Viruses: Structure and Replication

Basic Viral Structure

• All viruses possess two essential structures:

  • Genetic material (DNA or RNA)

  • Capsid (protein coat surrounding the genetic material)

• Optional structures include:

  • Envelope: A lipid membrane derived from the host cell, helps the virus evade the immune system by mimicking host membranes.

  • Spike proteins: Glycoproteins protruding from the envelope, function as attachment factors (fake cell receptors) to facilitate entry into host cells and increase virulence.

Viral Replication Pathways

• Primary method: Lytic cycle

• Goals of viral replication:

  • Copy genetic material

  • Assemble new viral particles

• Lytic cycle steps:

  1. Attachment: Virus binds to host cell surface.

  2. Uncoating: Viral capsid is removed, and genetic material enters 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.

• Alternative pathway: Lysogenic cycle (Lysogenesis)

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

  • Allows the virus to remain dormant and evade the immune system.

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

Viral Genomes and Replication Enzymes

• Types of viral genomes:

  • Double-stranded DNA (dsDNA)

  • Single-stranded (+) DNA

  • Single-stranded (-) DNA

  • Double-stranded RNA (dsRNA)

  • Single-stranded (+) RNA

  • Single-stranded (-) RNA

• 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 human cells, making it a good antiviral target)

  • RDDP: RNA-dependent DNA polymerase (reverse transcriptase; makes DNA from RNA, used by retroviruses)

Viral Replication Pathways by Genome Type

• dsDNA virus: Uses host DDRP to make mRNA from (-) DNA strand; DDDP copies DNA genome.

• ss(+) DNA virus: DDDP makes (-) DNA, DDRP makes (+) RNA (mRNA), DDDP copies DNA.

• ss(-) DNA virus: DDRP makes mRNA, DDDP copies DNA strands.

• ss(+) RNA virus: (+) RNA acts as mRNA, translated by ribosomes; RDRP makes (-) RNA, which is then used to make more (+) RNA.

• ss(-) RNA virus: RDRP (carried in the virion) makes (+) RNA (mRNA), which is translated; RDRP also makes more (-) RNA.

• dsRNA virus: (+) RNA is translated; RDRP copies RNA genome.

• ss(+) RNA retrovirus: RDDP makes (-) DNA, DDDP makes (+) DNA, viral DNA integrates into host genome, mRNA is made and translated.

Antiviral Drug Targets

• RDRP is an ideal antiviral target because it is not present in human cells.

Antimicrobial Drugs

General Principles

• All antibiotics have risks and side effects.

• Common bacterial targets:

  • Cell wall synthesis

  • Bacterial ribosomes

  • DNA replication

Cell Wall Synthesis Inhibitors

• Example: Beta-lactams (e.g., penicillin)

• Mechanism: Inhibit peptidoglycan synthesis

• Effective against: Gram-positive bacteria

• Side effects: Allergic reactions, gastrointestinal upset (due to disruption of gut flora)

Ribosomal Inhibitors

• Bacterial ribosomes: 70S; Eukaryotic ribosomes: 80S

• Examples: Tetracycline (targets 30S), Azithromycin (targets 50S), Gentamycin (targets 30S)

• Side effects:

  • Tetracycline: Tooth discoloration (especially in children, binds calcium in developing teeth)

  • Azithromycin: Blocks Ca2+ channels, may cause arrhythmia

  • Gentamycin: Nephrotoxicity, hearing loss (due to mitochondrial targeting)

• Note: Mitochondria evolved from prokaryotes, so ribosomal inhibitors may affect mitochondrial function.

DNA Replication Inhibitors

• Bacteria use DNA gyrase; humans use topoisomerase

• Example: Ciprofloxacin

• Mechanism: Inhibits DNA gyrase

• Side effects: Tendon rupture, agitation, seizures (due to GABA receptor blockade)

Metabolic Inhibitors

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

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

• Side effects: Stevens-Johnson Syndrome (severe skin reaction), arrhythmia

Antibiotic Risk Ranking (Lowest to Highest)

• Penicillin < Tetracycline < Azithromycin < Gentamycin < Ciprofloxacin < Sulfa drugs

Antibiotic Development

• Pharmaceutical companies are reluctant to develop new antibiotics due to limited profitability and shelf life compared to chronic medications.

Microbial Growth and Quantification

Plate Count Assays

• Used to estimate the number of viable bacteria in a sample.

