Transcript fungal_immunity_mod

Lecture 2 Fungal immunity
By
Prof Dr. M. Refai
Local defence mechanisms
against mucosal infection
Salivary proteins
such as lactoferrin, beta-defensins,
histatins, lysozyme, transferrin,
lactoperoxidase, mucins, and
secretory immunoglobulin A.
These impair adhesion and growth
of fungi in the oropharyngeal cavity.
Healthy oral epithelial cells
These inhibit blastoconidia and/or
hyphal growth of fungi
if
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The fungus succeeds in bridging any of the
body surfaces either as a result of
 burns, trauma or maceration or
 surgical interference or
 application of catheter etc.,
The fungus will face
 non-specific defenses
 humoral defenses
 cellular host defenses.
In general, man and animals are
resistant to fungus infection
 The intact skin and mucous
membranes are not easily invaded
by fungi.
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None specific defence
 Secretory Ig A
+
Complement
Opsonization
 Neutrophils, Monocytes
 Alveolar macrophages
Phagocytosis
Phagocytosis
Immune cells
 Neutrophils
 Macrophages
 Dendritic cells,
 Natural killer cells
 T cells
 Epithelial and endothelial cells
Specific
pathogenassociated
molecular
patterns
(PAMPs)
patternrecognition
receptor
(PRR) immune
cells.
immune cells
Pattern recognition of fungi
Signaling pathways
are activated
to release
Transicription
factor
to
regulate
expression
of Inflammatory cytokines
Strategies employed by fungal pathogens
to evade host defense mechanisms
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1. Shielding of stimulatory PAMPs,
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2. Modulation of inflammatory signals,
3. Shedding of decoy components,
4. Persistence in intracellular environments,
5. Complement evasion.
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surface proinflammatory b-(1,3)-glucans or
non-stimulatory a-(1,3)-glucan cell wall
Shielding of stimulatory PAMPs
detection of the
invading
pathogen
C. albicans & dectin-1
dectin-1 recognizes the ß-glucans
at the level of budding scars in the
C. albicans, but
 it cannot recognize the ß-glucans
in the hyphae, where they are
shielded by a layer of mannans
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Histoplasma capsulatum
the α -(1,3)-glucan present in its
outer layer cell wall contributes to
pathogenesis by shielding its
immunostimulatory β -glucans.
Paracoccidioides brasiliensis
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Paracoccidioides brasiliensis transforms into
its pathogenic yeast form, a change in its cell
wall glucan polymer linkage occurs from β (1,3) to α -(1,3)-glucan.
Cryptococcus neoformans
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Cryptococcus neoformans masks its surface
PAMPs through the production of an
extracellular capsule of glucuronoxylomannan
(GXM)
Coccidioides immitis
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Coccidioides is recognized by the host by its
immunodominant spherule outer wall
glycoprotein (SOWgp).
During endospore differentiation, the fungus
secretes a metalloproteinase (Mep1) which
digests SOWgp
This enables the fungus to evade
phagocytosis and killing at its
vulnerable endospore stage of
development
Strategies employed by fungal pathogens
to evade host defense mechanisms
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1. Shielding of stimulatory PAMPs,
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2. Modulation of inflammatory signals,
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3. Shedding of decoy components,
4. Persistence in intracellular environments,
5. Complement evasion.
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Modulation of inflammatory signals
cytokine
production
humoral
response
Preferential activation of TLR2 over TLR4 by certain fungal
morphotypic PAMP stimulates a Th2 humoral response over a Th1dependent antifungal proinflammatory cytokine production.
C. albicans
C. albicans induce immunosuppression
through
TLR2-mediated IL-10 release,
 this leads to
generation of CD4+CD25+ T-regulatory
cells
with immunosuppressive potential
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Aspergillus fumigatus
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A. fumigatus evades immune recognition by
germination into hyphae with subsequent loss of
TLR-4 recognition,
The TLR2-mediated IL-10 pathways remain
intact, thus shifting the balance towards a
permissive Th2-type profile
Strategies employed by fungal pathogens
to evade host defense mechanisms

1. Shielding of stimulatory PAMPs,
2. Modulation of inflammatory signals,
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3. Shedding of decoy components,
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4. Persistence in intracellular environments,
5. Complement evasion.
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Shedding of decoy components
Shedding of surface protein gpA by the fungus as decoy
to competitively inhibit mannose receptor (MR).
Strategies employed by fungal pathogens
to evade host defense mechanisms
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1. Shielding of stimulatory PAMPs,
2. Modulation of inflammatory signals,
3. Shedding of decoy components,
4. Persistence in intracellular environments,
5. Complement evasion.
Persistence in intracellular environments
Upon internalization by immune or non-phagocytic cells,
Candida or Aspergillus can resist intracellular killing,
germinate and eventually escape
Aspergillus fumigatus
Alveolar macrophages
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The first defensive cells that inhaled conidia
The conidia are internalized by the macrophages and
prevented from growth for several hours until the
macrophage begins to destroy them.
At 24 h after internalization, 90% of the conidia
are killed.
