Vaccination: An approach to the control of infectious diseases

B A ALLSOPP, L A BABIUK AND S L BABIUK

Introduction

It must have been observed for centuries that people exposed to a pathogen were often resistant to reinfection by the same or a related pathogen, but the first record of anyone acting on this observation is that of the English physician Edward Jenner.⁵² On 14 May 1796 Jenner inoculated James Phipps, a healthy eight-year-old boy who had never been exposed to smallpox, with cowpox virus, and challenged him 48 days later with a virulent dose of smallpox virus. Phipps did not react, and with this momentous medical discovery was born the era of immunology and vaccinology. Since then smallpox has been eradicated by vaccination, and human and animal losses and deaths due to other infectious and parasitic diseases have been dramatically reduced by the development of a range of vaccines. Even so both animals and humans continue to suffer from many common diseases, as well as new and emerging diseases. Among many reasons for this are:

• sub-optimal management conditions;
• exposure to pathogens at a young age before vaccination is recommended;
• unusual or unexpected patterns of the pathogenesis of certain organisms;
• the emergence of new forms of old diseases;
• the emergence of totally new pathogens and diseases (e.g. Nipah virus); and
• the relative ineffectiveness of some existing vaccines.

Despite the current shortcomings of vaccination it seems likely that it is, and will continue to be, a more durable and cost-effective method of combatting bacterial and viral infections than all other therapeutic and prophylactic methods combined. Micro-organisms have survived for millions, even billions, of years in the face of a continuously changing chemical environment, and they are only going to be briefly inconvenienced by the human introduction of new chemicals into their surroundings. The same is true for arthropod vectors of disease. These facts rule out any long-term disease containment strategy based solely on the use of pharmaceuticals, acaricides or insecticides, however successful these may be in the short term. The development of new vaccines is the only long-term strategy that can be foreseen at present for the control of diseases of animals or humans. Mammalian immunological defence mechanisms against infectious agents have also evolved over millions of years, and must therefore be high up on any arbitrary scale of optimality. Finding new ways to specifically stimulate the immune system has to be the most likely way to achieve the lasting control of infectious disease.

Vaccines licensed for use in humans and animals at present are primarily either killed vaccines or attenuated vaccines, produced by one of two conventional methods. In the case of killed vaccines the pathogen is grown in large quantities and then inactivated by some method that prevents its replication but does not significantly alter the antigenic properties. Attenuated vaccines employ a live mutated agent that has been selected in vitro for reduced pathogenicity but that is still able to replicate in the animal. Each of these conventional vaccine types has inherent advantages and disadvantages, and there is a need to find new ways to make safer, cheaper and more effective vaccines. Advances in genomics, biotechnology, immunology, and understanding of the pathogenesis of diseases have resulted in the development of many new types of vaccines, such as subunit vaccines, live genetically modified vaccines, and nucleic acid (or ‘genetic’) vaccines.

Principles of vaccination

The primary mammalian immune response to an invading agent, involving the clonal expansion of small populations of virgin T and B cells, typically takes two weeks to develop, and this is often too slow to prevent serious disease occurring. The secondary response to any subsequent reintroduction of the same agent, however, involves the stimulation of a population of specific memory cells that were produced during the primary response. This is a larger population than that of the original virgin cells, so the secondary response is faster than the primary response, typically being less than one week.^{17, 45, 57, 66} The secondary response will usually clear the pathogen before overt disease occurs. Successful vaccination depends upon exploiting this mechanism by specific stimulation of the primary response with an agent inherently unable to cause disease. During any subsequent exposure to a virulent form of the pathogen the secondary immune response prevents the occurrence of disease. This seems to be a simple concept, but mammalian immune systems involve inordinately complex cascades of molecular signals and cellular responses. For effective immunization we need to know which antigens elicit the appropriate specific protective response, and how best to deliver those antigens, for example, administration of vaccines via the most commonly used routes, either intramuscular or subcutaneous, may induce good systemic immunity but may not induce mucosal immunity, thus failing to protect at the site of entry of many pathogens.^{51, 72, 73} If the virulence factor of a bacterium is an extracellular toxin, it will be inappropriate to use whole cell bacterins to protect against this particular infection. Until recently few of the infectious agents encountered in veterinary medicine were characterized at the molecular level, nor had the host immune responses to them been well characterized. The molecular genetic advances made in the last decade have begun to produce significant changes to this situation, and we may expect that the next decade will see many important advances in vaccinology.

