Cell-mediated immunity for medical, biochemistry and biology undergraduates

1. Immunochemistry
Cell-mediated Immunity
R. C. Gupta
Professor and Head
Dept. of Biochemistry
National Institute of Medical Sciences
Jaipur, India

2. Cell-mediated immunity operates against
those pathogens/antigens that have
entered some self cell
Cell-mediated immunity is the function of
T lymphocytes

3. T lymphocytes (T cells)
are of four types:
Cytotoxic (Killer) T cells
Helper T cells
Suppressor T cells
Memory T cells

4. T cells possess receptors for antigens
T cell receptor (TCR) is in many ways
similar to antibodies
It can recognize a specific antigen

5. Each chain (a and b) of TCR has:
Two extra-cellular domains
A trans-membrane region
A short cytoplasmic tail
TCR is a protein made up of an a chain
and a b chain joined by a disulphide bond

6. T cell
chain
b chain
T cell
T cell receptor (TCR)
a
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7. The a-chain has got two extra-cellular
domains – a1 and a2
The b-chain has got two extra-cellular
domains – b1 and b2
The a1 and b1 domains are on the
periphery

8. The a1 and b1 domains are similar to the
variable regions of antibodies
These are also known as Va and Vb
domains
The a2 and b2 domains are similar to the
constant regions of antibodies

9. Like antibodies, TCRs can recognize and
bind antigens
Antigen-recognition is the function of
variable regions
The antigen-binding site (or cleft) is
formed by the a1 and b1 domains

10. T cell receptor

11. T cell receptors have a diversity similar to
antibody diversity
TCR diversity also arises from gene re-
arrangement
TCR diversity

12. One of the 70 V segments
Thousands of different combinations
are possible
The gene for the variable region of a
chain (Va) is constructed from:
One of the 61 J segments
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13. One of the 52 V segments
One of the 2 D segments
One of the 13 J segments
Thousands of different combinations
are possible
The gene for the variable region of b
chain (Vb) is constructed from:
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14. When an a chain combines with a b chain,
millions of combinations are possible
A given T cell has one combination, and is
specific for one particular antigen
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15. MHC proteins and MHC genes
The T cell receptor cannot bind a free
antigen
The antigen must be attached to a cell, and
it should be a self-cell
If a T cell is transferred from one person to
another, it will not recognize the antigen
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16. The T cell receptor recognizes a foreign
antigen combined with a self-molecule
The self-molecules are Major Histo-
compatibility Complex (MHC) proteins
MHC proteins are present on all cells and
are unique to each individual
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17. MHC genes
MHC proteins are encoded by MHC
genes
MHC genes can be divided into:
MHC class
I genes
MHC class
II genes
MHC class
III genes

18. MHC class I genes encode MHC I proteins
MHC I proteins are present on the surface of
all the cells
Foreign antigens combined with MHC I
proteins are recognized by cytotoxic T cells

19. MHC class II genes encode MHC II
proteins
MHC II proteins are present on
macrophages, B cells and follicular
dendritic cells
Foreign antigens combined with MHC II
proteins are recognized by helper T
cells
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20. EMB-RCG
MHC class III genes
encode:
Complement
proteins
Some other
plasma proteins

21. MHC proteins
MHC proteins are trans-membrane proteins
They are synthesized on the endoplasmic
reticulum
They get inserted in the cell membrane
They have a peptide (antigen) binding cleft
on their extracellular surface

22. While on their way to the cell membrane,
MHC proteins pick up peptide fragments
formed by degradation of:
The fragments are bound firmly and
displayed on the surface of the cell
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Endogenous
proteins
Exogenous
proteins

23. Endogenous peptides bound to MHC
proteins are ignored by the immune
system
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If a T cell finds a foreign peptide bound
to an MHC protein, cell-mediated immunity
comes into operation

24. MHC class I proteins are made up of a
large a subunit and a smaller b subunit
(b2-microglobulin)
The a subunit has three extra-cellular
domains (a1, a2 and a3), a trans-
membrane region and a cytoplasmic tail
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25. The a1 and a2 domains form the
peptide binding cleft

26. MHC I protein – View from above
‒‒‒ Peptide

27. MHC I proteins bind fragments of proteins
degraded by cytosolic pathway
In cytosolic pathway, the proteins are
degraded by 26S proteasome

