Micro II Exam 1 Poxviruses: Structure, Replication, and the History of Smallpox

Introduction to Poxviruses

• Virus Classification and Types:     * Variola: Known as the causative agent of Smallpox.     * Molluscum contagiosum: Causes localized skin lesions.     * Zoonoses: Poxviruses that can be transmitted from animals to humans.     * Vaccinia virus: Primarily used in the production of the smallpox vaccine.

Structure of Poxviruses

• Size and Shape:     * These are the largest known viruses.     * They are characterized by a complex, brick-shaped structure.

• Genome and Composition:     * Enveloped virus.     * Contains a large, double-stranded linear DNA genome.

• Detailed Internal Architecture (Figure 52-1):     * Core: Contains viral DNA and protein; assumes a dumbbell shape due to the presence of lateral bodies.     * Lateral Bodies: Large structures located between the core and the outer membrane.     * Membranes: Includes a core membrane and an "outer membrane" that assembles around the core within the cytoplasm.     * Surface Components: Surface tubules are visible on the exterior.

Poxvirus Replication Cycle

• Location: Replication takes place entirely within the host cell cytoplasm.

• Enzymatic Activity: The viral genome is self-sustained to a high degree, encoding its own mRNA and DNA polymerases.

• Steps of Replication (Figure 52-2):     * Adsorption and Penetration: The virus attaches to and enters the cell.     * Uncoating:         1. The outer membrane is lost.         2. Early Transcription: Virion enzymes initiate the transcription of early mRNA.         3. Uncoatase: An early gene product (a viral-encoded enzyme) triggers the release of viral DNA from the core.     * DNA Replication: Occurs in specific cytoplasmic inclusions referred to as "factories." This process typically takes place between $2$ and $5$ hours post-infection.     * Late Transcription and Translation: Structural proteins (late virion proteins) and enzymes are produced.     * Morphogenesis: Assembly of DNA and protein into cores occurs over a period of $4$ to $20$ hours. Spherical immature particles progress into mature virions.

• Viral Yield and Release:     * Viral Load: Approximately $10,000$ viral particles are produced per infected cell.     * Release Mechanism: Virions leave the cell either via exocytosis or upon cell lysis (disruption).

Hybrid Vaccines and Expression Vectors

• Mechanism: Vaccinia virus is used as an expression vector (cloning vector) to produce live recombinant vaccines.

• Process (Figure 52-3):     1. A foreign gene (the infectious material/immunizing antigen) is placed into a recombinant plasmid vector.     2. The plasmid is introduced into a host cell via transfection.     3. The cell is then infected with a "rescuing virus" (poxvirus).     4. The foreign gene is incorporated into the poxvirus genome through recombination of vaccinia sequences.     5. The recombinant vaccinia DNA replicates containing the foreign gene.     6. The maturation and release of recombinant vaccinia virus occur.

• Target Applications: Research and development for vaccines against Rabies, HIV, Hepatitis B, and Influenza.

Pathogenesis and Immunity

• Smallpox Pathogenesis:     * Primary infection begins in the upper respiratory tract.     * The virus spreads through the lymphatic system.     * Results in a simultaneous eruption of pocks across the body.

• Molluscum Contagiosum Pathogenesis:     * Remains localized at the site of infection.     * Causes wart-like lesions.

• Host-Virus Interaction:     * Poxvirus proteins stimulate host cell growth.     * Immune Response: Cell-mediated immunity is required to resolve the infection.     * Evasion Techniques:         * Utilization of cell-to-cell spread to avoid antibodies.         * Impairment of interferon, antibodies, and the complement system.         * Prevention of cell-mediated immune responses.

Spread of Smallpox Within the Body (Pathogenic Timeline)

• Initial Entry (Day 0): Inhalation of virus or transmission via scabs (less effective).

• Local Multiplication: Virus multiplies in the upper respiratory tract without causing symptoms.

• Systemic Spread:     1. The virus infects macrophages, which travel to regional lymph nodes.     2. Primary Viremia: Virus enters the bloodstream from the lymphatics.     3. Secondary Viremia: The virus spreads to internal organs including the spleen, liver, bone marrow, and additional lymph nodes.

• Prodrome (Approx. Day 12): Onset of fever and rash.

• Clinical Eruption (Day 16): Pustules develop as small vessels in the dermis hemorrhage, leading to the characteristic "pox" rash.

• Outcomes: Recovery (associated with lifelong immunity) or death.

Epidemiology of Poxviruses

• Strictly Human Viruses: Smallpox (Variola) and Molluscum contagiosum.