Conventional vaccines

The majority of vaccines used today in veterinary medicine are either live attenuated or killed vaccines, produced according to principles first developed by Jenner and Pasteur over 200 and 100 years ago, respectively.¹⁰⁴ Both of these types of vaccines have been effective in reducing or eliminating clinical disease following exposure to virulent field strains of various pathogens. Equally important has been their role in curtailing the spread of pathogens to other animals by reducing the quantity of pathogen shed into the environment. These effects result in what is known as ‘herd immunity’ to a pathogen¹¹⁶ and if the transmission is reduced sufficiently the pathogen is eventually eliminated from the herd.²⁸

Live attenuated vaccines

Live attenuated vaccines are based on pathogens which have mutated sufficiently not to cause disease,andthe two most important considerations are whether the level of attenuation is sufficient to make the vaccine safe and whether the potential exists for reversion to virulence. Attenuation has conventionally been achieved by passaging the agent in vitro, either in the presence of mutagens or under specific culture conditions, and then selecting variants that show reduced virulence but which still induce a protective immune response. Over-attenuationmayresult in such limited replication that theimmuneresponse to a virulent challenge is not protective, and underattenuation will result in the vaccine itself causing clinical disease. Achieving the required fine balance between over- and under-attenuation is, however, purely empirical and there are many practical problems which make this conventional approach both tedious and expensive. Each mutant must be tested in vivo under a wide range of circumstances to ensure that it cannot cause disease, while still stimulating effective protection against virulent challenge. Special factors may have to betaken into account, for example,someanimalsmayreact adversely to vaccination if they are immunosuppressed due to stress, or pregnant animals may be induced to abort or foetal defects may develop when animals are vaccinated particularly during the first trimester of pregnancy with, for example, the attenuated Rift Valley fever and bluetongue vaccines (see Rift Valley fever and Bluetongue) ¹⁰⁰

Most important, however, is the fact that in the past there has been no information about which genes have been mutated and there has long been a fear that the mutated genes could back-mutate, or recombine in the field with virulent alleles, and cause the pathogen to revert to virulence.^{3, 74} Recent advances in the genetic analysis of attenuated vaccine agents haveshown that these fears were well founded.^{65, 77}

Disadvantages of live vaccines are the possible presence of contaminating extraneous viruses in vaccines grown in cell culture, for example bovine viral diarrhoea (BVD) into vaccines grown in bovine cells,^{56, 103, 114} that must replicate the vaccine candidate in order to induce a protective response, and that the presence of passive antibody, such as that from colostrum, will limit this replication. In many instances the level of passive antibody sufficient to interfere ith vaccination is below that which will prevent infection with wild types of the pathogen.⁵⁷ In such cases vaccination can only be performed after passive antibody has decreased to a predetermined level, which may leave young animals vulnerable to disease for long periods. It may also lead to many animals becoming susceptible at once, at the same time that they are exposed to multiple diseases simultaneously. Animals are therefore often vaccinated with multiple vaccines at the same time, but this provides the opportunity for interference to occur among the responses to different components of the cocktail.^{39, 47, 92}

Live vaccines also require appropriate storage conditions between manufacturing and administration, and the maintenance of a cold chain is often a critical factor. This is a serious difficulty in remote areas, particularly in rural Africa, although it is a problem which also affects inactivated vaccines.

