MICR 420

1. MICR 420 Emerging and Re-Emerging Infectious Diseases Lecture 11: HIV Dr. Nancy McQueen & Dr. Edith Porter

2. Overview • Brief history • Nomenclature • Morphology and nature of the genome • Viral replication cycle • Pathogenesis and clinical symptoms • Diagnosis • Treatment • Prevention • Threat

3. Brief History • Late 1970’s physicians in L.A., San Francisco, and New York observe clusters of patients with similar symptoms • Severe fungal pneumonia (Pneumocystis jirovecii – formerly P. carinii) • Sudden weight loss • History of recurrent infections • Bluish-purple spots on the skin (Kaposi’s sarcoma) - caused by human herpesvirus type 8 • All of the patients were • Young • Male • Homosexual

4. Pneumocystis jirovecii DIF of P. jirovecii

5. Kaposi’s sarcoma

6. Brief History • The term AIDS (Acquired Immune Deficiency Syndrome) was first coined in 1981 and used in a CDC report in 1982: • A condition characterized by a marked depletion of CD4+ T lymphocytes • Opportunistic infections • Due to a transmissible agent acquired though • Sexual contact • Blood exchange • In 1982-83, a retrovirus was isolated from the blood of individuals by groups in Paris, France, and Maryland, US, lead by Luc Montagnier and Robert Gallo, respectively. • The virus, subsequently named human immunodeficiency virus - type I (HIV-1), was demonstrated to be the cause of AIDS.

7. Nomenclature • There are two types of HIV • HIV-1 - is the most common cause of AIDS worldwide • HIV-2 - is the most common cause of AIDS in West Africa and is rarely found outside this region • Both types have many subtypes (clades) • The infection caused by HIV-2 is less severe than that caused by HIV-1.

9. Worldwide prevalence of HIV infection

10. Where different HIV clades are found (HIV1)

11. 2009

12. Where did AIDS come from? • There is no definitive answer to this question • Evidence indicates HIV-1 and HIV-2 arose from different evolutionary lines • HIV-2 appears to have arisen from a mutation in a simian immunodeficiency virus (SIV) of mangabey monkeys in West Africa • HIV-1 is genetically similar to SIV carried by chimpanzees in Central Africa • The chimpanzee virus appears to be a hybrid of two mangabey SIVs.

13. Where did AIDS come from? • It has been suggested that HIV-1 jumped the species barrier in small villages around 1930 by • Humans eating infected monkeys • Humans being bitten by infected monkeys • The end of European colonialism and the subsequent disruption of the social structure of sub-Saharan Africa resulted in • Urbanization • Increased prostitution • Growth of highway transportation • Increased sexual promiscuity • Spread of the disease

14. Classification of HIV • HIV belongs to the family Retroviridae, sub-family Lentiviridae • Retroviruses • The name comes from “backward” nucleic acid synthesis • Retroviruses convert genomic viral (+)RNA to dsDNA that then integrates into the host cell DNA (provirus) • Use RT (reverse transcriptase), RNA-dependent, DNA polymerase for making DNA from an RNA template

15. Morphology and Nature of the Genome • Enveloped with glycoprotein spikes • Icosahedral • 2 identical strands of + sense single stranded RNA • Although the RNA is of the positive sense, expression of the genome requires that the virus is copied into cDNA and that the cDNA is integrated into the host cell DNA • Genome complexed with RT, integrase and protease gp 120 gp 41

16. Replication cycle of HIV Fusion at the plasma membrane Budding from plasma membrane Note that reverse transcription takes place inside a preintegration complex in the cytoplasm mRNA synthesis and genomic RNAsynthesis take place inside nucleus using host cell enzymes

17. HIV Attachment is Mediated by gp120 and fusion by gp41 Fusion peptide gp120 gp41 CD4 - receptor CXCR4 (lymphocytes) or CCR5 (macrophages) co-receptor

18. HIV Attachment • Requires cleavage of gp160 into gp120 and gp41 • Attachment to CD4 receptor • Attachment to chemokine co-receptors • receptors of chemical messages involved in cellular communication • Individual HIV strains are classified as being either lymphotropic or macrophageotropic based on which chemokine co-receptor the virus recognizes. • Rare individuals who are resistant to HIV • may have genetic defects (32-nucleotide deletion) in their R5 chemokine receptors • 1% of whites of European descent are homozygous • 20% are heterozygous • Protection from plague (Yersinia pestis)? • Defect only protects against HIV-1, subtype B • transmitted sexually • prevalent in the U.S. and Europe.

