Environmental factors (causes - nutrition, mechanical (cut gets infected)/thermal (overheat and get heat stress) injury, toxins) and affect on pathogens (temperature, humidity, degree of moisture in environment, grazing characteristics, prevailing wind direction, population density)
Mode of transmission (direct - droplet or aerosol, oral, contact; or indirect - foodborne, waterborne, vectorborne, fomite (bedding or things that the animal comes into contact with, that is passed on when another comes into contact with the object), environmental)
Human influences - on nutrition (over/under feeding), environment (improper housing), physical (inury or overcrowding), transportation (strees, hygiene), genetics (in-breeding)
What is phagocytosis
Where one cell can engulf organic material and/or whole cells, break it down and then expel the broken down contents
What are the two types of immune mechanisms?
Soluble factors (Humoral immune response - when antibodies are produced)
Immune cells (Cell-mediated immune response)
What are the two types of immune responses?
Innate: Non-specific recognition. No memory of previous assaults. First line of defence. Fast acting. Stops most potential threats. Present at birth.
Adaptive (Acquired): Specific recognition. Memory of previous response. Slower and more powerful in attacking the antigen. Develops over time (mostly).
What are the 6 steps in phagocytosis?
Pseudopodia surround microbes
Microbes are engulfed into cell
Vacuole/phagosome containing microbes forms
Vacuole and lysosome fuse to become a phagolysosome
Toxic compounds and lysosomal enzymes destroy microbes
Microbial debris is released by exocytosis
What are the roles of macrophages in the immune response?
Inflammation and fever
What is vasodilation?
Expansion of veins/capillaries to increase capillary permeability
What is chemotaxis?
The process involving macrophages and neutrophils that causes attraction of phagocytes to infection site
What is extravasation?
Phagocytes migrate capillaries to site of tissue damage
What are the two distinct phases of the inflammatory barrier?
Acute: Pain, swelling, heat, redness, loss of movement/function. Vasodilation, oedema (build up of fluid)
Chronic: 24-48hrs. Monocytes need to go through activation to become macrophages.
What are the two types of antibodies?
B cells - surface bound antibody that binds to the b cell receptor
Plasma cells - secrete antibody free into the serum
How can you test for antibodies/antigens?
Protein A/G or Strepavidin/Biotin binding
Enzyme-linked Immunosorbent Assays (ELISAs)
How are ELISA's conducted?
Indirect ELISA method: Antigen coated on a plate. Antibody added (1st Ab). Labelled secondary (2nd) Antibody (binds to 1st Ab) (DIRECT SKIPS THIS STEP). Chromogenic substrate added. Optical Density measured.
Sandwhich ELISA: Developed antibody that binds to antigen, this is added to the plate, followed by the antigen, and then the antibody again, then add antibody with label, then substrate added, then measure OD
Lymphoid cells and myeloid cells are collectively known as what?
Leukocytes/White blood cells
What CDs are expressed by which cells?
All T cells – CD3+, CD28+, CD45+
T helper cells – CD4+
T cytotoxic cells – CD8+
Treg cells CD4, CD25 (majority)
Natural killer cell – CD16+, CD56+, CD3-
B cells - CD19, CD 20, CD40
Macrophages - CD1a-c, CD11b, CD40, CD64
Dendritic cells - CD1a-e, CD35, CD40
NB: ‘+’ expressed by cell, ‘-’ not expressed by cells
What are the 3 types of T cells?
T Helper (Th) cells: Express CD4 – recognise MHC Class II. Help B cells divide, differentiate & produce ab. Can be further divided into Th1 and Th2 cells.
T cytotoxic (Tc) cells: Express CD8 – recognise MHC Class I. Bind & kill pathogens & infected/cancer cells.
Regulatory T (T reg) cells:Suppress T & B cell activity
What are the characteristics of a B cell?