Despite all these caveats there are some real advantages to attenuated vaccines as compared to killed vaccines. The live agent replicates in the host, so much less material has to be incorporated into each dose of vaccine than is the case for inactivated vaccines, making attenuated vaccines less expensive to produce. The immunity engendered by live vaccines is likely to be similar to that induced by natural infection, and of a longer duration, and broader, than that induced by a killed vaccine. This is especially true if the live vaccine is delivered via the mucosal route, which is the natural mode of infection for more than 90 per cent of all pathogens. The resulting mucosal immunity provides better protection at the likely site of entry of the pathogen, and should reduce shedding of the pathogen into the environment. This results in better herd immunity than that provided by killed vaccines, which do not generally induce mucosal immunity.

Inactivated vaccines

Inactivated, or killed, vaccines are produced by inactivating the infectious agent, so that it cannot replicate in the host, while still retaining the immunogenicity of the specific protection- inducing proteins. Most inactivating agents do, however, have an impact on protein immunogenicity and a balance must be sought between inactivation and reduction in immunogenicity.^{31, 34} Outbreaks of disease have occurred as a consequence of incomplete inactivation, a problem especially with pathogens that aggregate and thus limit the penetration of the inactivating agent.²⁰ Better inactivating agents are being developed, but testing the level of inactivation during the quality assurance phase is still vital. Even testing, however, would not have prevented the introduction of scrapie into sheep following immunization with an inactivated looping ill vaccine.⁴⁴ Obviously one cannot test for extraneous agents unless one has some information about what might be present.

A problem with inactivated vaccines, especially those produced from bacteria, is that some protective antigens are either not produced in vitro at all, or require special culture conditions for their expression. For example, some ironregulated outer membrane proteins of Mannheimia (Pasteurella) haemolytica, known to stimulate protection in vivo, are only produced under restricted iron growth conditions. ^{26, 27} Bacterial preparations may also lack sufficient concentrations of proteins which are secreted into the growth medium rather than being retained inside the bacterial cells.⁹⁵ An example of new ways of overcoming this deficiency will be dealt with under ‘subunit vaccines’ below.

The main disadvantage of inactivated vaccines is that they are relatively ineffective at generating protective immunity, particularly against intracellular bacteria and protozoa. This deficiency may be mitigated by the use of strong adjuvants⁶ (see further on), but in some cases it is the result of fundamental differences between the immune responses against live and killed organisms. For example, vaccination with heat-killed Listeria monocytogenes is not protective because it stimulates the production of memory T cells which fail to develop into effector CD8 T cells.⁶¹ This type of problem will be addressed by advances in the understanding of how antigenic epitopes bind to MHC receptors, and the development of algorithms which can predict potentially protective peptide epitopes.¹⁰²

Modern vaccines

Fundamental changes in biology are being brought about by the era of genomic data, and this is bound to impact powerfully on vaccine development. Currently in the public domain are 860 complete viral genomes, and 65 complete and 264 ongoing bacterial genomes. Volumes of genetic data such as these were unimaginable as recently as when the first edition of this book was published, and we will give just one example of the sort of work which is being facilitated by these changes. Bacillus Calmette-Guérin (BCG) is a live attenuated mixture of Mycobacterium bovis strains, originally developed at the Institut Pasteur in Lille, around the turn of the twentieth century, by Albert Calmette and Camille Guerin. After extensive testing, beginning in Paris in 1919, Calmette began to use BCG as a vaccine against tuberculosis in France in 1921²² and it has been, and still is, used for this purpose around the world. The effectiveness of BCG in any area is roughly inversely proportional to the distance of that area from the equator, and this is thought to be because in the tropics there are widespread infections with other, relatively benign, mycobacteria. It is surmised that these generate enough cross-immunity in the exposed human population that BCG does not develop sufficiently to stimulate protective immunity against tuberculosis. In order to unravel the molecular genetic fundamentals of this problem a comparative DNA micro-array expression analysis was performed on 13 strains of BCG, and on virulent strains of M. bovis and M. tuberculosis.¹¹ Subsequent genomic sequencing revealed that 91 open reading frames (probably genes) of virulent M. tuberculosis were absent from one or more virulent strains of M. bovis, and an additional 38 open reading frames that were present in virulent strains of M. bovis were absent from some or all of the 13 strains of BCG. This establishes a firm molecular genetic base from which to begin to analyse the reasons for the variable effectiveness of BCG vaccination, and to design better vaccine strains, in a way that Calmette and Guerin would not have begun to conceive.^{9, 10}

