Immune Responses To Infectious Agents: Immunology & Immunopathology

Immunology & Immunopathology
IMMUNE RESPONSES TO INFECTIOUS AGENTS
• Tc cells (CD8+) are one of the fastest secretors of cytokines & IFNs. They come in almost straight after an
innate immune response.
COMPLEMENT ACTIVATION
C3a, C3b & C5a have actions other than MAC formation.
• C5a, C3a: Inflammation
• Recruitment & activation of leukocytes
• Destruction of microbes by leukocytes
• C3b: Phagocytosis
• Recognition of bound C3b by phagocyte C3b receptor
• Phagocytosis of microbe
INTERFERON PRODUCTION
• IFNs were originally discovered in the context of virus protection mechanisms, but itʼs now known theyʼre
induced by other types of Ag
• Although produced by pretty much any cell, macrophages & dendritic cells are the most important
producers
• IFNs are being used in certain treatment scenarios against neoplasia
• They also have a big role in virus infections, as they act upon infected cells
ACUTE, INNATE RESPONSE TO INFECTION

MAST CELLS
• “MASTERMINDS” which regulate a large majority of the immune response
PREFORMED EFFECT
HISTAMINE Vasodilation, incr. capillary permeability,
chemokinesis, bronchoconstriction
GRANULE RELEASE INTERLEUKINS 3, 4, 5 Multiple, including macrophage activation, trigger
&6 acute phase proteins, etc.
GM-CSF, TNF
NEWLY SYNTHESISED EFFECT
LIPOXYGENASE LEUKOTRIENES C4, D4 Vasoactive, bronchoconstriction, chemotaxis
PATHWAY (SRS-A), B4
CYCLO-OXYGENASE PROSTAGLANDINS Affect bronchial muscle, platelet aggregation &
PATHWAY THROMBOXANES vasodilation
PAMPs & TLRs

• Can be monomers, dimers, heterodimers, homodimers, etc.
• Often involve other molecules e.g. CD4T catches LPS & gets it in contact with TLR4
• NF kappa B is a very important transcription factor that activates cytokines after PAMP binds to TLR
Th1 Th2
Main Macrophage B-cell
partner
cell type
Cytokines IFN-ϒ & TNF-β IL-4, IL-5, IL-6, IL-10, IL-13
produced
Immune Cellular immune system. Maximises killing Humoral immune system. Stimulates B-cell division,
stimulation efficacy of macrophages & CD8+ Tc-cells. B-cell antibody class switching & increase
promoted Also produces opsonising antibodies. neutralising antibody production
Other IFN-ϒ increases IL-12 production by DCs & IL-4 acts on Th cells to promote Th2 cytokines (auto-
functions macrophages via positive feedback. regulatory).
IL-12 stimulates IFN-ϒ production in Th IL-10 inhibits Th1 cytokines in DCs & macrophages.
cells. Combined action of these two cytokines suggests
IFN-ϒ also inhibits production of Type 2 that once T-cell has decided to produce these
cytokines (e.g. IL-4), preserving its cytokines, that decision is preserved (& encourages
response. other T-cells to do the same).
TYPES OF
ADAPTIVE
IMMUNE
RESPONSES
• Viral infection
= Th1 = IgG 2a;
this is the only
type of Ig
isotype that is
most effective
in neutralising
viruses.
• Other
organisms =
IgG 1 or IgA
• In most cases,
IgE is
undesirable as
it causes
allergic
reactions. IgE is
induced by
cytokines from
the Th2
pathway.
STRUCTURE OF BACTERIAL CELL WALLS

• Gram-Negative bacteria possess more layers for the immune system to deal with, but also more
recognition molecules.
• Mycobacteria are masters of persistent infections, requiring a complex immune response to eliminate.
IMMUNE RESPONSE TO BACTERIA - OFTEN SUFFICIENT TO ELIMINATE

1. Killing of bacteria by complement
2. Opsonisation of bacteria by phagocytic cell & their secretions e.g. Ab, C3b, etc.
3. Neutralisation of bacterial toxins by Abs
a. Basis of toxoid vaccines e.g. tetanus
4. Direct killing of bacteria by NK cells
a. Happens at least in a petri dish
5. Destruction of intracellular bacteria by activated macrophages & Tc cells
a. If macrophages & Tc cells arenʼt truly activated, the bacteria can persist
BACTERIAL OPSONISATION & PHAGOCYTOSIS

• Opsonising particles (e.g. IgM + C3b, IgG + C3b, IgA) send signals into the phagocyte, coordinating the
cytoskeletal mechanism.
• After killing & digestion, degradation products are released by phagocyte.
ANTIBODY DEFENSES AGAINST BACTERIAL INVASION
(Bacterial
adhesion
molecules)
Produced by - Could limit bacterial metabolism

immune - A number of other mechanisms less
system e.g. important in the bacteria compared
tetanus with viruses & protozoa
AVOIDANCE STRATEGIES (EVASION) BY EXTRACELLULAR BACTERIA

1. Antigenic change => “escape mutants”
2. Block phagocytosis
3. Inhibit destruction within phagocytes e.g. Listeria
4. Interfere with activation of complement
5. Destruction of or interference with inflammatory & immune cells
a. Some bacteria e.g. Staph aureus produce toxins that lyse neutrophil membranes
b. Gram-negative bacteria secrete leukotoxins
BACTERIAL TOXINS
• Neutralisation of bacterial toxins by antibodies
• Some bacteria produce exotoxins which the immune system must neutralise in addition to bacteria
• Antibodies directed against the toxin prevent the toxinʼs effect on host
• If toxin has already bound to receptor, antibodies are ineffective
• Production of toxin-neutralising antibodies forms the basis of vaccination against diseases such as
tetanus & anthrax, which are notorious for producing exotoxins
IMMUNE RESPONSES & EVASION STRATEGIES BY INTRACELLULAR BACTERIA

• Some bacteria survive & grow inside macrophages by e.g.
• Cell wall resistance to lysosomal enzymes
• Prevention of fusion of phagosome-lysosome => no exposure to enzymatic degradation
• Escape from phagosome into host cell cytoplasm (resident macrophages) e.g. Listeria,
Mycobacterium, TB
•Macrophages must be activated from activated Th1 & NK cells
• Th1 response => containment of intracellular bacteria & lesions with few bacteria
• Th2 response => antibody production => lesions of masses of bacteria (AAMs)
• Animals may have an MHC type skewed for Th2 vs. Th1 & vice versa.
• Host cells infected lysed by Tc cells
• Bacterial antigens present in MHCI
CAMs (APC1) AAMs (APC2)

DCs IL-4 induced, AAMs
Effector macrophages IL-10 induced, immature DCs
Activated by IFN-gamma, TNF- Activated by IL-4, IL-10 or PGE2 (Th2-derived cytokines)
alpha (Th1-type signals)
Pro-inflammatory Anti-inflammatory
Kills intracellular pathogens Kills parasites
Mediate induction & maintenance of tolerance towards allergens &
environmental substances, participate in healing processes by
enhancing angiogenesis
Abuse of AAMs in support of infectious susceptibility or tumor immune
escape is counteracted by the classical pathway.
Have receptors of innate immunity with broad specificity for foreign
antigen & anti-inflammatory cytokines.
Express cytokines, arginase 1, RELM-α, Ym1, etc.
Acute Chronic
ALTERNATIVELY ACTIVATED MACROPHAGES IN HELMINTHIC INFECTIONS

• Not a terminal differentiation stage; can ʻswitch backʼ to CAM (activated by Th1-type signals e.g. IFN-ϒ)
• Microbial destruction via NO, etc.
• L-arginine -(arginase 1)-> L-ornithine & Urea
• L-ornithine --> Proline & Polyamines
• Collagen production & cell proliferation -> Tissue repair & remodelling
• Excessive fibrosis = immunopathology (because causes permanent damage)
IMMUNE RESPONSES TO VIRAL INFECTIONS

• Immunopathology results due to “synergism” between virus & bacteria
EFFECTOR MECHANISMS INVOLVED IN CLEARING PRIMARY INFECTIONS & PROTECTION

AGAINST RE-INFECTION
• Inhibition of virus replication
• Interferons: Absolute key players
• TNF-a: Interferon-like activities, activates other leukocytes
• Destruction of virus infected cells
• Apoptosis, complement, NK cells, T-cells, PMN
• Neutrophils: There are a number of viral infections where theyʼre important, e.g. the innate
response to Herpesvirus. They can also produce IFN-like cytokines. They DO play a role, but arenʼt
seen in the numbers we see lymphocytes & macrophages.
• Neutralisation of virus particles: Similar components as in destruction of virus-infected cells
• Coordination of response
• Needs a kickstart from INF-b (This is a cytokine. Other cytokines include chemokines, GFs & ILs)
• APCs
• Recruitment of effector cells (CTL, NK cells, etc.)
THE INTERFERON RESPONSE: FIRST LINE OF DEFENSE
• Type I interferon (IFNα/β)

• Produced by lots of different cells, including virus-infected cells
• Type II interferon (IFN-ϒ)
• Produced by immune effector cells
• Soluble proteins (cytokines)
• Viruses take part or all of the cellʼs machinery fo its replication. INF comes in & blocks some of those steps.
It interacts with specific cell surface receptors (IFN-R) to trigger gene activation.
• Direct inhibitory effect on virus replication: block protein synthesis
• Species specific
• Not virus specific
• “Cross-protection” e.g. animal encounters BVD so IFNs are produced. Upon encountering the
second virus, they are more resistant to it due to the presence of these IFNs.
RECOGNITION OF RNA VIRUSES THROUGH INTERACTION OF

DSRNA WITH MDA5 & RIG-1 HELICASES (left)
• dsRNA activates a number of other important molecules (RIG-1 &
MDA5). Subsequent steps involve IFN-type
intermediates,culminating in IFN response factors (IRFs).
• The result is production of IFNs as well as other cytokines via NF-
κB.
• This mechanism can also result from caspases (in the context of
apoptosis).
THE INTERFERON RESPONSE: FIRST LINE OF DEFENCE

