Veterinary · Para-clinical · also MSVE VMC-221
Innate and adaptive immunity, antibodies and complement, hypersensitivity, vaccines and the cold chain, and the serological tests used in diagnosis and surveillance.
In the exams: ICAR AICE-JRF core group 06; ICAR AIEEA-PG 13 (Microbiology); NAVLE health maintenance and prevention 35%
Veterinary immunology is the study of how animals defend themselves against infection and how that defence can be measured, strengthened or turned against the animal's own tissues. It covers the innate defences that act within minutes, the adaptive responses of T and B lymphocytes that take days but leave memory, the antibodies and complement that do much of the work in blood and on mucosae, and the major histocompatibility complex that decides what a T cell can see. Serology, the use of antibody–antigen reactions in the laboratory, is the practical arm of the subject: most herd screening, export certification and disease surveillance in the world rests on an ELISA, an agglutination test, a complement fixation test or an agar gel immunodiffusion test.
The subject matters in practice for three reasons. First, vaccination is the single most common procedure in veterinary medicine, and every decision about a vaccine type, a schedule, a storage failure or an apparent vaccine breakdown is an immunological decision. Second, the neonate of every farm species is born almost without circulating antibody and depends on colostrum; failure of passive transfer is one of the largest preventable causes of calf, foal, lamb and piglet death. Third, many diseases that owners bring to a clinic, such as flea allergy, atopic dermatitis, immune-mediated haemolytic anaemia and neonatal isoerythrolysis, are immunological diseases, and the tuberculin test that underpins bovine tuberculosis control is itself a delayed hypersensitivity reaction.
Worldwide the subject is examined as a core paper. In India it is part of the microbiology group of the ICAR postgraduate and JRF/SRF examinations and of state veterinary officer papers; in China microbiology and immunology form the largest single block of the preventive paper; the NAVLE, the RCVS statutory membership examination and the Australasian examination test it through vaccination, test interpretation and immune-mediated disease in clinical cases. Numbers that vary by country, species, laboratory or product are flagged as such in this manual; always use the cut-offs and schedules your national authority, laboratory and product label specify.
Innate immunity is the set of defences present before an animal meets a particular pathogen. It acts within minutes to hours, needs no prior exposure, and recognises broad classes of microbe rather than single antigens. Its first layer is physical and chemical: keratinised skin, mucus, ciliary clearance, gastric acid, bile, antimicrobial peptides such as defensins and cathelicidins, and the normal microbiota, which competes with pathogens. Breaching a barrier opens the way to infection; in bovine respiratory disease, viral damage and transport stress impair the mucociliary escalator so that Mannheimia haemolytica can colonise the lung. Innate cells detect pathogen-associated molecular patterns (PAMPs), conserved microbial molecules that host cells lack, and damage-associated molecular patterns released by injured cells, through pattern recognition receptors (PRRs). Among the Toll-like receptors (TLRs), TLR4 recognises lipopolysaccharide, TLR5 flagellin, TLR2 (with TLR1 or TLR6) bacterial lipoproteins, TLR3 double-stranded RNA, TLR7 and TLR8 single-stranded RNA, and TLR9 unmethylated CpG DNA. Cytoplasmic sensors include RIG-I-like receptors for viral RNA and NOD-like receptors, some of which form inflammasomes that release interleukin-1β and IL-18. PRR signalling drives inflammatory cytokines and, after viral sensing, type I interferons (α and β), which put neighbouring cells into an antiviral state. Many adjuvants act on these same receptors.
Inflammation brings cells and plasma proteins to a site of infection through vasodilation and increased vascular permeability, producing heat, redness, swelling, pain and loss of function. Neutrophils arrive first: they roll on endothelium using selectins, bind firmly through β2 integrins (CD11/CD18) and migrate along chemokine gradients. This adhesion step fails in leucocyte adhesion deficiency (Chapter 8). Systemically, IL-1, IL-6 and tumour necrosis factor-α cause fever and switch hepatic protein synthesis, the acute phase response. The major positive acute phase protein differs by species: haptoglobin and serum amyloid A (SAA) in cattle; SAA and fibrinogen in the horse; C-reactive protein (CRP) and SAA in the dog; SAA and α1-acid glycoprotein in the cat; CRP, haptoglobin and pig major acute phase protein in the pig. Albumin is a negative acute phase protein and falls. Reference ranges depend on the assay and laboratory, so results are interpreted against the laboratory's own values and the clinical picture.
Neutrophils kill by phagocytosis followed by the respiratory burst, in which NADPH oxidase generates superoxide and myeloperoxidase makes hypochlorite, and by granule enzymes and extracellular DNA traps; they are short-lived and form pus. Macrophages come from blood monocytes and tissue-resident populations such as Kupffer cells and alveolar macrophages. Interferon-γ activates them classically to kill intracellular bacteria such as mycobacteria, whereas IL-4 and IL-13 give alternatively activated macrophages that favour repair. Natural killer (NK) cells kill virus-infected and some tumour cells that have lost MHC class I, the 'missing self' principle, and secrete interferon-γ early in infection. Mast cells release histamine at skin and mucosae; eosinophils attack helminths. Dendritic cells capture antigen in tissues, mature after PRR signalling, and migrate to lymph nodes to activate naive T cells. They are the bridge to adaptive immunity: without innate activation of dendritic cells, a purified antigen tends to induce tolerance rather than immunity, which is why most non-living vaccines need an adjuvant.