Develop in bone marrow. CD19, CD20, CD21. Express surface immunoglobulin (Ig) - Antigen receptor. Most divide and differentiate into plasma cells which secrete Ig. Some become memory cells, which have high affinity surface Ig, recognise repeat antigen and increase in numbers for stronger attack (Clonal expansion).
What are plasma cells?
Differentiated B cells:
↑ endoplasmic reticulum (ER) & Golgi apparatus
Secrete ↑ Ig
No longer APCs (lack MHC Class II)
Important for Humoral Immunity (antibodies)
Cannot switch Ig class once differentiated from B cells
Produce high affinity Ig
Found in bone marrow (long lived plasma cells) and secondary lymphoid organs (spleen, lymph nodes)
What are Null cells?
Non-T/Non-B Cells that don’t express CDs found on B or T cells. Lack conventional antigen receptors. Go through differentiation to become NK cells
What are NK cells?
CD16, CD56 (CD3-)
Kill virally infected cells/cells that have lost MHC Class I expression
2nd line of defence against viruses (get what Tc cells miss)
Similar mechanisms to Tc cells - granules (perforin & granzymes)
What are granulocytes?
A type of myeloid cell that attacks other cells. 4 types (eosinphils, neturophils, basophils and mast cells).
What are the 3 roles of macrophages?
Tissue repair, inflammatory response (chronic) and adaptive immunity activation (APCs)
What granulocyte is first at the inflammation site?
Can be phagocytic but main function is exocytosis (release granules)
What are platelets?
React with Fibrinogen
Clot to prevent blood loss
What are the primary and secondary lymphoid organs?
Primary (immune cell production): Bone marrow (humans, rodents and all others)/ileal Peyer’s patches (ruminants and dogs)/Bursa of Fabricius (birds) and Thymus
Secondary (cell storage and antigen contact): Lymph nodes, spleen and mucosal associated lymphoid tissue
What happens in the bone marrow?
Early cell differentiation in all animals (haemopoietic stem cells): T cell precursors and B cell precursors
B cell maturation in Rabbits, Rodents and Humans
What is the bursa of fabricius?
A lymphoepithelial organ located in the Cloaca/intestine, it is the site of Maturation of B cells. It is Larger in young birds
What are Ileal Peyer's patches?
Organised lymphoid nodules in the Intestinal tract (ileum) with a Thickened epithelium. It is the site of B cell maturation in ruminants, pigs, dogs
Happens in germinal centres
What is the thymus?
The site of T cell maturation, after T-cell precursors have migrated to here. The thymus selects cells that can distinguish self and foreign proteins, those cells that cannot distinguish are destroyed. The thymus is made up of lobules, with the outer cortex containing rapidly dividing pre-T cells and the inner medulla containing mature T cells and APCs.
What is lymph?
Circulating institial fluid that transports immune cells and antigens from body tissues to the lymph nodes (which filter lymph fluid are is the site of antigen/immune cell interaction). It is connected to the blood supply.
What is the spleen?
A secondary lymphoid organ that is made up of red pulp (destruction of old red blood cells and storage of current red blood cells) and white pulp (the peri-arteriolar lymphoid sheath - T and B cells, which doesnt not have lymph supply)
What is Mucosal Associated Lymphoid Tissue (MALT)
Tonsils, Peyer's patches, appendix, small aggregates of lymphoid tissue found in mucosal tissues. It diffuses aggregates of lymphoid tissue and is the main entry point of pathogens.
What is the immune cell circulation pattern?
Development in Bone Marrow then enter blood. Lymphocyte circulation/recirculation through blood and lymph. Increased chance of antigens to contact specific lymphocytes. Lymphocytes leave the lymph node and go to specific tissues, E.g. gut, skin, lung.
What is haematopoiesis?
Blood cell differentiation into lymphoid, myeloid and red blood cells that occurs in the bone marrow. Recognition and attack cells.
What are the general functions of immunoglobulin?
