HIV

This is our presentation slide on HIV presented as a part of seminar in B.Sc, Human Biology

1. 07/20/19
HIV/AIDS
Prepared By:
Akash Adhikari (02)
Bikram Adhikari (03)
Nikita Basnet (04)

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Introduction
•First indication came in summer of 1981, with reports from Newyork and
Los Angeles (USA), of a sudden unexplained outbreak of two very rare
diseases, Kaposi’s sarcoma and Pneumocystis carinii pneumonia in young
adults who were homosexuals or addicted to injected narcotics.
•They appeared to have lost their immune competence, rendering them
vulnerable to overwhelming and fatal infections with relatively avirulent
microorganisms, as well as to lymphoid and other malignacies named
AIDS.

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Introduction
•In 1983, Luc Montagnier and colleagues from Pasteur Institute, Paris,
isolated a retrovirus form a West African patient with persistent
generalized lymphadenopathy, which is manifestation of AIDS, and called
it lymphadenopathy associated virus(LAV).
•In 1984, Robert Gallo and colleagues from NIH, USA, reported isolation
of retrovirus from AIDS patients and called it human T cell lymphotrophic
virus- III or HTLV-III.
•International Committee on Virus nomenclature in 1986 decided the
generic name human immunodeficiency virus (HIV).
•In 1985, serological tests(ELISA) became available for detection of anti-
HIV antibodies.

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WHO report on HIV
•HIV continues to be a major global public health issue, having claimed
more than 34 million lives so far. In 2014, 1.2 [1.0–1.5] million people
died from HIV-related causes globally.
•There were approximately 36.9 [34.3–41.4] million people living with HIV
at the end of 2014 with 2.0 [1.9–2.2] million people becoming newly
infected with HIV in 2014 globally.
•Sub-Saharan Africa is the most affected region, with 25.8 [24.0–28.7]
million people living with HIV in 2014. Also sub-Saharan Africa accounts
for almost 70% of the global total of new HIV infections.
•In 2014, 14.9 million people living with HIV were receiving antiretroviral
therapy (ART) globally, of which 13.5 million were receiving ART in low-
and middle-income countries. The 14.9 million people on ART represent
40% [37–45%] of people living with HIV globally.

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Prevalence of HIV in Nepal
First seen in Nepal in 1988

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Acquired Immunodeficiency Syndrome (AIDS)
•Disease of the human immune system caused by the HIV
•Infection by HIV progressively reduces the effectiveness of the immune
system and leaves individuals susceptible to opportunistic infections and
tumors.
•AIDS is a condition that appears as the last stage in HIV infection
•HIV person is diagnosed as having AIDS, if he/ she has:
•any AIDS defining condition, or
•CD4 count less than 200 cells/ mm3

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Acquired Immunodeficiency Syndrome
(AIDS)
•Some AIDS defining conditions:
•Candidiasis
•Cytomegaloviral disease
•Kaposi’s sarcoma
•Mycobacterium avium complex
•Pneumocystis jiroveci Pneumonia
•Recurrent pneumonia
•Pulmonary Tuberculosis
•invasive Cervical Cancer
•Wasting syndrome etc

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Causative agent: HIV
•HIV: Human Immunodeficiency Virus
Classification:
•Group: VI (ssRNA-RT)
•Family: Retroviridae (Retrovirus)
•Subfamily: Orthoretrovirinae
•Genus: Lentivirus
Picture:
Scanning electron micrograph of HIV-1 budding (in green) from cultured
lymphocyte. This image has been colored to highlight important features;
from the original black and white view of this image. Multiple round bumps
on cell surface represent sites of assembly and budding of virions.

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Scanning electron micrograph of HIV-1 budding from cultured lymphocyte
Multiple round bumps on cell surface represent sites of assembly and budding of virions.

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Morphology: HIV
•Shape:
•Spherical enveloped virus
•about 90- 120 nm in size
•Envelop:
•Outer envelop of lipoprotein (lipid & viral protein), with spikes on the surface
•Core:
•Central icosahedral core of viral capsid proteins surround the genome
•HIV virion core looks like a truncated cone
•Genome:
•consists of 2 identical copies of single stranded positive sense RNA
•in association with viral RNA is the reverse transcriptase (DNA dependent RNA
polymerase) enzyme
•Integrase enzyme and protease are also present

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2 strains of HIV
•HIV-1
•The original isolates of HIV & the related strains prevalent all over the
world belong to HIV type-1
•HIV-2
•HIV strains first isolated from West Africa in 1986, which react with
HIV type 1 antiserum very weakly or not at all have been termed as
HIV-2
•Less virulentrarely causes full blown AIDS probably does not spread
as widely & as rapidly as HIV-1

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Viral genes & antigens
•Genome of HIV contains three structural genes
•gag
•pol
•env
•Product of these genes, both structural and nonstructural, act as
antigens.
•Sera of infected persons contain antibodies to them
•Detection of these antigens and antibodies is useful in the diagnosis and
prognosis of HIV infection.