Adaptive immunity is specific, diverse, slow on first exposure and remembers. Each lymphocyte carries one receptor specificity, created by random rearrangement of V, D and J gene segments by the RAG1 and RAG2 enzymes. A lymphocyte that meets its antigen with co-stimulation proliferates into a clone (clonal selection) and leaves long-lived memory cells. The primary response has a lag of one to two weeks and is mainly IgM; the secondary response is faster, larger, longer-lasting and dominated by higher-affinity IgG, the basis of booster vaccination and paired serology. T cells mature in the thymus in all species; B cell development differs. In primates and rodents it occurs in bone marrow. In birds the bursa of Fabricius, a dorsal outgrowth of the cloaca, diversifies B cells mainly by gene conversion, which is why infectious bursal disease virus leaves young chicks immunosuppressed. In sheep, cattle and pigs the ileal Peyer's patch acts as a primary B cell organ early in life, and in rabbits the appendix. Among secondary organs, pig lymph nodes are 'inverted', with lymphocytes leaving mostly through blood; ruminants have haemal nodes filled with blood; and birds lack true lymph nodes, relying on the spleen, caecal tonsils, Harderian gland and diffuse lymphoid tissue.
T cells recognise peptide on MHC through the T cell receptor, associated with CD3. Helper T cells carry CD4 and see MHC class II; cytotoxic T cells carry CD8 and see MHC class I. Naive helper cells differentiate according to local cytokines. Th1 cells, induced by IL-12, secrete interferon-γ, activate macrophages and support cytotoxic responses, the protective pattern against intracellular bacteria such as Mycobacterium and Brucella and many viruses. Th2 cells, induced by IL-4, secrete IL-4, IL-5 and IL-13, promote IgE and eosinophils, and dominate responses to helminths and allergens. Th17 cells secrete IL-17 and recruit neutrophils at mucosae. Regulatory T cells (Tregs), marked by the transcription factor FoxP3, suppress responses through IL-10, TGF-β and contact, and prevent autoimmunity. Cytotoxic T cells kill infected cells with perforin and granzymes and through Fas ligand. Most T cells use an αβ receptor; γδ T cells are a minority in humans and mice but form a large fraction of blood T cells in young ruminants and pigs and are abundant in epithelia.
B cells recognise intact antigen through surface immunoglobulin. For protein antigens, a B cell presents peptide on class II and receives help from a follicular helper T cell through CD40–CD40 ligand and cytokines. This T-dependent response forms germinal centres where class switch recombination changes the heavy chain from IgM to IgG, IgA or IgE with the same specificity, and somatic hypermutation with selection raises affinity over weeks (affinity maturation). The products are long-lived plasma cells, many resident in bone marrow, and memory B cells. Repetitive antigens such as bacterial polysaccharides can activate B cells without T cell help; these T-independent responses are mostly IgM with little memory and are weak in the very young, which is why polysaccharides are conjugated to protein carriers. Most pathogens enter across mucosae. M cells over Peyer's patches sample luminal antigen, and activated lymphocytes home back to mucosae. The main mucosal antibody in most species is secretory IgA, a J-chain-linked dimer carried across epithelium by the polymeric immunoglobulin receptor and protected by secretory component; it neutralises and blocks attachment without inflammation. Mucosal vaccines given by nose, mouth, eye drop or drinking water, as with many poultry vaccines, exploit this.
An antigen is any molecule that can be bound by an antibody or a T cell receptor. An immunogen is an antigen that can also provoke an immune response on its own; all immunogens are antigens, but not all antigens are immunogens. Immunogenicity rises with foreignness, molecular size (proteins above roughly 10 kDa tend to be good immunogens), chemical complexity and the ability to be processed by antigen-presenting cells. Proteins are the best immunogens, polysaccharides are moderate and largely T-independent, and lipids and nucleic acids are poor on their own. A hapten is a small molecule that is antigenic but not immunogenic unless coupled to a carrier protein; drugs such as penicillin can act as haptens by binding host proteins, one route to drug allergy. An epitope is the small part of an antigen actually bound. B cell epitopes may be linear or conformational; T cell epitopes are short linear peptides displayed on MHC. Dose, route and adjuvant also shape the response, and very high or very low doses can induce tolerance.
Antibody binds antigen by non-covalent, reversible forces: hydrogen bonds, electrostatic attraction, van der Waals forces and hydrophobic interactions, all effective only at short range, so a close fit is needed. Affinity is the strength of one binding site for one epitope; avidity is the overall strength of a multivalent antibody binding a multivalent antigen, so pentameric IgM has modest affinity but high avidity. Cross-reactivity, binding to a different antigen with a similar epitope, explains many false-positive serological results, for example Yersinia enterocolitica O:9 antibodies reacting in Brucella tests because the two share a similar O-polysaccharide. Antibodies protect by neutralising viruses and toxins, opsonising microbes for phagocytes bearing Fc or complement receptors, activating classical complement, and directing NK cells to kill antibody-coated cells (antibody-dependent cell-mediated cytotoxicity). Antibody works best against extracellular agents and toxins; once a pathogen is inside cells, cell-mediated immunity matters more. Tetanus is prevented by a toxoid that induces neutralising antibody, whereas protection against tuberculosis depends on Th1 cells and activated macrophages.
A natural immune response is polyclonal: many B cell clones make antibodies to many epitopes of an antigen. A monoclonal antibody comes from a single clone and recognises one epitope. It is made by fusing antibody-producing B cells with a myeloma cell line to form an immortal hybridoma, or by recombinant methods, and it gives competitive ELISAs and typing reagents their specificity. When soluble antigen and antibody are mixed in varying proportions, precipitate follows the precipitin curve: little in antibody excess, maximum at equivalence where large lattices form, and little again in antigen excess, where small soluble complexes remain. The prozone effect is a false-negative agglutination or precipitation result in great antibody excess; further dilution of the serum reveals the reaction. The small soluble complexes formed in antigen excess are the ones that deposit in vessel walls and glomeruli in type III hypersensitivity, and the equivalence zone is what makes a precipitin line visible in agar gel immunodiffusion.