• Transmission Routes:     * Smallpox: Respiratory route (airborne) or contact with lesions/scabs.     * Molluscum contagiosum: Direct contact with lesions.     * Zoonoses: Direct contact with animal lesions (e.g., Orf, Monkeypox).

• Factors Leading to Smallpox Eradication:     * Limited host range (humans only).     * Possesses only a single serotype.     * Consistent and identifiable disease presentation.     * Effective immunization using related animalpox proteins.     * Stable vaccine with a visible scar as proof of vaccination.     * Efficient global program managed by the World Health Organization (WHO).

Clinical Syndromes: Smallpox

• Classifications: Variola major and Variola minor.

• Symptom Progression:     * Incubation period followed by high fever, exhaustion, headache, and backache.     * Vesicular rash begins in the mouth followed by a simultaneously erupting rash (pocks) on the skin.     * Variola Major Severity: Can involve vomiting, diarrhea, and excessive bleeding.     * Centrifugal Pattern: Rash is more dense on the face and extremities than on the trunk.

• Historical Mortality:     * In the 18th century, Smallpox caused $7\% - 12\%$ of all deaths and $1/3$ of all childhood deaths.     * Total historical death toll is estimated at $300$ to $500$ million people.

• Societal Impact:     * Inca Empire: Pizarro and fewer than $200$ Spaniards faced $10$ million Incas; at least $50\%$ of the Inca population died of smallpox.     * Aztec Empire: Cortez's arrival led to the death of $15$ million Aztecs ($80\%$ of the population).

• Fatalities in Children: Approximately one-third of infected individuals died from bleeding, cardiovascular collapse, or secondary infections.

Clinical Syndromes: Other Poxviruses

• Vaccinia:     * Used as the live virus for smallpox vaccination.     * Administered by scratching the virus into the skin; successful vaccination is marked by the development of pustules.     * Past Complications: Encephalitis and progressive infections in immunocompromised patients.

• Molluscum Contagiosum:     * Lesions: Pink papules that progress to pearl-like nodular, wart-like lesions with a central plug that can be squeezed out.     * Target Populations: Children, sexually active individuals, and the immunocompromised.

• Zoonotic Poxes (Orf, Cowpox, Monkeypox):     * Exposure through direct animal contact.     * Symptoms: Usually a single lesion at the point of contact that regresses in $25$ to $35$ days.

History of Control and Eradication

• Variolation: An early, risky method (dating to the 1600s-1700s) involving smearing smallpox pus into opened veins. Associated with a $1$ in $50$ chance of death.     * Key figures: Lady Mary Wortley Montagu (Ottoman Empire/England), Cotton Mather, and Onesimus (USA).

• Vaccination: Edward Jenner introduced the use of cowpox pus in $1796$.

• Notable Survivors: Abraham Lincoln developed smallpox one day after the Gettysburg Address.

• Global Eradication Effort:     * WHO Resolution: Passed in $1959$.     * Intensified Program (1967): Headed by Dr. Donald A. Henderson; involved tracking every case, quarantining infected individuals, and immunizing all contacts.     * Milestones:         * Last Natural Variola Minor Case: Ali Maow Maalin in Merka, Somalia ($1977$).         * Last Accidental Case: Janet Parker, a photographer in Birmingham, UK ($1978$).         * Official Declaration: Eradicated worldwide in $1980$.

Modern Status and Biosecurity

• Official Repositories: Smallpox reference stocks are officially held in only two locations:     1. Vector in Koltsovo, Novosibirsk, Russia.     2. CDC in Atlanta, Georgia, USA.

• Unconfirmed Locations: Suspected stocks in France, Iraq, Lebanon, Israel, Turkey, Syria, Jordan, Saudi Arabia, Kuwait, Iran, and North Korea.

• Current Concerns:     * Potential use as a bioweapon.     * The world now has a huge population with no immunity.     * Mortality rate remains high ($30\% - 40\%$) and spread is airborne (Level 5 concern).

Diagnosis, Treatment, and Prevention

• Diagnosis:     * Smallpox: Historically required isolation and growth of the virus or identification of inclusion bodies; PCR/DNA probes were not available during the eradication era.     * Molluscum Contagiosum: Histological examination to find "molluscum bodies" (epidermis filled with viral inclusions); cannot be cultured in tissue.

• Treatment:     * Smallpox: New antivirals have been developed post-eradication.     * Molluscum Contagiosum: Often self-resolving; lesions can be removed via freezing or burning.     * Animal Poxes: Generally self-resolve in humans.

• Prevention:     * Smallpox: General vaccination has been discontinued.     * Molluscum Contagiosum: No specific prevention beyond avoiding lesion contact.     * Animal Poxes: Basic hygiene when handling animals.