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UNIT 7: Immunology and Vaccinology Terminologies: Antigen: an antigen is a substance that elicits a specific immune response when introduced into the tissue of an animal. The response may consist of antibody production, cell-mediated immunity, or immunologic tolerance. Haptens: a hapten is a simple chemical or a portion of an antigen that cannot induce in immune response, but can react demonstrably and specifically with an appropriate antibody or cell. Antibodies: an antibody is a modified blood globulin which is fored in response to an antigenic stimulus. It is capable of combining specifically with the corresponding antigen. What is immunity? Immunity is the resistance of animals to disease causing organisms. It is made possible by the ability of the immune system to recognize foreign agents or tissues and eliminate them. On encountering the same foreign agent again, the immune system eliminates it more rapidly and effectively. There are different types of immunity. Active immunity develops in response to direct exposure to an infectious agent. This can occur naturally by exposure (natural immunity) or artificially by the process of vaccination (vaccine- induced immunity). Passive immunity is the temporary immunity conferred on an animal when it receives antibodies from another animal. In nature this occurs when new born animals absorb antibodies from the colostrums of the mother. These antibodies are taken up into the blood stream and are in the circulation for a few months. This gives the young animal time to develop its own active immunity. In the case of some diseases the presence of passive antibodies can suppress the formation of active immunity and this must be taken into consideration when vaccinating young animals. Passive immunity can be given artificially in the form of antisera or feeding colostrums to a new born animal. The immune system: The immune system is made up of specific types of cells which are found in various organs. Organs of immune system The organ called thymus regulates the production and maturation of the lymphoid cells which are produced in the bone marrow. The lymphoid cells circulate via the lymphatic system to the spleen and the lymphnodes where they filter out any invading organisms. The immune cells The immune or lymphoid cells are found in the circulation as well as in the organs discussed above. They form the component in the blood known as the white blood cells. The white blood cells consist of a number of different types of cells: Neutrophils (pus cells): they move to the site where micro organisms invade the body, engulf and destroy them with digestive enzymes. The neutrophils die in the process and do not play any role in the memory of the immune system. Macrophages: these are amoeba- like cells which take up micro organisms and break them into small parts which they then “offer” to other white blood cells called lymphocytes. Lymphocytes: they respond to the fragments offered to them by macrophages. There are two types, B lymphocytes and T lymphocytes. Macrophages and lymphocytes are the cells which are involved in the process of recognizing and remembering specific organisms. The immune process How does the immune process take place? It is a complex process but can be simplified by distinguishing two different types of reaction, although both may occur in response to a particular disease. Let us look at the sequence of events if we inject bacteria under the skin of an animal. 1. Neutrophils and macrophages migrate to the site of injection, and engulf bacteria. The bacteria taken up by bacteria are killed. The macrophages are carried to the nearest lymph node where they break down the bacteria in to smaller particles and then exhibit these on their cell surface. 2. Lymphocytes in the lymph node respond to these particles or antigens which are offered. They can respond in two different ways. 3. Lymphocyte response: if T- lymphocytes respond to the antigen presented they activate other lymphocytes called “killer cells “or cytotoxic lymphocytes which kill the invading micro organisms by direct contact. This is called cellular immunity. If B lymphocytes respond, they stimulate the production of antibody from a set of lymphocytes called plasma cells. This is called humoral or antibodymediated immunity. What is antibody and how does it work? Antibodies are protein molecules which are arranged in a Y-shape. At the tip of each branch of Y are sites which are able to recognise and attach to antigens or parts of infectious agents. Antibody fights infections by binding to an infectious organism and therefore immobilising it. This allows it to be caught and processed by macrophages. Specific antibodies are produced for specific disease-causing organisms. The memory of immune system Animals become immune to disease causing organisms after an initial encounter because their immune system has a “memory”. The “memory” is in fact a set of lymphocytes which recognises a disease-causing organism that has been encountered before. These memory cells may remain in an animal’s body for the rest of its life. Primary immune response On exposure to a disease-causing organism for the first time, antibody or cellular response is stimulated. The response gradually decreases with time. Secondary immune response When the animal is exposed to the same organism for a second time, the antibody or cellular response that is activated is greater and more rapid because of the