▪ Live attenuated vaccines:
· insufficient attenuation
· reversion to wild type
· no safe administration to immunodeficient patient
· persistent infection
· contamination by other microbes
· fetal damage
▪ Non-living vaccines:
· contamination by live organisms
· contamination by toxins
· allergic reactions
· autoimmunity
▪ Genetically engineered vaccines:
·? inclusion of oncogenes
v LIVE ATTENUATED VACCINES. The most effective vaccines are those that actually multiply in the host, mimicking the early stages of natural infection. In these vaccines, the pathogens are attenuated —alive but weakened in their capacity to cause severe disease.
These attenuated microbes are called vaccine strains. Such strains are produced by different methods of attenuation:
▪ Serial passages in cells cultured in vitro
▪ Serial passage in nutrient culture media
▪ Adaptation to low temperatures
▪ Selection of spontaneous mutants
▪ Chemical mutagenesis
▪ Required genetic changes (by gene engineering)
It is rather difficult to achieve satisfactory attenuation of a “wild” microbial strain and to prepare a safe and stable vaccine strain. Because of that, there are not many effective live vaccines. Furthermore, attenuated microbes have been known to regain virulence and have caused serious disease in vaccinated persons. Only those attenuated organisms unlikely to revert can be safely used in vaccines. In general, living vaccines induce stronger and more lasting immunity than non-living vaccines.
NOTE: Vaccines containing attenuated organisms are especially effective if they can be introduced by the same routes the corresponding pathogen uses for entering the body. The attenuated microbes harmlessly propagate in the body to provide a greater and more prolonged antigenic stimulation. Furthermore, the attenuated pathogens may be shed by the immunized individuals and vaccinate other susceptible persons.
The mostly important live vaccines are as follows:
· Sabin poliovaccine. Attenuated trivalent preparation of poliovirus administered orally to induce immunity against poliomyelitis.
The attenuated Sabin polio vaccine is administered orally. It harmlessly replicates in the intestine, similar to the early stages of poliomyelitis. Unlike virulent poliovirus, the attenuated variant causes no paralysis. The Sabin vaccine stimulates the formation of secretory IgA, antibodies that neutralize viruses in the intestine before they invade the bloodstream.
Sabin vaccine virus is shed in the feces of vaccinated persons. Since polioviruses are naturally transmitted by the fecal-oral route, people may be inadvertently vaccinated by ingesting attenuated viruses shed by vaccinated individuals.
● BCG vaccine (The Bacille Calmette-Guérin). Vaccine to provide immunity against tuberculosis. It is made from attenuated bovine tubercle bacillus. The attenuation have been achieved by passage for 10 years in glycerol-bile-potato medium.
● Other vaccines that use attenuated versions of pathogens include those that help protect against influenza, yellow fever, measles, mumps, rubella, plaque,tularemia, brucellosis, etc.
v KILLED (inactivated) VACCINES. A pure culture of the highly immunogenic and virulent pathogen is exposed to an agent that will kill the microbe without altering the surface antigens. Injection of these killed organisms into a host provides safe initial exposure to the pathogen and subsequent resistance to disease.
A variety of methods are available for inactivation: formaldehyde inactivation, phenol, acetone, ultraviolet light, simple heating, β-propiolactone and various ethylenimines, prosalens, etc.
Killed vaccines have the advantage of non-infectivity and therefore relative safety, but disadvantage of generally lower immunogenicity, high risk of hypersensitivity complications and the consequent need for several doses. Most of these vaccines require several booster shots before protection is adequate.
NOTE: Booster – is an immunization given to enhance the memory response to an antigen
This group of vaccines includes the following:
· Typhoid fever vaccine, is a formalin-killed preparation of the pathogen Salmonella typhi.
● Salk poliovaccine. Formalin -inactivated poliovirus preparation administered by injection to induce immunity against poliomyelitis. The inactivated Salk vaccine must be injected into muscle, inducing little protection in the intestine.
● Vaccines containing killed organisms or inactivated viruses have been used to prevent cholera, influenza, whooping cough, rabies, epidemic typhus, etc.
v SUBUNIT VACCINES (Purified Antigen Fractions ). Some vaccines contain only the antigens against which the protective immune reactions are directed (such antigens are called protective). Removal of all live infectious material is obviously a vital element in ensuring the safety of such vaccines.