28. MHC class II proteins are made up of
almost equal sized a subunit and b
subunit
Each subunit has two extracellular
domains, a trans-membrane region and
a cytoplasmic tail
The a1 and b1 domains form the peptide
binding cleft
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30. ‒ Peptide
MHC II protein – View from above

31. MHC class II proteins bind fragments of
proteins degraded by lysosomal pathway
These proteins first enter intracellular
vesicles called endosomes
The endosomes fuse with lysosomes to
form endolysosomes

32. The proteins present in endolysosomes
are degraded by lysosomal enzymes
The lysosomal proteolytic enzymes are
cathepsins

33. The proteins present in
endosomes include:
Proteins of extracellular pathogens
Proteins of pathogens that reside
in endosomes
Proteins taken up by the cells
by endocytosis

34. MHC Class II
Protein
Antigen
fragment
a2 b2
b1
Cell membrane
Newly synthesized MHC II proteins bind
peptide fragments, and go to the cell
membrane
MHC II proteins get inserted in the cell
membrane, displaying the bound peptide
on the surface of the cell

35. Function of cytotoxic T cells
Cytotoxic T cells destroy infected self-cells
along with the pathogen present inside
them
This prevents the spread of infection to
healthy cells
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36. Most viruses and many bacteria reside and
replicate in cytosol of the infected cells
Their proteins are degraded by 26S
proteasome
Peptide fragment are displayed on the
surface of the cell by MHC class I proteins
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37. The foreign peptide bound to a self
MHC I protein is recognized by a
particular cytotoxic T cell
The T cell receptor binds to the
peptide:MHC I complex
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39. A trans-membrane protein CD8, present in
cytotoxic T cells, acts as a co-receptor
CD8 is made up of an a and a b chain
linked to each other by a disulphide bond
The extracellular portion of CD8 binds to
the a3 domain of MHC class I protein
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40. The cytoplasmic tail of CD8 is associated
with Lck, a cytosolic tyrosine kinase
The cytoplasmic tail of T cell receptor is
associated with a trans-membrane protein,
CD3 complex

41. CD3 complex consists of a g chain, a d
chain, two e chains and two x chains
Cytoplasmic portions of these chains have
immunoreceptor tyrosine-based activation
motifs (ITAMs)

42. On binding of T cell receptor and CD8 to
MHC I protein and antigen fragment, Lck
becomes active
Active Lck phosphorylates the ITAMs of CD3
complex
Phosphorylated ITAMs act as a docking site
for ZAP-70 (zeta associated protein of 70 kD)
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43. ZAP-70 phosphorylates the tyrosine
residues of some target proteins in the
cytotoxic T cell
This results in release of stored granules
from the cytotoxic T cell targeted at the
infected cell
The granules contain perforin, granzyme
and granulysin
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44. Action of cytotoxic T cell

45. EMB-RCG
Several perforin molecules get inserted in
the cell membrane of the infected cell
These are polymerized to form trans-
membrane pores
Granzyme and granulysin can enter the cell
through the pores

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Cell contents can leak out of the pores
Granzyme is a serine protease
It hydrolyse the proteins of the infected cell
and the pathogen
Granulysin activates apoptosis of the
infected cell

47. EMB-RCG
Thus, combined action of perforin,
granzyme and granulysin results in:
Destruction of the infected cell
Destruction of the pathogen
present in the infected cell

48. Function of helper T cells
Helper T cells share some common
features with cytotoxic T cells
But they also differ in some respects

49. Helper T cells and cytotoxic T cells
share the following common features:
• Their TCRs are made up of a and b
chains
• Cytoplasmic tail of TCRs is associated
with CD3 complex having ITAMs
• They have co-receptors associated with
Lck

50. Helper T cells and cytotoxic T cells
differ in the following respects:
• The co-receptor in helper T cells is
CD4 instead of CD8
• Their TCR recognizes a foreign
peptide bound to MHC class II
protein

51. CD4:
Is made up of a single
polypeptide chain
Has four domains – D1,
D2, D3 and D4

52. D3
D4
D1
D2
Four domains in
CD4 protein

53. Antigen-presenting cells
MCH II proteins are present on
macrophages, follicular dendritic cells
and B lymphocytes
These cells are known as antigen-
presenting cells (APCs)
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54. Antigen-presenting cells
Dendritic cell
Macrophage
B lymphocyte