Generation of oxidents
Activation of complement
Virus and toxin neutralisation
Antibody-dependent cell cytotoxicity (ADCC)
Direct anti-microbial activity
Reduced damage to host from inflammatory response
What is the nature of the antigen-antibody interaction?
Lock and key concept: antibody-antigen complex. Compatibility.
Non-covalent bonds: overall strength (affinity)
Reversability: bonds can be broken
Occurs in the Fab region: Antibody paratope binds to antigenic epitope
What is avidity?
Affinity x number of binding sides
What is cross reactivity and why is it important?
Ab recognises more than one antigen based on specificity and structure:
Same antigen, different pathogen
Similar antigen, different pathogen
Similar antigen, same pathogen (isomer)
Important because it can be used in immunisations to protect against multiple pathogens
What is the mechanism of antibody formation?
Antigen encounter with a B cell (primed)
Activation of the Th cell (APC)
B cell activation by Th cell
What does the Clonal Selection Model (Burnett 1959) say?
Each B cell - one type of antibody
Each B cell will place the antibody it makes into its cell membrane with antigen-binding side outward
Only B cells that bind to the antigen can complete their development into antibody-secreting plasma cell
The specificity of the antibody is exactly the same as that which was on the cell surface of the B cells
How are monoclonal antibodies produced?
Collect spleen cells
Fuse cells with myeloma cells - if they fuse the cells will survive
Select clonal population of B cells
Clone and purify monoclonal antibody
What are the main antibody classes that are transferred to neonates in pigs/humans, and cattle/sheep
IgA in pigs and humans
IgG in cattle and sheep
What is a nanobody?
The VH aka VHH aka V-NAR fragment of the Heavy chain antibody (VHH = variable domain of heavy chain of heavy-chain antibody)
Found in camelids and sharks
What are the key properties of nanobodies?
Like normal antibodies, Nanobodies show:
- high solubility
- high target specificity
- high affinity for their target
- low inherent toxicity
However, like small molecule drugs, they:
- inhibit enzymes
- rapid tissue penetration
- access receptor clefts, which is challenging with normal antibodies
- think of the nanobody as a drug
- are generally very stable
- easily produced in recombinant form
- recognize hidden antigenic sites
- can be administered by means other than injection
- are easy to manufacture
What are the uses of nanobody technologies?
Anti- fungal, viral, venom, inflammatory, cancer
How can nanobodies be used into treatment of trypanosomes?
Use NbAn33 to deliver the apoL-1 toxin to the trypanosome, as Apolipoprotein A1 in human serum or as a nanobody toxin is lytic for trypanosomes
What are the four types of tissue grafts, and what determines their success?
Autografts: Same individual, different site
Isografts: Between genetically identical individuals
Allografts: Closly matched, genetically dissimilar individuals – same species
Xenografts: From one species to another
Success is dependent on ‘tissue type’ and immunological acceptance by Major Histocompatibility complexes (MHC - donor and recipient must have the same). Skin biopsy from mouse genotype A can be given to another type A. Skin biopsy from mouse type B is rejected by type A, due to passenger leukocytes
How can tolerance of tissue grafts be induced?
Glucocorticoids - Suppress Ig production of B cells. Suppress IL genes (T cell proliferation). Prevent inflammation.
Cryostatics - Inhibit cell division (T and B cells).
Drugs acting on immunophilins - Block functions of immune cells
Using antibody treatments: target immune cells
Total body irradiation: Haemopoetic stem cell transplant. Lungs shielded. Removes residual cancer cells.
What are the three classes of MHC molecules?
Membrane surface molecules (Class I & II): Involved in antigen presentation to T cells. Identify self and non-self cells
Diverse secreted factors/proteins (Class III): Not on cell membranes but expressed by cells: Complement, cytokines, enzymes, heat shock proteins and more…
What is the function of MHCs?