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Viral genes & antigens
•Genes coding for structural proteins
•Gag gene determines the core and shell of the virus and is expressed as a
precursor protein, p7, p17 & p24 which make up the viral core and shell.
•The major core antigen is p24 which can be detected in serum during the
early stages of HIV infection before antibodies appear.
•In late stage there is decline of anti-p24 antibody and reappearance of
p24 antigen in circulation point to exacerbation of illness.

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Viral genes and antigens
•Env determines the synthesis of envelope glycoprotein gp160, which is
cleaved into the two envelope components: gp120 which forms surface
spikes and gp41 which is the transmembrane anchoring protein.
•Spike glycoprotein gp120 is the major envelope antigen and antibodies
to gp120 are present till the terminal stage of illness.
•Pol gene codes for reverse transcriptase, protease, integrase and
ribonuclease.

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Viral genes and antigens
•Nonstructural and regulatory genes:
•tat (trans activating gene) enhancing the expression of all viral genes.
•net (negative factor gene) down regulating viral replication.
•rev (regulator of virus gene) enhancing expression of structural proteins.
•vif (viral infectivity factor gene) influencing infectivity of viral particles.
•vpu (only in HIV-I) and vpx (only in HIV-II) enhances 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-1 and 2)
•vpr stimulating the promotor region of the virus.
•LTR (long terminal repeat) sequences, one at either end, containing sequences giving
promotor, enhancer and integration signals.

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Viral genes and antigens

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Viral genes and antigens

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Resistance
•HIV is thermolabile, being inactivated in 10 mins at 60ºC and in seconds
at 100ºC.
•At room temperature (20-25ºC) in dried blood it may survive for upto 7
days.
•It withstands lyophilisation.
•Is inactivated in 10 mins by treatment with 50% ethanol, 35%
isopropanol, 0.5% paraformaldehyde, 0.3% hydrogen peroxide.
•Standard recommendation is a hypochlorite solution at a conc. of 0.5%
available chlorine.
•For treatment of contaminated medical instruments, a 2% solution of
glutaraldehyde is useful.

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Life cycle of HIV

21. Modes of Transmission

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Through Bodily Fluids
•Blood products
•Semen
•Vaginal fluids
•Cervical Secretions
•Breast milk

23. 23
Sexual Transmission
•Unprotected Intercourse
•Oral
•Anal
•vaginal
•Probability of transmission per sexual
intercourse:0.0003-0.0015
•Higher Risk in those:
•Females
•Having multiple sexual partners
•Practicising anal intercourse

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Parenteral Transmission
•Transfusion of Blood products:
•Whole Blood
•Plasma
•Cellular fraction of Blood
•Clotting Factors
•Organ transplantation
•Sharing of needles by i.v. drug
users
•Transmission to health worker :
rare with occurrence of <1%
Note : Window period: risk of HIV transmission via this route

25. Perinatal Transmission
•Infected mother have 15-40%
chance of transmitting disease
•During Birth: Transplacentally
•After Birth: Breast Feeding

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How is HIV not transmitted?
•Bites of insects

27. 27
Mode of transmission in World wide

28. Clinical Stages of
HIV/AIDS

29. Clinical Stages of HIV/AIDS
The clinical picture of HIV infection has been divided into 5 groups:
•Group I: Acute HIV infection
•Group II: Asymptomatic or latent infection
•Group III: Persistent generalized lymphadenopathy (PGL)
•Group IV: AIDS related complex (ARC)
•Group V: AIDS

30. Group I: Acute HIV infection
•Within 3-6 weeks of infection with HIV, approx. 50% experience:
•Low grade fever,
•Malaise,
•Headache,
•Lymphadenopathy,
•Sometimes Rash and artharlgia
•Rarely Acute Encephalopathy
•Spontaneous resolution occurs within weeks
•Infected individual can transmit disease

31. Acute HIV infection
•Tests for HIV Ab :
•Negative  At onset of the illness
•Positive During its course.
•Hence, this syndrome is called seroconversion illness
•HIV antigenemia (p24 antigen) can be demonstrated at the beginning of
this phase.
•Pathogenesis: Immune complexes + viral multiplication
Note ::::::primary infection:::::........Window Period.....::::::::::::detectable Ab::::::

32. 32
Group II: Asymptomatic or latent infection
•Clinical Latency may lasts up to several years(approx 10yrs.)
•They show positive HIV antibody tests during this phase and are
infectious
•This period of clinical latency however does not mean microbiological
latency as virus multiplication goes on throughout

33. 33
Group III: Persistent generalized
lymphadenopathy
•Defined as the presence of enlarged lymph nodes, at least 1 cm in
diameter, in two or more extrainguinal sites, that persist for at least
three months, in the absence of any current illness or medication that
may cause lymphadenopathy
•This by itself is benign but the cases may progress to ARC or AIDS.