• Like with any cytokine, they are a double-edged sword
• For example, IFNs canʼt be used in treatment as there is no dose
which provides efficiency without side-effects
IMPORTANT INTERFERON
SIGNALLING PATHWAYS
• IFN binds to INF-R on cell
• Activates JAK/STAT pathway,
producing factors which interact
with protein synthesis
INTERFERON STIMULATED
GENE 15 (ISG15)
• Member of IFN signalling
pathway
• Ubiquitin-like protein
• Shows direct antiviral
activity
• Acts as a cytokine upon IFN
stimulation
• Elevated during early
pregnancy in response to
conceptus-derived IFN-τ as it
helps conceptus implantation
VIRUS-MEDIATED BLOCKING OF INTERFERON SIGNALLING

• Interferon receptor homologues
• Secreted viral proteins homologous to cellular receptors binding & inactivating IFN
• Disruption of IFN mediated signalling
• e.g. Nipah & Hendra virus V proteins bind & sequester STAT
VIRUS NEUTRALISATION BY ANTIBODY

• Blocking of virus binding to cells: Interferes with receptor binding
• Blocking of virus entry into cells: Ab binds & inhibits viral protein required for membrane fusion
• Prevention of virus uncoating: Ab binds virion protein required for release of viral nucleic acid into cell
• Prevention of cell-cell “migration” of virus: Some viruses pass through tight junctions
• Prevention of budding of virus
WAYS TO KILL A VIRUS-INFECTED CELL

• Virus-specific Tc cells can be turned into memory cells which will be there next time the virus is
encountered
• Abs can form & bind to virus, then NK cells kill the infect cells via the Fc receptor mechanism (ADCC)
VIRUS ANTI-APOPTOTIC STRATEGIES

• Induce apoptosis in Tc cells (stops that “kiss of death”)
• Subversion of apoptotic enzymes: Escape from death row
• Adenoviruses regulate apoptosis at several steps in cascade so infected cells survive to the end of
replication cycle
• “The virus insurance policy”
T-HELPER CELL ACTIVATION OF CTLS

• Activation of CD4+ Th cells by DCs involves a CD40-CD40L co-stimulatory signal & recognition of MHCII
• If Th & Tc are present at DC cell surface at same time, Th-derived cytokines stimulate differentiation of
CTL-precursors into activated, MHCI restricted CTL
• Also ʻlicenceʼ DC for future interaction with CD8+ cell: upregulation of co-stimulatory molecules (CD80,
CD86) & production of IL-12
• Pathway A) is most common
ANTIBODY-DEPENDENT CYTOTOXICITY (ADCC)

• Mechanism of cell-mediated immunity where effector immune cell actively lyses target cell bound by
specific Abs
• One of the mechanisms through which Abs, as part of the humoral immune response, can act to limit &
contain infection
• Mediated by NK cells; neutrophils & eosinophils can also mediate

• Eosinophils can kill helminths; their Fc receptor recognises IgE, the antibody which can coat the
integument of helminths
• An NK cellʼs Fc receptor recognises the Fc portion of an Ab bound to a pathogen-infected target cell
surface. Once bound, the NK cell releases cytokines & cytotoxic granules which enter the cell & trigger
apoptosis. This is similar to, but independent of CTLs (Cytotoxic T-cells).
VIRAL IMMUNO-EVASION
• Hide from IR effectors
• Selection of “antibody escape mutants”
• Expression of highly immunogenic ʻdecoyʼ molecules or epitopes => e.g. antigen overload
VIRAL PEPTIDE PRESENTATION BY MHCI

• Virus protein gets targeted by proteasome
• Peptide fragments generated & translocated into ER
• Peptide/MHCI/β2 microglobulin complex formed & trafficked to cell surface
VIRAL IMMUNO-EVASION: VIRAL INTERFERENCE WITH ANTIGEN PROCESSING & PRESENTATION

BY MHCI
VIRAL IMMUNO-EVASION: MODULATION OF CYTOKINE RESPONSES

• Interruption of cytokine production (previous examples given for blocking IFN induction)
• Interference with cytokine signalling pathways (examples of disruption of IFN-mediated signalling)
• Homologues of cytokines & receptors
• IL-10 homologues which mimic IL-10ʼs immunosuppressive effects
• IFN, TNF-α & IL-1 receptor homologues which inhibit cytokine binding
LYMPHOID TISSUE RXNS DURING LENTIVIRUS INFECTION
“DIRECT” IMMUNOPATHOLOGY (viral replication): HPAI INFLUENZA

MAREKʼS DISEASE
FIV/HIV/SIV IMMUNOPATHOLOGY
IMMUNE RESPONSES TO PARASITES & FUNGI

• Chronically infected host resistant to re-infection with fresh organisms = concomitant immunity
IMMUNE EFFECTOR CELLS IN HELMINTH INFECTION

• AAMs: Express a whole bunch of cytokines.
• Eosinophils: Arenʼt as effective as neutrophils, occur in response to parasite surface molecules
• Alongside elevated IgE, one of the main characteristics of a helminth infection (even peripheral
eosinophils)
• Occur particularly when parasites invade or migrate through tissues
• Eosinophils & high levels of IgE, driven by elevated IL-5 levels, play a critical role in killing
multicellular parasites
• Less phagocytic; damage parasita via secretory products
• Activity enhanced by mast cells
• Basophils: Minor component; skews immune response towards a Th2.
NEUTROPHILS IN PARASITE INFECTIONS

• Produce ROS, including NO & superoxide radicals
• Possess Fc- & Complement-receptors
• Some parasites activate the complement cascade. Once theyʼre covered in antibodies &
complement, neutrophils bind to them via Fc receptors & exert Antibody &/or Complement-
Dependent Neutrophil Cytotoxicity.
• May also respond to the tissue damage caused
IMMUNE RESPONSES TO HELMINTHS

• Recognition of helminth-derived products mediated by recognition receptors (e.g. TLRs, lectins)
• Secretion of effector molecules e.g. cytokines, enzymes
• “Alarmins” & Dendritic Cells conditioned with some helminth products promote Th2 differentiation
• May also require changes in expression of co-stimulatory molecules e.g. OX40L, CD40, Notch-ligands
CD4+ T-CELLS IN HELMINTH INFECTION

• IL-4 & IL-13
• Act on smooth muscle for increased contractility
• Act on epithelial cell for increased permeability
• => “Weep & sweep” response
• Act on goblet cell => mucus secretion
• Much peristalsis hopefully leads to explusion of parasite!
EPITHELIAL CELL-DERIVED CYTOKINES IN PARASITE INFECTIONS

• TSLP expression is induced following helminth infection, tissue damage & exposure to cytokines
• IL-25 expression induced by helminths & exposure to common allergens
• IL-33 expression is upregulated by helminths
IMMUNE RESPONSES TO HELMINTHS - EXPULSION FROM THE GUT

• Parasite first damaged by IgG passing into gut lumen - consequence of IgE-mediated inflammation & aided
by cytotoxic (ADCC) cells.
• Cytokines stimulate proliferation of goblet cells & mucus secretion.
• Mucous coats damaged worm & facilitates explusion by:
• Increased gut motility induced by mast cell mediators (e.g. leukotriene, D4)
• Diarrhea from inhibition of glucose-dependent Na+-absorption by mast cell-derived histamine &
PGE2
IMMUNE RESPONSES TO ECHINOCOCCUS spp.

• Follicle-Associated Epithelium (FAE) contains M cells that take up ingested parasite antigen & transfer it to
local Dendritic Cell (DC lying under M cells saying, “Feed me! Feed me!”)
• Dendritic Cells present antigen to B- & T-cells.
• Epithelial cells of guy express polymeric Ig-receptors (pIgR) which mediates transcellular transfer of Abs
• Inflammation & damage to gut epithelium may ensue.
IMMUNE RESPONSES TO PROTOZOAL INFECTIONS

• Protozoa change their surface molecules like people change clothes every day, & can escape the immune
response in this way.
• Plasmodium (malaria): Antibodies play some role => hemolysis with cells containing protozoa.
• Trypanosomes & Leishmania: Antibodies donʼt play much of a role. Immune response is similar to viral
immune response because they live intracellularly, so macrophages need a Th1, NOT Th2 response.
IMMUNO-EVASIVE MECHANISMS
• T. cruzi
• Escapes from confines of phagosome into cytoplasm
• Pre-emption of Complement-mediated defences by expressing a DAF-like
molecule which accelerates decay of C3b (e.g. T. cruzi)
• Expression of molecules which bind C9 => inhibition of MAC formation
(e.g. S. mansoni)
• Malaria in rbcs express protein which binds CR1 on other rbcs => rosette
formation -> facilitate spread of parasite with minimal IR exposure
• Release surface antigens = decoy proteins mobbing antibody (e.g. T.
brucei)
• T. gondii inhibits phagosome-lysosome fusion
• Leishmania spp.
• Surrounded by lipophosphoglycan - protects from oxidative burst by
scavenging oxygen radicals
• Downregulate MHC, CD80 & CD86 expression - diminish T-cell stimulation
• Molecular mimicry:
• Homology to host antigens (e.g. Ascaris & human collagen)
• Parasite cover surface in host protein (e.g. adult schistosomes take up
host rbc glycoproteins, MHC & IgG => protection despite circulating
antibodies meant to prevent re-infection)
• Antigenic variation
• Trypanosome is shed out of the host but becomes a recalcitrant persistent infection.
• CD80 & CD86 = co-stimulatory molecules affecting the way B- & T-cells interact with each other.
IMMUNITY TO ECTOPARASITES
• Defences - Grooming, hide thickness
• Adaptive responses
• Langerhans cells - the SALT DCs - present antigen to T-cells
• Immediate type I hypersensitivity (IgE & mast cells) stimulates grooming
• Delayed type IV hypersensitivity (CAMs & Th1) affects tick survival
• Responses mainly directed against saliva, which inhibits alternative complement pathway &
decreases lymphocyte responses (ectoparasites have Histamine-Binding Proteins)
• Immunosuppression of host enhances feeding & pathogen transmission
PARASITE-INDUCED IMMUNOSUPPRESSION
• Classical feature of African trypanosomiasis in cattle, sheep & humans
• Pathogenesis related to macrophage over-activation & uncontrolled production of harmful substances
• TNF-α produced by host; has a global effect on body, including the liver. Sustained levels exhaust
hepatocyte function.
• Molecules shed by trypanosomes
• Treatment & prevention strategies focused on “antidisease vaccines” i.e. immunity to molecules causing
anemia, thrombocytopenia, etc.
• Alternatives include selection of trypanotolerant breeds
• Trypanosomes change clothes on a regular basis!
PARASITE-INDUCED IMMUNOPATHOLOGY
• Chronic parasite infection accompanied by immune response frequently leads to tissue-damaging reactions
• Trypanosomiasis: sustained TNF-α response
• T. cruzi (Chagasʼ disease): auto-immunity (cross reaction b/w parasite & self) - possible cause of
cardiomyopathy
• Schistosoma mansoni: hepatic cirrhosis, macronodular regeneration, chronic granulomatous
lymphadenitis, bilharzial pigmentation
• Cryptosporidiosis: chronic biliary hyperplasia
VACCINES AGAINST PARASITES