19. HIV Penetration by Fusion

20. HIV - Latent versus Productive Infection • After integration of proviral DNA into the host cell genome, there are two possible outcomes • Productive infection in which new viruses are made and released • Latent infection in which no new virus is made and released • do not contribute to disease • represent a significant hurdle to treatment and virus eradication • not recognized by the immune system because they do not express viral proteins • Latently infected T cells and macrophages can start producing virus following • Activation of T cells via the T cell receptor • Virus infections • Treatment with cytokines

21. HIV Infection of T cells

22. HIV Infection of Macrophages

23. An infected macrophage. Viral particles are within the vacuoles.

24. How do you get AIDS? • Sexual contact • Homosexual • Heterosexual • Anal sex causes more damage with a greater chance of transmission • STIs increase infectivity of HIV • Transfusions with infected blood or blood products (hemophiliacs) • Sharing blood contaminated needles and syringes • Mother to child • Placental • During birth • Through infected breast milk

25. Transmission Modes

27. Male versus Female Transmission Modes

28. Progression of HIV Infection • HIV-1 strains are sub-classified into CCR5-requiring (R5) and CXCR4-requiring (X4) depending upon their co-receptor requirements for infection. • A single point mutation can change an R5 virus into an X4 virus • R5 viruses - the actual transmitters of infection • Infect macrophages, monocytes, and dendritic cells • X4 variants are generated via reverse transcriptase mistakes made during the RT activity of the virus in the macrophage • Appearance of X4 viruses is associated with disease progression • Infect T cells in lymph nodes and other tissues • Both R5 and X4 can infect T4 cells circulating in the blood

29. Progression of HIV Infection • The initial target of HIV infection at mucosal surfaces appears to be dendritic cells containing a DC-SIGN receptor (a lectin). • dendritic cell migrates to a lymph node • transfer of virus to naïve CD4+ T cells. • Systemic spread occurs very early in the infection.

30. HIV’s Favorite Cellular Targets Infection of several types of brain cells probably contributes to lethargy and HIV dementia – Crosses in macrophages Infection of gut intestinal epithelium and lymphoid tissue creating memory cells carrying CCR5+ and CD4+ receptors Dendritic cells- Are relatives of macrophages that reside throughout the tissues and are responsible for processing foreign matter and presenting it to lymphocytes CD4+ macrophages, dendritic and other antigen-presenting cells harbor HIV, but are not usually killed by it. The cells are a continuing source of the virus and can carry it to the brain Naïve CD4+ inflammatory cells are normally responsible for macrophage activation and cell-mediated immunity through CD8+ cytotoxic T cell activation Naïve CD4+ helper cells that normally control antibody production by B cells. Some of these become CCR5+ CD4+ memory cells

31. Stages of HIV Infection • The CDC has classified HIV infection into four stages • Prodromal stage (primary, acute infection) • Latency period (asymptomatic stage) • Persistent generalized lymphadenopathy (early and medium stage HIV symptomatic disease) • Full blown AIDS (Late-stage HIV infection)

32. Prodromal stage (primary, acute infection)of HIV Infection • Characterized by fever, rash, diarrhea, aches, headaches, lymphadenopathy, and fatigue (flu-like symptoms) • The CD4+ cell count drops, but not enough to impair immune function • HIV actively replicates and releases new viruses into the bloodstream so the viral load is high • Seroconversion and appearance of anti-HIV antibodies occurs at the end of this stage

33. Latency period (asymptomatic stage) of HIV infection • No symptoms or only swollen lymph nodes • Virus is hiding in inactive T cells, including T memory cells and in macrophages in lymphoid tissues • A major reservoir for HIV during latent infection is the lymphatic system. • Antibody against HIV can be found • CD4+ cells increase, but do not reach pre-HIV infected level • Viral load in the blood decreases and stabilizes