Subunit vaccines

Subunit vaccines are those which contain one or more pure or semi-pure antigens, and they can only be developed on the basis of an understanding of the mechanisms of pathogenesis and a molecular characterization of the gene products which stimulate protective immunity. Most of the gene products in any pathogen are irrelevant to the induction of protective immunity, and others may be immunosuppressive or even enhance the disease process, so the critical components of the pathogen’s complete translation repertoire must be identified.

Subunit vaccines can either be produced by in vitro expression from DNA recombinant constructs or by purifying a specific component from a conventionally produced vaccine, and the products may be used alone as vaccines or may be used to ‘spike’ conventional vaccines to increase the concentration of a particular protective component. For example, an extracellular leukotoxin of M. haemolytica is involved in pathogenesis and causes tissue damage in the lungs of cattle,⁹⁵ and immunity to this toxin is critical for inducing protection against the organism. It is possible to isolate and purify the leukotoxin from conventional cultures for incorporation into vaccines, but it is more economical to prepare the leukotoxin by recombinant biotechnology.⁴⁷ Subunit vaccines are safer than whole killed vaccines, and may be more effective. There is reduced antigenic competition between the constituents, one may target the vaccine to the site where immunity is required, and there is the potential for differentiating vaccinated from infected animals (see Marker vaccines, below).

A novel application of subunit vaccines has been their use to reduce disease transmission by targeting the vaccine against the arthropod vector.^{79, 115}Whenanimals are immunized against a specific glycoprotein present in the midgut of ticks, the antibody interacts with the protein in the tick midgut when the tick takes a blood meal. This interferes with the digestive process, thereby reducing both reproductive capacity and the viability of the ticks and therefore disease transmission.

Yeasts, mammalian, plant, and insect cells have all been used for the production of subunit vaccines1^{3, 69, 70, 105} and of particular interest currently is the possibility that genetic modifications can be made to food plants that can then be fed directly to animals for the induction of protective immunity. ⁴ Although theoretically attractive much work will need to be done before such a procedure can be demonstrated to be biologically safe and effective.

Live vaccines

The conventional method of producing live attenuated vaccine mutants had no molecular genetic basis. That situation has now completely changed, and it is now possible to pinpoint which genes are involved in virulence and stimulation of protective immunity. The degree of attenuation can therefore be controlled by deleting or mutating the appropriate genes.^{33, 35, 80, 84} The deletion of an entire gene makes it unlikely that the pathogen can revert to virulence, although recombination could theoretically occur with a virulent wild strain in the field. If an essential gene is deleted or inactivated from a virus for example, it is necessary to complement the gene function in vitro to allow the vaccine to be grown to high titres. After introduction of the vaccine into an animal an infection is initiated and protection is stimulated as the immune system sees almost the full complement of viral proteins. The lack of a replication cycle, however, means that the virus is not shed into the environment. ^{24, 33, 35, 81} A similar effect can be achieved for bacterial vaccines by modifying the organism to require a specific concentration of a compound to allow it to replicate.³⁰ This compound can either be introduced with the vaccine or fed to the vaccinated animals to allow the bacterium to replicate for a short time. They will thus be able to stimulate immunity but will not be viable when excreted into the environment.