Antigen fractions are too small to activate the adequate immune response (for example, polysaccharide antigens fail to stimulate T cells and therefore induce only primary responses). Many adjuvants will enhance the immune response when administered with antigen. Adjuvants are such substances that were shown to be safe, convenient, and effective to enhance the consequent immune response when administered simultaneously with antigen. A variety of foreign and endogenous substances can act as adjuvants, but only aluminum and calcium salts are routinely used in clinical practice (aluminum hydroxide, aluminum phosphate, and calcium phosphate). Alum-precipitated antigens are especially effective at inducing antibody responses, and help to induce cell-mediated immunity.
The subunit vaccines are as follows:
● Pneumovax. Purified extracts of the capsular antigens from virulent Streptococcus pneumoniae stimulate the host to produce opsonizing antibodies that effectively protect most immunized persons. This vaccine contains capsular antigens of the 14 most common virulent strains of S. pneumoniae. About 80 percent of all cases of pneumococcal pneumonia are caused by one of these 14 strains.
● Typhim Vi. Polysaccharide typhoid vaccine that is composed of purified Vi capsular polysaccharide of Salmonella typhi.
● Antigen subunits are also used in vaccines against influenza, cholera, some types of meningitis, etc.
v TOXOIDS. Active immunity against toxemic diseases can be induced by injecting a toxoid, an inactivated form of bacterial exotoxin that remains antigenically unaltered but has been chemically treated to destroy its poisonous properties. Exotoxin inactivation is usually by formaldehyde (0,4 per cent solution) for 4 weeks at 40˚C. Toxoid antigens are then purified and adsorbed on the adjuvants. Toxoid stimulates production of antitoxic antibodies (antitoxins), thereby inducing immunity against the corresponding disease.
The commonly used toxoids are the following:
· Tetanus toxoid. Prior immunization with a toxoid prepared from the toxin that the bacterium Clostridium tetani releases into the bloodstream from an infected wound site protects against the post-wound disease tetanus. After three boosters, protection against tetanus lasts at least 10 years, explaining why physicians treating injuries always inquire about the patient’s last tetanus shot. If it has been more than 10 years, the injury victim gets a booster shot.
● Toxoids have also been made against theneurotoxin of Clostridium botulinum and some other toxins of Clostridium sp.
There are also some combined preparations where toxoids are utilized with another types of vaccines:
● DPT vaccine (diphtheria, pertussis, tetanus toxoids). Triple vaccine which is combination of two toxoids –tetanus and diphtheria- and killed Bordetella pertussis. This vaccine contains two different types of vaccines - toxoid and killed vaccines.
● Cholera combined vaccine. This vaccine contains two types of vaccines – toxoid plus killed Vibrio cholerae.
The newest types of vaccines are:
v SYNTHETIC PEPTIDES. Through the use of sophisticated laboratory technology, the amino acid sequence of purified protein antigens can be precisely determined. Once the sequence is known, it is possible to synthesize peptides of approximately 20 amino acids that may represent antigenic determinants against which protective immunity is directed.
Vaccines have been created using peptides of hepatitis В virus surface antigen, Streptococcus pyogenes M protein, diphtheria toxin, influenza virus, and a number of other proteins. The advantage of synthetic vaccines is that they ensure exclusion of contaminating materials that might harm the host. The widespread use of synthetic vaccines awaits the development of high-yield production systems.
v GENETICALLY ENGINEERED VACCINES. Several vaccine components are manufactured through genetic engineering using easily cultured bacteria or yeast to synthesize the protein antigen. Genetically engineered vaccines against influenza and hepatitis В are cheaper and safer than conventional whole-virus vaccines.
Another technological advance in vaccine production is the creation of piggyback vaccines. The genes for the desired antigens are inserted into the genome of an infectious, but harmless, virus. The engineered virus is inoculated into the host. As the virus replicates, it produces the "vaccine" protein along with its own products, The host then launches immune responses against the viral products and the extra antigen as well. Vaccinia virus, successfully used in smallpox eradication, is the most commonly used vector for piggyback vaccines.
v ANTI-IDIOTYPE VACCINES. Antibodies themselves can be used as antigens. They stimulate production of other antibodies targeted at their variable regions, the binding sites specific for the corresponding antigenic determinant. Each antibody for a distinct epitope is called an idiotype.