Immune reactions and recognition of self:
Class I MHC - monitor self cells (healthy/unhealthy cells)
Class II MHC - recognition of foreign molecules
Bind antigens & present to T cells:
Class I to T cytotoxic cells (CD8)
Class II to T helper cells (CD4)
MHC combinations associated with different diseases & conditions E.g. Malaria:
Predict the likelihood of contracting certain disease
Breed animals for resistance to certain disease E.g. Scrapie (SCR: αA, A4, B6)
What are the functions of complement?
Lyse cells/bacteria (Membrane Attack Complex)
Opsonize bacteria to enhance phagocytosis
Recruit (chemotaxis) and activate various cells: granulocytes and macrophages
Regulation of antibody response
Clearance of immune complexes and apoptotic cells
Detrimental effects for the host: Inflammation and tissue damage. Trigger anaphylaxis. Lack of leads to increased bacterial infections.
What is the function of C3a (Anaphylatoxin)?
Can lead to anaphalyxsis due to:
Mast cell degranulation (histamine)
Smooth muscle contraction
Vasodilation: Dilation of the venules. Leukocyte extravasation.
Release of inflammatory cytokines. Increasing amount of acute phase proteins: anti-bacterial effects. TNFα: increase in body temperature or fever
C4a does the same but is weaker
What is the function of C5a?
Chemotaxis: Attracts leukocytes (mostly neutrophils) to the site of invasion
Vasodilation: Allows for cells to cross venal membrane
What is the function of C3b (opsonin)?
A recognition molecule for macrophages, granulocytes and cytotoxic lymphocytes
Coat surfaces of bacteria or immune complexes
Phagocytes have receptors for C3b - CR1 and CR3
Activate phagocytes for expression of receptors
Phagocytose coated pathogen/complex - Clearance from serum
What is the function of C2a (Prokinin)?
Cleaved by plasmin to yield kinin, which results in edema
What are the two types of pathogens?
Extracellular eg liver fluke
Intracellular eg malaria
How does extracellular protection (antibody-mediated) occur?
Antibodies are the primary defence: Secreted IgG, IgA, IgE most common forms
What is the relationship between F hepatica and bovine TB tests?
BTB diagnosis measures skin test response to TB antigen
Response measures Th1 cell response and IFN-γ
Fh suppresses Th1 responses
Fh compromises BTB diagnosis
How can worms be used as therapy?
Apply worms to treat IBD and Crohn’s Disease in humans
Via Th2 response - Helminth antigens can suppress Th1 inflammatory response in the gut
What is hypersensitivity?
Sensitisation to an antigen (Ag) by previous exposure - often repeated exposure over time
Re-exposure to Ag leads to an excessive response - Termed a “hypersensitivity” response which results in collateral damange as the immune system misdirects its response and attacks its own tissues.
Give an example of each type of hypersensitivity reaction
Type I: (immediate - mucosal surface) Hayfever; Asthma; Eczema; flea allergy dermatitis in dogs
Type II: autoantibodies to red blood cells; Kidney damage
Type III: Farmer’s lung disease
Type IV: Skin contact reactions: TB, leprosy, parasites
What is the process of type I hypersensitivity?
In the first exposure, IgE is produced, which binds to mast cells, basophils.
In the second exposure to the antigen, the IgE cross links causing mast cells to degranulate.
The release of Mediators eg histamine result in Local tissue Responses (vasodilation, oedema, bronchoconstriction, skin itching (pruritis)). This all happens rapidly (within 20 mins) so it is considered immediate. It is mast-cell mediated.
What is the process of type II hypersensitivity?
Type II is antibody mediated hypersensitivity. In the initiating mechanism, antigen is bound to host tissues. The IgG/IgM then binds to the antigen on the tissue's cells which recruits NK cells and macrophages or activates a complement cascade, which lyse the cell. This can cause severe damage, long term illness and possibly death. It takes 5-8 hours so is considered intermediate.
What is the process of type III hypersensitivity?
Type III is immune-complex mediated. Immune-complexes bind to tissue as insoluble complexes or as a result of their increased size.