The basic immunoglobulin unit has two identical heavy chains and two identical light chains joined by disulphide bonds into a Y shape. Each chain has an amino-terminal variable domain and constant domains; three hypervariable loops (complementarity-determining regions) in each variable domain contact the epitope. Papain cuts the molecule at the hinge into two Fab fragments, each with one binding site, and one Fc fragment carrying effector functions such as complement activation and Fc receptor binding. Pepsin yields one F(ab')2 fragment that still cross-links antigen. Light chains are kappa or lambda, in ratios that differ by species; lambda predominates in cattle, horses and cats, kappa in mice. The heavy-chain constant region defines the class: IgM (μ), IgG (γ), IgA (α), IgE (ε) and IgD (δ). Clinically, IgG in neonates is measured by radial immunodiffusion, the reference method, or by ELISA and turbidimetric assays, and estimated by refractometry. On serum protein electrophoresis, a broad polyclonal rise in γ globulin suggests chronic antigenic stimulation, while a narrow monoclonal spike suggests a plasma cell or B cell tumour, although some infections, such as canine ehrlichiosis, occasionally give a narrow peak.
IgM is the first antibody of a primary response and the first on naive B cells. Secreted IgM is usually a pentamer with a J chain; its ten binding sites give high avidity, it is the most efficient activator of classical complement and a strong agglutinator, and being large it stays mainly in blood. A high specific IgM suggests recent infection. IgG is the main antibody of serum and of secondary responses, a monomer that diffuses into tissues, neutralises and opsonises, and in most mammals is the class passed from mother to young. IgA is the chief antibody of secretions, often a dimer carrying secretory component, and in serum a monomer in humans, while domestic species differ, with much serum IgA polymeric in several of them. IgE is present in very low serum concentration but binds tightly to the high-affinity Fc epsilon receptor on mast cells and basophils; cross-linking by antigen triggers degranulation, the basis of type I hypersensitivity, and IgE helps expel helminths. IgD serves mainly as a surface receptor on naive B cells, and its full function is still not settled.
Immunoglobulin genetics vary among domestic animals, with clinical consequences. Cattle have IgG1, IgG2 and IgG3; IgG1 is selectively transported from blood into colostrum and milk, so bovine colostrum is dominated by IgG1, and in ruminants IgG1 rather than IgA remains the main immunoglobulin of mature milk. In non-ruminants such as the pig and horse, colostrum is rich in IgG but mature milk becomes dominated by IgA. Horses have several IgG subclasses, encoded by seven heavy-chain genes, that differ in complement fixation and Fc receptor binding. Pigs, dogs and cats also have several IgG subclasses. Birds have IgM, IgA and IgY. IgY is the functional avian equivalent of IgG, the main serum antibody, transferred from hen to chick through the egg yolk and harvested from yolk as a laboratory reagent; birds have no IgE. Camels, llamas and alpacas make, besides conventional antibodies, heavy-chain-only IgG lacking light chains; its single variable domain can be produced as a very small binding fragment (a nanobody or VHH) for research and diagnostics.
Complement is a set of more than thirty plasma and membrane proteins, mostly made in the liver, that act as a proteolytic cascade. Three pathways converge on cleavage of C3. The classical pathway starts when C1q binds the Fc regions of antigen-bound antibody: one surface-bound IgM pentamer is enough, whereas IgG must be clustered. C1r and C1s are activated, C1s cleaves C4 and C2, and C4b with the larger C2 fragment forms the classical C3 convertase, written C4b2a in older texts and C4b2b in current nomenclature. The lectin pathway starts when mannose-binding lectin or ficolins bind microbial carbohydrate; associated serine proteases (MASPs) cleave C4 and C2 to form the same convertase without antibody. The alternative pathway depends on slow spontaneous hydrolysis of C3 in plasma ('tick-over'). C3b deposited on a microbial surface binds factor B, which factor D cleaves, giving the alternative C3 convertase C3bBb, stabilised by properdin. Because every convertase makes more C3b, the alternative pathway amplifies whichever pathway started the process.
Cleavage of C3 is the central event. C3b coats surfaces and binds complement receptors on phagocytes, making it the major opsonin in most infections; its product C3d, bound to antigen, engages the B cell co-receptor and lowers the threshold for B cell activation, linking complement to antibody responses. The small fragments C3a and C5a are anaphylatoxins that degranulate mast cells and increase vascular permeability, and C5a strongly attracts neutrophils. When C3b joins a C3 convertase it forms a C5 convertase; C5b then assembles with C6, C7, C8 and several C9 molecules into the membrane attack complex, a pore that lyses Gram-negative bacteria, enveloped viruses and cells such as erythrocytes. Complement also solubilises immune complexes and helps clear them; defective clearance favours immune complex disease. Complement proteins are heat labile, so heating serum at 56 °C for 30 minutes inactivates it; this is done before complement fixation and neutralisation tests so that the serum's own complement does not interfere. Guinea pig serum, with high and stable haemolytic activity, is the conventional complement source in the complement fixation test.
Because complement can damage host cells, it is tightly regulated. C1 inhibitor controls C1r, C1s and the MASPs. Plasma factor H and factor I inactivate C3b. Membrane regulators, including decay-accelerating factor (CD55), membrane cofactor protein and CD59, which blocks assembly of the pore, protect host cells. Some microbes evade complement by binding factor H or mimicking regulators. Two inherited deficiencies in domestic animals are standard examples. Hereditary C3 deficiency in Brittany spaniels causes recurrent bacterial infections and membranoproliferative glomerulonephritis. Factor H deficiency in Yorkshire pigs allows uncontrolled alternative pathway activation, consumption of C3 and complement deposition in glomeruli, causing fatal membranoproliferative glomerulonephritis in young piglets. In the laboratory, some sera are 'anticomplementary', destroying complement without antigen because of bacterial contamination, haemolysis, immune complexes or poor storage; such sera cannot be read in the complement fixation test and are reported as anticomplementary rather than positive or negative. Bovine serum also contains conglutinin, a lectin that binds fixed complement, exploited in the conglutinating complement absorption test.