55. EMB-RCG
TCR of a helper T cell binds to an antigen
fragment displayed by MHC II protein of APC
The D1 domain of CD4 binds to b1 domain of
MHC II protein
After this binding, intracellular Fyn is activated
by CD45

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Fyn phosphorylates ITAMs of CD3
complex
Phosphorylated ITAMs are the docking site
for ZAP-70
ZAP-70 binds to phosphorylated ITAMs and
is phosphorylated by Lck

57. ZAP-70 phosphorylates the tyrosine
residues of some target proteins
This results in the release of cytokines
(lymphokines) from the helper T cell
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58. Action of helper T cell

59. Lymphokines include:
• Interleukins (ILs) e.g. IL-2, IL-3, Il-4,
IL-5, IL-6 etc
• Granulocyte colony stimulating
factor (G-CSF)
• Granulocyte-macrophage colony
stimulating factor (GM-CSF)
• Tumour necrosis factor (TNF)
• Interferon-g (IFNg)

60. Actions of lymphokines
IL-2 stimulates multiplication of helper T
cells themselves
IL-3 stimulates the proliferation of bone
marrow stem cells
IL-4 stimulates the proliferation of B cells, T
cells and mast cells
Continued

61. IL-5 stimulates the growth of eosinophils
IL-10 stimulates growth of mast cells and
synthesis of MHC II proteins in B cells
GM-CSF stimulates growth of granulo-
cytes and macrophages
Continued

62. IFN-g activates macrophages and natural
killer (NK) cells
TNF activates macrophages
Some lymphokines attract the phagocytic
cells to the site of infection
Continued

63. Thus, release of lymphokines leads to:
• Increase in the number of helper T cells
• Increase in the number of cytotoxic T
cells
• Increase in the number of B cells and
plasma cells
• Increase in the number of antigen-
presenting cells
• Increase in the number of phagocytic
cells
• Activation of phagocytic cells

64. Thus, both innate and adaptive immunity
are geared up to fight against the pathogen
A co-ordinated and effective immune
response is mounted against the pathogen
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65. Helper T cells respond to calls for help
The call is given by antigen-presenting
cells
The call is in the form of an antigen
fragment displayed on MHC II protein
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66. Helper T cells have no direct role in
destroying the antigen
They activate different arms of the defense
system and co-ordinate their actions
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67. Helper T cells are described as the
Commanding Officer of the defense force
If helper T cells are depleted or debilitated,
the entire immune system is crippled
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68. Function of suppressor T cells
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Also known as regulatory T cells
Shut down the immune response after the
invading organisms are destroyed
Play a regulatory role in immune response
Possess TCR and CD8

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Suppressor T cells release some
lymphokines after an immune response
has achieved its goal
Suppressor T cells also prevent immune
response against self-molecules
This signals all other immune system
participants to cease their attack

70. Functions of memory T cells
Retain memory of antigen after first exposure
Spread throughout the lymphoid tissue
Are released quickly into circulation if the
same antigen enters again
Help in mounting a quick immune response
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71. Integrated immune response
Different arms of defense system do not act
in isolation
Innate immunity acts in co-ordination with
adaptive immunity
Humoral immunity and cell-mediated
immunity also act in co-ordination
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72. Many molecules having antigenic
properties are present in our body also
Our immune system doesn’t act against
these self-antigens
Tolerance to self-antigens is known as
self-tolerance
Self-tolerence
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73. Suppressor T cells have some role in self-
tolerance
They suppress immune reaction against
self-antigens
But there is a second and more important
mechanism
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74. Newly-formed lymphocytes are processed
during foetal life
The purpose of this processing is two-fold:
To allow only the immunologically
competent lymphocytes to mature
To eliminate those lymphocytes
that can react with self-antigens

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Developing T cells express the genes
for TCR, CD4 and CD8
Some of the TCRs are incapable of
recognizing any self-MHC protein
Such T cells would be functionally
useless, if allowed to mature
Processing of T cells