memory of the immune system. Primary and secondary antibody response can be demonstrated for both antibody and cellular immunity. Antibody-mediated immunity: with the first exposure to an antigen the antibody response is slow and relatively weak while the second exposure is rapid and strong. Cellular immunity: this can be demonstrated by tissue transplants. A skin graft from mouse A to mouse B. Mouse B will be rejected eventually because the cells of its immune system recognise that the tissue is foreign. If the transplant is repeated, the rejection process will take place rapidly, as the immune system of mouse B has “remembered the transplant from A. Vaccinology A vaccine is a preparation containing an infectious agent which when administered to an animal will produce resistance to that specific agent. The animal is protected on a subsequent exposure to the same organism. There are various types of vaccines. 1. Live vaccine: live vaccine usually contain a live micro organism which has been weakened (attenuated) in some way, or a naturally occurring avirulent strain. Most live vaccines are in a freeze-dried form. Freeze-dried vaccine is presented in the form of a compact, dry pellet which must be mixed with sterile water before it can be injected. In this form the micro organisms in the vaccine can be successfully stored provided the vaccine is kept in a refrigerator and is not exposed to direct sunlight. 2. Inactivated vaccines: there are three types of inactivated or non-living vaccines. a. Killed vaccines: this is a vaccine made of micro organisms that have been killed using chemicals or heat. Killed bacterial vaccines are sometimes referred to as bacterins. b. Toxoids: these are toxins made of bacterial toxins which have been rendered non-toxic by chemical treatment. c. Subunit vaccines: subunit vaccines contain only certain parts or structures of a micro organism. Inactivated vaccines are usually less effective inducers of immune response than live vaccines and so they have chemicals added to them which improve the immune reaction. These chemicals are called adjuvants. The adjuvants may be mineral oils or potassium or aluminium salts. In some adjuvanted vaccines that have been standing for a while sediment may settle out to the bottom of the vaccine bottle. Vaccines containing adjuvants must be shaken well before use to mix all the components. Vaccine and cold chain maintenance The ‘cold chain’ is the system of transporting and storing vaccines within the temperature range of 35°F (2°C) to 45°F (8°C). The cold chain begins when vaccine is manufactured, moves through to the Dzongkhags distribution center and ends with the local immunization provider at the time of administration. Vaccine procurement and storage Vaccines after production or import are stored in cold chain at a temperature range of 2-8ºC. All the vaccines are initially stored at the central store at Serbithang. The vaccines are then distributed to different Dzongkhags according to the demand. The Dzongkhag in turn distribute to Livestock Extension Centres (LECs) for vaccination. During all this period from the manufacture to vaccination, cold chain has to be maintained. Essential equipment required to keep vaccines cold are: 1. refrigerator for storage, 2. Cold box for transport, ice packs to keep cool and separator for vaccine and ice pack. Placing vaccine in refrigerator Freezer sensitive vaccine in middle shelf. Diluents and emergency drugs in lower shelf Vaccine should be kept in refrigerator until just before administration. The vaccine should return into fridge after each dose. Cold chain Management Anyone handling vaccines is responsible for their potency, at each step in transport, storage and administration of vaccines. Vaccines are delicate biological substances that can become less effective or destroyed if they are: Frozen Allowed to get too hot Exposed to direct sunlight or fluorescent light Vaccines should be maintained within the recommended temperature range of 35°F (2°C) to 45°F (8°C). The loss of vaccine effectiveness is cumulative and cannot be reversed. Equipment for Transporting and Storing Vaccines The essential cold chain equipment needed to transport and store vaccines within a consistent safe temperature range include: A refrigerator for storing vaccines cold boxes for transporting and storing vaccines Ice packs to keep vaccines cool Material to separate ice packs from the vaccines when using cold boxes (e.g. shredded paper, cardboard, bubble wrap or Styrofoam). Refrigerators and Freezers It is recommended that one refrigerator should be dedicated to store vaccines. At least 50% of the space needs to be filled at all times to allow for adequate circulation of cold air, and to stabilize the refrigerator temperature. [Vaccines should be maintained with the recommended temperature range of 35°F (2°C) to 45°F (8°C)]. Maintenance of the vaccine refrigerator Safe vaccines storage is possible with refrigerators if the following procedures or modifications are carried out. Report breakdowns immediately so that repairs can be made. Regularly check refrigerator seals to ensure cold air cannot leak out. If they are brittle or torn arrange for replacement. Defrost refrigerators are required to prevent ice build up which reduces efficiency. Ensure the area around (including behind and under) the refrigerator is clean and dust free. Arrange for regular maintenance inspections by the manufacturer or technician. Inspections may need to be more frequent as the