The major histocompatibility complex (MHC) is a gene cluster whose products present peptides to T cells. It was discovered through graft rejection, but its real job is antigen presentation. MHC class I molecules consist of a polymorphic α chain with β2-microglobulin and are expressed on almost all nucleated cells, including the nucleated erythrocytes of birds but not mammalian erythrocytes. MHC class II molecules have α and β chains, both MHC-encoded, and are expressed constitutively on dendritic cells, macrophages and B cells; in pigs, dogs, cattle, horses and cats, resting T cells can also express class II. The class III region encodes other immune proteins, including some complement components and tumour necrosis factor. Each species' MHC has its own name: HLA in humans, BoLA in cattle, OLA in sheep, SLA in pigs, ELA in horses, DLA in dogs and FLA in cats. The chicken MHC, the B complex on microchromosome 16, is compact and has been called a minimal essential MHC.
Class I presents peptides from proteins made inside the cell, including viral proteins. The proteasome degrades them, the TAP transporter moves peptides into the endoplasmic reticulum, and loaded class I molecules go to the surface for CD8 cytotoxic T cells. Class II presents peptides from proteins taken up from outside the cell and degraded in endosomes; the invariant chain blocks the class II groove until its fragment CLIP is exchanged for an antigenic peptide with the help of a chaperone (HLA-DM in humans), and the complex is seen by CD4 helper T cells. Dendritic cells can also cross-present exogenous antigen on class I, which is how CD8 responses start against viruses that do not infect dendritic cells, and how some non-replicating vaccines induce cytotoxic T cells. A T cell recognises its peptide only on its own host's MHC molecules, MHC restriction. Superantigens, such as some staphylococcal and streptococcal toxins, bind outside the groove to class II and to the variable part of the T cell receptor β chain, activating a large fraction of T cells and causing a cytokine storm.
MHC genes are the most polymorphic genes known. Each animal expresses the haplotypes inherited from both parents, and a population carries many alleles, so a pathogen is unlikely to escape presentation in every individual. Some alleles are statistically associated with resistance or susceptibility, and the associations vary with breed and pathogen strain. The best-known example is resistance to Marek's disease in chickens carrying the B21 haplotype. In cattle, certain BoLA-DRB3 alleles have been associated with proviral load and persistent lymphocytosis after bovine leukaemia virus infection. Cheetahs, with little MHC variation, accept skin grafts from unrelated cheetahs. Selecting breeding stock on one MHC allele risks narrowing diversity. Self-tolerance is taught in the thymus, where T cells reacting strongly to self are deleted, helped by the transcription factor AIRE, and maintained in the periphery by anergy and regulatory T cells. A classic veterinary demonstration of acquired tolerance is the bovine freemartin: placental anastomoses between heterosexual twins exchange blood-forming cells, so each twin carries the other's blood cells for life without rejecting them. Exposure before immune maturity also explains why cattle infected with bovine viral diarrhoea virus early in fetal life become persistently infected and immunotolerant.
Hypersensitivity is an immune response that damages the host. The Gell and Coombs scheme divides it by mechanism: type I is IgE and mast cells, type II antibody against cell-surface antigen, type III immune complexes, and type IV T cells. Real diseases often mix types. In type I, allergen cross-links IgE on sensitised mast cells and basophils, which release histamine within minutes, followed by prostaglandins, leukotrienes and cytokines that cause a late-phase reaction with eosinophils hours later. The shock organ of anaphylaxis differs by species: the lungs in cattle, sheep, horses and cats; the liver and portal circulation in dogs, which pool blood in the viscera; lungs and intestine in pigs. Adrenaline (epinephrine) is the treatment of choice because it reverses vasodilation and bronchoconstriction, and it should be at hand wherever vaccines are given. Veterinary type I diseases include vaccine or drug anaphylaxis, atopic dermatitis, food allergy, flea allergy dermatitis (which also has delayed components), insect bite hypersensitivity of horses to Culicoides, feline asthma, and milk allergy in cows reacting to their own casein when milking is delayed or at drying off. Allergen-specific IgE serology and intradermal tests select allergens for immunotherapy; they do not diagnose atopy, which is a clinical diagnosis.
In type II, antibody binds cell-surface antigen and destroys the cell by complement lysis, opsonisation and removal by macrophages, or antibody-dependent cytotoxicity. Transfusion reactions are the simplest example. In neonatal isoerythrolysis, a dam lacking a red cell antigen, sensitised by earlier pregnancies or transfusion, concentrates antibody in colostrum; a newborn that inherited the antigen from its sire absorbs it and haemolyses within one to a few days. In horses the Aa and Qa antigens are mainly involved, and mule foals are more often affected. In cats no prior sensitisation is needed because type B cats carry strong natural anti-A antibodies, so type A kittens of a type B queen are at risk. Prevention is by typing or cross-matching and giving at-risk newborns safe colostrum instead of the dam's for the first day. In type III, small complexes formed in moderate antigen excess deposit in vessels, glomeruli, joints and uvea, activate complement and attract neutrophils. The local Arthus reaction follows intradermal antigen in an animal with high IgG. Examples include hypersensitivity pneumonitis (farmer's lung) from mouldy hay; purpura haemorrhagica, a vasculitis after Streptococcus equi infection or vaccination in horses; 'blue eye' (corneal oedema and uveitis) after canine adenovirus type 1 infection or old CAV-1 vaccines, avoided by using CAV-2 vaccines; and glomerulonephritis in chronic infections such as leishmaniosis, dirofilariosis and pyometra, and in systemic lupus erythematosus.
Type IV reactions are mediated by sensitised T cells, mainly Th1 cells releasing interferon-γ that recruit and activate macrophages, with help from cytotoxic T cells. They take 24 to 72 hours to develop and can be transferred by cells but not by serum. The tuberculin skin test is the classic example: purified protein derivative (PPD) of Mycobacterium bovis injected intradermally produces a firm swelling in infected cattle, read at about 72 hours. The comparative cervical test uses bovine and avian PPD at separate sites to distinguish M. bovis infection from sensitisation by environmental mycobacteria or M. avium subsp. paratuberculosis; the caudal fold test is used as a screen in some countries. The injection site, PPD dose and thresholds for reactors or inconclusive results are set by each national programme and differ between countries. The same principle underlies the mallein test for glanders and johnin testing. The interferon-γ release assay measures the same T cell response in blood and is used alongside the skin test. Allergic contact dermatitis and granulomas around persistent antigens are also type IV reactions.