Once deposited, complexes bind complement and trigger CDCC, ADCC and inflammatory reactions as in Type II over a 4-10h period.
What is the process of type IV hypersensitivity?
Type IV hypersensitivity is cell mediated.
During the sensitisation phase:
- Antigen is taken up by APC and presents it on its surface in MHC class II. This allows Th cells to recognise the antigen - this is the "first exposure"
- Memory Th1 cells specific for this antigen are generated
- Th1 memory cells re-encounter antigen and then become activated.
- This causes Th1 to secrete cytokines (IFN-y - activates macrophages) and chemokines (attract neutrophils, basophils etc.) to recruit pro-inflammatory cells and amplify inflammation.
- More T cells (CD4 and CD8) may also be recruited to the area
This reaction tends to happen in the skin - so when an inflammatory response occurs, redness, swelling and a bump may be present, which will represent the accumulation of T cells to the site. The reaction time is commonly 21-28 days.
What is the role of Treg cells?
Mediate immune suppression by:
Targeting dendritic cells
What are the two types of regulatory T cells?
T regs and Tr1, which are both derived from CD4+ T cells.
Treg is termed “natural” as it is always active in the body. It suppresses via direct contact with Th1 or Th2 cells.
Tr1 cells are induced to release IL10 which suppresses Th1 responses. Tr1 recognise peptides via MHC
What does the Hygeine hypothesis say?
Our modern “hygienic” lifestyle in childhood has led to rise in allergy and autoimmune disease. The modern 'hygienic' lifestyle due to our indoor lifestyle with smaller families, early childhood vaccinations, and less exposure to pathogens. Exposure to pathogens is needed to drive Treg expansion. Certain helminth antigens may 'set' the Treg clock.
The hygiene hypothesis also says that there is a reduced rate of allergy due to growing up on a farm, having pets in the household, and having diverse gut microbiota.
What is self tolerance?
The negative selection of self-reactive cells. Failure is known as an autoimmune response.
What are the two types of autoimmune disease?
Systemic- the auto-immunity is directed against an antigen that is present at many different sites and can include involvement of several organs (e.g. rheumatoid arthritis)
Organ specific - Organ specific means the auto-immunity is directed against a component of one particular type of organ. (e.g. Type 1 diabetes, Multiple sclerosis)
What is the difference between primary and secondary autoimmune disease?
Primary is due to genetic susceptibility. There is no trigger factor required.
Secondary requires a trigger factor.
What are some options for treatment of autoimmune disease?
Corticosteroids regulate the expression of many genes, with a net anti-inflammatory effect. First, they reduce the production of inflammatory mediators, including cytokines, prostaglandins, and nitric oxide. Second, they inhibit inflammatory cell migration to sites of inflammation by inhibiting the expression of adhesion molecules. Third, corticosteroids promote the death by apoptosis of leukocytes and lymphocytes. They are used in transplant chemotherapy.
How was the vaccine for smallpox created?
Edward Jenner inoculated James Phipps, a 13-year-old boy, with the vaccinia virus obtained from a young woman named Sarah Nelmes who had been accidentally infected by a cow
James Phipps was then found to be “secure” (immune) to smallpox - unsuccessful challenge with variola virus “some months afterwards”.
What is the difference between immunisation and vaccination?
Immunisation is the administration of an antigen to a live host with the purpose of inducing an immune response.
Vaccination is immunisation for veterinary or public health purposes.
What are the aims of vaccination?
Sterile immunity- prevent infection
Prevent disease eg malaria vaccine: patient infected but not ill
Cure current infection (therapeutic)
Alter physiological state (e.g. to alter hormones levels and prevent boar taint)
How does the body respond to a vaccination?
Priming 1st Dose: a foreign substance is recognised by the immune system leading to a primary IgG immune response. This may be inadequate and slow. Memory cells are produced.
Booster/ 2nd dose: a second, faster immune response as more IgG follows the booster dose. Memory cells from the primary response are triggered to produce IgG forming cells. It is known a prime-boost vaccine.