• Vaccines against parasitic diseases are difficult to develop
• Focus on: immunity to infectious stages; immunity to blood cell stages; immunity to saliva; immune
responses blocking laying & viability of parasite eggs/development of larvae
• Blocking of transmission cycles have greater impact
• Majority of vaccines against parasites are live, attenuated formulations (little known about mechanism
therefore hard to improve)
• Subunit vaccines = main focus (molecular approach with antigen identification)
• Problems:
• Complexity of antigens
• Complexity of life cycles
• Difficulty maintaining parasite cultures in laboratory
• Inaccessibility of key stages of life cycle
• Immunosuppression
• Antigenic variation
• Examples of live attenuated vaccines:
• Tick Fever (cattle): Targets Babesia bovis (main pathogen), B. bigemina, Anaplasma marginale.
Long lasting immunity (at least 4 years). Cons are cold chain requirement (may result in co-
transmission) & non-responders.
• Coccidiosis (poultry)
• Toxoplasmosis (sheep)
• Lungworm (cattle)
• Examples of sub-unit vaccines: Taenia saginata, T. ovis, T. solium, Echinococcus granulosus
• TickGARDPLUS: Antibody mediated; ticks ingest antibodies with blood meal. Reduce pasture
contamination.
IMMUNE RESPONSES TO FUNGI

• Infection often associated with immuno-suppression of the host or commensal flora are upset by prolonged
broad-spectrum -> systemic infection
• Phagocytosis, following Th1-mediated activation of macrophages
• Some fungi reside in macrophages!
• ROS = fungicidal; counteracted by some fungi by catalase, mannitol & melanin production -> inhibits
respiratory burst of macrophages & granulocytes
• NK cells may have constitute anti-fungal activity
• Antibodies against fungal antigens may be protective in some infections
• Antigenic changes & changes from filamentous moulds to unicellular yeasts can = virulence factors
IMMUNISATION - VACCINES
• Consider specificity & memory
• The Ideal Vaccine:
• Induces strong, long-lived, “sterilising” immunity

• Prevents replication of agent & disease
• No side effects
• Cheap
• Easy to store
KILLED VACCINES
• Meant to destroy infectivity BUT retain immunogenicity
• Predominantly elicits humoral response
Pros Cons
Usually safe to use in preg & very young animals Immunity not as strong or long-lasting c.f. live vaccines
Easier to store & apply vs. live vaccines Usually poor cell mediated immunity
No contaminating organisms (assuming proper Cost of production often high
inactivation)
Need multiple doses (boosters)
Usually requires adjuvant (may cause severe local rxn)
Problems with improper inactivation & quality control
LIVE VACCINES
• Organism replicates in host - mimics natural infection, giving strong immune response
• Attenuation achieved by:
• Using naturally occurring avirulent/low virulence variant
• Adapted/selected in lab
• Genetic engineering
• e.g vaccines against parasites
• Virulent (unmodified) organisms donʼt cause disease because theyʼre:
• Inoculated by abnormal route (e.g. scabby mouth)
• Given to an “abnormal” host e.g. turkey herpesvirus to protect chickens against Marekʼs disease
• Live recombinant organisms are when the antigen is inserted into another organism which is then
inserted into the host
• Protective immune responses to “gene-donor” organism
• Many of the advantages of live vaccines
• e.g. pox virus expressing G-protein of rabies virus; fowlpox virus expressing Newcastle disease
virus proteins; lactobacillus expressing Chlamydia spp. antigens
Pros Cons
Strong immunity Constraints on storage & administration
Better inducer of cell-mediated immunity Canʼt be used in very young/pregnant animals
One dose needed Contamination with pathogenic organisms*
Lower cost of production Reversion to virulent strains*
Better at overcoming maternal immunity * Unlikely due to strict quality control
Good mucosal immunity standards
SUBUNIT VACCINES
Protein/Peptide-based
• Purified
• Immunogenic portion of organism
• e.g. footroot virus antigens, herpesvirus envelope proteins
• Recombinant
• Intact protein
• Part of protein
• Post-translational modifications (glycosylation, folding)
• Usually need adjuvant protein carrier to elicit adequate immune responses
• In general, similar advantages & disadvantages as killed vaccines
• e.g. vaccines against parasites
Nucleic Acid-based
• Immunogenic epitope(s) [vaccine antigen] inserted into plasmid w/ strong promoter [adjuvant effect]
• Adjuvant: agent that may stimulate the immune system & increase response to vaccine, without
having any specific antigenic effect in itself
• DNA taken up into cells - vaccine antigen & adjuvant molecule expressed
• If injected into muscle => prolonged expression of protein -> taken up by dendritic cells & presented to T-
cells
• e.g. Melanoma DNA vaccine for dogs
Pros Cons
Induces cell-mediated & humoral immunity Public concern re: injection of foreign DNA into food
Can include multiple antigens in a single plasmid animals/humans
Low cost of production & easy to store
TOXOID
• Purified toxin from bacterial cultures -> inactivated/modified (usually with formaldehyde)
• Given after adsorption to alluminium hydroxide, an adjuvant
• Some mixture of bacteria & toxoid e.g. some Pasteurella & Haemophilus vaccines. Toxoid provides Th-
epitopes for such vaccines i.e. adjuvant effect.
• T-cell epitopes are presented on the surface of an APC, where they are bound to MHC molecules.
• Epitopes are an antigenic determinant; the part of an antigen recognised by T-cells, B-cells or antibodies.
Adjuvants
• Pharmacological or immunological agent that modifies the effects of other agents while having few direct
effects when given by itself
• Non-living organisms & subunit vaccines require adjuvant to activate innate immune response via Pattern
Recognition Receptors (PARRs e.g. TLR) on accessory cells
• Often included in vaccines to enhance the recipientʼs immune response to a supplied antigen while keeping
the injected foreign material at a minimum
Mode of Action **NOT IMPORTANT?**

• Depot effects: Delay release of antigen & enhance uptake by macrophages
• Activation of APCs: Induce a granulomatous inflammation & activation of APCs (express co-stimulatory
molecules)
• Lymphocytic effects: APC cytokines co-localise with antigen-sensitised lymphocytes => enhanced cell
activation & antigen presentation
New Generation Adjuvants

• Liposomes
• Cytokines
• ISCOMS: Hydrophobic matrix of saponin bark of Amazonian tree
• Virus-like particles: Self-reassembling components (structural proteins) of certain viruses e.g. rotavirus
• CpG oligodeoxynucleotides (ODNs): Bind to TLR9 & stimulate innate response. Potent Th1 immune
enhancer.
VACCINE DELIVERY ROUTES: MUCOSAL VACCINES

• Constantly bombarded & vulnerable
• Immune responses most efficiently induced by administration of vaccines onto surfaces
• Injected vaccines = poor inducers, so poor/suboptimal efficacy of respiratory & enteric disease vaccines
• Lots of Ab-producing cells in intestinal mucosa
• Mucosal tissues in state of constant alert, but adapted to foreign micro-organisms & their products
• Vaccines that would produce vigorous immune responses if injected into a sterile environment (e.g. muscle)
might be “ignored”
• “Correlates of mucosal protection” (concentrations, epitopes specificities & locations of immune
effectors likely to protect hosts during mucosal exposure to disease) are difficult to define
• Vaccines given orally or deposited directly face hostʼs defences:
• Dilution in mucosal secretions, capture, attack, exclusion
• Relatively large doses that mimic successful mucosal pathogens required: multimeric &/or
particular, stimulate innate responses & evoke adaptive immune responses
VACCINE DELIVERY ROUTES: MUCOSAL VACCINE-DESIGN CHALLENGES

• Breaching epithelial barrier
• Adherence is key
• Achieved with live attenuated virus or bacteria
• Increased by incorporation into microparticles
• Alerting mucosal immune system
• Live attenuated vaccines better due to activation of multiple immune pathways
• Microparticles designed to contain innate immune responses-activators (e.g. bacterial CpG
sequences)
CpG sites: methylation leads to silencing.
Unmethylated CpG motifs are prevalent in bacterial DNA but not vertebrate genomic DNAs.
CpG motifs in bacteria rapidly induce lymphocytes to secrete IL-6, IL-12 & IFN-ϒ.
PASSIVE IMMUNISATION (not a type of vaccine)
• e.g. Tetanus antitoxin immune globulin: Raised in horses, host had suspected exposure to Clostridium
tetani
• e.g. Anti-rabies immunoglobulin: Pooled human immunoglobulin - give to people following suspected
bite
ADVERSE REACTIONS TO VACCINES