Primary immunodeficiencies are inherited and usually show as repeated or unusual infections early in life, often after maternal antibody wanes. Severe combined immunodeficiency (SCID) of Arabian foals is an autosomal recessive defect in the DNA-dependent protein kinase needed for V(D)J recombination; affected foals lack functional T and B cells, absorb colostrum normally, and die of infections, classically adenoviral pneumonia, within a few months. A DNA test identifies carriers. X-linked SCID in Basset hounds and Cardigan Welsh corgis results from mutations in the common gamma chain of several interleukin receptors. Bovine leucocyte adhesion deficiency (BLAD) of Holstein cattle is an autosomal recessive defect of CD18; neutrophils cannot leave the blood, so calves show marked persistent neutrophilia with recurrent infections, periodontitis and poor healing. A similar canine disease occurs in Irish setters. Chédiak–Higashi syndrome, in some cattle breeds, Persian cats and other species, causes giant granules in leucocytes and melanocytes, impaired neutrophil and NK function, partial albinism and bleeding. Others include selective IgA deficiency in some dog breeds, trapped neutrophil syndrome in Border Collies, cyclic haematopoiesis in grey collies and Fell pony immunodeficiency syndrome.
Acquired immunodeficiency is far more common than inherited. Failure of passive transfer (Chapter 9) is the most important in farm neonates. Feline immunodeficiency virus depletes CD4 T cells; feline leukaemia virus causes immunosuppression, cytopenias and lymphoma; infectious bursal disease virus destroys the bursa and chicken anaemia virus the thymus and marrow of young chicks; canine distemper virus and porcine circovirus type 2 deplete lymphoid tissue. Bovine viral diarrhoea virus is immunosuppressive in acute infection and, after fetal infection in roughly the first four months of gestation, produces persistently infected, immunotolerant calves that shed virus for life. Malnutrition (protein, zinc, selenium, vitamin E), stress and glucocorticoids, cytotoxic drugs, old age and heavy parasitism also suppress immunity, often showing as vaccine failure and opportunistic infection. The immune system also gives rise to tumours. Lymphoma is common in dogs and cats and, in cats, strongly associated with FeLV. Enzootic bovine leukosis is caused by bovine leukaemia virus. Marek's disease is a herpesvirus-induced T cell lymphoma of chickens, and avian leukosis a retrovirus-induced B cell neoplasm arising in the bursa. Multiple myeloma produces a monoclonal immunoglobulin, sometimes with urinary Bence Jones light chains. Reactive AA amyloidosis, deposition of serum amyloid A fibrils after chronic inflammation, is familial in Shar-pei dogs and Abyssinian cats.
Autoimmune disease follows a breakdown of self-tolerance. Contributors include genetic background and breed, molecular mimicry by microbial antigens, release of hidden antigens after injury, drugs acting as haptens, and loss of regulatory control. Immune-mediated haemolytic anaemia (IMHA) is the commonest in dogs: regenerative anaemia, often with spherocytes and autoagglutination; a direct antiglobulin (Coombs) test detects antibody or complement on red cells, though persistent saline autoagglutination makes it unnecessary. Immune-mediated thrombocytopenia causes petechiae and mucosal bleeding. Systemic lupus erythematosus involves antinuclear antibodies (ANA) and immune complexes; diagnosis needs several compatible signs, such as polyarthritis, glomerulonephritis, skin lesions or cytopenias, plus a positive ANA, because ANA alone is non-specific. Pemphigus foliaceus, the most common autoimmune skin disease of dogs and cats, targets desmosomal adhesion proteins, mainly desmocollin-1 in dogs, producing pustules and crusts. Acquired myasthenia gravis is caused by antibodies to the acetylcholine receptor, presents as exercise-induced weakness often with megaoesophagus, and is diagnosed by the serum anti-receptor antibody test. Lymphocytic thyroiditis causes most canine primary hypothyroidism, and immune-mediated adrenalitis underlies many cases of hypoadrenocorticism. Treatment relies on immunosuppressive drugs such as glucocorticoids — confirm drugs and doses against current specialist guidance and your national formulary.
Whether maternal antibody crosses the placenta depends on how many tissue layers separate maternal and fetal blood. In the epitheliochorial placenta of the pig and horse, and the synepitheliochorial placenta of ruminants, no immunoglobulin crosses; the newborn calf, foal, lamb, kid and piglet is born essentially agammaglobulinaemic and depends entirely on colostrum. In the endotheliochorial placenta of the dog and cat a small fraction of IgG, of the order of 5–10% of the neonate's eventual level, crosses before birth. In the haemochorial placenta of primates, IgG crosses via the neonatal Fc receptor (FcRn); rodents and rabbits receive IgG before birth largely through the yolk sac. In birds the hen transfers IgY from serum into the yolk, absorbed by the chick in the last days of incubation, while IgA and IgM from the oviduct enter the albumen and are swallowed by the embryo. The lesson for practice: the neonate of every farm mammal has no humoral protection until it has sucked.
