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HIV Infection- A summary


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Structure of Virus, modes of transmission, pathogenesis, clinical features, biochemical basis of clinical symptoms, laboratory diagnosis, treatment and prevention.

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HIV Infection- A summary

  1. 1.
  2. 2. Learning objectives To understand: • Etiology • Pathogenesis • Laboratory diagnosis • Treatment, and • Prevention of HIV infection 30-Mar-18 Biochemistry for medics 2
  3. 3. What is AIDS ? o Acquired (acquired during lifetime, not inherited) o Immuno- (related with immune system) o Deficiency (related with deficient immune response) o Syndrome (collection of symptoms) 30-Mar-18 3Biochemistry for medics
  4. 4. Syndrome ? ❖A collection of symptoms ❖due to underlying infections and malignancies ❖resulting from specific damage to immune system ❖caused by human immunodeficiency virus (HIV). 30-Mar-18 Biochemistry for medics 4
  5. 5. Incidence • HIV (human immunodeficiency virus) infection has now spread to every country in the world. • Approximately 40 million people are currently living with HIV infection, and • An estimated 25 million have died from this disease. 30-Mar-18 5Biochemistry for medics
  6. 6. History of AIDS • The first indication of this new syndrome came in 1981 in homosexual drug addict males; • two things were common- Pneumocystis pneumonia and Kaposi’s sarcoma. • Both these are markers of collapsed immune system, 30-Mar-18 6Biochemistry for medics
  7. 7. AIDS Markers Pneumocystis pneumonia and Kaposi’s sarcoma are considered AIDS markers, since they reflect the underlying collapsed immune system30-Mar-18 Biochemistry for medics 7
  8. 8. AIDS Immune incompetence Infections and malignancies Collection of symptoms 30-Mar-18 Biochemistry for medics 8
  9. 9. Etiological agent of AIDS Retrovirus (retroviridae family) Latency Persistent viremia Infection of nervous system Weak host immune responses Lentivirus 30-Mar-18 Biochemistry for medics 9
  10. 10. Nomenclature of virus • In 1986, The International Committee on virus Nomenclature decided on the generic name of the causative virus as the : • Human Immunodeficiency Virus. 30-Mar-18 10Biochemistry for medics
  11. 11. Human Immunodeficiency Virus • HIV has high affinity for CD4 T lymphocytes and monocytes. • HIV binds to CD4 cells and becomes internalized. The virus replicates itself by generating a DNA copy by reverse transcriptase. • Viral DNA becomes incorporated into the host DNA, enabling further replication. 30-Mar-18 11Biochemistry for medics
  12. 12. Immune deficiency in HIV Infection Decreased Number of T-Helper cells Decreased activity of monocytes and macrophages Decreased activity of B lymphocytes Decreased cell mediated immune response Decreased activity of NK cells 30-Mar-18 12Biochemistry for medics
  13. 13. Structural Characteristics of HIV (Envelop) • Shape- Spherical • Size- 90-120 nm in diameter • Envelop- Host lipoprotein and viral glycoproteins • Viral glycoproteins consist of: o Surface projecting knob like spikes (gp 120) o Anchoring transmembrane pedicles(gp 41) 30-Mar-18 Biochemistry for medics 13
  14. 14. Structural Characteristics of HIV (Nucleocapsid core) • Interior to the envelope is an outer icosahedral nuclear capsid shell(p17) and • an inner cone-shaped core containing ribonucleoproteins (p24). 30-Mar-18 14Biochemistry for medics (The proteins and glycoproteins are indicated by their mass expressed as kilo Daltons)
  15. 15. Components of nucleocapsid core • The enzymes integrase p32, • protease p10, • reverse transcriptase p55/66 and • 2 copies of single stranded genomic RNA are present inside the core.30-Mar-18 Biochemistry for medics 15
  16. 16. Functions of structural components of HIV Structural component Function gp 120 Constitute the major surface component of the virus which binds to the cell CD4 receptors on susceptible host cells. gp41 causes cell to cell fusion p17 Protects nucleocapsid core p24 First diagnostic structural component p10 (protease) Cleaves viral precursor proteins P32(integrase) Integrates viral DNA to host genome P55/66(reverse transcriptase) Catalyzes conversion of viral RNA to DNA 30-Mar-18 Biochemistry for medics 16
  17. 17. HIV Genome 30-Mar-18 Biochemistry for medics 17
  18. 18. HIV Genome 30-Mar-18 Biochemistry for medics 18 • gag • pol • envStructural genes • LTR • Vif • Vpr • Vpu/vpx • tat • nef • rev Regulatory genes
  19. 19. HIV Genome 30-Mar-18 Biochemistry for medics 19
