Virology & Vaccines

Virology & Vaccines

Lecture on virus structure (including HPV as case study)

Virology

Journal paper discussions

Vaccines

These notes were prepared by Diana Hardie for Virology Lectures to 3rd Year Medical Students in the Department of Medical Microbiology, University of Cape Town. http://web.uct.ac.za/depts/mmi/jmoodie/welcome1.html

The introduction of vaccination has been one of the most decisive advances leading to the dramatic downward trend in the incidence of many viral diseases.

The principle of vaccination is to induce a "primed" state in the vaccinated subject so that, following exposure to a pathogen, a rapid secondary immune response is generated leading to the accelerated elimination of the organism and protection from clinical disease. Success depends on the generation of memory T and B cells and the presence in the serum of neutralizing antibody.


Attributes of a good vaccine

  1. Ability to elicit the appropriate immune response for the particular pathogen:
    • Tuberculosis - cell mediated response
    • most bacterial and viral infections - antibody
  2. Long term protection - ideally life-long
  3. Safety - vaccine itself should not cause disease
  4. Stable - retain immunogenicity, despite adverse storage conditions prior to administration
  5. Inexpensive

Types of Vaccine

Vaccines in general use include: LIVE vaccines and KILLED vaccines. Vaccines are available for:


A. Live Vaccines

A.1. Types of Live Vaccines

  1. Live attenuated organisms

    Organisms whose virulence has been artificially reduced by in vitro culture under adverse conditions, such as reduced temperature. This results in the selection of mutants which replicate poorly in the human hostand are therefore of reduced virulence. Replication of the vaccine strain in the host reproduces many ofthe features of wild type infection, without causing clinical disease.Most successful viral vaccinesbelong to this group.

    The immune response is usually good - when the virus replicates in the host cells, both antibody as well ascell mediated immune responses are generated and immunity is generally long lived. Often, only a singledose is needed to induce long term immunity.

    Potential drawbacks to these vaccines include:

    • the danger of reversion to virulence and
    • the possibility of causing extensive disease in immunocompromised individuals.
  2. Heterologous vaccines

    Closely related organism of lesser virulence, which shares many antigens with the virulent organism. The vaccine strain replicates in the host and induces an immune response that cross reacts with antigens of the virulent organism. The most famous example of this type of vaccine is vaccinia virus: Both cowpox virus and vaccinia virus are closely related to variola virus, the causative agent of smallpox. The eighteenth century physician, Edward Jenner observed that milkmaids who had been infected with cowpox virus were immune to smallpox. Widespread use of vaccinia virus as a vaccine has lead to the world-wide eradication of smallpox.

  3. Live recombinant vaccines

    It is possible, using genetic engineering, to introduce a gene coding for an immunogenic protein from one organism into the genome of another (such as vaccinia virus). The organism expressing a foreign gene is called a recombinant. Following injection into the subject, the recombinant organism will replicate and express sufficient amounts of the foreign protein to induce a specific immune response to the protein.


A.2 Attributes of live vaccines

    1. Good immune response
      • Both Cell Mediated Immunity and antibody responses.
      • Immunity is long lived
      • Single dose
    2. Safety
      • Danger of reversion to virulence, or
      • Severe disease in immunocomprised
    3. Stability
      • Organisms in the vaccine must remain viable in order to infect and replicate in the host
      • Vaccine preparations are therefore very sensitive to adverse storage conditions
      • Maintenance of the cold chain is very important.
    4. Expense
      • Cheap to prepare

B. Killed (inactivated) vaccines

When safe live vaccines are not available, either because attenuated strains have not been developed or else because reversion to wild type occurs too readily, it may be possible to use an inactivated preparation of the virulent organism to immunize the host.

The organism is propagated in bulk, in vitro, and inactivated with either beta-propiolactone or formaldehyde. These vaccines are not infectious and are therefore relatively safe. However, they are usually of lower immunogenicity and multiple doses may be needed to induce immunity. In addition, they are usually expensive to prepare.


B.1 Types of Killed Vaccines

  1. Subcellular fractions

    When protective immunity is known to be directed against only one or two proteins of an organism, it may be possible to use a purified preparation of these proteins as a vaccine. The organism is grown in bulk and inactivated, and then the protein of interest is purified and concentrated from the culture suspension. These vaccines are safe and fewer local reactions occur at the injection site. However, the same disadvantages of poor immunogenicity and the need for multiple boosters applies.