In ruminants IgG1 is selectively transported from blood into the mammary gland in late pregnancy, and first-milking colostrum may contain several tens of grams of IgG per litre, falling rapidly at later milkings. Colostrum also supplies leucocytes, cytokines, energy and fat-soluble vitamins. The newborn gut absorbs intact macromolecules non-selectively for a limited time, aided by low proteolytic activity and colostral trypsin inhibitors. Efficiency declines from birth and absorption ceases ('gut closure') at around 24 hours in calves, foals and lambs, with a steep fall after 6 to 12 hours. The apparent efficiency of absorption (AEA) is serum IgG × plasma volume (about 9% of body weight in calves) ÷ IgG fed; values of 20 to 35% are typical for promptly fed calves, lower with delay or bacterial contamination. Management follows quality, quantity and quickness, plus cleanliness. Good bovine colostrum is conventionally at least 50 g IgG/L, commonly estimated as a Brix reading of 22% or more; the correlation varies between herds. The first feed is about 10 to 12% of birth weight, roughly 4 litres for a 40 kg Holstein calf, within the first hours, by nipple bottle or oesophageal feeder. Controlled heat treatment (about 60 °C for 60 minutes is often cited) reduces bacteria without major IgG loss; pooled colostrum risks spreading Johne's disease, bovine leukosis and Mycoplasma. A colostrum supplement is not a substitute for a replacer. For foals, mare colostrum is checked by refractometer and frozen banked colostrum used when needed.
Failure of passive transfer (FPT) is inadequate transfer of maternal immunoglobulin; it is not a disease but raises the risk of septicaemia, diarrhoea, pneumonia and death. In calves it has long been defined as serum IgG below 10 g/L (1000 mg/dL) at 24 to 48 hours. Indirect estimates are serum total protein by refractometer, with cut-offs of about 5.2 to 5.5 g/dL in healthy calves, and serum Brix, with cut-offs near 8.1 to 8.7% reported; dehydration falsely raises both. Because 10 g/L still leaves many calves under-protected, a US consensus proposed herd categories of at least 25 g/L (excellent), 18 to 24.9 (good), 10 to 17.9 (fair) and below 10 g/L (poor); national recommendations vary. In foals, IgG below 400 mg/dL is FPT, 400 to 800 mg/dL partial FPT and above 800 mg/dL adequate; stall-side ELISA and turbidity tests are used at 12 to 24 hours. Before closure, extra colostrum by tube is the remedy; after closure only intravenous plasma raises serum IgG, and the response is checked by re-measuring IgG. Maternal antibody also neutralises many vaccines while it persists. Because the age at which it wanes varies between individuals, puppy and kitten schedules repeat doses until about 16 weeks of age or later to cover the window between loss of maternal protection and a vaccine take.
Live attenuated (modified-live) vaccines contain organisms weakened by passage, by selecting natural low-virulence strains, or by deleting virulence genes. Because they replicate, they induce antibody and cytotoxic T cells, often protect after one dose, and can be given by natural routes. Their risks are residual virulence or reversion, adventitious contaminants, sensitivity to heat and disinfectants, disease in immunosuppressed animals, abortion or fetal infection with some products in pregnancy (check the label), and interference by maternal antibody. Inactivated vaccines contain whole organisms killed with agents such as formaldehyde or binary ethyleneimine, the latter favoured for foot-and-mouth disease virus because it destroys nucleic acid while preserving the capsid. They cannot revert and are stable, but usually need an adjuvant, two priming doses and boosters, and induce mainly antibody. Toxoids are formalin-inactivated toxins, as in clostridial and tetanus vaccines. Subunit vaccines contain purified antigens, such as the F4 (K88) and F5 (K99) fimbrial adhesins of enterotoxigenic E. coli given to dams to protect newborns through colostrum; virus-like particles, such as porcine circovirus type 2 capsid vaccines, contain no genome. Vectored vaccines put a protective gene into a harmless vector: canarypox vectors for canine distemper, feline leukaemia and equine influenza; vaccinia and raccoonpox vectors in oral wildlife rabies baits; and herpesvirus of turkeys carrying Newcastle disease, infectious bursal disease or avian influenza genes. DNA vaccines have been licensed for West Nile virus in horses and for infectious haematopoietic necrosis in farmed salmon, and RNA-based vaccines for some uses in some countries. Licensed products differ between countries.
An adjuvant increases or shapes the response to a vaccine antigen by forming a depot, delivering antigen to antigen-presenting cells as particles, and triggering innate sensors so dendritic cells mature. Aluminium salts (hydroxide and phosphate) are the most widely used; they adsorb antigen, activate the NLRP3 inflammasome, favour antibody and Th2-type responses, and are destroyed by freezing. Water-in-oil, oil-in-water and double emulsions give strong, lasting antibody responses and are used in many inactivated poultry, fish and foot-and-mouth disease vaccines. Freund's complete adjuvant, a water-in-oil emulsion with killed mycobacteria, is powerful but causes granulomas and tuberculin sensitisation and is not used in food animals or clinical vaccines. Saponins, especially Quil A from Quillaja saponaria bark, promote cell-mediated and cytotoxic responses; with cholesterol and phospholipid they form immune-stimulating complexes (ISCOMs). CpG oligonucleotides act on TLR9. The cost of adjuvants is local reaction, causing carcass trim, so cattle injections go in the neck. Accidental self-injection of oil-adjuvanted vaccine can severely damage a finger or hand and needs urgent surgical assessment even if the wound looks trivial. In cats, injection-site sarcoma can arise months to years after vaccines or other injections; guidance is to inject where wide excision is possible, such as low on a limb or the tail, record the site, and investigate a mass that persists three months, exceeds 2 cm, or grows one month after injection.
Ordinary serology cannot separate vaccinated from infected animals. A DIVA strategy pairs a marker vaccine lacking an antigen with a test for antibody to that antigen. Glycoprotein E (gE)-deleted infectious bovine rhinotracheitis vaccines leave animals gE-antibody negative, so a gE ELISA detects field infection; gE-deleted vaccines served Aujeszky's disease eradication in pigs. Purified inactivated foot-and-mouth disease vaccines contain mainly structural proteins, so antibody to non-structural proteins such as 3ABC indicates infection. In avian influenza, a vaccine with a different neuraminidase subtype from the field virus, or unvaccinated sentinels, allows detection. Brucella abortus strain RB51 is rough and lacks O-polysaccharide, so it does not induce antibody in standard tests, whereas strain 19 can leave persistent titres, especially in adults; both are live, can cause abortion and infect people, and national programmes set age and dose. At population level, transmission fails when more than 1 − 1/R0 of animals are immune; required coverage is that threshold divided by vaccine effectiveness. Trial efficacy is (attack rate unvaccinated − attack rate vaccinated) ÷ attack rate unvaccinated. WSAVA guidelines class as core, for dogs, canine distemper virus, canine adenovirus and canine parvovirus type 2; for cats, feline panleukopenia virus, feline herpesvirus-1 and feline calicivirus; and rabies where endemic or legally required. Apparent failure has many causes: maternal antibody, immunosuppression, incubating infection, an uncovered strain, wrong dosing interval or route, damaged vaccine, or an animal never actually vaccinated. Report suspected adverse events to the national pharmacovigilance scheme.