Genetically engineered vaccines can also be used as vectors of genes expressing protective antigens from other pathogens. One of the first vectored vaccines to be tested was recombinant vaccinia virus expressing rinderpest genes¹¹⁸ and further developments have led to the development of a recombinant vaccinia virus vaccine that expresses both the fusion and haemagglutinin genes of rinderpest virus under the control of strong synthetic vaccinia virus promoters.¹¹¹ This vaccine provides cattle with long-term sterilizing immunity against rinderpest, it is safe, heatstable, inexpensive, and easily administered, and it allows serological differentiation between vaccinated and naturally infected animals. Another vaccinia-based vaccine, containing the gene which expresses the immunizing glycoprotein of rabies virus, has been incorporated into baits and distributed in areas of Europe where the fox population serves as a reservoir for rabies.^{15, 19, 82, 83} The use of this vaccine has resulted in the reduction or elimination of wildlife rabies from large areas of Europe. A number of recombinant vaccines based on avian pox viruses are also being tested or marketed for use in mammals.⁹¹ Since avian pox viruses cannot replicate in mammals these vaccines deliver the required antigens while undergoing an abortive infection cycle in vivo. A comprehensive demonstration of the immunostimulatory effectiveness of such vaccines comes from a recent study with an experimental HIV vaccine consisting of a canarypox vector containing Env, Gag, and Pro genes of HIV, which was used in combination with booster injections of recombinant HIV gp¹²⁰ subunit protein resulted in neutralizing antibodies in 91 per cent of subjects and CD8(+) T cell responses in 62 per cent of subjects.²

Another live vaccine vector technique which is being developed is the use of attenuated suicide strains of intracellular bacteria carrying recombinant eukaryotic expression plasmids, or naked DNA vaccine vectors, into host cells. The technique has been most highly studied using Listeria monocytogenes, and the vector bacteria have been shown to be lysed after entry into host macrophages and to release the recombinant plasmids.^{38, 98, 113} This is an alternative way of delivering DNA vaccine constructs (see Nucleic acid immunization, below).

Nucleic acid immunization

Nucleic acid immunization is the title recommended by the World Health Organization for a procedure which has also been called third generation vaccination, genetic immunization, naked DNA immunization, and polynucleotide immunization. The basis of this approach is to vaccinate with recombinant expression plasmids, containing protection stimulating genes, directly into host cells.^{29, 50, 117} Various delivery methods have been used to introduce these plasmids into cells, including injection, particle bombardment (‘gene gun’) and direct application to mucosal surfaces, all of which result in transfection of cells and expression of the foreign protein, which then induces an immune response.

There are many inherent advantages to nucleic acid vaccines. As a result of the endogenous production of the protein the immune response resembles that observed following a natural viral infection. Typical responses have been shown to include both cellular and humoral immunity, production of memory cells, and protective immunity of long duration.^{89, 90} The DNA plasmids naturally contain specificCpGsequences and this helps to create a cytokine microenvironment which enhances the immune response against the novel protein.^{25, 54, 59, 60, 99} By delivering the plasmids to mucosal surfaces, it is possible to induce mucosal immune responses and immunological memory.^{5, 36, 58, 64} Since a number of plasmids can be introduced simultaneously it should be possible to immunize animals against a variety of diseases at once without the concern of interference that is often observed with conventional vaccines.^{18, 47} Also of importance is the fact that any essential post-translational modification of the immunizing protein which may be required to induce specific immunity will occur naturally.

DNA vaccines can induce immunity in neonatal animals, even if they have maternal antibodies to the immunizing agent,¹⁶ and it is even possible to immunize foetuses.^{40, 41} If animals were vaccinated at birth they would first be protected by passive antibody acquired from the mother, and as this protection wanes active immunity would develop as a result of vaccination. This will ensure that there is no ‘window of opportunity’ for the pathogen to establish itself.^{33, 107}

Even with these many advantages no nucleic acid vaccines are licensed for any animal species and the main reason for this is the inability to deliver the vaccines effectively. DNA vaccination is very inefficient at present, with as much as 95 per cent of the plasmid being degraded before it enters the cells, and there is unlikely to be any major commercial uptake unless better delivery systems are developed to target the DNA into the nucleus of antigen-presenting cells.