Anti-antibody antibodies recognize and bind to the antigen binding sites of the antibody idiotype that stimulated their production. These antibodies are aptly known as anti-idiotype antibodies. Because the original antigen and the anti-idiotype antibody have identical antigenic determinants, both can bind to the same antibody molecule, so anti-idiotype antibodies can replace a potentially harmful pathogen or toxin as the antigen in a vaccine. The advantage of anti-idiotype vaccines is that there is no pathogen present and therefore no possibility of reactivating the disease. Currently several such vaccines are under investigation.
Active immunoprophylaxis can be considered under two headings:
▪ Routine immunization of children which forms part of basic health care of communities
▪ Immunization of individuals or selected groups exposed to risk of particular infections.
Routine immunization schedules have been developed for different countries, based on the prevalence of infectious diseases, their public health importance, availability of suitable vaccines, their adverse reactions, etc.
The duration of response is of prime importance. For short-term protection (e.g. a tourist about to visit a disease area), the presence of antibody arising from the vaccine itself may be perfectly adequate and memory cells may not be strictly necessary. On the other hand, for protection against exposure at some time in future, the induction of memory is essential. Memory is often naturally boosted by periodic outbreaks of disease in the community.
¨ PASSIVE IMMUNITY
Antibodies produced by active immunity in one individual can be transferred in serum to a non-immune recipient. The antibody recipient is then said to have passive immunity. This type of immunity is characterized as follows:
1. Although these antibodies provide immediate protection, they are eventually depleted and are not replaced by the body.
2. Passive immunization, therefore, provides only temporary, short-term protection, usually lasting no more than a few weeks. This is commonly used for immunotherapy. Cell-mediated immunity can be passively transferred by immune lymphocytes derived from a sensitized donor.
Natural passive immunity helps to protect newborns from infectious disease. The fetus, which is incapable of producing antibodies, becomes passively immunized by maternal antibodies that cross the placenta from the mother to the fetus. Maternal antibodies persist for many weeks following birth and help protect the baby until he or she can actively produce antibodies. (Immunologic competence begins during the third to sixth month after birth and may not be adequate for 2 or 3 years.) Additional protection is provided by maternal antibodies that are passively transferred in breast milk. Breast milk also contains 25 percent of the mother’s daily production of monocytes. Such protection cannot be acquired from a commercial infant formula or cow's milk.
Artificial passive immunity is a valuable therapeutic tool when immediate protection is required, with no time to wait for active immunity to develop. Antibodies or immune T lymphocytes produced by hyperimmunized individual (human or animal) are injected into a recipient to provide temporary protection.
Antibodies of known specificity for passive immunization are administered as hyperimmune serum (antiserum), or specific immunoglobulins obtained from persons who have been actively immunized with an appropriate vaccine (see Table 15-6 and Table 15-7).
The main features of preparations for artificial passive immunity are as follows:
· Antiserum of human origin is called homologous, and antibodies that are taken from hyperimmunized animal arenamed heterologous.
· The antiserum (or immunoglobulin) may be specific (immune) or non-specific (normal). Hyperimmune human antisera are available for use against rabies, measles, pertussis, and tetanus. Pooled normal human immunoglobulin contains non-specific antibody to several pathogens and it is prepared from batches of plasma from 1000-6000 healthy donors after screening for hepatitis B, C and HIV.
· Immune antisera may contain antimicrobial or antitoxic antibodies, so such types of antisera may develop antimicrobial or antitoxic passive immunity.
· Protective antibodies can be prepared against several exotoxins that are lethal unless quickly neutralized in the body. Antiserum is available for use against botulism,tetanus, and the poisonous bites of spiders and snakes. If the antiserum contains antibodies that neutralize toxin, it is called antitoxin.
· Antitoxic immunoglobulins are produced by actively immunizing a horse with tetanus toxoid and concentrating the gamma-globulin fraction from blood collected from the animal several weeks later. Patients in danger of developing tetanus are injected with this antitoxin preparation, which rapidly neutralizes the potentially lethal exotoxin; these persons could otherwise die of the disease before they produced their own antibodies.
