HIV/AIDS

HIV/AIDS
Dr. Suprakash Das
Assist. Prof.

Introduction
 Infection with human immunodeficiency virus type 1 (HIV- 1) is prevalent
throughout the world and is characterized by a progressive deterioration of the
immune system that is usually fatal if untreated.
 AIDS was first recognized as a clinical entity in 1981.
 The syndrome was identified by clusters of unusual diseases including Kaposi’s
sarcoma and Pneumocystis jiroveci pneumonia in young homosexual men who
had immunodeficiency due to depletion of CD4+, helper T cells.
 AIDS cases were subsequently reported in intravenous drug users (IVDU),
hemophiliacs, and in infants born to mothers with AIDS, suggesting a blood-borne
as well as sexually-transmitted pathogen.
 In 1983, HIV-1 was isolated and this novel human retrovirus was proposed as the
cause of AIDS.
 In 1985, a sensitive enzyme-linked immunosorbent assay (ELISA) was
developed.

Introduction
 The current CDC classification system for HIV-infected adolescents and adults
categorizes persons on the basis of clinical conditions associated with HIV infection and
CD4+ T-lymphocyte counts.
 The system is based on three ranges of CD4+ T-lymphocyte counts and three clinical
categories and is represented by a matrix of nine mutually exclusive categories.
 Any HIV-infected individual with a CD4+ T-cell count of <200/μL has AIDS by
definition, regardless of the presence of symptoms or opportunistic diseases.
 Once individuals have had a clinical condition in category B, their disease
classification cannot be reverted back to category A, even if the condition resolves;
the same holds true for category C in relation to category B.
 The definition of AIDS is indeed complex and comprehensive and basically for
surveillance purposes.
 HIV disease has spectrum ranging from primary infection, with or without the acute
syndrome, to the asymptomatic stage, to advanced disease.

Virology
 HIV family (Retroviridae) subfamily Lentiviruses.
 The four recognized human retroviruses: The human T-lymphotropic viruses (HTLV)-I
and HTLV-II, which are transforming retroviruses
 The human immunodeficiency viruses, HIV- 1 and HIV-2 cause cytopathic effects either
directly or indirectly.
 The most common cause of HIV disease throughout the world is HIV-1 Several
subtypes in geographic distributions.
 HIV-2 was first identified in 1986 in West Africa.
 The Pan troglodytes troglodytes species of chimpanzees has been established as the
natural reservoir of HIV-1.
 HIV-2 is more closely related to the simian immunodeficiency virus (SIV) found in sooty
mangabeys.

Sooty mangabeys
Pan troglodytes troglodytes species of chimpanzees

Virology
 HIV virion (100 to 150 nm) Icosahedral structure containing numerous external
spikes formed by the two major envelope proteins, the external gp120 and the
transmembrane gp41.
 The virion buds from the surface of the infected cell and incorporates a variety of host
proteins, including MHC class I and II antigens.
 Mature viral particles are characterized by an electron-dense, conical core.
 The core is surrounded by a lipid envelope that is acquired as the virion buds from the
infected cell.
 The virion core contains two copies of single-stranded, positive sense, genomic RNA,
each of which encodes the complete viral repertoire of structural, enzymatic, and
regulatory proteins.
 HIV genome is approximately 10kb in length.

Virology
 The 5/ LTR contains the enhancer/promoter sequences for viral transcription
 The 3/ LTR contains the polyadenylation signal.
 From 5/ to 3/ the viral genome contains the gag gene, which encodes the virion
structural components;
 the pol gene, which encodes the viral enzymes; and
 the env gene, which encodes the envelope glycoproteins.
 The primate immunodeficiency viral genomes contain six additional genes: vif,
vpr, tat, rev, nef, and
 vpx in the case of HIV-2 and SIV, or
 vpu in the case of HIV-1.
 The products of these genes function via interactions with host cell proteins to
optimize viral replication by a variety of mechanisms.

HIV genes and gene products
Gene Protein Size Function
S
T
R
U
C
T
U
R
A
L
G
E
N
E
S
gag Matrix (MA) p17 Structural protein (recruits envelope glycoprotein)
Capsid (CA) p24 Structural protein (forms conical core)
Nucleocapsid (NC) p7 Binds viral RNA to encapsidate genome
p6 p6 Budding
pol Protease (PR) P12 Viral enzyme: cleavage of polyprotein precursors
Reverse transcriptase
(RT)
P66/P
51
Viral enzyme: reverse transcription and RNase H
activities
Integrase (IN) P32 Viral enzyme: integration of viral cDNA into host
chromosomes
env Surface glycoprotein
(SU)
gp 120 Viral envelope glycoprotein: receptor binding
Transmembrane (TM) gp 41 Viral envelope glycoprotein: fusion