34. Group IV: AIDS related complex
•The typical constitutional symptoms are :
•Fatigue,
•Unexplained fever,
•Persistent diarrhea
•Marked weight loss or more than 10% of body weight
•Generalized lymphadenopathy and splenomegaly are usually present.
•ARC patients  severely ill + may progress to AIDS

35. 35
Group IV: AIDS related complex
•Opportunistic Infection Associated to HIV/AIDS
•It do not occur until CD4 T cell counts have dropped from 1000 cells/uL
to less than 200 cells/uL.
•Common:
•Candidiasis
•Herpes zoster
•Hairy cell leucoplakia
•Salmonellosis
•Tuberculosis

36. 36
Group V: AIDS
•AIDS is the end-stage disease representing the irreversible breakdown
of immune defense mechanism, leaving the patient prey to
progressive opportunistic infection and malignancies
•The median time period is 10 years from primary HIV infection to
Development of AIDS
•About 5-10% HIV infected patients escape AIDS for 15 years

37. Clinical Manifestation of AIDS
•Common: Dry cough, Dsypnea, fever, recurrent pneumonia
•GIT: Thrush, gingivitis, Dysphagia, Intestinal infection, chronic colitis
•CNS: Lymphoma of CNS, Toxoplasmosis, cryptococcosis
•Cutaneous: Herpes lesions, Candidiasis, xeroderma, impetigo etc.
•Malignancies: Kaposi’s Sarcoma, lymphomas
•Dementia
•Pediatric AIDS

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Opportunistic Infections
Organisms
Fungi Candida albicans, Cryptococcus neoformans, Coccidioides immitis, Histoplasma
capsulatum, Pneumocystis jiroveci.
Bacteria Mycobacterium avium-intracellulare, Mycobacterium tuberculosis, Listeria
monocytogenes, Nocardia asteroides, Salmonella sps, Streptococcus sps
Viruses Cytomegalovirus, herpes simplex virus, varicella-zoster virus, adenovirus,
polyomavirus, JC virus, hepatitis B virus, hepatitis C virus
Protozoa Toxoplasma gondii, Isospora belli, Cryptosporidium sps

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CD4<500
•Tuberculosis (TB)
•Herpes Simplex
•Pneumococcal pneumonia
•Herpes Zoster
•Vaginal candidiasis
•Hairy leukoplakia
•Kaposi’s sarcoma
Opportunistic Infections associated with
AIDS
Extensive tumor lesions of Kaposis’s sarcoma in AIDS
patient.
Source: AIDS, 1997
Chronic Herpes Simplex infection with lesions on tongue and lips.
Source: Atlas of Clinical Oral Pathology, 1999.
oral Hairy Leukoplakia

40. CD4<200
•Pneumocystic carinii
•Toxoplasmosis
•Cryptococcosis
•Coccidiodomycosis
•Cryptosporiosis
•Non hodgkin’s lymphoma
Opportunistic Infections associated with
AIDS
Non-Hodgkin’s Lymphoma & ascites in AIDS patient
Source: Tropical Medicine and Parasitiology, 1997

41. Opportunistic Infections associated with
AIDS
CD4 <50
•Disseminated mycobacterium avium complex (MAC) infection
•Histoplasmosis
•CMV retinitis
•CNS lymphoma
•Progressive multifocal leukoencephalopathy
•HIV dementia

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Pattern of opportunistic infections
associated with declining CD4+ cell
counts.

