Docstoc

HIV and AIDS - PowerPoint

Document Sample
HIV and AIDS - PowerPoint Powered By Docstoc
					          HIV and AIDS

Acquired Immunodeficiency Syndrome
• Disease caused by an infectious agent:
              a retrovirus




                                           1
                          HIV and AIDS
                           an infectious agent
             In Los Angeles 1967-1978 only two cases of
             Pneumocystis carinii pneumonia

• 1979 - 5 cases of Pneumocystis carinii         All Homosexual


                                                     With giemsa stain at high
                                                      magnification, the faint
                                                           bluish dot-like
                                                       intracystic bodies of
                                                     Pneumocystis carinii in
                                                       lung are seen in this
                                                       cytologic preparation
                                                     from a bronchoalveolar
                                                               lavage

                                                                        2
                          HIV and AIDS




With dissemination to extrapulmonary sites, Pneumocystis carinii tends to
produce foci with prominent calcification, as seen in the kidney here
grossly.                                                              3
                           HIV and AIDS
                 an infectious agent – Kaposi’s Sarcoma



Early 1981 MMWR: 5 cases of
        Kaposi’s sarcoma
              Hitherto: rare


• 1981 - 26 cases of Kaposi’s sarcoma
       • Young
       • San Francisco and New York
       • All Homosexuals
                                                          4
             HIV and AIDS
Two rare diseases in the gay community linked to

          IMMUNOSUPPRESSION


      OPPORTUNISTIC INFECTIONS
                     Also
               Lymphadenopathy
             Hodgkin’s Lymphoma
 • Gay-Related Immune Deficiency
 • Acquired Immune Deficiency Syndrome (AIDS)
                                                   5
HIV and AIDS




   Candida infections are common with
   AIDS, but most often appear as oral
   thrush, which is a nuisance but not life-
   threatening. Disseminated infections
   are uncommon, but here is a rare
   Candida pneumonia, which resembles a
   bacterial bronchopneumonia.         6
      HIV and AIDS
      Little in common but:
            • Young
             • White
             • Male
         • Large towns
   • Homosexual community

But not all gay men got the disease
                                      7
                   HIV and AIDS
              Distinguishing characteristics

            • Clusters of infected men
• Apparent concentration within sexually interactive
                      groups
          • High numbers of sex partners

         Suggests an infectious agent

                                                 8
                  HIV and AIDS
      More evidence for an infectious agent:
      • Different ways of getting a similar syndrome
                   • Blood transfusions
               • Intravenous drug use
             • Hemophilia (clotting factor)

Female sex partners of AIDS-positive IV drug users and
                    hemophiliacs

               Not just in the Gay community           9
           HIV and AIDS

     Obvious agent:
     A virus……that is now in
     the blood supply
     Primary route of
     transmission: Sex


AIDS is a sexually-transmitted disease
                                         10
                  HIV and AIDS
               The Cellular Picture
Loss of one cell type throughout the course of the disease
                  CD4+ T4 helper cells
     A fall in the CD4+ cells always precedes disease


    In advanced disease the loss of another cell type
                CD8+ cytotoxic killer cells

          Suggests an infectious agent
                        A virus                         11
                     AIDS Definition
• AIDS is currently defined in persons older than 13 years as
the presence of one of 25 conditions indicative of severe
immunosuppression         or
• HIV infection in an individual with a CD4+ cell count of <200
cells per cubic mm of blood.

• AIDS is therefore the end point of an infection that is
continuous, progressive and pathogenic

• With the prevalence of HIV in the developing world, HIV and
its complications will be with us for generations
                                                            12
                  AIDS Statistics
Approximately 8500 new HIV infections occur daily around the
world
Over 90% of these are in developing countries
1000 are in children less than 15 years of age.
Of adult infections, 40% are in women and 15% in individuals 15-
25 years.

As of December 1999, 733,374 Americans reported with AIDS. At
least 430,441 of them have died.

Prior to the introduction of combination therapies for HIV, AIDS
incidence was increasing at a rate of just under 5% each
year.                                                      13
                         AIDS Statistics
                       Sub-Saharan Africa
• About 1 million new cases of AIDS per year

• 24 million people with HIV infection

• AIDS is responsible for a decrease in life expectancy and increase in
child mortality. Child mortality rates in East Africa will double by 2010 and
adult life expectancy has already declined by 2 years in that region.