  20. 20. HIV Genome There are two types of genes analyzed- a) Structural genes encode for products which participate in formation of functional structure of virus 1) gag gene • encodes for core and shell of virus. • The gene product is a precursor protein p55, which is cleaved into p17, p24 and p15. • The p24 antigen (major core antigen) can be detected in serum during the early stages of infection till the appearance of the antibodies. 30-Mar-18 20Biochemistry for medics
  21. 21. Genome of HIV (Structural genes) 2) pol gene • Encodes for the polymerase reverse transcriptase and other viral enzymes such as protease and integrase. • It is expressed as a precursor protein, which is cleaved in to components like p64 which has reverse transcriptase and RNAse activity: p51 which has only reverse transcriptase activity: p10 is a protease that cleaves gag precursor and p32 is an integrase. 30-Mar-18 21Biochemistry for medics
  22. 22. b) Non structural and Regulatory genes 1) vif - (Viral infectivity factor gene) influences infectivity of viral particles. 2) vpr-stimulates promoter region of the virus 3) vpu (in HIV-1 ) and vpx (in HIV-2) enhance maturation and release of progeny virus from cells. Detection of the type specific sequences vpu and vpx is useful in distinguishing between infection by HIV type 1 and 2. 30-Mar-18 23Biochemistry for medics
  23. 23. HIV Types HIV1 Original isolates of HIV Present all over the world HIV2 First isolated from West Africa 40 % genetic similarity 30-Mar-18 Biochemistry for medics 26
  24. 24. Antigenic variations in HIV Antigenic variations Different persons Same person sequential isolates Same person- different sites Different races 30-Mar-18 Biochemistry for medics 27
  25. 25. Modes of transmission Modes of Transmission Sexual contact Sharing of needles Needle prick injury Organ transplantation Mother to child Blood transfusion 30-Mar-18 Biochemistry for medics 28
  26. 26. Modes of transmission 30-Mar-18 Biochemistry for medics 29
  27. 27. Age for HIV Infection Age for HIV Infection Adults (25-49) 70% Adolescent (13-24) 25% Young children (<13 years) Less than 5%% 30-Mar-18 Biochemistry for medics 30
  28. 28. Pathogenesis • Infection is transmitted when virus enters the blood or tissues of a person and comes in to contact with a suitable host cell, principally the CD4 lymphocytes. • The virus may infect any cell bearing the CD4 antigen on the surface. • Primarily these are the CD4 + helper T lymphocytes. 30-Mar-18 31Biochemistry for medics
  29. 29. CD4 presenting cells CD 4 cells T Helper cells Monocytes B Lymphocytes Macrophages 30-Mar-18 Biochemistry for medics 32
  30. 30. CD 4 presenting cells • Some other immune cells possessing CD4 antigens are also susceptible to infection, like B lymphocytes, monocytes and macrophages including specialized macrophages such as Alveolar macrophages in the lungs and Langerhans cells in the dermis. • Glial cells and microglia cells are also susceptible. 30-Mar-18 Biochemistry for medics 33
  31. 31. Steps of viral entry in to the host cell 1) Attachment of virus to the host cell –Specific binding of the virus to the CD4 receptors is by the envelop glycoprotein gp120. 30-Mar-18 Biochemistry for medics 34
  32. 32. Steps of viral entry in to the host cell 2) Cell to cell fusion – For infection to take place the cell fusion is essential. o This is brought about by the transmembrane glycoprotein gp 41. 30-Mar-18 35Biochemistry for medics
  33. 33. Steps of viral entry in to the host cell • HIV-1 utilizes two major co-receptors along with CD4 to bind to, fuse with, and enter target cells; • these co-receptors are CCR5 and CXCR4, which are also receptors for certain endogenous chemokines. 30-Mar-18 Biochemistry for medics 36
  34. 34. Steps of viral entry in to the host cell o Strains of HIV that utilize CCR5 as a co-receptor are referred to as macrophage tropic viruses (M –tropic viruses) o Strains of HIV that utilize CXCR4 are referred to as T - tropic viruses. o Many virus strains are dual tropic in that they utilize both CCR5 and CXCR4. 30-Mar-18 Biochemistry for medics 37
  35. 35. Steps of viral entry in to the host cell 3) Uncoating of the viral envelope and entry of nuclear capsid core into the cell o After fusion of virus with the host cell membrane, HIV genome is uncoated and internalized in to cell. o Viral RNA is released into the core cytoplasm 30-Mar-18 38Biochemistry for medics
  36. 36. Steps of viral entry in to the host cell 4) Viral transcription o viral reverse transcriptase mediates transcription of its RNA; o RNA-DNA hybrid is formed. o Original RNA strand is degraded by ribonuclease H, followed by o synthesis of second strand of DNA to yield double strand HIV DNA30-Mar-18 39Biochemistry for medics