The conidia then germinate to hyphae
The hyphae are too large to be engulfed
Aspergillus fumigatus
neutrophils
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Resting conidia are
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relatively resistant to killing by either reactive oxygen
intermediates or neutrophil cationic peptides and
their ingestion triggers neutrophil degranulation and
the respiratory burst only weakly.
Resting conidia
 activate the alternative pathway
 induce neutrophil chemotaxis
Aspergillus fumigatus
Polymorphonuclear neutrophils
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bind the surface without the need for
complement or immunoglobulin
This binding triggers secretion of reactive
oxidative intermediary agents that rapidly
damage the hyphae;
50% of hyphae are destroyed in 2 h
Histoplasma capsulatum
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Macrophages
represent the first line of defense during
infection with H. capsulatum,
they rapidly phagocytose the inhaled conidia
and transforming yeast cells,
the infected macrophage subsequently activate
effector T cells and enhance the release of
Th1-associated proinflammatory cytokines
(IL-12, IFN-γ ,and TNF-α(
Histoplasma capsulatum
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the main cytokines involved in Histoplasma
clearance from the host are IL-12, IFN-γ, and
TNF-α
IL-12 through its ability to regulate IFN-γ
production is critical in inducing a protective
immune response in primary infection with the
pathogen.
IFN-γ is pivotal for the host's innate resistance
to systemic infection with H. capsulatum.
Strategies employed by fungal pathogens
to evade host defense mechanisms
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1. Shielding of stimulatory PAMPs,
2. Modulation of inflammatory signals,
3. Shedding of decoy components,
4. Persistence in intracellular environments,
5. Complement evasion.
Complement evasion
Binding of complement-inhibitory C4 binding protein
(C4BP)and Factor H on fungal surface.
Immune response to fungi:
HI
The main recognized functions of
antibodies in fungal infections include
 prevention of adherence,
 toxin neutralization,
 antibody opsonization and
 antibody-dependent cellular
cytotoxicity
CMI
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The type of CMI induced is critical in determining
resistance or susceptibility to fungal infection.
In general,
Th1-type CMI is required for
clearance of a fungal infection,
 Th2 immunity usually results in
susceptibility to infection or allergic
responses
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CMI
Th1 cells produce predominantly
cytokines such as IFN-g, and promote
cell-mediated immunity and
phagocyte activation.
 Th2 cells produce predominantly
cytokines such as interleukins 3 and 4
(IL-3 and IL-4) and tend to promote
antibody production
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Cell mediated immunity
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Resistance to several mycoses is associated
however with cell mediated immunity,
particularly phagocytosis and killing of invading
fungi by
peripheral blood monocytes,
 alveolar and peritoneal macrophages
 polymorphonuclear leukocytes.
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i. Alveolar macrophage defense
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The first phagocytic cells to encounter the
inhaled or aspirated fungal particles
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have the capacity to kill Aspergillus conidia
by both oxidative and non-oxidative
mechanisms.
have relatively weak oxidative candidicidal
activity
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fungi characterized by hyphal forms, are too
large to be ingested completely by phagocytic
cells.
the alveolar macrophages
Histoplasma capsulatum:
 Macrophages are the primary host cells
 The yeast form survive within
macrophages and multiplies
intracellularly.
 Thus the macrophage, which usually
eliminates organisms from the primary
site of infection, becomes a site of
continued fungal growth and aids in
dissemination of the yeast to other tissues.
ii. Peritoneal macrophages
can phagocytize, inhibit intracellular
growth or inhibit germination of
conidia of Aspergillus or Candida.
 Activated macrophages appear to play
a role in resistance against
disseminated candidosis.
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iii. Normal peripheral blood monocytes
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Monocytes can damage and apparently kill
Aspergillus hyphae, even though hyphae
are too large to be ingested completely.
Monocytes and macrophages have been
shown to be important cells in host defense
against dimorphic fungi. When activated
they have increased phagocytosis and
decrease growth of intracellular yeasts.
iv. Polymorphnuclear leukocytes (PMN)
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PMN play a major role in host defense
against fungi.
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PMN can kill Candida blastoconidia,
Coccidioides immitis arthroconidia and
Blastomyces conidia.
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can also damage the hyphal forms of
Aspergillus and Candida.
The important role of the PMN in the
host defense against fungus infection
becomes obvious after the fungus escapes
control by macrophage and monocyte
cell lines and start mycelial growth,
where an impressive ability of PMN to
damage these organism is evident
Application of immune response
Vaccination
 Serodiagnosis
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Important points to consider in
antifungal immunity
and its relevance to vaccination
usually fungi display only moderate
virulence
 antifungal immune responses are
usually redundant.
 almost all pathogenic fungi have
mechanisms to evade or intoxicate
immune responses
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Most fungal diseases pose obstacles to
the concept and practice of vaccination
coccidiomycosis, histoplasmosis,
blastomycosis, and paracoccidioidomycosis
 are geographically limited, and
 low-incidence deep-seated diseases
 widespread illnesses e.g. aspergillosis,
cryptococcosis, and candidiasis
 typically occur in the immunocompromised
 theoretically—ineligible for active
immunisation
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Antibodies used for passive
immunity
Fungal vaccines
Fungal vaccines
Dermatphyte vaccines
The inactivated vaccines stimulate the
CMI, as demonstrated by skin tests
and leukocyte migration
inhibition tests.