The cold chain is the system of people, equipment and procedures that keeps a vaccine within its labelled temperature from manufacturer to animal. Potency lost to heat, freezing or light is irreversible, and a damaged vaccine usually looks normal. Most veterinary vaccines are labelled for 2 to 8 °C. Live freeze-dried vaccines are the most heat-sensitive; many tolerate freezing when dry, but the label decides. Aluminium-adjuvanted and oil-emulsion vaccines must never be frozen: the aluminium gel clumps and releases antigen, and emulsions break. Cell-associated Marek's disease vaccines are stored in liquid nitrogen at about −196 °C and thawed and diluted exactly as directed. A dedicated vaccine refrigerator should hold 2 to 8 °C throughout. Store vaccines on middle shelves in their boxes, not in the door or against the cooling plate, and not with food or samples; water bottles stabilise temperature. Read and record temperature at least twice daily with a maximum–minimum thermometer or data logger. In the field, ice-lined and solar direct-drive refrigerators, cold boxes and carriers are used. Ice packs straight from a freezer can freeze vaccine, so condition them until the ice moves inside the pack, or use cool packs for freeze-sensitive products. Vaccine vial monitors darken with cumulative heat; once the inner square matches or is darker than the outer ring, the vaccine should not be used.
The shake test checks whether an aluminium-adsorbed vaccine has been frozen. A test vial is shaken alongside a control vial of the same product and batch that has been deliberately frozen solid and thawed, and both are left to stand. If the test vial sediments as fast as or faster than the frozen control, it has been frozen and must be discarded. The test applies only to adsorbed vaccines and does not detect heat damage. At the point of use, live vaccines are reconstituted with the supplied diluent, itself cool, and used within the labelled time, often one to two hours, kept cool and out of sunlight. Syringes and needles must be free of disinfectant or alcohol residues, which inactivate live vaccines; sterile disposable or boiled equipment is used. Drinking-water vaccination of poultry needs water free of chlorine and other sanitisers, often stabilised with skimmed milk powder, and birds slightly thirsty so the water is consumed within the labelled time.
In hot climates and pastoral systems the cold chain is often the weakest point of a programme. Thermostable vaccines reduce dependence on it. Rinderpest eradication, declared by WOAH and FAO in 2011, was helped by a thermostable freeze-dried formulation of the live vaccine that could be used for days without refrigeration. The I-2 strain of Newcastle disease vaccine was developed as a thermotolerant vaccine for village chickens and is widely used in Asia and Africa, and thermostable peste des petits ruminants vaccine formulations have been developed and field-tested. When a cold chain failure is found, quarantine the affected vaccine in the refrigerator, record the minimum and maximum temperatures and the duration, and contact the manufacturer or supplier for advice on that product. If potency cannot be assured, discard it and revaccinate the animals that received it. A disease breakdown in a vaccinated herd should always prompt a review of storage and handling records before the vaccine or strain is blamed.
Serology detects antibody (or sometimes antigen) through specific antigen–antibody binding made visible by precipitation, agglutination, complement consumption, neutralisation or an enzyme or fluorescent label. A titre is the reciprocal of the highest serum dilution still positive; in a doubling series a last positive well of 1:256 gives a titre of 256, or 8 on a log2 scale. A one-dilution difference is within test error; a fourfold or greater rise between acute and convalescent sera taken 2 to 4 weeks apart indicates recent infection. Groups are summarised by the geometric mean titre. A single positive titre shows exposure or vaccination, not necessarily current disease, and antibody appears one to three weeks after infection. Agglutination occurs when antibody cross-links particles; IgM agglutinates best. The Rose Bengal plate test for brucellosis uses stained antigen buffered to about pH 3.6–3.7, which reduces non-specific IgM reactions; it is a sensitive screen, confirmed by complement fixation or ELISA. The milk ring test detects antibody in bulk milk as a coloured cream ring. The microscopic agglutination test (MAT) for leptospirosis uses live leptospires of a serovar panel under dark-field microscopy and is the reference test, though cross-reactions are common. Whole-blood plate agglutination is used on farm for Salmonella Pullorum/Gallinarum and Mycoplasma in poultry. The Coombs (antiglobulin) test detects antibody or complement on red cells. Haemagglutination inhibition (HI), with 4 haemagglutinating units of antigen per well and chicken red cells, is the standard antibody test for Newcastle disease and for avian influenza haemagglutinin subtypes.
The complement fixation test (CFT) has two stages. Heat-inactivated test serum is incubated with antigen and a measured amount of guinea pig complement; then an indicator system of sheep red cells sensitised with anti-sheep red cell antibody (haemolysin) is added. If antibody was present, complement was fixed and the indicator cells stay intact: no haemolysis is a positive result. Without antibody, free complement lyses the cells. The CFT is laborious and troubled by anticomplementary sera, but specific; it confirms brucellosis and is used for glanders, dourine and contagious bovine pleuropneumonia in trade and control. In agar gel immunodiffusion (AGID), antigen and antibody diffuse towards each other from wells in agar and form a precipitin line at equivalence; a line of identity with a known positive control confirms specificity. AGID is simple and specific but relatively insensitive. It is the basis of the Coggins test for equine infectious anaemia and has been used for bluetongue, enzootic bovine leukosis, avian influenza group antibody and Marek's disease antigen in feather tips. Virus neutralisation (VN) mixes serum dilutions with a fixed dose of live virus and inoculates cell culture or eggs; antibody that prevents cytopathic effect is neutralising. VN is serotype-specific, correlates well with protection and is often the reference method, but needs live virus, cell culture and biosafety facilities, and takes several days.