• Several countries in sub-Saharan Africa report infection rates of 20-25%,
especially urban areas.

•Botswana: 35.8% of adult population infected
• In Zambia, 1 in 5 urban girls is HIV-positive by the age of 20       14
    AIDS Statistics

South and East Asia
• In south and east Asia there are
more than 200,000 new cases per
year and 7.3 million people with HIV


Worldwide

• Current estimates are that in
2000, 34.3 million people have
HIV infection
                                       15
HIV and AIDS




               16
HIV and AIDS




               17
                             HIV and AIDS
                               The Virus
The virus only grows on T4 cells that are proliferating in response to
an immune stimulus
Therefore difficult to grow in culture
• Robert Gallo : reverse transcriptase in
activated T4 cells in blood of patients
with AIDS : HTLV-3

• Luc Montagnier: LAV



Human Immunodeficiency                      Human immunodeficiency viral particles are seen at medium
                                            magnification in this electron micrograph. Note the central core
                                            and the outer envelope (CDC)

Virus (HIV)                                                                                     18
                      HIV and AIDS
The cellular and immunological picture - The course of the disease




                                                              19
                       HIV and AIDS
The cellular and immunological picture - The course of the disease




                                                               20
                              HIV and AIDS
 The cellular and immunological picture
           The course of the disease
                 1. Acute Infection
• High virus titer
• Mild symptoms
• Fall in CD4+ cells but recovers
• Rise in CD8+ cells but recovers
• A high virus titer (up to 10 million viruses per ml
blood)
• Macrophages infected
                                                        21
                      HIV and AIDS
             2. A strong immune response


Virus almost disappears from circulation
• Good cytoxic T cell response
• Soluble antibodies appear later against both surface
and internal proteins
• Most virus at this stage comes from recently activated
and infected CD4+ cells (half life about a day)
• CD4+ cell production compensates for loss due to
lysis of cells by virus production and destruction of
infected cells by CTLs                                     22
                             HIV and AIDS
                             3. A latent state


Latency of virus and of symptoms
• Virus persists in extravascular tissues
• Lymph node dendritic cells
• Resting CD4+ memory cells (last a very
long time - a very stable population of cells)
• Resting CD4+ cells carry provirus



                                                 23
               HIV and AIDS
       • 10 billion HIV particles per day
           • Virus half life 5.7 hours
• 100-10 million virions per ml blood (set point)
   • Small minority of T4 cells are infected
         • Virus found in lymph nodes


                                                24
     HIV and AIDS
                   4. The beginning of disease
Massive loss of CD4+ cells
• CD4+ cells are the targets of the virus
• Cells that proliferate to respond to the
 virus are killed by it: Clonal deletion
• Dendritic cells present antigen and virus
 to CD4 cells just as they are activated
• Epitope variation allows more and more HIV to
escape from immune response just as response wanes
• Apoptosis of CD4+ cells

• HIV patients with high T4 cell counts
                                                     25
 do not develop AIDS
     HIV and AIDS
  5. Advanced disease - AIDS
CD8+ cells destroy more CD4+ cells
• CD4 cell loss means virus and infected
 cells no longer controlled
• As CD4+ cells fall below 200 per cu mm
 virus titer rises rapidly and remaining
immune response collapses
• CD8+ cell number collapses
• Opportunistic infections
• Death in ~2 years without intervention
Good correlation between loss of
                                           26
CD4+ cells and onset of symptoms
HIV and AIDS



               Good correlation between
               number of HIV particles
               measured by PCR and
               progression to disease




                                 27
HIV and AIDS


               Viral load predicts
               survival time




                                     28
HIV and AIDS



               CD4 cell count is not a
               good predictor of
               progression to disease




                                  29
               HIV and AIDS
                  Cofactors
Not all cases of Kaposi’s are associated with HIV
Not all HIV infected persons suffer from Kaposi’s
  20% of homosexual HIV+ males get Kaposi’s
Few IV drug users or hemophiliacs get Kaposi’s
  Kaposi’s sarcoma associated herpes virus
           Human herpes virus-8                30
                     HIV and AIDS
                     Three Views of AIDS