1. Local reaction at site: Reaction to inactivating agent or adjuvant
2. Transient immunosuppression
3. Hypersensitivity reactions
• Type I: Within min to hours e.g. urticaria (raised area of skin often warm, red & itchy) following
rabies vaccination
• Type III: Intense local inflamm rxn or gen vascular disturbance
• Immune complex uveitis (inflammation of uvea - iris, ciliary body, choroid - of eye) after
canine adenovirus vaccination.
• Type IV: granuloma formation at vaccination site
• Eosinophilic granulomas with necrotic centers (horses)
4. Autoimmune disease
• Autoimmune encephalitis with use of rabies vaccine
5. Soft tissue sarcomas
• Following subcutaneous delivery of several feline vacines
• Pathogenesis thought to involve Ag/adjuvant-induced granulomatous reaction (Type IV)
VACCINE FAILURE: DEVELOPMENT OF DISEASE DESPITE VACCINATION

• Vaccine factors
• Improper storage of vaccine
• Improper administration of vaccine (wrong route or dose)
• Reversal to virulence
• Using vaccine past expiry date
• Host factors
• Animal already incubating disease at vaccination or infected immediately after
• Maternal Ab interference
• Immunosuppression (health/nutrition, drugs or infectious agents)
• “Non-responder” (genetic factors) - small proportion of animals mount poor immune response
SEROLOGICAL TESTS
• Based on Ag-Ab interactions
• Diagnose infection, allergy, cancer etc. by:
• Detection of Ag-specific Ab in serum
• Detection of Ags e.g. infectious agents, cancer cell-specific components, toxins, cytokines, etc. in
blood/tissues
REAGENTS USED IN SEROLOGICAL TESTS

• Polyclonal antibodies: Animal immunised with Ag preparation - Ab of different classes & subclasses
develop in response to multiple epitopes of Ag - blood collected & serum separated = polyclonal antiserum
• Monoclonal antibodies
• Anti-globulins:
• Immunoglobulins purified from serum (Ag) -> injected into another species -> production of anti-
globulin polyclonal antiserum (e.g. goat-anti-rabbit-IgM or IgG)
• Can be isotype specific
PRODUCTION OF MONOCLONAL ANTIBODIES

• Selected hybridoma (fusion of lymphocyte & tumor cell) grown to
high density in tissue culture -> transferred to serum-free medium ->
secreted Abs precipitated out or purified on affinity columns =
monoclonal Abs
• Cell culture supernatant can also be used directly
ENZYME-LINKED IMMUNOSORBENT ASSAY (ELISA)

• Requires a pure preparation of known Ag or Ab (or both) as standard
• Detection of antibody uses indirect ELISA:
• Microwells coated with antigen
• Dilution series of control & test serum added to wells - Ag-specific Abs
bind to Ag
• Wash away unbound & add tagged antiglobulin
• Add enzyme substrate - measure colour development
• Detection of antigen uses “sandwich” ELISA = antigen-capture
ELISA:
• Microwells coated with antibody = “capture antibody”
• Test sample added to wells
• Wash away unbound & add second antibody (also specific for
antigen)
• Second antibody may be tagged with enzyme or add tagged
antiglobulin
• Add enzyme substrate - measure colour development
AFFINITY CHROMATOGRAPHY
• Antigen-antibody binding purifies antigens or antibodies
• Other proteins cleaned out & eventually => purified antibodies

• Protein G-sepharose & other “inert” materials bind immunoglobulins in a non-specific manner
COMPETITIVE INHIBITION ASSAY (right)

• Unlike ELISA, does allow direct measure of antigen/antibody in unknown sample
• Determined by antigenʼs ability to compete with labelled reference antigen for antibody attached to
microtitre plate wells
• Determined by antibodyʼs ability to inhibit binding of labelled antibody
LUMINEX ASSAY (above)

• Detection & quantitation of proteins & DNA
• Requires less starting material than ELISA; faster & more sensitive
• Can be multiplexed i.e. detection of diff proteins/DNA molecules in
unknown sample
• Assays for detection of >50 proteins in body fluids now available for some
species
• Multiplex assays developed for measurement of selected cytokines in
plasma & milk from horses, cattle & dogs
RAPID IMMUNOASSAY DEVICES (left)

• Quick tests used in vet clinics
PRECIPITIN REACTION-BASED TESTS

• Visible precipitate formed when sufficient quantities of Ab are
mixed with soluble macromolecular Ags
• Precipitate = large aggregates of Ag cross-linked by Ab molecules
• Ag or Ab excess will hinder immune complex formation
• Precipitin reaction affected by # of binding sites & valence of Ab &
Ag (max. # of Abs that can be bound by an Ag)
• Often replaced by ELISA
• Layer of agar that permits radial diffusion
• Includes:
• Double gel diffusion assay - Ag & Ab are placed in diff wells. Used
to determine Ag relationships.
• Immunoelectrophoresis
• If EQUIVALENCE is reached, there will be a white line. In this
example, the presence/absence of Ag or Ab have been
measured. If itʼs the sample Ag in sample M as in V, result
mimics B.
AGGLUTINATION ASSAYS
• Interaction between antibody & particulate antigen resulting in visible clumping
HAEMAGGLUTINATION ASSAYS
• Typing of blood groups
• Identification of viruses or detection of antibodies to a virus
• Some viruses can agglutinate rbcs (e.g. from chicken) = virus HA assay
• Antibodies to virus can inhibit this agglutination = HI assay
COMPLEMENT FIXATION TEST

• Detect antigen-specific antibody in serum
• Based on principle that when Ab is bound to Ag, complement is activated & MAC produced
• If Ag bound to rbc, MACs lyse rbcs
• Assay protocol:
• Inactivate complement in serum
• Mix serum with Ag of interest, then add complement
• Add Ab-coated sheep rbcs
• If all complement had been “fixed” in initial rxn due to presence of Ag-specific Ab, no lysis will
occur
• If no specific Ab is present, then complement binds to Ab-coated sheep rbcs => haemolysis
WESTERN BLOTTING
• Goal: ID Ags in complex mixture of proteins or measure Ab responses to particular Ag(s) in
mixture of proteins
• Protein mix first separated on acrylamide gel by electrophoresis
• Protein bands transfered onto nitrocellulose membrane by electrophoresis (“blotting”)
• Protein-containing membrane (“blot”) is immuno-labelled after same principles as for an
ELISA
• Specific protein bands are visualised with chromogen or enhanced chemiluminescence
(ECL)
Quick Refresh: ELISA

Antigen Capture - “Sandwich” ELISA
Indirect: Antigen-specific primary antibodies (if present in serum) bind to antigenic well
coating.
• Antiglobulins act as detection antibodies.
ECL WESTERN BLOTTING

• Common technique for detection assays.
• HRP (HorseRadish Peroxidase) tethered to molecule of interest (usually through labelled Ig).
• Enzyme complex catalyses peracid -> oxidant.
• Combined with luminol, further oxidation by H2O2 => triplet (excited) carbonyl which emits light when it
decays to singlet carbonyl.
IMMUNOLABELLING OF CELLS & TISSUES: IMMUNOFLUORESCENCE MICROSCOPY

• Direct immunofluorescence
• Fluorochrome bound to 1° antibody
• Used to detect Ag in tissue sections or culture cells (e.g. confirm presence of virus)
• Indirect immunofluorescence
• Fluorochrome bound to antiglobulin
• Antiglobulin: Ab directed against gamma globulin, as used in antiglobulin test
• Used to detect Ag in tissue sections (i.e. viral Ag) or Ab in serum
IMMUNOLABELLING OF CELLS & TISSUES: IMMUNOHISTOCHEMISTRY

• Provides a permanent sample, c.f. immunofluorescence microscopy (but similar use)
• Direct or indirect labelling - latter more sensitive
• Double-labelling also possible
• Enhancer can be used to create stronger staining labelled antibody
IMMUNOLABELLING OF CELLS & TISSUES: IMMUNO-ELECTRON MICROSCOPY

• Used to ID bacteria & viruses in tissues, faecal samples & body effluents - especially those difficult or
impossible to grow in culture
• Superseded by PCR or RT-PCR
• Still some usage as research tool
FLOW CYTOMETRY
• Uses principles of light scattering, light excitation & emission of
fluorochrome molecules to generate data from particles & cells
1.Sample is injected into center of sheath flow

2. Combined flow is reduced in diameter, forcing cell into center of
the stream
3. Cells are lasered one at a time
4. As cells of interest intercept light source, they scatter light &
fluorochromes are excited to a higher energy state
5. Energy is released as a photon of light with specific spectral
properties unique to different fluorochromes
• MEASURE FLUORESCENCE PER CELL OR PARTICLE

(c.f. spectrophotometry in which percent absorption & transmission
of wavelengths is measured for a bulk volume of sample)
NEUTRALISATION ASSAYS
• Detection of Ab by its ability to neutralise
biological activity of Ag or organism
• Cytopathic: Degenerative change in cell (in this
case, caused by virus suspension)
TITRATION OF ANTIBODIES
• Serum is diluted in series
• Each dilution is added to serological test
• Highest dilution giving a positive result = titre (positive result being presence of Ab)
INTERPRETATION OF SEROLOGICAL TESTS

• Must recognise limitations of testing for Ab
• Other interpretations if animal is Ab positive (past exposure, vaccination, maternal Ab, cross-reactivity to
related organism)
• Other interpretations if animal is Ab negative (sample taken too early in course of infection,
immunosuppression, presence of neutralisation-interfering Abs, congenital persistent infection e.g. border
disease virus)
• Need to take paired blood samples to diagnose infections

• First taken at acute stage of disease
• Second taken at convalescent stage (2-3 weeks after disease period)
• If thereʼs a 4-fold or greater increase in titre, conclude infection did occur.
• Single sample useful for:

• Surveys of infection in populations
• Confirming vaccination responses
• Determining infection status of animal to
persistent pathogen
• Presence of Abs in face of
agentʼs presence suggests
persistence
• Lack of Abs in face of agentʼs
presence may also suggest
persistence e.g. BVD virus
SENSITIVITY & SPECIFICITY

Sensitivity = infected animals with a positive result / total # of infected animals
• In a highly sensitive test, nearly all infected animals are detected (so few false negatives)
Specificity = non-infected animals with a negative result / total # of non-infected animals

