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Comprehensive Study Notes: Immunology, Virology, Bacterial Growth, and Antimicrobial Agents

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Viral Replication Pathways

Overview of Viral Structure and Replication

Viruses are acellular infectious agents that require host cells for replication. They possess genetic material (DNA or RNA) and a protein coat (capsid), with some viruses also having an envelope and spike proteins that enhance virulence.

  • All viruses contain: Genetic material and a capsid.

  • Optional structures: Envelope and spike proteins (increase virulence).

  • Main goals of viruses: Replicate genetic material and assemble new viral particles.

General Steps of the Lytic Cycle

  1. Attachment: Virus binds to host cell surface.

  2. Uncoating: Viral capsid is removed, releasing genetic material into the host cell.

  3. Assembly: New viral genomes and proteins are synthesized and assembled into new virions.

  4. Lysis: Host cell bursts, releasing new viruses.

Lysogenic Cycle (Lysogenesis)

  • Viral genome integrates into host chromosome and replicates with the cell.

  • Allows the virus to evade the immune system.

  • Can revert to the lytic cycle under certain conditions (e.g., weakened immunity).

Viral Genome Types and Replication Pathways

  • Double-Stranded DNA (DS DNA) Virus:

    • DS DNA enters cell; host DNA-dependent RNA polymerase (DDRP) transcribes (-) strand to mRNA.

    • mRNA is translated by ribosomes to make viral proteins.

    • DNA-dependent DNA polymerase (DDDP) replicates viral DNA.

  • Single-Stranded (+) DNA Virus:

    • DDDP synthesizes (-) DNA from (+) DNA.

    • DDRP transcribes (-) DNA to mRNA.

    • mRNA is translated; DDDP replicates both DNA strands.

  • Single-Stranded (-) DNA Virus:

    • DDRP transcribes (-) DNA to mRNA.

    • mRNA is translated; DDDP synthesizes (+) DNA, then (-) DNA.

  • Double-Stranded RNA (DS RNA) Virus:

    • (+) RNA is translated to make viral proteins and RNA-dependent RNA polymerase (RDRP).

    • RDRP replicates viral RNA.

  • Single-Stranded (+) RNA Virus:

    • (+) RNA acts as mRNA; translated to make viral proteins and RDRP.

    • RDRP synthesizes (-) RNA, which is then used to make more (+) RNA.

  • Single-Stranded (-) RNA Virus:

    • RDRP (carried in the viral capsid) transcribes (-) RNA to (+) RNA (mRNA).

    • mRNA is translated; RDRP synthesizes more (-) RNA from (+) RNA.

  • Single-Stranded (+) RNA Retrovirus:

    • Reverse transcriptase (RDDP) synthesizes (-) DNA from (+) RNA.

    • DDDP synthesizes (+) DNA, forming DS DNA.

    • Viral DNA integrates into host genome; mRNA is later transcribed and translated.

Key Enzymes:

  • DDDP: DNA-dependent DNA polymerase (makes DNA from DNA)

  • DDRP: DNA-dependent RNA polymerase (makes RNA from DNA)

  • RDRP: RNA-dependent RNA polymerase (makes RNA from RNA; not found in host cells)

  • RDDP: RNA-dependent DNA polymerase (reverse transcriptase; makes DNA from RNA)

Example: Targeting RDRP in RNA viruses is ideal for antiviral drugs, as humans lack this enzyme.

Microbial Metabolism: Cellular Respiration and Fermentation

Overview of Cellular Respiration

Cellular respiration is the process by which cells extract energy from glucose. It can be aerobic (using oxygen) or anaerobic (using other electron acceptors).

  • Glucose is the primary sugar broken down.

  • In eukaryotes: Mitochondria are the site of aerobic respiration (originated from bacteria).

Anaerobic Respiration Pathway

  • Glycolysis (cytoplasm):

    • Input: 2 ATP, glucose

    • Output: 2 NADH, 4 ATP (net 2 ATP), 2 pyruvate

  • Pyruvate Oxidation (cytoplasm):

    • Input: 2 pyruvate

    • Output: 2 CO2, 2 NADH, 2 acetyl-CoA

  • Citric Acid Cycle (cytoplasm):

    • Input: Acetyl-CoA

    • Output: 6 NADH, 2 FADH2, 2 ATP, 4 CO2

  • Electron Transport Chain:

    • NADH pumps 3 electrons; FADH2 pumps 2 electrons.

    • Final electron acceptor: Oxygen (aerobic); NO3- or SO42- (anaerobic bacteria).

Equation for Aerobic Respiration:

Fermentation

  • Occurs when oxygen is absent.

  • Purpose: Regenerate NAD+ for glycolysis.

  • Outputs: Acids/alcohols, 2 ATP, regenerated NAD+.

  • Less efficient than respiration (only 2 ATP produced).

Example: Lactic acid fermentation in muscle cells and some bacteria.

Microbial Growth and Quantification

Direct Count Method

  • Uses a special slide with a grid to count bacteria in a sample.

  • Advantages: Cost-effective.

  • Disadvantages: Cannot distinguish live/dead cells, phase of growth, or sample representativeness.(represent which phase the bacteria are in)

Flow Cytometry

  • Cells in a liquid sample pass through a laser; each cell is counted.

  • Advantages: Fast, accurate, distinguishes live/dead cells.