HIV genes and gene products
Gene Protein Size Function
R
E
G
U
L
A
T
O
R
Y
G
E
N
E
S
tat Tat P14 Transactivates viral transcription
rev Rev P19 Transports unspliced mRNA to cytoplasm
vif Vif p24 Promotes virion infectivity by degrading cellular cytidine
deaminases in the APOBEC3 family.
nef Nef P27 Down-regulates class I MHC and CD4; enhances viral
infectivity; facilitates T-cell activation
vpu Vpu P16 Promotes release of viral particles by counteracting
BST2/tetherin; induces degradation of CD4
vpr Vpr p15 Arrests cell cycle in G2/M; facilitates viral replication
in macrophages
vpx Vpx p14 Facilitates viral replication in dendritic cells and
macrophages by degrading the cellular enzyme SAMHD1

REPLICATION CYCLE OF HIV
 HIV is an RNA virus whose hallmark is the reverse transcription of its
genomic RNA to DNA by the enzyme reverse transcriptase.
 The replication cycle of HIV begins with the high-affinity binding of the
gp120 protein to its receptor on the host cell surface, the CD4 molecule.
 The CD4 molecule is found predominantly on a subset of T lymphocytes
that are responsible for helper function in the immune system.
 It is also expressed on the surface of monocytes/ macrophages and
dendritic/Langerhans cells.
 Once gp120 binds to CD4, the gp120 undergoes a conformational change
that facilitates binding to one of a group of co-receptors.
 The two major co-receptors for HIV-1 are CCR5 and CXCR4 An
important determinant of the cellular tropism of the virus.

 Certain dendritic cells express DC-SIGN (C-type lectin receptors) on their
surface that also bind with high affinity to the HIV gp120 envelope protein,
allowing the dendritic cell to facilitate the binding of virus to the CD4+
T cell upon engagement of dendritic cells with CD4+ T cells.
 Binding of GP120 to the CD4 molecule Conformational change in the
viral envelope gp120 Fusion with the host cell membrane occurs via the
newly exposed gp41 molecule Penetrating the plasma membrane of the
target cell.
 After fusion, the pre-integration complex, composed of viral RNA and
viral enzymes and surrounded by a capsid protein coat, is released into
the cytoplasm of the target cell.

 Pre-integration complex traverses the cytoplasm to reach the nucleus Viral
reverse transcriptase enzyme catalyzes the reverse transcription of the
genomic RNA into DNA Protein coat opens to release the resulting double-
stranded HIV-DNA.
 At this point in the replication cycle, the viral genome is vulnerable to cellular
factors that can block the progression of infection (e.g, Clyclophilin A &
APOBEC family of cellular proteins).
 Activation of the cell The viral DNA accesses the nuclear pore Exported
from the cytoplasm to the nucleus Integrated into the host cell chromosomes
through the action of another virally encoded enzyme, integrase.
 HIV provirus (DNA) may remain transcriptionally inactive (latent) or it may
manifest varying levels of gene expression, up to active production of virus.

 Cellular activation plays an important role in the replication cycle of HIV and
is critical to the pathogenesis of HIV disease.
 After initial binding and internalization of virions into the target cell,
incompletely reverse-transcribed DNA intermediates are labile in quiescent
cells and do not integrate efficiently into the host cell genome unless cellular
activation occurs shortly after infection.
 Some degree of activation of the host cell is required for the initiation of
transcription of the integrated proviral DNA into either genomic RNA or
mRNA.
 After transcription, HIV mRNA is translated into proteins that undergo
modification through glycosylation, myristylation, phosphorylation, and
cleavage.
 The viral particle is formed by the assembly of HIV proteins, enzymes, and
genomic RNA at the plasma membrane of the cells.

 Budding of the progeny virion occurs through specialized regions in the
lipid bilayer of the host cell membrane known as lipid rafts, where the core
acquires its external envelope.
 The virally encoded protease then catalyzes the cleavage of the gag-pol
precursor to yield the mature virion.
 Progression through the virus replication cycle is profoundly influenced by
a variety of viral regulatory gene products.
 Likewise, each point in the replication cycle of HIV is a real or potential
target for therapeutic intervention.
 Thus far, the reverse transcriptase, protease, and integrase enzymes as
well as the process of virus–target cell binding and fusion have proven
clinically to be susceptible to pharmacologic disruption.

Pathogenesis
 The hallmark of HIV disease is a profound immunodeficiency resulting
primarily from a progressive quantitative and qualitative deficiency of the
subset of T lymphocytes referred to as helper T cells.
 This subset of T cells is defined phenotypically by the presence on its surface of
the CD4 molecule, which serves as the primary cellular receptor for HIV.
 Co-receptors (CCR5 and CXCR4) must also be present together with CD4 for
efficient fusion and entry of HIV-1 into its target cells.
Mechanisms of cellular depletion and/or immune dysfunction of CD4+ T cells
Direct infection and destruction of these cells by HIV
Immune clearance of infected cells
Indirect effects such as immune exhaustion due to aberrant cellular
activation and activation-induced cell death.