  37. 37. Steps of viral entry in to the host cell 4) Integration into the host DNA as provirus o The double stranded DNA is integrated in to the genome of the infected host cell through the action of the viral integrase enzyme, causing a latent infection. 30-Mar-18 40Biochemistry for medics
  38. 38. Fate of provirus o From time to time, lytic infection is initiated releasing progeny virions, which infect other cells. o The long and variable incubation period of HIV is because of the latency. o In an infected individual the virus can be isolated from the blood, lymphocytes, cell free plasma, semen, cervical secretions, saliva, urine and breast milk 30-Mar-18 Biochemistry for medics 41
  39. 39. Steps of viral exit from host cell • Transcription back into RNA o The viral DNA is transcribed into RNA and multiple copies of viral RNA are produced. o There are only nine genes in HIV RNA, and these code for the production of structural proteins, accessory proteins, and enzymes essential for the virus's replicative cycle. 30-Mar-18 42Biochemistry for medics
  40. 40. Steps of viral exit from host cell • Virion assembly - With the help of viral protease, the new virions are assembled into the polypeptide sequences needed for HIV virion formation and infectivity. • Cell lysis. The infected cell is made to burst open, presumably by the action of cellular proteins. 30-Mar-18 43Biochemistry for medics
  41. 41. 30-Mar-18 44Biochemistry for medics
  42. 42. Clinical Manifestations • The center for disease control (USA) has classified the clinical course of HIV infection under various groups. • Acute HIV infection • Asymptomatic or Latent infection • Persistent generalized lymphadenopathy (PGL) • AIDS related complex • Full blown AIDS (Last stage) 30-Mar-18 45Biochemistry for medics
  43. 43. 1. Acute HIV infection • Symptoms- A flu-like illness with fever, sore throat, headache, tiredness, skin rashes and enlarged lymph nodes in the neck within several days to weeks after exposure to virus. • These symptoms usually disappear of their own within a few weeks. • Some patients do not develop symptoms after they first get infected with HIV. 30-Mar-18 46Biochemistry for medics
  44. 44. 1. Acute HIV infection • This phase is also called window period or phase of Seroconversion. • The test for HIV antibodies appears negative while HIV antigenemia (p24 antigen) and viral nucleic acids can be demonstrated at the beginning of the phase. 30-Mar-18 Biochemistry for medics 47
  45. 45. 2. Asymptomatic or Latent infection • All persons infected with HIV, pass through a phase of symptomless infection (Clinical latency), which may last up to several years. • Even though the person has no symptoms, he or she is contagious and can pass HIV to others. • This state may last from a few months to more than 10 years. 30-Mar-18 48Biochemistry for medics
  46. 46. 2. Asymptomatic or Latent infection (contd.) • The virus continues to multiply actively and infects and kills the cells of the immune system • The virus destroys the CD4 cells that are the primary infection fighters. • The patients show positive antibody tests during this phase. 30-Mar-18 49Biochemistry for medics
  47. 47. 2. Asymptomatic or Latent infection (contd.) • The median time between primary HIV infection and development of AIDS has been stated as approximately 10 years. • About 5-10 % percent of the infected appear to escape clinical AIDS for 15 years or more. • They have been ‘long term survivors” or “long term non progressors”. 30-Mar-18 Biochemistry for medics 50
  48. 48. 3. Persistent generalized lymphadenopathy (PGL) • This has been defined by presence of: • enlarged lymph nodes, • at least I cm in diameter, • in two or more non contiguous extra inguinal sites, • that persist for at least three months, • in the absence of any current illness or medication that may cause lymphadenopathy. 30-Mar-18 51Biochemistry for medics
  49. 49. 3. Persistent generalized lymphadenopathy (PGL) These are diagnostic of HIV when blood tests are positive for antibodies. 30-Mar-18 Biochemistry for medics 52 PGL Enlarged lymph nodes (>1cm) Two or more non -contiguous extra- inguinal sites Persistence for at least three months Absence of any current illness or medication
  50. 50. 4. AIDS related complex AIDS related complex Weight loss Persistent fever Diarrhea Generalize d fatigue Signs of other opportuni stic infections 30-Mar-18 Biochemistry for medics 53 The common opportunistic infections are oral candidiasis, herpes zoster, salmonellosis or Tuberculosis and hairy cell leucoplakia.