 Vaccines containing T. verrucosum
conidia
inactivated with formalin
have been described for use in cattle
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inactivated vaccine plus adjuvant
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An inactivated vaccine plus adjuvant
containing conidia and mycelium of
two T. equinum strains has been used
in the immunization of horses
The vaccine does not prevent the
disease, but the lesions are less severe
in vaccinated animals in compared to
non-vaccinated animals.
Insol Dermatophyton
inactivated vaccine
effective in horse, dog and cat,
 can be used as treatment of the disease,
improving the clinical outcome.
 It contains strains of T. verrucosum, T.
mentagrophytes, T. sarkisovii, T.
equinum,M. canis,M. canis var. distortum,
M. canis var. obesum, and M. gypseum.
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The inactivated vaccine Dermatovac-IV
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It contains an adjuvant and an
optically standardized inactivated
suspension of conidia and
mycelium of the fungi M. canis, T.
equimun, M. gypseum and T.
mentagrophytes
The Ringvac bovis LTF1301 live vaccine
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The most effective and widely used
LTF-130 strain of T. verrucosum,
 has a characteristic high level of
immunogenicity,
 low virulence and great stability
 has been used effectively in Russia and
Norway
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Vaccine-induced protection against 3
systemic mycoses(Coccidioides posadasii ,
Histoplasma capsulatum , and Blastomyces
dermatitidi endemic to North America
Wüthrich et al. J Clin Invest
568–554:)2(121;2011
Serological diagnosis
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Serological tests are important in the
diagnosis of systemic mycoses
The standard methods of serology are
commonly used, i.e. agglutination,
precipitation, Complement fixation,
immunoflourescence and ELISA.
Serological diagnosis
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the results often permit differentiation
between colonization by fungi and a
true mycotic infection.
In all serological methods only the titre
dynamics are meaningful
A single serological result is of no
diagnostic value.
Serological diagnosis
The choice of the test
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Candida infection
Antibody developed against the polysaccharide component
of surface glycoprotein
Antigen: extracted polysaccharides from C. albicans
serotype A
Antibody type: -IgM in early infection.
the indirect haemagglutination test
-IgG in late infection
indirect immunoflourescence.
Antibodies developed against protein antigens of the cell
content are detected by precipiation tests.
Serological diagnosis
The choice of the test
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Candida infection
Because of the ubiquitous nature of Candida species ,
these serologic tests are limited in discriminating
between normal and disease levels of antibodies.
More specific tests are used for detection of
circulating C. albicans surface antigens and
cytoplasmic proteins.
Serological tests commonly used are latex
agglutination, immunoflourescence
Serological diagnosis
The choice of the test
Cryptococcus infections
 Antibodies to Cr. neoformans are usually not
detected at early infection, but later or on recovery
 Antibodies can be detected using. indirect
immunoflourescent technique, haemagglutination
inhibition test or ELISA.
If an infection of the CNS is suspected, cerebrospinal
fluid must be submitted for serological testing.
Serological diagnosis
The choice of the test
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Cryptococcus infections
During infection, capsular material is solubilized
in the body, and being an antigen, it can be titrated
with a specific rabbit anti-Cr. neoformans antiserum
using latex agglutination test.
A positive agglutination is diagnostic for
cryptococcosis.
This test is very useful in the early
infection.
Serological diagnosis
The choice of the test
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Aspergillosis
Complement fixation test with conidia
suspension,
Indirect immunoflourescence with intact fungal
elements,
Agglutination tests with extracted
polysaccharides on latex particles or erythrocytes.
Precipitation test (Double immunodiffusion and
immunoelectrophoresis )
Serological diagnosis
The choice of the test
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Aspergillosis
Detection of Aspergillus antigenaemia in
patients with invasive aspergillosis is more
sensitive and specific.
ELISA using monoclonal antibodies is
increasingly used in diagnosis nowadays.
Serological diagnosis
The choice of the test
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Blastomycosis
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Serological tests are not useful in the diagnosis or
prognosis.
Cross-reactions are common with other diseases like
histoplasmosis and coccidioidomycosis.
Paracoccidioidomycosis
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Immunoduffusion test is extremely useful.
The CF test is quantitative and useful in assessing
prognosis, but cross reactions occur
Serological diagnosis
The choice of the test
Histoplasmosis
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1. Latex agglutination, 2. Complement Fixation, 3.
Immunodiffusion, 4. ELISA
Cross-reactions with blastomycosis and other fungus
infections are seen.
Serology for histoplasmosis is a little more complicated
than for other mycoses, but it provides more information
than blastomycosis
Coccidioidomycosis
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Complement fixation, immunodiffusion and latex
agglutination tests
Serological diagnosis
The choice of the test
Skin tests
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Histoplasmin.
Histfarcin
Coccidioidin
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intradermal injection of 0.1 ml
evoke in infected individual a positive delayed
hypersensitivity reaction larger than 5 mm in
diameter after 24-96 hours
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