76. Developing T cells are first subjected to
positive selection
If the TCR and CD8 bind to some
MHC I protein, the cell is allowed to
mature into a cytotoxic T cell
Expression of CD4 gene is suppressed
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Positive selection

77. If the TCR and CD4 bind to some
MHC II protein, the cell is allowed to
mature into a helper T cell
Expression of CD8 gene is suppressed
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78. The positively selected cells are subjected
to negative selection
If the TCR binds some self-antigen:self-MHC
complex, the T cell is pushed into apoptosis
Thus T cells capable of acting against self-
antigens are eliminated
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Negative selection

79. Receptor editing
Clonal deletion
Anergy
Immunological ignorance
Developing B cells recognizing self-
antigens can have the following fates:
Processing of B cells

80. Receptor editing
Developing B cells that can react with
multivalent self-antigens undergo further
light chain gene re-arrangement
Re-arrangement continues until the
antigen receptor is so modified that it
cannot bind the self-antigen
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81. Multivalent antigen
Antigen

82. Clonal deletion
If self-reactivity is not lost by
receptor editing, the developing
B cell is pushed into apoptosis
Thus, the entire clone of self-reacting
B cells is deleted
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83. Anergy
Developing B cells that encounter
self-antigens of low valency
become anergic
Their antigen receptors remain
within the cell and signal
transduction is impaired
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84. Antigen of low valency
Antigen
Epitope Epitope

85. Immunological ignorance
Some sites in our body are
immunologically privileged
Immune cells and molecules
cannot reach these sites
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86. Immunologically privileged sites
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Brain Eye
Testes Foetus

87. Self-antigens present in these areas do not
come in contact with immune system
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The immune system remains ignorant of
such self-antigens

88. This results in self-tolerance
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Thus, potentially self-reacting
T cells and B cells are:
Deleted or
Modified or
Incapacitated

89. Autoimmune diseases
Self-tolerance is sometimes lost,
resulting in self-reactivity
When the immune system reacts
against a self-antigen, it results in
an autoimmune disease
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90. • Some self-molecules
have a structural
resemblance with
foreign antigens
• These may be
treated as foreign
antigens by the
immune system
Autoimmune diseases can occur because:
• Some self-molecules
combine with
microbial proteins to
form new molecules
• These molecules are
treated as foreign
antigens by the
immune system

91. Some diseases resulting from auto-
immunity are:
• Hashimoto’s thyroiditis
• Myasthenia gravis
• Systemic lupus erythematosus
• Rheumatoid arthritis
• Some cases of type 1 diabetes
mellitus, male infertility and pernicious
anaemia

92. Occurs due to impairment of immune
system
May be inherited (primary) or acquired
(secondary)
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Immunodeficiency

93. Both
Cell-mediated immunity or
Humoral immunity or
Inherited immunodeficiency
may affect:

94. Examples of immunodeficiency diseases
Type of immunity Example
Humoral Agammaglobulinaemia
Cell-mediated DiGeorge syndrome
Both humoral
and cell-
mediated
Severe combined
immuno-deficiency
disease

95. Congenital agammaglobulinaemia is a
disease in which only humoral immunity
is impaired
It is an x-linked disease in which gamma-
globulins are totally absent

96. In DiGeorge syndrome, only cell-mediated
immunity is impaired
DiGeorge syndrome occurs due to lack of
development of thymus

97. Both humoral and cell-mediated immunity
are impaired in severe combined immuno-
deficiency disease (SCID)
SCID is most commonly caused by a
deficiency of adenosine deaminase

98. Acquired immunodeficiency
may result from:
Severe malnutrition
Irradiation
Immunosuppressive drugs
Viral infections
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Acquired immunodeficiency

99. A very sinister form of acquired immuno-
deficiency is caused by human immuno-
deficiency virus (HIV)
The disease caused by HIV is known as
acquired immuno deficiency syndrome
(AIDS)
HIV and AIDS

100. HIV is a retrovirus
Retroviruses have an RNA genome
They also possess reverse transcriptase

101. After infection by a retrovirus:
Its genomic RNA enters the
infected cell
Reverse transcriptase also
enters the infected cell

102. Reverse transcriptase prepares a DNA
copy of the viral genome
This copy is incorporated in the DNA of
the infected cell
Viral genome becomes a part of the host
cell genome