  • Disadvantages: Expensive.

Plate Count Assay

  • Serial dilution and plating to count colony-forming units (cfu).

  • FDA convention: Count plates with 30-300 colonies.

Calculation Example:

  • Milk sample: 1 ml diluted in 1:10 series, 1 ml of third dilution plated, 54 colonies observed.

  • Original concentration: cfu/ml

Antimicrobial Drugs and Mechanisms

Targets of Antibiotics

  • Cell wall synthesis (e.g., peptidoglycan)

  • Bacterial ribosomes (protein synthesis)

  • DNA replication

  • Metabolic pathways (e.g., folate synthesis)

B-Lactam Antibiotics

  • Example: Penicillin

  • Mechanism: Inhibits peptidoglycan synthesis (cell wall)

  • Effective against: Gram-positive bacteria

  • Side effects: Allergic reactions, GI upset (kills gut flora)

Ribosomal Inhibitors

Drug

Target

Side Effects

Tetracycline

30S subunit

Teeth discoloration (especially in children)

Azithromycin

50S subunit

Blocks Ca2+ channels, arrhythmia

Gentamycin

30S subunit

Nephrotoxicity, hearing loss

Note: Similarity between bacterial and mitochondrial ribosomes can cause side effects.

DNA Replication Inhibitors

  • Example: Ciprofloxacin

  • Mechanism: Inhibits DNA gyrase (bacteria-specific)

  • Side effects: Tendon rupture, agitation, seizures

Metabolic Inhibitors

  • Example: Sulfa drugs (e.g., trimethoprim)

  • Mechanism: Inhibits folate synthesis (not used by humans)

  • Side effects: Stevens-Johnson Syndrome, arrhythmia

Antibiotic Risk Ranking (Lowest to Highest)

  1. Penicillin

  2. Tetracycline

  3. Azithromycin

  4. Gentamycin

  5. Ciprofloxacin

  6. Sulfa drugs

Immune System: Structure and Function

Overview and Components

  • Function: Protects against bacteria, viruses, parasites, fungi, allergens, and chemicals.

  • Main components: White blood cells, T-cells, platelets, antibodies, skin, mucous membranes, bone marrow, lymph nodes, thymus.

  • First line of defense: Skin

Innate vs. Adaptive Immunity

Feature

Innate Immunity

Adaptive Immunity

Response Time

Immediate

Delayed (but faster on second exposure)

Specificity

Non-specific

Highly specific (memory)

Main Cells

Macrophages, dendritic cells, mast cells, complements, cytokines

B-cells, T-killer cells, Helper T-cells

Cells of the Immune System

  • Plasma cells: Produce antibodies.

  • Macrophages: Phagocytize pathogens, present antigens to other immune cells.

  • Dendritic cells: Antigen-presenting cells (APCs), activate naive T-cells.

  • Mast cells: Release histamine, cause vasodilation and allergic responses.

  • Complements: Proteins that form membrane attack complexes (MACs) to lyse bacteria.

  • Cytokines: Signaling proteins that mediate inflammation and fever.

  • B-cells: Produce antibodies (adaptive immunity).

  • T-killer cells (CD8): Destroy infected or abnormal cells.

  • Helper T-cells (CD4): Coordinate immune response, link innate and adaptive immunity.

  • Natural Killer (NK) cells: Kill tumor and infected cells without prior activation.

Immune System Development and Stem Cells

  • Pluripotent stem cells: Found in bone marrow; differentiate into lymphoid or myeloid progenitors.

  • Lymphoid progenitors: Give rise to B-cells, T-cells, NK cells (adaptive immunity).

  • Myeloid progenitors: Give rise to erythrocytes, platelets, monocytes, neutrophils, eosinophils, basophils (innate immunity).

Immune Response and Memory

  • First exposure: Innate immunity responds; adaptive immunity is delayed but highly effective.

  • Second exposure: Adaptive immunity responds rapidly due to memory cells.

  • Allergies: Occur on second exposure due to immune memory (e.g., anaphylaxis).

Major Histocompatibility Complex (MHC)

  • MHC I: Present on all nucleated cells (except RBCs); presents endogenous antigens.

  • MHC II: Present on macrophages and dendritic cells; presents exogenous antigens to Helper T-cells.

Activation and Regulation

  • Helper T-cells are essential for adaptive immunity; their loss (e.g., HIV infection) leads to immune deficiency.

  • B-cells and T-cells must both recognize an antigen to initiate a full immune response (prevents autoimmunity).

  • Thymic selection ensures T-cells can distinguish self from non-self; failure leads to autoimmune diseases.

Inflammation and Immune Response

  • Symptoms: Edema, redness, heat, pain (due to increased blood flow and vascular permeability).

  • Diapedesis: White blood cells move from blood vessels to tissues.

  • Lymphatic system: Filters blood, houses immune cells, returns filtered fluid to circulation.

Examples and Clinical Applications

  • Anaphylaxis: Severe, rapid allergic reaction caused by the immune system.

  • Leukemia: Mutation in stem cells leads to overproduction of one blood cell type; treated with chemotherapy and bone marrow transplant.

  • Ibuprofen: Reduces inflammation by blocking cytokine production.

Additional info: The immune system's memory is protective but can also cause harm (e.g., allergies, autoimmune diseases). The double-check system between B-cells and T-cells helps prevent inappropriate immune responses.

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