• Procedure:

  • Serial dilution of sample (e.g., 1:10 dilution series)

  • Plating a known volume on agar

  • Counting colonies after incubation

• Calculation: $\text{CFU/mL} = \frac{\text{Number of colonies}}{\text{Volume plated (mL)} \times \text{Dilution factor}}$

• FDA convention: Count plates with 30–300 colonies

Immunology: Innate and Adaptive Immunity

Overview of the Immune System

• Innate immunity: First line of defense, rapid but non-specific

• Adaptive immunity: Slower, highly specific, and has memory

• Functions: Protects against pathogens (bacteria, viruses, fungi, parasites), allergens, and chemicals

Primary Immune Organs and Cells

• Bone marrow: Produces pluripotent stem cells

• Lymph nodes: Filter blood and dead cells; sites for lymphocyte activation

• Thymus: Site of T-cell maturation

Innate Immune System Components

• Macrophages (APCs): Phagocytize pathogens, present antigens

• Dendritic cells (APCs): Capture antigens, migrate to lymph nodes to activate T cells

• Mast cells: Release histamine, cause inflammation and vasodilation

• Complement proteins: Form membrane attack complex (MAC) to lyse bacteria

• Cytokines: Mediate inflammation and fever

Adaptive Immune System Components

• B cells: Produce antibodies

• T-killer (cytotoxic) cells: Destroy infected or flagged cells

• Helper T cells: Coordinate immune response, link innate and adaptive immunity

• Memory cells: Provide long-term immunity

Immune Cell Development

• Pluripotent stem cells (in bone marrow) differentiate into:

  • Lymphoid progenitors (adaptive immunity): Immature B cells, T cells, Natural Killer cells

  • Myeloid progenitors (innate immunity): Erythroblasts (RBCs), megakaryoblasts (platelets), myeloblasts (WBCs), immature dendritic cells

Major Histocompatibility Complex (MHC)

• MHC I: Present on all nucleated cells (except RBCs)

• MHC II: Present on macrophages and dendritic cells

• Critical for self/non-self recognition and T cell activation

Antigen Presentation and Immune Activation

• APCs present antigens to Helper T cells (CD4+)

• Helper T cells activate B cells and cytotoxic T cells (CD8+)

• Double-check mechanism prevents autoimmunity

Antibody Structure and Function

• Antibodies (immunoglobulins, Ig) are proteins with:

  • Variable region: Determines antigen specificity

  • Constant region: Determines antibody class (IgG, IgM, etc.)

• Classes and functions:

  • IgM: Pentamer, agglutination, complement activation

  • IgD: Membrane-bound, function unclear

  • IgE: Triggers histamine release, allergic responses

  • IgA: Dimer, agglutination, antiviral properties

  • IgG: Most abundant, crosses placenta, opsonization, neutralizes toxins

Hypersensitivity Reactions

Types of Hypersensitivity

Type I Hypersensitivity (Immediate)

• First exposure: Sensitization, IgE produced and binds mast cells

• Second exposure: Antigen cross-links IgE, mast cell degranulation, histamine and leukotriene release

• Symptoms: Anaphylaxis (bronchoconstriction, hypotension, hives)

Type II Hypersensitivity (Cytotoxic)

• Antibodies bind to antigens on cells (e.g., RBCs, platelets), activate complement, cause cell lysis

• Examples: Penicillin-induced hemolytic anemia, Rh incompatibility in pregnancy

Type III Hypersensitivity (Immune Complex)

• Immune complexes (antigen-antibody) deposit in tissues, activate complement, cause inflammation

• Examples: Serum sickness, post-strep nephritis, dust-induced alveolitis

Type IV Hypersensitivity (Delayed, Cell-Mediated)

• Mediated by T cells and macrophages, not antibodies

• Examples: Contact dermatitis (poison ivy), TB skin test, latex allergy

• Symptoms develop 24–72 hours after exposure

Key Terms and Concepts

• Opsonization: Enhanced phagocytosis by coating pathogens with antibodies or complement

• Agglutination: Clumping of pathogens by antibodies, facilitates phagocytosis

• Antibody-Dependent Cell-Mediated Cytotoxicity (ADCC): Immune cells kill antibody-coated target cells; safer than chemotherapy/radiation for some therapies

• Diapedesis: Movement of white blood cells from blood vessels into tissues

• PAMPs (Pathogen-Associated Molecular Patterns): Molecules on pathogens recognized by innate immune cells (e.g., peptidoglycan, LPS)

Sample Calculations

• CFU/mL calculation example:

  • Plate has 54 colonies, plated from 1 mL of 1:1000 dilution: $54 \times 1000 = 54,000$ CFU/mL

  • Plate has 83 colonies, plated from 1 mL of 1:10,000 dilution: $83 \times 10,000 = 830,000$ CFU/mL

  • Plate has 191 colonies, plated from 0.1 mL of 1:100 dilution: $191 \times 10 \times 100 = 191,000$ CFU/mL

Additional Info

• Gene expression differences, not DNA sequence, determine cell type (e.g., nerve vs. muscle cell).

• Thymic deletion removes self-reactive T cells; failure can lead to autoimmune diseases (e.g., diabetes, multiple sclerosis).

• Memory in adaptive immunity ensures rapid and robust response upon re-exposure to the same antigen.