103. The structure is similar to that of other
retroviruses
Its core is made up of two copies of its
RNA genome and some enzymes
The genomic RNA and the enzymes are
surrounded by a conical capsid
Structure of HIV

104. The capsid is made up of core proteins
(p 24, p 7 and p 6)
The capsid is surrounded by a spherical
envelope
The envelope is made up of lipid bilayer
and some proteins

105. The envelope is formed when the virus
buds out of an infected cell
The virus takes a part of the host-cell
membrane with it

106. The envelope contains two viral glyco-
proteins, gp 120 and gp 41
The glycoprotein gp 41 is embedded in
the lipid bilayer
The other glycoprotein, gp 120 is attached
to gp 41 by non-covalent interactions

107. gp120 gp41
p6
Integrase
Protease
Reverse
transcriptase
p24
p17
p7
Genomic
RNA
Lipid bilayer

108. The HIV genome is made up of 9,749
nucleotides
Three genes, env, gag and pol, are
common to all retroviruses
There are six accessory genes unique to
HIV
Genome of HIV

109. The accessory genes in HIV are: (i) tat, (ii)
rev, (iii) nef, (iv) vif, (v) vpu and (vi) vpr
Fifteen proteins are synthesized from
these nine genes

110. The name of env gene is derived from
envelope
It encodes a poly-protein, gp160
After translation, is gp160 cleaved by
protease
The products of cleavage are gp 120
and gp 41

111. gag gene is named after group-specific
antigen
It encodes the core proteins of the virus
It is translated into a poly-protein

112. The poly-protein encoded by gag gene is
cleaved into – p 24, p 17, p 7 and p 6
p 17 forms the inner lining of lipid
bilayer
The other proteins are present in the
capsid

113. The name of pol gene is derived from
polymerase
It encodes the viral enzymes
It is translated into a poly-protein

114. The polyprotein encoded by pol
gene is cleaved into:
Reverse transcriptase
Integrase
Protease
Reverse transcriptase possesses
ribonuclease H activity also

115. HIV infection and immunodeficiency
HIV is present in infected persons in:
Some blood cells
Genital secretions
An infected person can transmit the virus
to other persons

116. HIV can be transmitted from an infected
person to an uninfected person through:
Sexual contact (vaginal or anal)
Transfusion of infected blood
Sharing of contaminated needles
Mother to foetus transmission in
pregnancy

117. The molecule exposed on the surface of
HIV is gp 120
This happens to have a structure comple-
mentary to that of CD4
Due to the complementarity, gp 120 binds
avidly to CD4
CD4 molecules are present on the surface
of helper T cells

118. HIV and helper T cell
gp 120 of HIV binds avidly to CD4 of helper T cells

119. Viral membrane fuses with the membrane
of helper T cell
The virus sheds its envelope and its
core enters the helper T cell
Viral reverse transcriptase synthesizes a
complementary DNA strand
A cDNA-RNA hybrid is formed

120. The ribonuclease H activity of the enzyme
hydrolyses the genomic RNA
The single DNA strand then acts as a
template
A second complementary DNA strand is
synthesized

121. Thus, a double-stranded DNA is formed
This is a DNA copy of the viral genome
This is incorporated in the host cell DNA
by the viral integrase
Thus, the viral genome becomes a
permanent part of the host DNA

122. The viral genome incorporated in host cell
genome is called proviral DNA
The proviral DNA is transcribed to mRNA
by the host cell
The mRNA is translated to form viral poly-
proteins
These are cleaved by protease

123. The mRNA and the viral enzymes are
surrounded by the core proteins
The core is surrounded by p 17 and
envelope proteins to form a new virus
The virus buds out of the cell
Thus, a new virus is released by the
infected cell

125. The new virus infects another uninfected
helper T cell
This cycle is repeated
New virus particles are formed and
released
More and more helper T cells are infected

126. Multiplication of HIV inside helper T cells
leads to rupture of cells
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127. Viral proteins are cleaved inside the
infected helper T cells
MHC I proteins of the infected cells can
display viral peptides on the cell
Killer T cells can destroy helper T cells
displaying viral peptides
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128. Rupture and destruction of helper T cells
decreases their number
Massive depletion of helper T cells cripples
the immune system
The affected person becomes extremely
prone to infections due to immunodeficiency