  51. 51. Opportunistic infections in AIDS 30-Mar-18 Biochemistry for medics 54
  52. 52. 4. AIDS related complex • The patients are usually severely ill and many of them progress to AIDS in few months. • The CD4 cell count decreases steadily when the count falls to 200, or less, clinical AIDS usually sets in. • For this reason the case definition by CDC includes all HIV infected cases with CD4 + T cell counts of 200 or less, irrespective of clinical condition. 30-Mar-18 55Biochemistry for medics
  53. 53. 5. Full blown AIDS • This is the end stage disease representing the irreversible break down of immune defense mechanisms. • In addition to the opportunistic infections the patient may develop primary CNS lymphomas and progressive encephalopathy, dementia and other neurological abnormalities. • Kaposi sarcoma and Pneumocystis pneumonia are almost always observed in a majority of patients. 30-Mar-18 56Biochemistry for medics
  54. 54. Kaposi’s sarcoma • Kaposi’s sarcoma- is an indolent, multifocal non metastasizing mucosal or cutaneous tumor probably of endothelial origin, represented in the form of purple spots in the skin. 30-Mar-18 57Biochemistry for medics
  55. 55. Laboratory diagnosis of HIV infection Non Specific Tests Total Leukocyte and lymphocyte count- T cell subset Assays IgA and Ig G levels Platelet count- Specific Tests HIV antigen Antibodies Isolation of virus Viral nucleic acids 30-Mar-18 Biochemistry for medics 58
  56. 56. Laboratory Diagnosis of HIV infection 1) Non Specific Tests- The following tests help to establish the immunodeficiency in HIV infection. a) Total Leukocyte and lymphocyte count- to demonstrate leucopenia and lymphopenia. The lymphocytic count is usually below 2000/mm3 b) T cell subset Assays- Absolute CD4+ cell count is less than 200 /L.T4 T8 ratio is reversed. The decrease in CD4 is the hall mark for AIDS. c) Platelet count- shows Thrombocytopenia. d) IgA and Ig G levels are raised e) Diminished cell mediated Immunity as indicated by skin tests f) Lymph node biopsy shows profound abnormalities. 30-Mar-18 59Biochemistry for medics
  57. 57. Laboratory Diagnosis of HIV infection 2.Specific Tests for HIV infection- These include demonstration of - • HIV antigen, • Antibodies, • Viral nucleic acids or other components and • Isolation of virus 30-Mar-18 60Biochemistry for medics
  58. 58. Laboratory Diagnosis of HIV infection i) Detection of antigen o The major core antigen p24 is the earliest virus marker to appear in blood. o The p24 Capture ELISA assay, which uses anti p24 antibody as the solid phase can be used for this. 30-Mar-18 61Biochemistry for medics
  59. 59. Laboratory Diagnosis of HIV infection • i) Detection of antigen (contd.) • Free p24 antigen disappears from circulation and remains absent during the long asymptomatic phase to reappear only when severe clinical disease sets in. • This test is positive in about 30% of the infected persons. • In the first few weeks after infection and in the terminal phase, the test is uniformly positive. 30-Mar-18 62Biochemistry for medics
  60. 60. Disease progression through different phases in HIV infected cases CD4 count comes down while the viral count goes high with the passage of time 30-Mar-18 63Biochemistry for medics
  61. 61. Laboratory Diagnosis of HIV infection • Detection of antibodies o It takes 2-8 weeks to months for the antibodies to appear in circulation o IgM antibodies appear first, to be followed by IgG antibodies o Once antibodies appear they increase in titer for the next several months o IgM antibodies disappear in 8-10weeks while IgG antibodies remain through out. 30-Mar-18 64Biochemistry for medics
  62. 62. Laboratory Diagnosis of HIV infection • Detection of antibodies (contd.) o ELISA- ELISA is the most frequently used method for screening of blood samples for HIV antibody. 30-Mar-18 65Biochemistry for medics
  63. 63. Laboratory Diagnosis of HIV infection Significance of ELISA • Antibodies can be detected within 6-12 weeks after infection using the earlier generation of assays. • Using the newer third generation ELISA, Antibodies can be detected within 3-4 weeks 30-Mar-18 66Biochemistry for medics