      Gallo: Infection by HIV is sufficient to cause AIDS

Montagnier: HIV may be harmless in the absence of other co-
factors
Duesberg / Mullis: HIV is too silent to be the etiologic agent of
AIDS. It is a much maligned by-stander
So far it seems that >50% of HIV-infected persons have
progressed to AIDS
There is NO strong evidence there is any other infectious
agent involved than HIV                                   31
                      HIV and AIDS
Despite possible co-factors associated with lifestyle HIV-infected
  persons progress to AIDS at a similar rate if not treated with
                         chemotherapy

            100                                                  HIV negative
                                                                 homosexual
% without         HIV+ hemophiliac
AIDS              >25 years                           HIV+ hemophiliac <20 years
            50
                                         HIV+
                                         homosexual



                                     6                   12                 32
                               years
                 HIV and AIDS
          Exposed uninfected persons
           Long term non-progressors
• People who have been infected with HIV for more
than seven years that have stable CD4+ cell counts
  above 600 per cu mm with no symptoms and no
                  chemotherapy
• Many have produced and immune response to the
                     virus
            •Co-receptors and disease        33
                 HIV and AIDS
          Co-receptors and HIV infection
            • CCR5 is a chemokine receptor
• Cells with homozygous mutant CCR5 molecules are not
                    infected by HIV
                 1 in 100 Caucasians
                     No Africans
 • Persons with heterozygous mutant CCR5 molecules
            progress to AIDS more slowly
                  17% Caucasians
                     2% Africans                  34
             HIV and AIDS
               Co-receptors
25% of long term survivors are CCR5 or CCR2
             mutants (deletions)
Others may make high levels of chemokines
      Many other chemokine receptors

                                        35
                   HIV and AIDS
                    • Nairobi prostitutes

                    • Rare HLA antigens
  • Associations between resistance to infection and their
         class I and class II MHC (HLA) haplotypes
 • May present epitopes that are highly conserved between
different HIV-1 variant strains. Epitope is presented by HLA-
A*6802 is in the protease. Protease may not be able to bear
                 much mutation in this region
                                                         36
          HIV - The Virus



           Membrane: host derived
Two glycoproteins: gp160      gp120 and gp41
   gp41 is fusogen that spans the membrane
            sugars: immunosilent vaccine problem
                                               37
                 HIV - The Virus
   Group-Specific Antigens

p17: inner surface - myristoylated

p24: nucleocapsid
p9: nucleocapsid associated with RNA



            GAG gene

                                       38
                    HIV - The Virus
   Enzymes

• Polymerase (reverse transcriptase)
            • Integrase

   • Protease (cuts polyproteins)



           • POL gene
                                       39
HIV - The Virus




                  40
HIV - Life History
                  A retrovirus
                     • Latency
        • Specific destruction of CD4+ cells


        •How does the virus enter the
                   cell?



                                        41
             HIV - Life History
                Entry into the cell
         T4 (CD4+) cells are major target


                         Human HeLa Cell
Human HeLa               transfected with CD4
   Cell                  antigen


NOT INFECTED                           INFECTED

                                                  42
HIV - Life History
               • Fusion at ambient pH
              • No need for entry into
                    lysosomes
                     • Syncytia

            Profound significance for
            AIDS progression
            Profound significance for
            therapy

                                    43
HIV - Life History
         Co-Receptors
           CD8+ Cells


MIP-1 alpha MIP-1 beta RANTES
           Chemokines


Block HIV infection of macrophages

                                     44
             HIV - Life History

               HIV



                             chemokine
                                               Mutant
                      CD4   CCR5               CCR5
      CCR5                               CD4
CD4


                     macrophage
                                                        45
                  HIV - Life History
                       Co-receptors
                       CXCR4 (Fusin)
                Co-receptor on CD4+ T4 helper cells
                    Another chemokine receptor
                       Similar to IL-8 receptor
                     G-protein coupled receptor

                  Thousands of similar receptors
                 May explain the change of tropism
May explain why at a given time only a fraction of T4 helper cells are
                                                                  46
                              infected
          CD4 antigen

           Co-receptor




                                       CD4+
                                       cell




HIV gp120 binds to CD4 / co-receptor


                                              47
CD4 antigen

Co-receptor




              CD4+
              cell




                     48
Entry of HIV into a CD4+ Cell

   Movie 1 Binding of HIV gp120 to CD4 cell

  Movie 2 Entry of HIV into a CD4 cell




                                              49
         HIV - Life History
               HIV carries with it:
                Reverse transcriptase
                • Integrase
                • Protease
                • tRNA primer