One improved delivery system that is being investigated involves an RNA replicon vector derived from an attenuated strain of Venezuelan equine encephalitis (VEE) virus and packaged into VEE-like particles. The genes of interest were expressed from recombinant replicon RNAs that also encoded the replicase function and were capable of efficient intracellular self-amplification.^{46, 49, 85–87} Recent developments in replicon technology, based on the fact that the genes can be delivered in an alphavirus coat, ensures that well over 90 per cent of the plasmids enter cells, and the gene is not only protected by an artificial viral coat/envelope, but the envelope can be engineered in such a way that it targets the antigen-presenting (dendritic) cells.

If the problems of delivery of nucleic acid vaccines are overcome there will still be regulatory issues as potential barriers to the adoption of this technology.⁹⁷ There is concern over the use of antibiotic-resistant markers in plasmid selection, but since other selection markers could be used this problem should be easy to overcome. Of greater concern is the possibility of integration of plasmids into the cell nucleus, which could lead to activation of endogenous retroviruses or to deactivation of tumour-suppressor genes.⁷⁶

To date, however, the frequency of integration is considered to be less than five plasmids per 150 000 cells,^{71, 76} which is two orders of magnitude lower than normal mutations. Another concern is the production of anti-DNA antibodies following immunization with DNA, although experiments suggest that this possibility is almost non-existent. The induction of anti-DNA antibodies against naked DNA is known to be very difficult^{42, 43} and it has been observed that milligrams of DNA injected into cattle on numerous occasions over a period of more than a year failed to produce any detectable anti-DNA antibodies.¹⁰⁸ Only when DNAis delivered in Freund’s complete adjuvant is it possible to induce anti-DNA antibodies, so regulatory agencies are beginning to accept that this should not be a concern, especially in the case of livestock, where their lifespan is relatively short. There is still a concern that humans consuming animal tissues derived from livestock immunized with nucleic acid vaccines may suffer some untold consequence, and future testing will have to be performed before using such vaccinated animals for food.

Marker vaccines

The ability to differentiate animals that have been vaccinated from those that are potential carriers of an infection is an important requirement, and marker (DIVA – Differentiate Infected from Vaccinated Animals) vaccines have been developed for this purpose.^{109, 110} Efforts are underway to eradicate Aujeszky’s disease from the Netherlands by using a DIVA vaccination strategy,³² and similar vaccines are proposed for the eradication of bovine herpesvirus-1. The concept of DIVA vaccination can be applied to any disease situation and such vaccines are certain to become important in the future.^{1, 12, 14, 55, 96} Marker vaccines can also be used to complement quarantine and slaughter-out policies in countries where an exotic disease is introduced. It is possible to vaccinate animals at the periphery of an outbreak area and then to differentiate the vaccinated animals from those that have been exposed to the ‘wild’ infection. This is a far more economic and humane method of eradicating a disease than the conventional slaughter-out policy.^{8, 48, 112}

The basis for DIVA vaccines is the development of a recombinant vaccine from which a gene or genes have been deleted, and then to develop a parallel diagnostic kit which detects and compares the antibodies developed as a result of infection or of immunization. For example, animals infected naturally with Aujeszky’s disease or bovine herpesvirus-1 develop antibodies to all of the proteins of the virus. However, if the animal is vaccinated with a single glycoprotein (gD) or with a gE gene-deleted live vaccine, they will not develop antibodies to gE. An animal with antibodies to gE would therefore be considered to be a latent carrier of the virus.⁵⁵ Currently this approach is used to differentiate between animals that have recovered from foot-and-mouth disease (FMD) and may be carriers and those that have merely been vaccinated. Modern FMD vaccines are inactivated and do not contain significant quantities of nonstructural proteins of FMD virus, so vaccinated animals do not have antibodies to the non-structural proteins of FMD virus, in contrast to infected animals^{.67, 68}