  64. 64. Laboratory Diagnosis of HIV infection Supplemental test Western blotting-Confirmatory test • Western blots are regarded as the gold standard • antibodies against both the env and the gag proteins are detected. 30-Mar-18 67Biochemistry for medics
  65. 65. Laboratory Diagnosis of HIV infection 3. Demonstration of viral Nucleic acid • probes or • PCR • The latter may be useful because of its extremely high sensitivity. 30-Mar-18 68Biochemistry for medics
  66. 66. Laboratory Diagnosis of HIV infection • PCR -In this the target HIV RNA or proviral DNA is amplified enzymatically in vitro by chemical reaction. • It is an extremely sensitive assay because a single copy of proviral DNA can be amplified. • Qualitative PCR is useful for diagnostic purposes. 30-Mar-18 69Biochemistry for medics
  67. 67. Laboratory Diagnosis of HIV infection 4. Virus isolation • Virus isolation is tedious and time-consuming (weeks) and is successful in only 70 to 90% of cases. • Therefore virus isolation is mainly used for the characterization of the virus. 30-Mar-18 70Biochemistry for medics
  68. 68. Laboratory Diagnosis of HIV infection 5. Alternative to classical tests a) Oral fluid (saliva) HIV tests b) Urine tests 30-Mar-18 71Biochemistry for medics
  69. 69. Treatment of HIV Infection • All treatment for HIV infection and AIDS today focuses on arresting the progression of the disease within the body as measured by T cell counts and tests for viral load. • There are two principal approaches to treatment: immunotherapy and anti-HIV drug treatments 30-Mar-18 72Biochemistry for medics
  70. 70. Immunotherapy Immunotherapy Immunoglobulins Interleukins Altered immune cells 30-Mar-18 Biochemistry for medics 73
  71. 71. Treatment of HIV Infection b) Anti-HIV drug treatment • Treatment with anti-HIV drugs attempts to reduce viral load by blocking new infection in the host cell. • The drugs used target two major enzymes of HIV which are needed for the infection cycle: reverse transcriptase and protease. 30-Mar-18 74Biochemistry for medics
  72. 72. Anti HIV treatment Reverse transcriptase inhibitors Protease inhibitors Combination therapy (HAART) 30-Mar-18 Biochemistry for medics 75
  73. 73. Treatment of HIV Infection 30-Mar-18 76Biochemistry for medics
  74. 74. Prevention Lifestyle modifications 30-Mar-18 Biochemistry for medics 77
  75. 75. Development of vaccine Development of vaccine is fraught with several problems unique to this virus. These include- 1) HIV can mutate rapidly, thus, it is not possible to design antibodies against all antigens. 2) Antibody alone is not sufficient, cell mediated immunity may also be necessary. 3) Virus enters the body not as free virions but also as infected cells, in which the virus or the provirus is protected against antibody or cell mediated lysis. 4) Virus readily establishes life long latent infection hiding from antibodies. 30-Mar-18 78Biochemistry for medics
  76. 76. Failure to develop Vaccine HIV can mutate rapidly Antibody alone is not sufficient Virus readily establishes life long latent infection Virus enters the body not as free virions but also as infected cells 30-Mar-18 Biochemistry for medics 79
  77. 77. Summary • Acquired immunodeficiency syndrome, is a disease caused by Human Immunodeficiency Virus and is characterized by marked Immunosuppression. • The mode of transmission is by sexual contact, blood transfusions, needle prick injury or from mother to child. • About 5-10 % percent of the infected appear to escape clinical AIDS for 15 years or more. They have been ‘long term survivors” or “ long term non progressors”. 30-Mar-18 80Biochemistry for medics
  78. 78. Summary • ELISA is the method most commonly employed for the screening purpose and western Blotting is used for the confirmation of the diagnosis. • Treatment is imparted simply to check the progression of the disease, there is no cure of AIDS and there is no vaccine available for prophylaxis of this disease. 30-Mar-18 Biochemistry for medics 81