                 HIV genes
              GAG POL ENV

HIV has no oncogene but could still be oncogenic

              vaccine problem                      50
                 HIV - Life History
                                                       RT RNA
                                                          genome
     R       U5 GAG         POL   ENV    U3    R
                                                   DNA
                                               pol
U3   R       U5 GAG         POL   ENV    U3 R U5 provirus
                  splicing                                 RNA genome
         R    U5 GAG POL           ENV    U3       R

                  spliced                                 GAG and POL
                                                          proteins


               ENV glycoprotein                                    51
                     HIV - Life History
                           Latency - Cellular
Only activated T4 cells can replicate virus
Most infected T4 cells are rapidly lyzed but are replaced
Some T4 cells revert to resting state as memory cells which are long-lived
Memory T4 cells cannot replicate the virus
Macrophages do not show latency

                            Clinical Latency
           HIV infection is not manifested as disease for years
                                                                        52
  During apparent clinical latency, virus is being replicated and cleared
      Dynamics of CD4 T cells in an HIV
      Cell death
                 infection       Chronically-
    apoptosis etc                                                   infected
                                                                 memory T cells
                                                                  with provirus
                                                  Return to
                       Infection                   resting
Uninfected                                          state
activated
  T cell
                    Long lived!
                                                 Reactivation


               Uninfected          Cell death                                  Long lived!
               unactivated          immune
                memory                                   Adapted from Saag and Kilby
               T cell pool
                                   destruction           Nat Med 5: 609, 1999          53
Dynamics of CD4 T cells in an HIV
           infection




                     Infection spreads

                                         54
             The Genome of HIV




                     Three structural genes
                             LTRs
         Extra open reading frames are clue to latency
These ORFs code for small proteins - antibodies in AIDS patients 55
              The Genome of HIV
                    Small non-structural proteins
         mRNAs made by multiple splicing of genomic RNA
                  (c.f. mRNA for structural proteins)


                 EARLY
   TAT: TransActivator of Transcription                 TAT and REV
REV: Regulator of Virion Protein Expression             are essential for
                                                        HIV replication
     NEF: Negative Regulatory Factor

                  LATE
       VIF: Virion Infectivity Factor
           VPU: Viral Protein U
           VPR: Viral Protein R                                       56
                              TAT
    RNA Pol II



     U3     R U5                            U3     R    U5
                 TAR




          TAT


When TAT         All proteins (ENV, POL, GAG, TAT, REV etc)

                                                              57
                                  REV
                          REV modulates splicing
REV
level        REV regulates ratio of structural to control proteins
                                                                        DNA
        U3    R    U5                           U3     R     U5
High                                                                   Genomic RNA

                      single
High                                                          ENV glycoprotein
                      splice



                      Multiple                               Mini-proteins (TAT, REV)
Low
                      splices

                                                                            58
          Latency

In the absence of any activating
           stimulus:
          Homeostasis




                                   59
                           Latency
 Homeostasis leads to latency of T4 cells but cell faces two problems:
          • Superinfection by another HIV via CD4 antigen
 • Gp120 attached to CD4 antigen at cell surface invites immune attack
                     HIV

            CD4
                                 Anti-Gp120
    HIV



             Free
T4 cell      Gp120

      Answer: Get rid of surface CD4 antigen on infected cell
                                                                    60
                       Latency              NEF binds
                                             to CD4
   Co-receptor                               antigen

   CD4 antigen
                     NEF
                           NEF protein is
                           early function




                     NEF

NEF cause internalization of CD4 and digestion in lysosomes
                                                              61
             NEF




                   NEF
  lysosome




 CD4 antigen is
internalized and
    digested             62
                     Latency Breaks
     Antigen binds

                     Immune response



T4 resting                                       T4 activated

       HIV LTR has usurped
        activation control
            sequences           HIV production
                                                                63
        Another problem for HIV
In the activated cell gp120 will be made –so will CD4 antigen



 Endoplasmic
 reticulum                          Gp120/CD4 complexes will be
            ribosome                  targeted for destruction

                       Gp160
                                  VPU causes specific proteolysis of
                                  CD4 in the endoplasmic reticulum
               CD4                       of HIV-infected cell
                                                                64
          Inexorable decline of CD4+ T4
                      cells
 Why do all of
 the T4 cells
 disappear?