• In a highly specific test, nearly all positively-tested animals are truly positive (so few false positives)
TUMOR IMMUNOLOGY
• Tumor cells are negative for vascular endothelial & smooth muscle cell markers.
• Many malignant lymphomas are multicentric c.f. metastatic (i.e. they spread to more than one center)
TUMOR DEVELOPMENT
• Initiation: Irreversible fixation of a genetic alteration (e.g. hyperplasia)
• Promotion: Reversible expansion (e.g. dysplasia)
• Progression: Irreversible transition to malignant & invasive tumor (e.g. neoplasia, wit preneoplasia being
point of no return?)
DEFINING PROPERTIES OF PRE-NEOPLASIA

1. Associated with an increased risk of cancer
2. Some chance of progressing to cancer & resulting cancer arises from cells within precancer
3. Different from normal tissue from which it arises
4. Different from cancer into which it develops. May have some, but not all, of the molecular & phenotypic
properties that characterise cancer.
5. Must be a method by which it can be diagnosed.
CANCER: AN INFLAMMATORY LINK

• During the inflammatory process, lots of GFs are produced. As cells replicate faster during inflammatory
process, 2 things can happen:
1. DNA repair processes may not be able to keep up & work fast enough
2. A single cell has its cancer gene switched on; replication of this cell during inflammation leads to
increased numbers of cancerous cells
• For all we know, there are a lot more preneoplastic cells arising that our immune system handles.
• In a chronic inflammatory process, everything is sped up & the immune system may not be able to handle
it.
TUMOUR-ASSOCIATED SURFACE CHANGES

• A whole bunch of things can be up- or down-regulated on the
tumour cellʼs surface which confuzzle the immune system.
• Ideal tumor antigen would be expressed by tumor cells, but not
normal cells & would be required for tumor growth, preventing the
tumor from losing expression of this antigen.
• It might also be acceptable to target antigens highly expressed
by the tumor but also expressed by a restricted range of normal
non-transformed cells, depending whether the potential damage to
normal tissue can be kept in an acceptable range. For the most
part, tumors antigens are non-mutated proteins or other molecules
aberrantly expressed by that specific tumor.
IDENTIFICATION OF TUMOR-SPECIFIC AGS

• Artificial processes
1. Identification of tumor-specific gene using tumorspecific cytotoxic T-cell (Tc) clones derived from mixed
tumor-lymphocyte culture
• A cosmid library incorporating the tumor DNA is transfected into an autologous target cell (cell derived from
wild-type tumor but antigen-negative)
• Small pools of transfected cells are tested against the tumor-reactive Tc.
• Expansion of T-cells in response to cells transfected with a particular cDNA identifies the protein encoded
by this cDNA as a candidate tumor antigen
• A positive pool is cloned by limiting dilution & the tumor-specific gene cloned from the antigen-positive
well(s).
Cosmid Library
• Used for cloning genes with large introns & sequencing larger chunks of the genome.
• Cosmid vectors are hybrids of plasmid & bacteriophage lambda DNA.
• Because of a COS sequence, cosmid recombinants can be packaged into virus particles (allowing high
efficiency of transformation).
• Cosmids donʼt form viral particles because the recombinant DNA doesnʼt encode any phage proteins but
rather forms large circular plasmids.
• The colonies that arise can be selected on antibiotic plates, like other plasmid DNA transformants.
• The likelihood of obtaining a DNA clone containing the entire gene sequence is increased significantly
when using a cosmid library.
2. Using serological ID of Ags expressed by recombinant cloning (SEREX)
IMMUNE RESPONSE TO TUMORS
TUMOR ANTIGENS: THE ROLE OF HEAT SHOCK PROTEINS

• Heat Shock Proteins = Upregulated during “stress” to form complexes with tumor Ags & increase their
MHCI presentation
• Possible vaccine target, despite being difficult to regulate
• These are up-regulated during the inflammatory response as well as being expressed by a number of
bacteria. Therefore, they can induce an autoimmune response.
IMMUNE SURVEILLANCE OF MALIGNANT CELLS

IMMUNE EVASION BY TUMOURS

• Regulatory/suppressor T lymphocytes prevents transplant rejection & allergies but likely promotes
cancer.
• In fetuses, to ensure no rejection by dam, there is an absence of “danger signals” on Ags (PAMPs &
molecules which activate DCs). T-cells which then meet the non-activated DC presenting the tumour-Ag may
become tolerant to the tumour-Ag. This mechanism is bad in relation to tumors!
CANCER IMMUNO-THERAPY
• Monoclonal Abs (Abs that are the same because
theyʼre made by the identical immune cells cloned
from a unique parent cell) that recognise tumour-
specific Ags are used to help eliminate tumours.
DEVELOPMENT OF THE IMMUNE SYSTEM IN THE FETUS & NEONATE

• In humans & most domestic animals, functional CD8+ (cytotoxic) cells present at birth
• Immune system requires antigenic stimulation usually encountered after birth to complete development
• In general, every exposure to Ags in neonates is a primary response
• Maternal humoral immunity transferred via colostrum, placenta or yolk sac.
• Ab transfer in ruminants, pigs & horses purely via colostrum.
• Horses & pigs predominantly contain IgA in milk & IgG in colostrum
COMPOSITION OF COLOSTRUM
• Accumulated secretions of mammary gland over last few weeks of pregnancy + proteins actively
transferred from blood
• Contains IgG & IgA, with smaller amounts of IgM & IgE, as well as lymphocytes & cytokines (regulatory)
• (Multiparous) Degree of passive immunity heterogeneous between litters & members of same litter
FORMATION OF COLOSTRUM
The entero-mammary immune link
• Gut-derived integrin expressing T-cells -> mammary gland via binding to addressin
• B-cells & IgA lymphoblasts require “milk chemokine” for extravasation (escape of fluid from containing
vessel)
• IgA lymphoblast crosses endothelium, develops into plasma cells & delivers IgA
• Same specificity as gut IgA
• Similar mechanisms probably applies to IgG plasmablasts?
ABSORPTION OF COLOSTRUM
• Ab in milk still plays role in protection (especially IgA)
• Time taken for serum antibody to decline to un-protective levels after colostral peak depends on initial conc
of Ab, half-life of Ab & catabolic rate in newborn (higher in fast growing spp./breeds)
MATERNAL IMMUNITY IN BIRDS

• Henʼs serum IgY (equivalent of IgG) transported into yolk while egg in ovary
• As egg moves down oviduct, IgM & IgA from oviduct secretions -> albumin -> amniotic fluid & ingested
• At hatching, chicks have IgY in circulation & IgM/IgA in intestinal tract
• Disappear ~10-20d of age
FAILURE OF PASSIVE TRANSFER

Causes
1. Production failure
a. Poor quality or insufficient colostrum produced (premature birth, low serum Ab levels in dam, genetic,
premature lactation)
2. Ingestion failure
a. Poor mothering
b. Weak, premature or deformed newborn with poor sucking
c. Litter too big so some miss out
3. Absorption failure (adequate intake of colostrum, but insufficient Ab absorbed)
a. Delay before first suckling; unknown reasons (esp. foals)
DIAGNOSIS & TREATMENT OF FPT

• Commercial assays for serum Ig concentrate used to measure specific gravity of colostrum
• If inadequate, supplement with oral colostrum or plasma transfusion if >15h old
NEONATAL IMMUNE MATURATION

• Inhibition of neonatal immune system by passively acquired maternal antibodies
• Binding of maternal Ab to FcR on neonate B-cells => inhibitory signal
• Ag is coated/blocked by maternal Abs => neonateʼs B-cells canʼt bind to Ag & respond
• Infection or vaccination during neonatal period must overcome:
• Inhibitory effects of maternal-derived Ab
• Natural Th2 regulatory environment
Early Life T-Cell Responses

• Secretion of significantly higher IL-5 & lower IFN-y levels by Ag-specific T-cells.
• This Th2 polarisation is thought to be linked to the persistence of maternal hormonal influences (e.g.
progesterone & prostaglandins) that protect the fetus against toxic effects of Th1 inflammatory cytokines
during pregnancy.
• Along with the dilution of this influence, the induction of Th1 & CTL responses after birth requires increased
exposure to antigens and progressive maturation of the cellular immune network.
• Neonatal T-cells lean towards a Th2 response due to crap interactions between APC, NK & T-cells.
• Lots of IL-4 produced by naive CD4+T-cells & other cells - could be enough to inhibit a Th1 response &
encourage a Th2.
NEONATAL IMMUNE RESPONSES & VACCINATION

• Series of vaccinations to young animals cover all levels of susceptibility/inhibition
Role of adjuvants in infant immunisation:

1. Rapidly induce strong Ab responses of the appropriate isotypes
2. Elicit sustained Ab responses extending beyond infancy
3. Induce efficient Th1 & CTL responses in spite of the preferential Th2 polarisation of early life responses
4. Escape from maternal antibody mediated inhibition of vaccine responses
5. Show acceptable reactogenicity in early life
6. Incorporate several vaccine antigens into one formulation to reduce the number of required injections
CAN WE CLOSE THE WINDOW OF SUSCEPTIBILITY?
IMMUNODEFICIENCY SYNDROMES
CHÉDIAK-HIGASHI SYNDROME
• Giant granules: Melanocytes, lysosomes, PMN
• Bleeding & recurrent infections
• Hypopigmentation
SEVERE COMBINED IMMUNODEFICIENCY DISEASE (SCID)

• Arabian or Arabian-cross bred horses: autosomal recessive, lymphopenic (low numbers of lymphocytes in
blood)
COMMON CONSEQUENCE OF IMMUNODEFICIENCY: INFECTIONS

• Granulocyte defects: Vulnerable to opportunistic environmental infections
• Complement defects: Pyogenic (pus-producing) bacterial infections
• T-lymphocyte defects: Bacterial
• B-lymphocyte defects: Viral; chronic recurrent gastrointestinal infections, meningitis
T-LYMPHOCYTE DEFICIENCY -> FUNGAL INFECTION (PNEUMOCYSTIS