  2. Recombinant proteins

    Immunogenic proteins of virulent organisms may be synthesized artificially by introducing the gene coding for the protein into an expression vector, such as E-coli or yeasts. The protein of interest can be extracted from lysates of the expression vector, then concentrated and purified for use as a vaccine. The only example of such a vaccine, in current use, is the hepatitis B vaccine.


B.2 Attributes of killed vaccines

  1. Immune response
    • poor;only antibody - no cell immediated immune response.
    • response is short-lived and multiple doses are needed.
    • may be enhanced by the incorporation of adjuvants into the vaccine preparation (see below)
  2. Safety
    • Inactivated, therefore cannot replicate in the host and cause disease.
    • Local reactions at the site of injection may occur.
  3. Stability
    • Efficacy of the vaccine does not rely on the viability of the organisms.
    • These vaccines tend to be able to withstand more adverse storage conditions.
  4. Expense
    • Expensive to prepare.

C. Adjuvants

Certain substances, when administered simultaneously with a specific antigen, will enhance the immune response to that antigen. Such compounds are routinely included in inactivated or purified antigen vaccines.

Adjuvants in common use

C.1. Aluminium salts

  • First safe and effective compound to be used in human vaccines.
  • It promotes a good antibody response, but poor cell mediated immunity.

C.2. Liposomes and Immunostimulating complexes (ISCOMS)

C.3. Complete Freunds adjuvant is an emulsion of Mycobacteria, oil and water

  • Too toxic for man
  • Induces a good cell mediatedimmune response.

C.4. Incomplete Freund's adjuvant as above, but without Mycobacteria.

C.5. Muramyl di-peptide

  • Derived from Mycobacterial cell wall.

C.6. Cytokines

  • IL-2, IL-12 and Interferon-gamma.

C.7. Possible modes of action

  • By trapping antigen in the tissues,thus allowing maximal exposure to dendritic cells and specific T and B lymphocytes.
  • By activating antigen-presenting cells to secrete cytokines that enhance the recruitment of antigen-specific T and B cells to the site of inoculation.

D. DNA Vaccines

DNA vaccines are at present experimental, but hold promise for future therapy since they will evoke both humoral and cell-mediated immunity, without the dangers associated with live virus vaccines.

The gene for an antigenic determinant of a pathogenic organism is inserted into a plasmid.This genetically engineered plasmid comprises the DNA vaccine which is then injected into the host.Within the host cells,the foreign gene can be expressed (transcribed and translated) from the plasmid DNA, and if sufficient amounts of the foreign protein are produced, they will elicit an immune response.

E. Vaccines in general use

Measles

Live attenuated virus grown in chick embryo fibroblasts, first introduced in the 1960's. Its extensive use has led to the virtual eradication of measles in the first world. In developed countries, the vaccine is administered to all children in the second year of life (at about 15 months). However, in developing countries, where measles is still widespread, children tend to become infected early (in the first year),which frequently results in severe disease. It is therefore important to administer the vaccine as early as possible (between six months and a year). If the vaccine is administered too early, however, there is a poor take rate due to the interference by maternal antibody. For this reason, when vaccine is administered before the age of one year, a booster dose is recommended at 15 months.


Mumps

Live attenuated virus developed in the 1960's. In first world countries it is administered together with measles and rubella at 15 months in the MMR vaccine. This is not a legal requirement in South Africa.


Rubella

Live attenuated virus. Rubella causes a mild febrile illness in children, but if infection occurs during pregnancy, the foetus may develop severe congenital abnormalities. Two vaccination policies have been adopted in the first world. In the USA, the vaccine is administered to all children in their second year of life (in an attempt to eradicate infection), while in Britain, until recently, only post pubertal girls were vaccinated. It was feared that if the prevalence of rubella in the community fell, then infection in the unimmunized might occur later - thus increasing the likelihood of infection occurring in the child-bearing years. This programme has since been abandoned in Britain and immunization of all children is the current practice.


Polio

Two highly effective vaccines containing all 3 strains of poliovirus are in general use:

  • The killed virus vaccine (Salk, 1954) is used mainly in Sweden, Finland, Holland and Iceland.
  • The live attenuated oral polio vaccine (Sabin, 1957) has been adopted in most parts of the world;its chief advantages being: low cost, the fact that it induces mucosal immunity and the possibility that, in poorly immunized communities, vaccine strains might replace circulating wild strains and improve herd immunity.Against this is the risk of reversion to virulence (especially of types 2 and 3) and the fact that the vaccine is sensitive to storage under adverse conditions.
  • The inactivated Salk vaccine is recommended for children who are immunosuppressed.