At early stages
  of infection
only 1 in 10,000
cells is infected
  Late 1 in 40


                                                                  65
                    Of great importance to therapeutic strategy
               Virus destroys the cell as a result of
                            budding

           But few cells are infected:
           Early stage of infection 1:10,000
           Late 1:40
           HIV could kill sub population of
           precursor cells
           People develop AIDS even when they
           have HIV that does not lyze cells
                                                        Why do all T4
                                                        cells
1. PUNCTURED
                                                        disappear?
                                                                66
  MEMBRANE
    Why do all T4 cells disappear? - 2
                                           But syncytia not
                                           common
Infected CD4                               Most T4 cells are
     cell          Cells Fuse              not HIV+
Gp120 positive                             Could “sweep up”
                                           uninfected cells

                                           Syncytia may be
                            Uninfected     poor or ineffective
  Killing of CD4 cells       CD4 cell      at immune
      2. Syncytium               Gp120     response
       Formation                negative               67
         Why do all T4
         cells
         disappear?


         Cytotoxi
         c T cell


    Killing of CD4 cells
3. Cytotoxic T cell-mediated
            lysis

                         68
Killing of CD4+ cells
4. Binding of free Gp120
 to CD4 antigen makes
 uninfected T4 cell look
   like an infected cell
 Complement-mediated
           lysis
Could account for the
loss of uninfected T4
cells

                           69
       Why do all T4 cells disappear?
5. AIDS related cytotoxic antibodies that bind to specific
antigen on surface of activated T4 cells
6. Involvement of a precursor T4 cell population
7. Autoimmunity. Gp120 binds to CD4 antigen. CD4 antigen
binds to type II MHC: similar sequences in MHC and Gp120.
Anti gp120 may be anti MHC - may be problem for a vaccine
8. Apoptosis of T4 cells. Apoptosis of T4 cells is normal in
clonal deletion to overcome autoimmunity

                                                             70
At a late stage of HIV infection, the
                                             Why do all CD8+
cytotoxic T cells disappear very rapidly       killer cells
These cells do not have CD4 antigen            disappear?
They are not infected by HIV
There is a correlation of control of virus
and presence of CD8+ cells
HIV subtypes present in late infection
cause mass apoptosis of CD8+ cells
Cytotoxic T cells do have CXCR4 co-
receptor. HIV binds but does not enter
cells
Signal from binding prompts mass
suicide of CD8+/CXCR4+ cells                               71
      CD8 cell                              Macrophage
       (no CD4 antigen)
                          gp120
HIV
                              chemokine

                                  CXCR4
                                                G protein

                  ?
                                                 signal

                                                    ?




                                               Binding to
        Binding to
                                             CXCR4 results
      CXCR4 results in                      in expression of
       expression of                          TNF-alpha on
        TNF-alpha                            the cell surface
         receptor II

                            Apoptosis of T cells
                                                                72
CD8 cell            Macrophage


   CXCR4


 Death




           CD8 T cell
           apoptotic
            bodies
                                 73
 Macrophage ingests
  CD8 cell apoptotic
       bodies

 Apoptotic bodies are
digested in lysosomes   74
              HIV and Auto-Immunity

            gp120                                    Class II MHC
                                CD4
                               antigen



Gp120 and MHC II both bind to CD4 antigen. Both have CD4 binding sites

                          Anti-HIV
                          (gp120) binds
                          MHC II also
                                                               75
  Macrophages may be infected by
           two routes
HIV   gp120
              CD4
                                 HIV gp120 binds to
                                 macrophage CD4 antigen


                                 Virus is opsonized by anti
                                 gp120 antibodies which
                                 bind to macrophage Fc
                                 receptors - an enhancing
                Fc receptor      antibody

                    Anti-gp120      vaccine problem
                                                        76
          HIV
     Macrophages - The Trojan Horse
  Early HIV isolated during infection are macrophage tropic (have a
             macrophage chemokine co-receptor (CCR5)
      Virus probably infects patient via macrophages in semen
Infection by HIV leads to altered cytokine production        slim disease”
   Slim disease very like Visna in sheep - also infects macrophages