refrigerator ages. Defrosting or Cleaning When defrosting or cleaning the refrigerator, move the vaccines to second refrigerator. This temporary storage refrigerator must also be monitored to ensure the correct temperature 35°F (2°C) to 45°F (8°C) is maintained. If you don't have a storage refrigerator, store the vaccines in a pre-cooler insulted container with icepacks. Continue to monitor the temperature inside the container until the normal vaccine refrigerator is ready for use again. Power Failures During a power failure of 4 hours or less, the refrigerator door should be kept closed. If the power failure continues for more than 4 hours, store vaccines in an insulted container containing ice packs. If power failures are a common occurrence vaccines could be transferred to another site with power. Maintaining and Monitoring Refrigerator Temperatures The thermometer needs to measure the refrigerator temperature close to the vaccine vials. Choosing a thermometer to monitor the vaccine refrigerator: Digital-type thermometers or mercury minimum-maximum thermometers are the reliable and easiest to read. Ordering Vaccines Keep vaccine stock to a minimum by regularly ordering only the quantity of Vaccine required for the period until the next scheduled delivery. At least 14-day supply of vaccine should be avialable in the refrigerator when you are re-ordering. Storing excess vaccines supplies in the refrigerator can increase the risk of wastage from vaccines. Equipment for Transporting Vaccines A number of issues should be considered when choosing equipment to transport vaccines. They should include: Space required for the amount of vaccine transported Number of icepacks needed to keep the vaccines at less than 50°F The length of time in transit How the cold box will be handled Criteria for selecting a cold Box It needs to be large enough to store vaccines and icepacks during transport External surface material needs to be durable and robust Lid needs to be tight fitting Strong handles for carrying the cold box Icepacks, Freezer Blocks, Ice Blocks Icepacks are used to cool insulated containers or cold boxes. Icepacks are flat plastic containers that are filled with water and frozen. Some commercial icepacks contain a chemical product that ensures the icepack remains colder than 23°F (0°) longer than water-filled icepacks. Unpacking Vaccines after Transport Unpack vaccines from the transport container and store them in their packaging regardless of their bulkiness. Removing vaccines from the original packaging exposes vaccines to room temperature and light. Check the temperature to ensure the vaccines have not been exposed to temperatures above 35°F or below 32°F. Placing Vaccines in Refrigerator All freezer sensitive vaccines should be stored on the middle shelves Diluents and emergency drugs can be stored on the lower shelves Vaccines should remain in the refrigerator until immediately before they are administered. They should be returned to the refrigerator immediately after drawing up each recommended dose. Vaccine break downs Some times in the field the disease outbreak occur soon after the vaccination and the farmers lose faith in extension worker. Such a condition is called vaccine break down but the cause could be many for example: break in cold chain, mishandling like exposure to heat or light, some time due to the expiry of vaccine if not checked properly at the time of procurement. NOTE: Take all precaution to keep vaccine at a required temperature for better protection of animal health. How to use a vaccine Vaccines may be available from different manufacturers for the same disease and the directions for use of a specific vaccine must always be followed carefully. As the directions for using a vaccine are changed from time to time vaccines should never be purchased without the package inserts which contain the directions for use. It is essential to read the package inserts with regard to the following important information: 1. Storage: the vaccine must be stored at the recommended temperature. With the exception of heart water and frozen red water vaccines, which are kept frozen on dry ice or in liquid nitrogen, all other vaccines should not be frozen and are usually stored at temperature range of 2-8º C. Freezing and thawing destroys certain components in vaccines and therefore a vaccine which has been accidentally frozen must be discarded. Fridges used for vaccine storage should be checked regularly for efficient functioning. Maximum-minimum thermometers or commercially available colour indicators can be used for this. 2. Dose: the dose recommended by the manufacturer has been determined by extensive and expensive research. Reducing the dose to save vaccine will result in toxic or allergic reactions. 3. Route: the route by which the vaccine is administered is very important and use of the wrong route of vaccination may cause problems. 4. Age: the recommended age of vaccination is usually based on the age at which the animal is exposed to or becomes susceptible to that particular disease. 5. Season: some diseases are seasonal and vaccination must be carried out before the exposure period in order to be effective. Time must therefore be allowed for an immune response to develop. It must be borne in mind that for some diseases (e.g., blue tongue in sheep) a number of vaccinations must be given before the animal is fully immune and sufficient time must be allowed for this. Precautions when using vaccines: 1. Needle and syringe hygiene: needles and syringes used for vaccination must be sterile. During outbreaks of disease a single needle must be used for each animal. Using unsterilised needles and syringes can cause abscesses or generalised infections. Syringes must be set aside for administering Brucella abortus vaccine and only used for this purpose. 