Hepatitis B

Two vaccines are in current use: a serum derived vaccine and a recombinant vaccine. Both contain purified preparations of the hepatitis B surface protein.

The serum derived vaccine is prepared from hepatitis B surface protein, purified from the serum of hepatitis B carriers. This protein is synthesised in vast excess by infected hepatocytes and secreted into the blood of infected individuals. A vaccine trial performed on homosexual men in the USA has shown that, following three intra-muscular doses at 0, 1 and 6 months, the vaccine is at least 95% protective.

A second vaccine, produced by recombinant DNA technology, has since become available. Previously, vaccine administration was restricted to individuals who were at high risk of exposure to hepatitis B, namely: infants of hepatitis B carrier mothers, health care workers, homosexual men and intravenous drug abusers. However, hepatitis B has been targeted for eradication, and since 1995 the vaccine has been included in the universal childhood immunization schedule. Three doses are given;at 6, 10, and 14 weeks of age. As with any killed viral vaccines, a booster will be required at some interval (not yet determined, but about 5 years) to provide protection in later life from hepatitis B infection as a venereal disease.


Hepatitis A

A vaccine for hepatitis A has been developed from formalin-inactivated , cell culture-derived virus. Two doses, administered one month apart, appear to induce high levels of neutralising antibodies.The vaccine is recommended for travellers to third world countries, and indeed all adults who are not immune to hepatitis A.


Yellow Fever

The 17D strain is a live attenuated vaccine developed in 1937. It is a highly effective vaccine which is administered to residents in the tropics and travellers to endemic areas. A single dose induces protective immunity to travellers and booster doses, every 10 years, are recommended for residents in endemic areas.


Rabies

No safe attenuated strain of rabies virus has yet been developed for humans. Vaccines in current use include:

  • The neurotissue vaccine - here the virus is grown in the spinal cords of rabbits, and then inactivated with beta-propiolactone. There is a high incidence of neurological complications following administration of this vaccine due to a hypersensitivity reaction to the myelin in the preparation and largely it has been replaced by
  • A human diploid cell culture-derived vaccine (also inactivated) which is much safer.

There are two situations where vaccine is given:

  1. Post-exposure prophylaxis, following the bite of a rabid animal:

    A course of 5-6 intramuscular injections, starting on the day of exposure. Hyperimmune rabies globulin may also administered on the day of exposure.

  2. Pre-exposure prophylaxis is used for protection of those whose occupation puts them at risk of infection with rabies:

    for example, vets, abbatoir and laboratory workers. This schedule is 2 doses one month apart ,and a booster dose one year later. (Further boosters every 2-3 years should be given if risk of exposure continues).


Influenza

Repeated infections with influenza virus are common due to rapid antigenic variation of the viral envelope glycoproteins. Antibodies to the viral neuraminidase and haemagglutinin proteins protect the host from infection. However, because of the rapid antigenic variation, new vaccines, containing antigens derived from influenza strains currently circulating in the community, are produced every year. Surveillance of influenza strains now allows the inclusion of appropriate antigens for each season.The vaccines consist of partially purified envelope proteins of inactivated current influenza A and B strains.

Individuals who are at risk of developing severe, life threatening disease if infected with influenza should receive vaccination. People at risk include the elderly, immunocompromised individuals, and patients with cardiac disease. In these patients, protection from disease is only partial, but the severity of infection is reduced.


Varicella-Zoster virus (Chickenpox, Shingles)

A live attenuated strain of varicella zoster virus has been developed. It is not licensed in South Africa for general use, but is used in some oncology units to protect immunocompromised children who have not been exposed to wild-type varicella zoster virus. Such patients may develop severe, life threatening infections if infected with the wild type virus.

Assessment

  • 15 min presentations on papers given to you to read.
  • Short background of the virus the paper deals with.
    • Structural unity among viral origin binding proteins: crystal structure of nuclease domain of Adeno-associated virus Rep (10.1016/S1097-2765(02)00592-0)
    • Architecture of herpes simplex major capsid protein derived from structural bioinformatics (10.1016/S0022-2836(03)00696-X)
    • Dynamics of herpes simplex virus capsid maturation visualised by time-lapse cryp-EM (10.1038/nsb922)
  • Presentation of the techniques, results and discussion.

Bibliography