34. Persistent Generalized Lymphadenopathy (early and medium stage HIV symptomatic disease) • Swollen lymph nodes, recurrent fevers, skin rashes, night sweats, persistent diarrhea, persistent cough, and extreme fatigue occur • Neurological symptoms of memory loss, confusion, and depression may occur. • Opportunistic infections, including oral and vaginal candidiasis (thrush) and herpesvirus outbreaks occur • CD4+ count decreases • Viral load increases

35. Full blown AIDS (Late-stage HIV infection) • CD4+ T cell count decreases to 200/mm3 or less • Lymph nodes degenerate • Severe, recurring opportunistic infections

38. HIV Disease Progression Rates • How quickly the disease progresses varies from individual to individual • Progression rate is determined by • The nature of the immune response • Slower progression occurs in those who develop a good CD8 T-cell response or those who recognize a diversity of epitopes • Level of basal RNA during latent stage • HIV subtype • Host genetic susceptibility • Co-infections - may enhance viral replication

39. Pathogenesis- Contributions to Clinical Immunodeficiency • Lysis of CD4+ T cells caused by • Virus releases gp120 that binds to CD4 receptors and cells become targets for destruction • gp120 cross-linking of CD4 receptors priming cells for apoptosis • down regulation of MHC class I molecules cause NK cells to destroy the infected cell • syncytium formation (giant, multinucleated cells formed by the fusion of the membranes of adjacent cells)

40. Syncytia formation

41. Pathogenesis- Contributions to clinical immunodeficiency • Functional impairment of immune system • Soluble gp120 blocks interaction of CD4+ T cells with Class II MHC on antigen presenting cells (APCs) • Endogenous gp120 prevents CD4 from being transported to the cell surface. • Nef and Vpu viral proteins downregulate CD4 • Impaired macrophage and natural killer cell function • Destruction of architecture of lymph nodes • Central nervous system damage • Release of inflammatory cytokines from HIV-infected CNS macrophages • Soluble gp120 may interfere with neurotransmitter action on neurons

42. Diagnosis of HIV Infection • ELISA – used for first screening • Western blot – used to confirm a positive ELISA result • RT-PCR – used for following viral load (serum viral concentration) during treatment • Detects the presence of viral RNA by making a cDNA copy of the RNA and amplifiying that copy by PCR. • Can also be used to detect specific resistant variants

43. Treatment of HIV infection • All anti-HIV drugs have numerous side effects • Common side effects include nausea, vomiting, headache, fatigue, weakness, and/or muscle pain. • Other side effects included changes in body fat distribution (lipodystrophy), inflammation, insomnia, and kidney disorders. • Reverse transcriptase inhibitors • Nucleoside analogues such as AZT • Prevent successful synthesis of cDNA by RT • act as chain terminators • best application of AZT is use during pregnancy • reduces the risk of HIV being passed from mother to child. • Non-nucleoside analogue RT inhibitors • Bind non-competitively to RT to block its polymerization function • Nucleotide analogues • Work similar to the nucleoside analogues • Protease inhibitors • Prevent cleavage of polyproteins (by viral protease) required to make mature virus

44. Examples for Lipodystrophy in AIDS Patients

45. Treatment of HIV infection • Fusion inhibitors • Integrase inhibitors • Combination therapy – helps prevent the development of resistant strains • HAART - highly active anti-retroviral therapy (3 or more different classes of drugs in combination) • serious long-term side effects • accumulation of lactic acid in the bloodstream • physical and metabolic changes that cause changes in fat distribution • cholesterol and glucose abnormalities that can lead to a risk of heart disease. • Long-term use of the drugs can also promote the development of drug-resistant strains of HIV.

46. HIV Anti-Viral Therapies

47. Nucleoside Analogues

48. Protease Inhibiters

49. Protease Inhibiters • This picture shows the HIV protease (purple and green) complexed with the inhibitor (spacefill). This prevents the normal viral substrate from reacting with the protease and thus, the viral polypeptides are not cleaved.

50. Antiviral Therapy - in the future? • Antisense RNA