What are the components of a vaccine?
Infectious agent or specific antigen, adjuvant, stabilizers, preservative, buffers and salts.
What are the characteristics of live attenuated vaccines?
Live bacteria or viruses are treated so they are no longer virulent - cannot cause severe disease.
The organism is modified so it mimics the virulent form - causes the body to mount an immune response
The organisms in the vaccine multiply in the recipient animal - gives better immunity than killed vaccines.
Used against bacteria, viruses (lung worm)
What are the advantages and disadvantages of live vaccines?
Strong, long lasting immunity.
Fewer doses required.
Adjuvants not necessary.
Fewer adverse reactions.
May cause a mild form of the disease.
May cause abortion.
Limited shelf life.
Needs cold storage.
What are the characteristics of killed/inactivated vaccines?
Killed infectious agents (eg using formalin) - still induce an immune response in the recipient.
Cannot cause disease and cannot replicate in the body - used for bacterial diseases and toxins.
What are the advantages and disadvantages of killed/inactivated vaccines?
Rarely cause disease
Safe in pregnant animals and immuno-compromised animals
Usually have a long shelf life
Give only short lived immunity
Require multiple doses
Hypersensitivity reactions are common
Adjuvants can cause severe localised tissue reactions
What are the characteristics of subunit vaccines?
It is not the whole organism that is used, only proteins - vaccines are safe, stable
Vaccines produced by genetic engineering - protein vaccines, DNA vaccines
What are the advantages and disadvantages of subunit vaccines?
Need for adjuvants
Organism can mutate away (lose efficacy)
What is an adjuvant?
A substance which results in a specific increase in the immunogenicity of a vaccine component - it enhances immunity. E.g. liposomes, proteosomes, aluminium salts
What is the cattle tick vaccine Bm86?
A subunit antigen from tick gut. Vaccination induces IgG that damages tick’s gut - reduces egg production, transmission
Describe the current vaccine status for Hydatid
Hydatid vaccine EG95 is a subunit antigen from oncosphere. It shows high (>95%) efficacy, blocks invasion of oncosphere, and is now being tested in the field
Describe the current vaccine status for Liver fluke
Liver fluke vaccines are subunit antigens from juvenile/adult flukes. Elicits <100% protection but may still induce production benefits
Blocks growth of juvenile flukes - flukes do not replicate in hosts
Threshold effects: <40 flukes, losses are minimal
Combination vaccines may be best approach: fluke vaccine is feasible- good news!
Name the barriers of the Innate immune system
Epithelial barrier - made up of skin and mucous membranes
Physiological barrier - made up of different soluble components (e.g. temperature, low pH in stomach, chemical mediators)
Phagocytic barrier - Cells e.g. dendritic cells and macrogphages that can phagocytose. Cellular ingestion of pathogens/foreign particles.
What are the five main types of antibody classes that can be produced in animals and which classes are produced in sheep?
5 main classes:
Immunoglobulin M (IgM) - µ - pentameric
Immunoglobulin G (IgG) - γ
Immunoglobulin A (IgA) - α - dimeric
Immunoglobulin E (IgE) - ε
Immunoglobulin D (IgD) – δ
No IgD found in sheep, all others are.
Name the 3 main types of cells that present antigen to T cells
Dendritic cells: Found in skin & other tissues. Ingest antigens, transport antigens to lymph nodes and spleen: T cell area. The most effective APCs: they present antigen to naive T cells. Present internalized antigens & MHC I or II.
Macrophage: Ingest antigen by phagocytosis. Very good in activating memory T cells.
B cells: Bind antigen via surface Ig and ingest antigen by pinocytosis. Very good in activating memory T cells when the antigen concentration is low.
Name 3 ways phagocytes kill pathogens
Cationic proteins (which destroy cell walls)
Lysosomal acidic enzymes (to digest the carbohydrate of the cell walls of bacteria)