CARINII)
• Fatal, special fungus reclassified from protozoa
• Fill the alveoli to the point where the animal canʼt breathe & the lungs begin to
look like liver
ACQUIRED IMMUNODEFICIENCY SYNDROME (AIDS)

• Lymphopenia (CD4+ T-cells), thrombocytopenia (few platelets), neutropenia/
neutrophilia, opportunistic infections, cancer
FELINE LEUKEMIA VIRUS

• Lymphopenia, neutropenia, impaired B- & T-cell function
• Lymphoma, fibrosarcoma, myeloproliferative disorders
• Aplastic anemia
OTHER ACQUIRED IMMUNODEFICIENCIES

• Demodectic mange
• Toxins: Immunosuppressive
• Drugs: Corticosteroids, antibiotics - inadvertently cause increased
susceptibility & immunodeficiencies
• Gumboroʼs disease (right)
• Knocks out avian bursa so B-cell component is gone
• Many viral inclusion bodies
BOVINE VIRAL DIARRHEA - MUCOSAL

DISEASE
• Immunologically incompetent bovine fetus
& persistent infection (virus seen as “self”)
• If virus mutates, animal develops mucosal
disease (fatal)
• Craters form on mucous membranes
AUTOIMMUNE DISEASE
IMMUNOLOGICAL TOLERANCE
Central
tolerance
• Arise during T-
lymphocyte development in the thymus
• Negative selection: T-cells expressing
TCRs capable of recognising self-
antigens under apoptosis (Fas-FasL)
Peripheral tolerance
• Three processes induce tolerance
induction, modifying the immune system
to prevent self-destruction
1. Anergy, lack of reaction by bodyʼs
defense mechanisms to foreign
substances, usually self-antigen,
consisting of a direct induction of
peripheral tolerance. Failure to
respond is accompanied by an
absence of co-stimulatory signals or
presence of inhibitory signals via
CTLA-4
2. Suppression: Tregs via IL-4, IL-10,
TGF-b
3. Clonal deletion: Caused by
persistent stimulation - Fas-FasL
mediated apoptosis = AICD
(Activation-Induced Cell Death)
• Antigen sequestration e.g. eye, brain,
testis, thyroid
PATHOGENESIS OF AUTOIMMUNITY
• Penetrance increases with age
• Environmental triggers (e.g. infection) often at play
• Sex hormones & defective pituitary-adrenal feedback loops may contribute
• Immune attack on self-antigens => tissue damage & disease
MECHANISMS OF AUTOIMMUNITY
Release of
sequestered antigens
Modification of auto- Provision of new carrier determinants i.e. self-antigen complexing with foreign
antigen antigen to which no self-tolerance has been established
Altered self-antigen
Cross-reactions with • Molecular mimicry
microbial antigens • Epitope spreading
Inflammation-induced • Pro-inflammatory cytokines induce de novo MHCI expression on cells - targets for
changes in antigen self-reactive CTL
processing & • Inflammation-activated DCs & macrophages may present self-antigens (e.g.
presentation nucleoprotein)
Genetic factors MHC type important
Failure of central or • Decreased suppression
peripheral tolerance • Increased proliferation
AUTOIMMUNITY DUE TO MODIFICATION OF SELF-ANTIGEN

• Can occur as a result of:
• Post-translational modification of the molecule
• Self-antigen combines with drug
• Self-antigen modified by virus-replication & -protein expression e.g. membrane proteins
AUTOIMMUNITY & THE ROLE OF INFECTIONS

Molecular Mimicry
• Similarity between epitopes on a micro-organism & a self-antigen
• T- & B-Cell Receptors specific for a microbial epitope (e.g. bacterial HSP) cross-react with similar epitopes
on other molecules, including self-antigens.
• Affinity highest for cognate antigen (whatever initially induced the reaction), but once the immune response
really gets going, there will be enough T- & B-Cells to bind to epitopes with lesser affinity.
EQUINE RECURRENT UVEITIS

• Common cause of blindness in horses
• Affected horses have antibodies to Leptospira interrogans
• Antigenic similarity between proteins in equine cornea & those of L. interrogans
MICROBIAL INDUCTION OF AUTOIMMUNITY
PARVOVIRUS B19 INDUCED AUTOIMMUNE THROMBOCYTOPENIA & ANEMIA
MICROBES & AUTOIMMUNITY

• Infection may also protect from autoimmunity!
• “Hygiene hypothesis”: Decrease of infections cause an increase in
immune disorders
• Bystander suppression: Hallmark of acquired tolerance. Response
against one Ag is suppressed when it is presented in the context of an
Ag to which tolerance is already established e.g. Ag-specific (adaptive)
T regulatory cells inhibit the T effector cell response both to specific Ag &
to a colocalised third-party Ag.
AUTOIMMUNE DERMATOSIS e.g. PEMPHIGUS FOLLEACEUS (left)

• Type II hypersensitivity response involving autoantibodies to proteins
responsible for keratinocyte cell-to-cell adhesion
• Targets desmoglein 1 in ~6% of affected dogs
SYSTEMIC LUPUS ERYTHEMATOSUS

• Results from loss of control of B-cells -> production of antibodies to a
wide variety of antigens
• Predominant autoantibody is antinuclear antibody (ANA), directed against nuclear antigens
• Genes involved: MHC class II, complement, additional unidentified genes
• Associated with defects in apoptosis

• Decreased percentage & absolute numbers of CD8+ lymphocytes & increased percentage but decreased
absolute numbers of CD4+ cells
CANINE FAMILIAL DERMATOMYOSITIS

• Inflammatory disease of skin, muscle & vasculature
• Alopecia & erythematous dermatitis involving face, ears & bony
prominences of distal extremities
• Poor vasculitis is the underlying lesion. Erosions & ulcers are common.
• Circulating immune complexes have been identified, but the auto-
antigen remains to be identified
• C4d (plays role in activation of classical pathway of complement
system) antibody deposition in affected vessels, suggestive of presence
of Ag-Ab-Cʼ complexes have been demonstrated.
SJOGREN-LIKE SYNDROME
• Systemic autoimmune disease characterised by dry eyes, dry mouth & lymphoplasmacytic adenitis
(glandular inflammation due to infiltration of lymphocytes & plasma cells)
• Adult dogs & cats
• Lymphocytic infiltration & fibrosis of lacrimal & salivary glands
• Affected dogs often hypergammaglobulinemic (presence of excess gamma globulins in blood e.g.
immunoglobulins), less frequently have identifiable antinuclear antibodies (ANAs), rheumatoid factors
(autoantibody most relevant in rheumatoid arthritis) & non-organ specific autoantibodies
• Autoantibodies may not be involved in pathogenesis, but they are biomarkers
INSULIN-DEPENDENT DIABETES MELLITUS

• Possibly canine (genetic predisposition?); β-cells of pancreatic islets
attacked by immune system
• Caused by CTL & Abs directed against one of the islet cellʼs enzymes
(Type IV hypersensitivity rxn)
• Unknown what triggers initial attack.
MYASTHENIA GRAVIS
• Disease of skeletal muscles in dogs & cats
• Manifests as abnormal fatigue & weakness after exercise -
progressive
• Acquired or congenital (Jack Russell, Springer spaniel, Smooth Fox terrier)
• Acquired = immune-mediated disorder caused by circulating auto-antibodies against skeletal muscle ACh
receptors -> receptor damage by:
1. Direct damage to the neuromuscular junction
2. Formation of cross-linked antibodies leading to receptor internalisation
• Origin of autoantibodies causing MG not always known; link between thymic abnormalities & MG
• Myoid cells in the thymus express skeletal m proteins; also involved in development of self-tolerance
• Thymoma (tumor originating from thymus) & thymic follicular hyperplasia => loss of self-tolerance to ACh-
R
REPRODUCTIVE IMMUNOLOGY
• The prenatal period holds by far the greatest risk to human life!
KEY FACTORS THAT INFLUENCE SUCCESSFUL CONCEPTUS GROWTH DURING PRE-

IMPLANTATION & PERI-IMPLANTATION PERIODS
FACTORS REGULATED DURING THE EARLY

STAGES OF IMPLANTATION
MATERNAL-EMBRYONIC INTERACTIONS
DURING PERI-IMPLANTATION DEVELOPMENT
• Initial adhesion conducted by adhesion molecules
& ECM
• Cross-talk b/w conceptus & epithelium
• Conceptus comes down & touches the epithelium,
then starts rolling
• Apoptosis of the epithelium is induced by the
conceptus
• Integrins => adhesion of conceptus; adhere
straight to ECM
INTEGRIN TRAFFICKING DURING TROPHOBLAST DEVELOPMENT

• Pre-implantation, the trophoblast expresses various integrins
• Expression of certain integrins leads to adhesion
• Recycling of these integrins => trophoblast migration, elongation & subsequent adhesion occurring
INTERFERON-TAU: ANTILUTEOLYTIC EFFECTS OF THE CONCEPTUS

• Not primarily produced as an antiviral component of the conceptus
• The conceptus doesnʼt have interferon receptors at this stage
• Therefore IFN-τ has anti-viral effects on the uterine wall & endometrium, but not the conceptus
• This may be perceived as detrimental for the conceptus but in actual fact, IFNs are VERY growth inhibitory
so conceptus growth would be limited if they responded.
• There is a brief period (14-15d) during which IFN-tau is produced that conceptus doesnʼt express interferon
receptors. After this period, the conceptus starts expressing them.
OSTEOPONTIN (OPN) IN THE REPRODUCTIVE TRACT

• An ECM molecule that mediates cell-cell & cell-matrix adhesion & communication
• A cytokine that can exhibit (i) pro-inflammatory properties, (ii) anti-inflammatory actions (later) & (iii) effects
on tissue repair at sites of inflammation
• An intracellular protein associated with CD44 & ERM that can profoundly influence implantation & placental
development
ERM crosslinks actin filaments with plasma membranes. They co-localise with CD44 at actin filament-
plasma membrane interaction sites. Highly conserved.
CD44 is a receptor for hyaluronic acid & also interacts with osteopontin. Participates in a wide variety of
cellular functions including lymphocyte activation, recirculation & homing, hematopoiesis & tumor metastasis.
Endometrial cells in women with endometriosis show increased expression of splice variants of CD44.
ACTIONS OF SEMINAL FLUID IN THE FEMALE REPRO TRACT
HLA-DR ON SPERM & SOLUBLE MHCI