43. Time course of HIV infection

44. LABORATORY DIAGNOSIS OF HIV

45. LABORATORY DIAGNOSIS OF HIV
A. NON-SPECIFIC TESTS
i.Blood test
- Blood count
- T-cell subset assay
- Hypergammaglobulinaemia
ii. Cell mediated immunity
B. SPECIFIC TESTS
i.Direct method
i.Indirect method

46. NON-SPECIFIC TESTS
i.Blood tests
•Blood count:
- Leucopenia, low lymphocyte count (<400/mm ), thrombocytopenia
ᶟ
•T-cell subset assay
- CD4+ lymphocytes < 200/mm
ᶟ
- Normal CD4: CD8 (2:1) ratio is reversed to 0.5-1: 1
•Hypergammaglobulinaemia
- IgG and IgA increases
ii. Cell mediated immunity
•Diminished CMI indicated by skin tests

47. SPECIFIC TESTS
i. DIRECT METHOD

ii. INDIRECT METHOD

48. DIRECT METHOD
a.Viral culture/Virus Isolation
•Can be isolated from the peripheral lymphocytes
•Isolated by cocultivation of the patient’s lymphocytes with uninfected
lymphocytes in the presence of IL-2
•Viral replication can be detected by the demonstration of reverse
transcriptase activity as well as antigens
•Not suitable as a routine diagnostic test
•Risky so performed only in laboratories with adqeuate containment
facilities

49. DIRECT METHOD
b.P24 Ag detection
•Earliest virus marker
•Can be detected before HIV antibody in newly infected individuals
•IgM antibodies appear in about 4-6 weeks, to be followed by IgG antibodies
•p24 Ag disappears for long time and is again seen in several clinical disease
•p24 Ag capture assay (ELISA) uses anti-p24 Ab
•The test is positive in about 30% of HIV infected persons

50. DIRECT METHOD
c.Polymerase Chain Reaction (PCR)
•The most sensitive and specific test in all stages of HIV infection
•DNA PCR and RNA PCR are two forms of PCR used
•In DNA PCR, peripheral lymphocytes from the subject are lysed and the
proviral DNA amplified
•RNA PCR can be used for diagnosis as well as monitoring the level of viremia
•Complex and costly
•Indicated only when other methods cannot give a definitive result

51. INDIRECT METHOD
Enzyme Linked Immnosorbant Assay (ELISA)

•HIV Ag is attached to a well of microtiter plate
•Test serum added, antibody present binds to the
coated Ag
•Washing away unbound serum
•Antihuman immunoglobulin linked to a suitable
enzyme is added
•Washed unbound serum
•Addition of appropriate colour substrate
•Detected by spectrophotometer

52. INDIRECT METHOD
Rapid test
•Screening test that produces results within 60 mins or less
•Is the method of choice when immediate information is necessary to
determine the need for prophylaxis as labor/delivery, newborn, etc
•Confirmatory test must be performed

53. INDIRECT METHOD
Western blot test

54. INDIRECT METHOD
Western blot test
•HIV proteins separated according to their electrophoretic mobility by
polyacrylamide gel electrophoresis
•are blotted onto strips of nitrocellulose paper
•These strips are reacted with test sera and then with enzyme conjugated
antihuman globulin
•A suitable substrate is then added which produces a prominent colour band in
reaction area
•Position of the bands on the strip indicate patterns of reactivity which can be
visibly read

55. INDIRECT METHOD
Western blot test
•Three major viral bands:
- p24 (gag gene, core proteins)
- p31 (pol gene, reverse transcriptase)
- gp41, gp120/160 (env gene, surface antigens)
•All three genes gag, pol & env are present in
positive HIV infection
•No detectable viral bands in nonreactive western
blot

56. INDIRECT METHOD
Immunofluorescent Antibody Assay
•Infected cells fixed to a microscope slide
•Patient’s serum containing HIV Abs added
•Slide is washed
•Addition of antiimmunoglobulin Abs with fluorescent label attached
•Reaction visualized using fluorescent microscope

57. TREATMENT
HAART

58. PREVENTION

60. REFERENCES
•P Chakraborty. (2010). A Text Book of Microbiology. Delhi: New
Central Book Agency (P) Ltd.
•R Anantanarayan & C K J Panikar, (2009). Textbook of
microbiology. Manipal Press Ltd.
•Richard A. Harvey. Pamela C. Champe. Lippincott's Illustrated Review
Microbiology.2nd edition,2007.Lippincott Williams and Wilkins
•http://www.hivguidelines.sorg/clinical-
guidelines/adults/diagnostic-monitoring-and-resistance-
laboratory-tests-for-hiv/

61. REFERENCES
•P Chakraborty. (2010). A Text Book of Microbiology. Delhi: New Central
Book Agency (P) Ltd.
•R Anantanarayan & C K J Panikar, (2009). Textbook of microbiology.
Manipal Press Ltd.
•Richard A. Harvey. Pamela C. Champe. Lippincott's Illustrated Review
Microbiology.2nd edition,2007.Lippincott Williams and Wilkins

62. THANK YOU !!