NOTE: This is why an injured patient will sometimes receive two tetanus shots, one a toxoid to boost active immunity for subsequent years of protection, the other an antitoxin preparation to provide immediate protection against the toxin. The two shots are given in different arms so they don't neutralize each other.
v monoclonal antibodies. Pure monospecific antibodies can be produced in cell culture in which all the cells in the population are descendants of a single parental antibody-producing cell. All the cells in this clone produce identical products. A culture grown from one isolated В cell produces immunoglobulin specific for a single antigenic determinant. Immunoglobulin manufactured by cloned cell cultures is called monoclonal antibody. A myeloma cell and an antigen-specific В lymphocyte can be fused to form a single cell, a hybridoma, which is committed to the production of the single antibody type specified by the В cell. Monoclonal antibodies are currently used in immunotherapy, diagnostic kits, and as a "guided missile" that delivers cytotoxic chemicals directly to tumor cells that bear the target antigen.
¨ Complications of Passive Immunotherapy
Antiserum can be raised in horses or other animals. Unfortunately, artificial passive immunization with animal antisera may cause severe complications, resulting from the immune response to the antibodies, which are of course foreign proteins. These include progressively more rapidelimination in 2-3 weeks (that reduced clinical effectiveness), and more severe complications as serum sickness and anaphylaxis. Serum sickness, a side effect that develops when the body eventually makes antibodies against the massive amounts of foreign protein found in the sera. Serum sickness (a model of type III hypersensitivity) follows repeated injections of foreign protein, leading to production of antibodies against the foreign (animal) immunoglobulins in antiserum used for passive immunization. Circulating immune complexes deposit in the kidneys, skin, and joints, which may be fatal. The danger of serum sickness increases with each successive exposure. Serum sickness may also follow passive immunization with human serum and human immunoglobulin preparations, but the danger is diminished because the proteins are less foreign. Passive immunization with a purified preparation of antibody specific for a single antigenic determinant further reduces the danger of serum sickness. Such uniformly specific antibody preparations are obtained by preparing monoclonal antibodies.
Another side effect of passive immunization with foreign immunoglobulins is sensitization – production of immunological memory following initial contact with an antigen (allergen), that stimulates the development of hypersensitivity (type 1). There are no symptoms, however until subsequent contact with the same allergen. Allergen exposure that elicits symptoms of hypersensitivity type 1 (anaphylaxis) in a sensitized host is termed the shocking dose.
Allergy desensitization (Bezredka’ method) is applied to prevent the development of anaphylaxis -generalized IgE-mediated allergic response. Allergy desensitization – introduction of small doses of allergen to eliminate IgE –mediated allergy.
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Besides MIBP for immunoprophylaxis and immunotherapy, there are preparations for diagnosis of infection. For diagnosis in vivo skin allergic tests (or allergological examination) are applied. MIBP for these tests are called allergens.
Skin allergic test – is an intradermal injection of antigen (allergen) to determine immunological sensitivity to a pathogen or its products. Allergen – is an antigen that elicits an allergic response (for example, PPD (purified protein derivative) from Mycobacterium tuberculosis cultures which is used for skin testing in tuberculosis). Skin test is an important tool for revealing the development of delayed type hypersensitivity and, therefore, for diagnosis of such infectious diseases where this type of allergic reactions develops.
The development of T effector cells to an antigen can often be revealed by intradermal challenge with that antigen. Such an intradermal challenge usually gives rise to erythema and induration, peaking at around 48 hours. This time course has led to the reaction being described as delayed-typehypersensitivity (type IV). In this type of hypersensitivity T lymphocytes, sensitized by an antigen (allergen), release lymphokines upon second contact with the same antigen. The lymphokines induce inflammation and activate macrophages. Delayed-type hypersensitivity is an allergic response produced by the cell-mediated immune system.
PRACTICAL WORK
The Vaccine Control
· Estimate vaccine safety by examination of its appearance (turbidity or transparency), if turbid – turbidity according to optical turbidity standard, morphology (by preparation of smear and its simple staining), sterility (growth on the nutrient media). For sterility estimation use the results of demonstration inoculations.
· Fill in Table 15-3 with the results of examination of two types of typhoid vaccines.
· Make a conclusion (whether the vaccines are standard or not).
Table 15-3