ANTIGENS MAY INDUCE MHCI OR
CLASS II-SPECIFIC TOLERANCE
• HLA-DR is a MHCII receptor which
constitutes a ligand for a T-cell receptor
• Typically found on APCs
• It induces MHCII-specific tolerance,
instead of rejection
• T-cells play a central role in induction &
maintenance of MHCII Ag-specific
tolerance
• Seminal fluid contains soluble MHCI
which induces MHCI-specific tolerance
• In this tolerance induction to MHCI, NK
cells play a central role
• Successful pregnancy requires a state of
maternal immune ʻtoleranceʼ to
accommodate antigens expressed by the conceptus. (Same goes for paternal side!)
ROLE OF GALECTIN-1 IN THE REGULATION OF IMMUNE CELL TOLERANCE

• Inhibits T-cell activation & differentiation
• Induces T-cell apoptosis
• Modulates cell-cell & cell-matrix interactions
• Regulates Th1/Th2 cytokine balance
FACTORS DETERMINING VIRAL TRANSMISSION TO THE FETUS

• Virus family, genus & strain
• Type of viremia: Cell-associated (Trojan horse effect makes them more successful in traversing placenta
e.g. in monocyte) vs. cell-free
• Placental structure
• Placental integrity: Some viruses wait for an ascending bacterial infection to occur & once the placenta is
damaged by e.g. cytokines, virus enters.
• Target cell(s): Receptor expression, intracellular support of virus replication
• Time in gestation
• Species characteristics of innate & acquired immunity (systemic & locally)
TOLERANCE & LIMITATION OF EXTRAVILLOUS TROPHOBLAST

• In order for conceptus to be accepted, it must stay w/n a certain zone - it canʼt move to the extra-villous
area of the placenta
• By various mechanisms, the EVT (ExtraVillous Trophoblast) undergoes apoptosis & those apoptotic bodies
are taken up by macrophages & dendritic cells
• Goal = protection against intra-uterine infections so investment in a lot of regulatory mechanisms is
important.
• Ideally, EVT is presented to Th1 cells w/o a co-stimulatory molecule. The T-cells are then anergised (no
response to the trophoblast).
• Alternative mechanisms:
• CTLA-4 may be activated by an APC, & another anergising signal sent to the T-cell.
• At the same time, the APC will start producing IFNy which activates other cells to produce IDO.
• IDO leads to tryptophan breakdown. Tryptophan is an absolutely essential molecule; without it
there is T-cell inactivation.
• Treg (T-suppressor) cells are also released.
IMMUNOREGULATORY MECHANISMS OF CD4+ CD25+ REGULATORY T-CELLS

• These Treg cells express CD4 (cluster of glycoproteins expressed on cell membrane surface that assists in
T-cell activation), CD25 (transmembrane protein that can form complex with high affinity for IL-2) & Foxp3
(master regulator in development & function of Treg cells).
INADEQUATE CYTOKINE PRODUCTION MAY CAUSE POOR ANGIOGENESIS OR REJECTION

RESULTING IN PRE-ECLAMPSIA
FUNCTIONS CARRIED OUT BY UTERINE EPITHELIAL

CELLS AS SENTINELS OF IMMUNE PROTECTION IN
THE FEMALE REPRODUCTIVE TRACT
• Ascending bacterial infections (not viral - no such thing
as ascending viral infection!) from the urinary tract are a
big problem
• Cytokine & chemokine production dampened initially but
once the motherʼs immune system becomes tolerant of the
implant, they become functional.
EXAMPLES OF IN UTERO VIRAL INFECTIONS

• Viruses doing it ʻcause theyʼre too school for cool: Pestiviruses, rubella virus, lentivirus
• Random (Stochastic) when mumʼs infected: BHV-1,
PHV-1, dengue virus, influenza viruses, HIV/FIV,
smallpox virus
• Ones that require a buddy: Enterovirus, morbillivirus.
INVOLVEMENT OF EXTRA-UTERINE INFLAMMATION

& IMMUNE ACTIVATION IN EMBRYONIC MORTALITY
• IFN-a: Increases body temperature, inhibits LH
secretion, reduces circulating progesterone.
• IFN-y: Toxic effect on CL.
• TNF-a: Toxic effect on CL, stimulates endometrial PG
synthesis, interferes with oocyte & blastocyst maturation
& with embryonic development.
• IL-1β: Increases body temperature, stimulates
endometrial PG synthesis, reduces endometrial cell
proliferation.
• NO: Interferes with oocyte maturation & embryonic
development
• PGF2α: Toxic effect on CL, interferes with oocyte
maturation & embryonic development.
MECHANISMS OF IMMUNOLOGIC TISSUE INJURY
Type Mediators Antigen Mechanisms Examples Pathologic

lesion
I IgE Allergens Immediate release of vasoactive Anaphylaxis Vascular dilation,
amines & other mediators from Atopy oedema, smooth
mast cells “armed” with IgE & muscle
cross-linked by allergen -> contraction,
recruitment of inflammatory cells mucus
production,
inflammation
II IgG, IgM Cell- or matrix- Cytotoxic Haemolytic Inflammation
associated Ags Abs bind to Ag on self -> anemia
Cell surface phagocytosis or lysis of target cell Pemphigus
receptors by activated complement or Fc
receptors. Recruitment of
leukocytes.
III IgG, IgM Soluble Ags Deposition of Ab-Ag complexes Glomerulonephrit Necrotising
(e.g. bacterial & -> complement activation -> is vasculitis
viral) recruitment of leukocytes (by (fibrinoid
complement products & Fc-Rs) -> necrosis);
release of enzymes & other toxic inflammation
molecules
IV T-cells, Soluble Ags Delayed TB, contact Perivascular
macrophages Contact Ags Activated T-cells: dermatitis cellular infiltrates;
Cell-associated (i) Release cytokines & activate Transplant oedema; cell
Ags (e.g. macrophages rejection destruction;
tumour Ags) (ii) CTL-activity granuloma
formation
TYPE I HYPERSENSITIVITY: IMMEDIATE HYPERSENSITIVITY

Primary exposure to allergen
• Allergen deposited on skin or mucosa
• Proteins diffuse in & captured by DCs
• DCs mature & migrate to LN -> present Ag
• Th2 lymphocyte response, IL-4 (helps naive T-cells differentiate into Th2 cells) leading to IgE (released by
B-cells), B-cell proliferation
• IgE secreted, bound by Fc fragment to mast cells, basophils, eosinophils
• Genetic constitution influences susceptibility
• Early reaction (minutes): Mast cell degranulation & preformed vasoactive substances -> vasodilation,
increased vascular permeability -> oedema of interstitial tissues
• Late phase (hours): Eosinophils, fewer lymphocytes & neutrophils
TYPE I HYPERSENSITIVITY: MAST CELL DEGRANULATION & ACTIVATION

• Mast cells are important not only as an instigator of the inflammatory response but also as immune
modulators.
• They are common & found in close proximity to blood vessels & nerves, where they can regulate vascular
permeability, effector-cell recruitment & neuro-immuno-endocrine circuitry.
• They spit out not only preformed substances, but also cytokines which attract leukocytes to the site.
• On top of that, they have receptors for a lot of things, not only Fc-R; TLRs also play a role in their
activation.
• Histamine: Vasodilation
• PAF (Platelet Activating Factor): Increased vascular permeability
• Prostaglandins: Smooth muscle cell spasm
• Leukotriene: Cellular infiltration
MECHANISMS BY WHICH MAST CELLS ENHANCE IMMUNE EFFECTOR-CELL RECRUITMENT

• Direct (TLR) or indirect (Ig/Fc-R) mechanisms => selective production of mediators for cell recruitment
• Mast cells keep pushing reaction towards a Th2 response
SYSTEMIC ANAPHYLAXIS
• Systemic delivery of an allergen
• Smooth muscle response in shock organs (organ that exhibits the most marked response to the Ag-Ab
interaction in hypersensitivity, as the lungs in allergic asthma or skin in allergic contact dermatitis)
• e.g. drugs such as penicillin, vaccines
BOVINE RESPIRATORY SYNCYTIAL VIRUS-ASSOCIATED PAROXYSTIC

RESPIRATORY DISTRESS SYNDROME
• Cow canʼt breathe so just lies there
• Genetic background predisposes disease
• Massive mast cell degranulation
• Due to unbalanced Th1/Th2 response to virus infection? In a virus infection, you
want a Th1 response to get IgG-2α produced which neutralises the virus. In some
animals, that switch hasnʼt been made & there is a Th2 response, with a
hypersensitivity reaction after vaccination or upon re-encounter.
• Or due to BRSV-activation of complement, or substance P release?
CANINE ATOPY (Allergic reaction that becomes apparent in minutes)

• Chronic pruritis caused by allergens (e.g. moulds, pollen, house dust mites)
• Common, with a genetic predisposition
• Dermal exposure
• Ingestion - food allergens (not proven)
• ? Abnormal suppressor T-cells, so there is a switch to a Th2 response
TYPE II HYPERSENSITIVITY: CYTOTOXIC HYPERSENSITIVITY

• Antibodies produced by the immune response bind to Ags on the patientʼs own cell surfaces
• Antigens can be exogenous agents that bind to cells of the body, Fc-Rs (there are many Abs against Fc-
Rs), molecules in rbc membranes (common) or nuclear Ags (common - DNA, histones, proteins associated
with DNA in the nucleus). In exogenous Ags, there is a hapten & the cell itself is called a carrier. In that
context, the immune system doesnʼt recognise the cell so attacks it.
• First phase: Immune complex formation