2. Exposure of vaccine to sunlight: the ultraviolet rays of the sun are damaging to all infectious agents and it is therefore essential that live vaccines are kept out of direct sunlight. All vaccines are degraded by heating and care must be taken to keep vaccines cool during transport and use in the field. A cool bag containing cool packs should be used for this purpose. 3. Mixing vaccines: different vaccines should not be mixed in the same syringe unless the manufacturer states that this can be done. Vaccines must also not be mixed in syringes with any other injectable preparations (vitamins, antibiotics, dewormers, etc) as chemicals contained in the latter may damage the vaccine components. 4. Vaccinating during outbreaks: vaccinating during outbreak is fraught with problems and should be avoided by implementing a well-planned vaccination program. Apart from the fact that animals may already be incubating disease when they are vaccinated, immunity takes time to develop and animals may sometimes be infected before they can develop immunity. Another consideration is that diseases can be transmitted from animals incubating disease to healthy stock, by inoculation needles. Needle transmission can occur with the following diseases: lumpy skin disease, anthrax, black quarter, anaplasmosis, etc. It is essential a use a separate needle for each animal during outbreaks of these diseases. Vaccine failure Vaccinating an animal does not necessarily protect it against disease. There are many reasons why a vaccine may fail to give protection and they can be divided into problems arising from: 1. the VACCINE itself 2. the ADMINISTRATION of the vaccine 3. the ANIMAL being inoculated 1. The Vaccine a. Damage to vaccines: damage caused to a vaccine by exposing to heat, direct sunlight, freezing or storage at room temperature for too long a time will result in a failure of effective immunization. b. Expired vaccines: Expired vaccines should never be used. c. Wrong vaccine used due to wrong diagnosis: vaccines are often used without a specific diagnosis being made. This usually happens when a vaccine is used based on the occurrence of one single symptom. d. Serotypes or strains not included in the vaccines: some micro organisms have more than one type of the same organism. These different types are called SEROTYPES. In some cases the different serotypes do not cross-protect against each other and if a certain serotype is not included in a vaccine there will be no protection. e. Sub-standard vaccine: a poor quality vaccine may have been issued from the manufacturer and if after eliminating all the other reasons for vaccine failure, this is suspected, the company in question should be contacted. 2. Administration of vaccine a. Antibiotics and live bacterial vaccines: live bacterial vaccines like anthrax, salmonella, brucella must not be used simultaneously with antibiotics because the bacteria will be killed off and no immune response will be produced. b. Transmission of disease during vaccination: some diseases can be transmitted during vaccination. c. Vaccinating at the wrong age: in many cases there is an optimal age for vaccination. Examples are pulpy kidney vaccine which does not stimulate immunity before two months of age in lambs, and Rev. I vaccine used to prevent brucellosis in rams does not give good protection if it is given after weaning age. d. Vaccinating too late: vaccinating animals during an outbreak will result in animals becoming infected before immunity can develop. e. Vaccinating during disease incubation: animals may sometimes be incubating a disease at the time of vaccination, and it may appear that the vaccine has failed to protect or that the vaccine itself has caused the disease. f. Failure to give boosters: booster vaccinations for killed vaccines are essential for the development of good immunity. The manufacturer will recommend when the booster must be given. g. Poor management: vaccines designed to prevent diseases of new born animals are given to the mothers to raise the antibody level of the colostrums or the “first milk”. If the new born animal does not receive colostrums for any reason, the vaccine will not prevent disease. h. The interaction of vaccines: live vaccines may interfere with each other, and more than one live vaccine should not be administered at the same time unless this is recommended by the manufacturer. i. Wrong dose: too little vaccine may be given deliberately to save vaccine or by accident if an automatic syringe is not calibrated correctly or is sticking, or if a freeze-dried vaccine is diluted too much. Administering insufficient vaccine will almost certainly result in poor immunity, or occasionally “sensitization” which results in exacerbation of disease. 