Adjuvants

Both subunit and inactivated vaccines generally require formulation with an adjuvant to enhance immune responses in order to be effective. Purified antigens, however, are generally inherently weaker immunogens than whole killed bacteria, since the latter contain components such as lipopolysacharides and CpG sequences which act as adjuvants. An effective adjuvant is therefore particularly important for successful subunit vaccination. Few experimental adjuvants have been shown to be effective and most of them are expensive and often lead to adverse side reactions and tissue damage. For example, mineral oil-based adjuvants are often effective, but they are not metabolized and remain at the injection site, and for these reasons a number of countries in Europe are threatening to ban them. Similarly, aluminum hydroxide adjuvants are associated with fibrosarcomas in cats. ^{21, 62} Finally, even if an adjuvant induces good humoral immunity there is frequently very limited, if any, cell-mediated or mucosal immunity. Since these latter types of immune responses are critical for protection against many infections this is a serious concern.

Although the molecular mechanisms by which adjuvants exert their effect are not fully known, it is believed that they help create a micro-environment where the cells of the immune system can interact and create an appropriate cytokine cascade for attracting those cells of the immune system which lead to immunity. Since most adjuvants help retain antigen at the injection site, some believe that maintaining a depot of antigen is critical for the establishment of the required microenvironment for cell and antigen interaction.

Many natural and synthetic immune modulators are used or have been tested as potential adjuvants and it is convenient to classify them into mineral oils, plant, bacterial, or chemical/ synthetic adjuvants, although many adjuvants contain a combination of these components.¹⁰⁶ For example, Freund’s complete adjuvant is a mixture of a mineral oil and bacterial components, and a new adjuvant is being tested which combines aluminum hydroxide with a synthetic oligonucleotide containing CpG sequences.²⁵ Adjuvants have to act as delivery vehicles for the antigen, and the vehicles not only affect the adjuvanting properties butmayalso have adverse effects at injection sites. It has been shown that injection site reactions in cattle resulting in tissue damage cause significant economic losses.¹⁰⁶ As a result efforts are being made to find alternative methods of immunization or to find adjuvants that do not cause adverse tissue reactions.

Readers are directed to specific reviews for a more indepth treatment on adjuvants.^{53, 78, 93}

Passive immunization

The new-born of livestock acquire passive antibody from the colostrum of their dams but this immunity wanes relatively rapidly, leaving the neonate fully susceptible to infection by a variety of infectious agents within a few weeks or months of birth. Neonatal immunity can be extended by hyper- immunization of the mothers, so that higher levels of passive antibody are transferred, a technique which has been used in cattle and pigs to increase the immunity of neonates to diarrhoea caused by Escherichia coli, rotavirus, or coronavirus.^{7, 75} The dams are immunized prior to parturition and the elevated levels of antibodies in the colostrum and milk bathe the lumen of the neonatal gut, thereby preventing attachment of E. coli, rotavirus, or coronavirus to the intestinal villi. It is also possible to feed animals specific antibody for a short period of time when they are most susceptible to disease, an example being the use of monoclonal or polyclonal antibodies to the K99 antigen of E. coli to protect calves from E. coli induced diarrhoea.⁹⁴ Since the infection only occurs in the first week or two of life such an approach is economically feasible.

Passive antibody to a particular infectious agent tends to prevent effective immunization against that agent, especially when using vaccines involving live organisms. Replication of the vaccine agent is inhibited, and a protective immunity fails to develop. In most cases the level of passive antibody which will interfere with vaccination is higher than that which will prevent infection with field strains of the pathogen. We have noted above the closure of this ‘window of susceptibility’ by using nucleic acid vaccines. Since antibody in animals with passive immunity is not present at mucosal surfaces it is also possible to immunize them in the presence of colostral antibody by vaccine delivery to mucosal surfaces and thereby induce mucosal immunity.¹¹⁹ Finally, in utero vaccination with nucleic acid vaccines has been mentioned, and in ovo vaccination is gaining more popularity.^{37, 88, 101}

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