The enzyme-linked immunosorbent assay (ELISA) uses an enzyme-labelled reagent whose bound amount is read as optical density (OD). In the indirect ELISA, antigen coats the plate, test serum is added, and bound antibody is detected with enzyme-labelled anti-species immunoglobulin, so each host species needs its own conjugate. Results are often expressed as a sample-to-positive (S/P) ratio: (OD sample − OD negative control) ÷ (OD positive control − OD negative control). In the competitive or blocking ELISA, test antibody competes with a labelled monoclonal antibody for antigen; a positive serum reduces colour, results are expressed as percent inhibition, and no anti-species conjugate is needed, which suits multi-species diseases such as bluetongue and foot-and-mouth disease. In the sandwich (antigen-capture) ELISA, a capture antibody binds antigen from the sample and a second labelled antibody detects it; this detects the agent, as in rapid tests for canine parvovirus antigen in faeces or feline leukaemia virus p27 antigen in blood. ELISAs are sensitive, automatable and can use serum, milk or meat juice. Cut-offs are set by the kit maker and validated for a purpose; a kit validated for herd screening may not be fit for individual certification.
Sensitivity (Se) is the proportion of infected animals testing positive; specificity (Sp) the proportion of uninfected animals testing negative. Positive predictive value (PPV) is the probability that a positive animal is infected: PPV = Se × P ÷ [Se × P + (1 − Sp)(1 − P)], where P is prevalence. With Se 95%, Sp 98% and 2% prevalence, 1,000 animals give 19 true and 19.6 false positives, a PPV of about 49%; at 30% prevalence PPV is about 95%. So late in eradication, many reactors are false positives and are confirmed with a more specific test. Series testing (positive only if both tests positive) raises Sp and lowers Se; parallel testing does the opposite. Apparent prevalence is corrected to true prevalence as (AP + Sp − 1) ÷ (Se + Sp − 1). Serology shows the host's response; direct tests show the agent. The polymerase chain reaction (PCR) detects pathogen nucleic acid within hours and before antibody appears, suits organisms that are hard to culture, and finds persistently infected, seronegative animals such as BVDV carriers. But it detects dead as well as live organisms, can be positive after live vaccination, is prone to contamination, and misses cleared infections or mutated targets. Serology shows past exposure and immune status, suits surveillance and proof of freedom, and assesses vaccine response, but misses infection before seroconversion and is confounded by maternal and vaccinal antibody: a young calf's positive BVDV antibody result may be colostral, whereas PCR or antigen testing identifies persistent infection.
As a student you need a clear mental model of how innate and adaptive immunity cooperate, because it explains why vaccines work, why neonates die without colostrum and why some diseases are immune-mediated rather than infectious. Learn the species peculiarities that exams love and practice uses: the bursa, IgY, bovine IgG1 in colostrum, the placental types, and the major acute phase proteins. Be able to explain in one sentence what each serological test detects and how a positive looks: no haemolysis in the CFT, a precipitin line in AGID, inhibition of haemagglutination in HI. Practise the predictive value calculation until you can do it on a 1,000-animal table in under two minutes.
Five years into practice, immunology is the daily work of vaccination consults, neonatal care and test interpretation. You decide when to start puppy and kitten series given maternal antibody, check refrigerator logs when a herd reports a vaccine breakdown, measure calf serum total protein on a sample of calves each month to audit colostrum management, and give plasma to a foal with IgG below 400 mg/dL. You recognise anaphylaxis and keep adrenaline to hand, choose injection sites in cats with sarcoma risk in mind, and advise owners that one positive ELISA in a low-prevalence setting needs confirmation before a clinical or trade decision is made.
With twenty-five years' experience, the questions are about programmes rather than animals. You design herd vaccination and colostrum protocols with measurable targets, set up cold chain audits across a district, and decide which vaccine to use in an outbreak, including whether a DIVA vaccine is needed to protect export status. You interpret national surveillance data knowing how predictive values collapse at low prevalence, choose series testing to protect farmers from false positives in eradication schemes, and advise authorities on emergency vaccination for FMD or avian influenza with NSP or sentinel-based surveillance. You mentor junior staff to report suspected adverse events and to investigate failures before blaming a product.
A researcher with long field experience works on the open questions: correlates of protection for diseases without them, thermostable and single-dose vaccines for pastoral systems, new adjuvants that give cell-mediated immunity without injection-site damage, mucosal and in ovo delivery, and the genetics of disease resistance, including MHC associations and the risk of narrowing diversity by selection. Test validation is another core area: estimating sensitivity and specificity without a gold standard with latent class models, fitness-for-purpose validation under WOAH standards, and pen-side assays for low-resource settings. Such work must separate what is established from what is promising and report negative results honestly.
This manual does not yet cover: detailed cytokine and chemokine families beyond the main ones; the immunology of fish, reptiles and bees; flow cytometry and cellular assays such as lymphocyte proliferation; the immunology of pregnancy beyond passive transfer; transplantation and tumour immunotherapy; vaccine registration law in individual countries; and product-specific storage times, vaccination schedules and drug doses, which must be taken from labels and national formularies. Several numbers are given as widely taught cut-offs that vary by country, laboratory, method and breed: calf FPT thresholds, colostrum Brix cut-offs, refractometer total protein cut-offs, tuberculin test thresholds and ELISA cut-offs. Species differences in light-chain ratios and IgG subclass functions are summarised, not tabulated. Blood group systems of each species, beyond those that cause neonatal isoerythrolysis, are left out.
Original practice questions written to the syllabus and pattern of the exams named in each question's examTags. They are not past papers and do not reproduce any board's questions.
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