3. The animal being vaccinated a. Animals in poor health: animals in poor health condition because of starvation, vitamin or mineral deficiencies, heavy parasite infestations or chronic diseases will not respond well to vaccination. b. Passive immunity: the antibodies in the colostrums of a mother are transferred to the new born animal and provide what is called PASSIVE or maternal immunity. This protection declines after a few months. The young animal must then develop its own active immunity. If certain vaccines are given to a very young animal, the presence of passive immunity will prevent the development of ACTIVE immunity. Therefore if a mother has been vaccinated with vaccine A and the passive immunity is to last for 3 months, the best time to vaccinate the young animal against A is roughly at 3 months of age. c. Exposure to severe challenge: the vaccine immunity can be neutralized or overwhelmed if there is massive exposure to an infectious agent. d. Population response: in a population of animals, the ability to develop an immune response follows follows a normal response curve. This means that most of the individuals in a population have the ability to to develop adequate immunity, while a small percentage develops poor immunity. Therefore in any population not all animals will be solidly immune. THE SIDE EFFECTS OF VACCINATION Manufacturers of vaccines are required to do extensive safety testing before a product can be sold for use in the field. In most cases vaccination is therefore an uneventful procedure but some side effects can occur even with highly purified products. Some of the side effects that can occur are discussed below. 1. allergic reactions: allergic reactions can be either immediate “shock” type reactions or local reactions at the site of injection 2. Local non-allergic reactions: some vaccines contain substances (adjuvants) which are included to improve the immune response. The adjuvants may produce irritation and swelling at the site of injection which takes a few days to develop and a few weeks to subside. This type of reaction when it occurs will be seen in a large proportion of the vaccinated animals. 3. Other post-vaccination problems: other adverse responses to vaccination (e.g., abortion, drop in milk production, etc) can occur with certain vaccines. 4. Accidental injection of humans: this should be avoided because although most veterinary vaccines are harmless to humans many can cause severe swelling and pain at the site of injection. Vaccination Schedule for Livestock Vaccine Species Dosage Route Primary Booster Anthrax Spore vaccine (Locally produced) Cattle buffalo & yak. Sheep, goat & pig Cattle buffalo & yak. Sheep, goat & pig 1 ml s/c 3 months Not required Revaccinatio n Annually in March/ April s/c 3 months Not required Annually in March/ April Every 6 months in April/may & Aug/Sept. Every 6 months Black Quarter Vaccine (locally produced) 0.5 ml 5 ml 2 ml Haemorrhagic septicaemia (locally produced) FMD tri-valent oil-adjuvant Vaccine (importedIndia) Cattle & buffalo 4 ml s/c 3 months 6 months Cattle buffalo & yak. Sheep, goat & pig 2 ml I/m 3-4 months 1 month Neonatal enterotoxicosis vaccine (Imported-UK) Pigs 2 ml i/m Gilts & sows 5-6 weeks Every 6 months Antirabies vaccine (importedIindia & France) Dogs & cats 1 ml s/c or i/m 1 month 1 month Classical swine fever vaccine (locally produced) E.coli (oral ) Vaccine (locally produced) Newcastle Disease thermostable I2 vaccine (locally produced) Pigs 1 ml s/c 45-60 days Not required Annually in endemic areas & once in 2 years in non-endemic areas. Annually 200 ml mixed with 1.5 kg feed. 1 drop Orally 75 days of gestation - - Eye drop All ages 3 months Every 3 months Pigs (pregna nt sows) Poultry 1 ml 2 months Remarks Vaccination not required if there has been no outbreak in a particular area for three years after the last outbreak. Vaccinate animals upto 3 years of age. In endemic areas healthy adults may be vaccinated. Primary vaccination should be done when the calf reaches 3 months of age & not wait for annual vaccination programme. Vaccination to be carried out in the risk areas only & prior to the on set of monsoon and migration. High risk & medium areas- twice a year vaccination. Low risk areas- annually. In pigs, breeding stock should be revaccinated at 6 months interval. Vaccinated at least 21 days before migration. All the breeding stock present in the farm may be vaccinated at the same time. Avoid vaccination of sows during two weeks prior to farrowing. - May produce hypersensititve reaction in animals. Adrenaline/antihistamine should be given immediately in such reactions. The vaccine should be given for 3 consecutive days starting from 75 days of gestation. If primary vaccination is given via drinking water, booster dose should be given after 3 weeks and then re-vaccination to be done every after 3 months. Throough eye-drop, onl;y re-vaccination to be done every 3 months. Poultry Vaccination Schedule in the Organised Farms Age Vaccine Route Dose Remarks Day 1 Mareks Disease s/c Life-long immunity Day 3 IBD Intermediate Strain (first dose) Newcastle Disease B1 strain Eye drop 0.2 ml /chick or as per manufacturer’s instruction 0.03 ml or as per manufacturer’s instruction 0.03 ml or as per manufacturer’s instruction Day 14 IBD intermediate strain (2nd dose) Day 28 or 30 IBD intermediate strain (3rd dose) Day 42 Fowl pox Eye drop or drinking water Eye drop or drinking water i/m Day 56 ND R2B strain (booster) s/c or I/m Day 98 Fowl pox (repeat) i/m Day 120 ND R2B strain s/c or i/m Day 7 Eye drop 0.03 ml or as per manufacturer’s instruction 0.03 ml or as per manufacturer’s instruction 0.02 ml or as per manufacturer’s instruction 0.05 ml or as per manufacturer’s instruction 0.02 ml or as per manufacturer’s instruction 0.05 ml or as per manufacturer’s instruction In case the locally produced I-2 ND vaccine is used it has to be repeated every 3 months@ 1 drop eye drop