• Second phase: Ag-Ab complexes become lodged in the vessel wall & activate inflammatory cells
• Third phase: Elicitation of tissue damage & an inflammatory response
IMMUNOHAEMATOLOGICAL
Name Mechanism
Transfusion reaction Blood group incompatibility
Antigenically different RBCs are recognised as foreign by their cell

surface Ags -> immune response
• Delayed
• Subsequent transfusion = rapid destruction
Erythroblastosis foetalis Rh incompatibility
Autoimmune Anti-RBC
• Anemia, spherocytosis, positive direct antiglobulin reaction
• Agranulocytosis associated with certain drugs
• Thrombocytopenia - idiopathic; can be associated with viral antigens
or drugs
• Hemorrhages often seen on mucosal surfaces.
Haemolytic reactions to drugs Drug binds to red cell as hapten & covalently bond to membrane
proteins, leading to Ab-Ag interaction with the cell-bound drug. This
“drug-adsorption” mechanism is the basis for penicillin-induced
hemolytic anemia.
Drugs may also initiate a true autoimmune hemolytic anemia by
somehow causing antibodies to form directly against red cell antigens.
Neonatal isoerythrolysis Blood group incompatibility
• Fetal RBCs -> dam -> immune response
• Passive transfer of Abs in colostrum -> haemolysis
• Rare, occasionally seen in vaccination with blood-containing vaccines
e.g. Babesia
=> Hb deposition in all tissues & jaundice
HAPTEN
• Small molecule that elicits an immune response when attached to a large carrier such as a protein.
• The carrier may be one that doesnʼt elicit an immune response by itself.
• Once the body has generated Abs to a hapten-carrier adduct, the hapten will also be able to bind to the Ab,
but it will usually not initiate an immune response; usually only the hapten-carrier adduct can do this.
• Sometimes the hapten can block the immune response to the hapten-carrier by preventing the adduct from
binding to the Ab.
IMMUNE COMPLEX GLOMERULONEPHRITIS

• Glomerular deposition of circulating soluble immune complexes: Determined by molecular charge of
the complex & avidity of Ag/Ab binding
• Between endothelial cells & glomerular basement membranes (GMB) - subendothelial
• Within the GMB
• Between GMB & epithelium - subepithelial
• Within the mesangium (inner layer of glomerulus, with BM surrounding capillaries)
• Formation of immune complexes in the glomerulus (in situ)
• Self antigens
• Exogenous antigens planted in glomerulus e.g. drugs, infectious agent antigens
• Membranous glomerulonephritis: Diffuse thickening of capillary walls without increase in cell number;
electron-dense deposits along epithelial side of BM with effacement of foot processes
• Membranoproliferative glomerulitis & glomerular sclerosis: Mesangial proliferation so glomeruli look

very cellular & busy, with a thickened BM. The filtration apparatus is definitely affected.
SYSTEMIC IMMUNE COMPLEX DISEASE
TYPE IV HYPERSENSITIVITY: DELAYED HYPERSENSITIVITY

• Tuberculosis is the most classical case (Mycobacterium spp. Ags).
• Johneʼs disease: Mycobacterium avium spp. with paratuberculosis Ags.
• Granulomatous enteritis: Multinucleated giant cells donʼt form nice granulomas. Fluid absorption
that should take place, doesnʼt.
• Components of insecticides in flea collars, sprays & dips; pollens; house plants; viruses, etc. may cause it.
TUBERCULOSIS
• Central necrotic zone marked yellow because caseous
• Zone of epithelioid macrophages (giant cells)
• Outer zone a mixture of lymphocytes & monocytes coming in
• Depending on what direction the macrophages are skewed, the disease may
be resolved (Th1) or chronic (Th2)
• Secondary lymphatic (lung, draining lymph nodes) or haematogenous (lung,
liver, spleen) lesions may form in the primary site or secondary sites
• The only tissue where Mycobacterium doesnʼt like to live is the kidneys, for
some odd reason!
TUBERCULIN REACTION
• Test performed under the tail or on eyelids on animals being imported or
exported for TB
• PPD (Purified protein derivative) is extracted from mycobacteria & injected
intradermally - painful!!!
• 2h - neutrophil infiltrate around venules
• 12h - CD4+, CD8+, monocytes, basophils & cytokine release
• 24 - 72h - swelling
• This is the reaction youʼd get if the animal was infected or in the past had been vaccinated against TB.
ALLERGIC CONTACT DERMATITIS

• Ags are haptens - bind to cells or matrix in epidermis
• House plants, pollen, formadehyde, insecticides
• Affects skin: Langerhans cells (dendritic cells in the epidermis) ultimately activate T-cells
• Repeat or continuous exposure
• Alopecia, hyperpigmentation
PRIMARY PATHWAYS OF GRAFT REJECTION

• Direct: Mediated by APCs within the graft
• Indirect: Mediated by graft Ags presented by APCs of the recipient -> type IV hypersensitivity rxn
THE IMMUNOPATHOLOGY OF STRESS

Stress perception results in release of central stress-response coordinators, which activate:
• Structural peripheral stress-response elements e.g. skin, which is under constant exposure to foreign Ags
• Important to have equilibrium between the systems which control stress
• Psychological stress associated with many skin diseases e.g. pruritis
• Neuropeptides bind to recetors on mast cells, causing them to degranulate. Mast cell-derived vasoactive,
pro-inflammatory & neurosensitising mediators increase vascular permeability & contribute to pathogenesis
of inflammatory skin disorders.
• Activated mast cells skew immune response to Th2 bias & Ab production (humoral response)
• Some animals react with diarrhea or vomitting if they are really stressed - the shock organs are the greatest
affected.
MAST CELLS
• Mediate both innate & Th2-induced immune responses. IL3 & IL-4 result in mast cell proliferation, &
perpetuation of Th2-type responses is facilitated by the activation of mast cells.

Immune Responses To Infectious Agents: Immunology & Immunopathology

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Immunology & Immunopathology

IMMUNE RESPONSES TO INFECTIOUS AGENTS

ACUTE, INNATE RESPONSE TO INFECTION

PAMPs & TLRs

STRUCTURE OF BACTERIAL CELL WALLS

IMMUNE RESPONSE TO BACTERIA - OFTEN SUFFICIENT TO ELIMINATE

BACTERIAL OPSONISATION & PHAGOCYTOSIS

ANTIBODY DEFENSES AGAINST BACTERIAL INVASION

Produced by - Could limit bacterial metabolism

AVOIDANCE STRATEGIES (EVASION) BY EXTRACELLULAR BACTERIA

IMMUNE RESPONSES & EVASION STRATEGIES BY INTRACELLULAR BACTERIA

CAMs (APC1) AAMs (APC2)

ALTERNATIVELY ACTIVATED MACROPHAGES IN HELMINTHIC INFECTIONS

IMMUNE RESPONSES TO VIRAL INFECTIONS

EFFECTOR MECHANISMS INVOLVED IN CLEARING PRIMARY INFECTIONS & PROTECTION

THE INTERFERON RESPONSE: FIRST LINE OF DEFENSE

• Type I interferon (IFNα/β)

RECOGNITION OF RNA VIRUSES THROUGH INTERACTION OF

THE INTERFERON RESPONSE: FIRST LINE OF DEFENCE

VIRUS-MEDIATED BLOCKING OF INTERFERON SIGNALLING

VIRUS NEUTRALISATION BY ANTIBODY

WAYS TO KILL A VIRUS-INFECTED CELL

VIRUS ANTI-APOPTOTIC STRATEGIES

T-HELPER CELL ACTIVATION OF CTLS

ANTIBODY-DEPENDENT CYTOTOXICITY (ADCC)

• Mediated by NK cells; neutrophils & eosinophils can also mediate

VIRAL PEPTIDE PRESENTATION BY MHCI

VIRAL IMMUNO-EVASION: VIRAL INTERFERENCE WITH ANTIGEN PROCESSING & PRESENTATION

VIRAL IMMUNO-EVASION: MODULATION OF CYTOKINE RESPONSES

LYMPHOID TISSUE RXNS DURING LENTIVIRUS INFECTION

“DIRECT” IMMUNOPATHOLOGY (viral replication): HPAI INFLUENZA

IMMUNE RESPONSES TO PARASITES & FUNGI

IMMUNE EFFECTOR CELLS IN HELMINTH INFECTION

NEUTROPHILS IN PARASITE INFECTIONS

IMMUNE RESPONSES TO HELMINTHS

CD4+ T-CELLS IN HELMINTH INFECTION

EPITHELIAL CELL-DERIVED CYTOKINES IN PARASITE INFECTIONS

IMMUNE RESPONSES TO HELMINTHS - EXPULSION FROM THE GUT

IMMUNE RESPONSES TO ECHINOCOCCUS spp.

IMMUNE RESPONSES TO PROTOZOAL INFECTIONS

VACCINES AGAINST PARASITES

IMMUNE RESPONSES TO FUNGI

• Induces strong, long-lived, “sterilising” immunity

Mode of Action **NOT IMPORTANT?**

New Generation Adjuvants

VACCINE DELIVERY ROUTES: MUCOSAL VACCINES

VACCINE DELIVERY ROUTES: MUCOSAL VACCINE-DESIGN CHALLENGES

ADVERSE REACTIONS TO VACCINES

VACCINE FAILURE: DEVELOPMENT OF DISEASE DESPITE VACCINATION

REAGENTS USED IN SEROLOGICAL TESTS

PRODUCTION OF MONOCLONAL ANTIBODIES

ENZYME-LINKED IMMUNOSORBENT ASSAY (ELISA)

• Other proteins cleaned out & eventually => purified antibodies

COMPETITIVE INHIBITION ASSAY (right)

LUMINEX ASSAY (above)

RAPID IMMUNOASSAY DEVICES (left)

PRECIPITIN REACTION-BASED TESTS

COMPLEMENT FIXATION TEST

Quick Refresh: ELISA

ECL WESTERN BLOTTING

IMMUNOLABELLING OF CELLS & TISSUES: IMMUNOFLUORESCENCE MICROSCOPY

IMMUNOLABELLING OF CELLS & TISSUES: IMMUNOHISTOCHEMISTRY

Mode of Action NOT IMPORTANT?