can kill sporinQ bacteria.