           Macrophages form a reservoir outside the blood

                 Carry virus into different organs (brain)
   Non-proliferating mature macrophages sustain HIV production for
        a long time without being killed by virus - - no latency
                                                                     77
             Population Polymorphism
                           HIV is a retrovirus
  Retroviruses use host cell RNA polymerase II to replicate
                       their genome
             Pol II has a high error rate 1:2,000-10,000
                   HIV genome 9749 nucleotides
         Therefore EVERY new virus has at least one mutation!
                 Every possible single mutation arises daily
               1% of all possible double mutations arise daily

The HIV that infects a patient is very different
  from that seen by the time AIDS appears          vaccine problem
                                                                 78
          Population Polymorphism
• Initial infecting virus is macrophage-tropic (has CCR5 as co-
                           receptor)
         • These are non-syncytium-inducing strains
 (Note: most vaccines have been made against syncytium-
              inducing T4 cell tropic strains)
• As virus mutates, it changes subtypes of cells that it infects
     as the ability to bind different co-receptors changes

                                                           79
Population Polymorphism
      Early in infection:
       • Macrophage-tropic
     • Non-syncytium-inducing
        • Slowly replicating
       Late in infection
          •T4 cell tropic      vaccine problem
      • Syncytium-inducing
         • High titer virus
                                          80
      Population Polymorphism
            • The most variable protein is gp120
• Amino acid sequence within a single patient varies by 1-6%
                 • Up to 30% in population

                   vaccine problem

            • Glycosylation masks conserved sites

                   vaccine problem

           Co-infection may result in recombination

                   vaccine problem                             81
           Population Polymorphism
• Variation in reverse transcriptase leads to resistance to
nucleoside analogs
                       drug problem

• Variation in protease leads to resistance to protease
inhibitors
                       drug problem

   Polymorphism due to high mutation rate as a result of
       lack of proof-reading in reverse transcriptase
       Sub-populations arise with altered cell tropism        82
  Other cells infected by HIV
             CD4-
  • Epithelial cells of   bowel and vagina
       • Endothelial cells of brain
• Brains cells : Astroglia, oligodendroglia


    Galactocerebroside receptor
                                              83
       Anti-HIV Strategies
                  • Education
               Sexually transmitted
               Not highly infectious



              • Chemotherapy
      Mutation selection            Resistance
                        but
Suppress replication          No capacity for mutation   84
                Anti-HIV Strategies
                              Highly
                              Active
                               Anti-
                            Retroviral
                             Therapy
HAART: Two nucleoside analog RT inhibitors and 1 protease inhibitor
Now also: Two nucleoside analog RT inhibitors and 1 non nucleoside
                                                              85
Towards an Anti-HIV Vaccine
  Problems for an anti-HIV vaccine
         HIV is a RETROVIRUS

               Latency
            • Genetic drift
                      reverse transcriptase
                       RNA polymerase II

            Polymorphism
            • Back mutation
         • Oncogenicity? - LTR                86
      Towards an Anti-HIV Vaccine
         Problems for an anti-HIV Vaccine 2
    • Syncytia             Cell to cell transmission
    (humoral antibody ineffective, need CTL)

                   • Hidden epitopes
      • Vaccine may be immunosuppressive
      (antibodies to MHC II)

• Enhancing antibodies (opsonizes HIV           Fc receptor)


                                                       87
 Towards an Anti-HIV Vaccine
   Problems for an anti-HIV vaccine 3

• What is the end point for success in trials
- What are the correlates of protection?

 • Vaccinee appears to be HIV positive

    • Will counseling alter the results?

                                                88
     Does HIV Cause AIDS?
    Single common factor between:
         • Gay San Franciscans
        • New York I.V. drug users
         • African heterosexuals
             • Hemophiliacs
• Spouses of hemophiliacs and drug users
• Children of hemophiliacs and drug users   89
        Does HIV Cause AIDS?

• HIV precedes AIDS in every population in which
                  AIDS occurs
           • Infection by cloned virus
              SIV              HIV




          Simian AIDS      Human AIDS         90
                   Remember!
  • Education led to leveling off   of rate of increase in
                          AIDS
        • HAART has greatly slowed death rate
•The fact that fewer people are dying per year from
the infection means that the number of HIV-infected
people in the population is rising!
• Unless education continues to be successful and
unless we can cure infected people of virus, the
problem of virus spread is and will continue to be
with us                                            91

				
DOCUMENT INFO