Pathogenesis
 Patients with CD4+ T-cell levels below certain thresholds are at high risk of
developing a variety of opportunistic diseases, particularly the infections and
neoplasms that are AIDS-defining illnesses.
 Some features of AIDS, such as Kaposi’s sarcoma and neurologic abnormalities,
cannot be explained completely by the immunodeficiency caused by HIV
infection, since these complications may occur before the development of severe
immunologic impairment.
 The combination of viral pathogenic and immunopathogenic events that
occurs during the course of HIV disease from the moment of initial (primary)
infection through the development of advanced-stage disease is complex and
varied.
 It is important to appreciate that the pathogenic mechanisms of HIV disease are
multifactorial and multiphasic and are different at different stages of the disease

Main mechanisms of HIV persistence in latent
reservoirs/ Pathogenesis
 HIV entry at the mucosal site Viral capture via DCs Migrate to lymph nodes
Present HIV antigens to CD4+ T helper cells.
 In this process, DCs may also facilitate the infection of activated CD4+ T cells. CD4+ T
cells ‘help’ CD8+ T cells and B cells in mounting cellular and humoral anti-HIV immune
responses.
 [Mechanisms of HIV immune evasion are listed in the figure. Black text indicates events
of the HIV-specific immune response; red text indicates events leading to HIV immune
evasion and dissemination.]
 Active infection of CD4+ immune cells followed by HIV provirus integration, mainly in
the genome of memory T cells, then by HIV production when cells are activated,
leading to the death of most HIV-producing cells.
 The escape of rare HIV-infected cells; post-integration latency in HIV reservoirs;
 establishment of latency in memory CD4 T cells that are either resting or exhausted
and negatively regulated by immune checkpoints.

Fate of reservoirs in latently infected cells
The long-term resting status allows long-term stable reservoirs, or
The homoeostatic proliferation allows proliferation of reservoirs with scarce HIV
production, or
The antigen-activation induces cell death and reseeding of new HIV reservoirs;
and
In exhausted cells where immune checkpoints block HIV production, thus
inducing persistence of HIV reservoirs.

The main therapeutic targets towards HIV cure include
 Antiretroviral therapy,
 Reversion of latency in resting cells mainly by epigenetic modifiers,
 Immune check points inhibitors restoring HIV production capacity,
and
 Anti-HIV immunity mediated either by cytotoxic T cells or antibodies
induced by therapeutic vaccines or
 Passive transfers of antibodies.

Stages of HIV Infection
Acute HIV Infection
 Acute HIV infection is the earliest stage of HIV infection, and it generally
develops within 2 to 4 weeks after infection with HIV.
 During this time, some people have flu-like symptoms, such as fever, headache,
and rash.
 In the acute stage of infection, HIV multiplies rapidly and spreads throughout the
body.
 The virus attacks and destroys the infection-fighting CD4 cells of the immune
system.
 During the acute HIV infection stage, the level of HIV in the blood is very high,
which greatly increases the risk of HIV transmission.
 A person may experience significant health benefits if they start ART during
this stage.

Chronic HIV Infection
 The second stage of HIV infection is chronic HIV infection (also called
asymptomatic HIV infection or clinical latency).
 During this stage, HIV continues to multiply in the body but at very low levels.
 People with chronic HIV infection may not have any HIV-related symptoms.
 Without ART, chronic HIV infection usually advances to AIDS in 10 years or
longer, though in some people it may advance faster.
 People who are taking ART may be in this stage for several decades.
 While it is still possible to transmit HIV to others during this stage, people who
take ART exactly as prescribed and maintain an undetectable viral load have
effectively no risk of transmitting HIV to an HIV-negative partner through
sex.

AIDS
 AIDS is the final, most severe stage of HIV infection.
 Because HIV has severely damaged the immune system, the body can’t fight
off opportunistic infections.
 Opportunistic infections Infections and infection-related cancers that occur
more frequently or are more severe in people with weakened immune systems than
in people with healthy immune systems.
 People with HIV are diagnosed with AIDS if they have a CD4 count of less than 200
cells/mm3or if they have certain opportunistic infections.
 Once a person is diagnosed with AIDS, they can have a high viral load and are able
to transmit HIV to others very easily.
 Without treatment, people with AIDS typically survive about 3 years.
•

CLINICAL CATEGORIES OF HIV INFECTION
Category A
 Consists of one or more of the conditions listed below in an adolescent or adult (>13 years) with
documented HIV infection.
 Conditions listed in categories B and C must not have occurred.
Asymptomatic HIV infection.
Persistent generalized lymphadenopathy
Acute (primary) HIV infection with accompanying illness or history of acute HIV infection.
Category B
 Consists of symptomatic conditions in an HIV-infected adolescent or adult that are not included
among conditions listed in clinical category C and that meet at least one of the following criteria:
(1) The conditions are attributed to HIV infection or are indicative of a defect in cell-mediated
immunity.
(2) The conditions are considered by physicians to have a clinical course or to require management
that is complicated by HIV infection

Examples include, but are not limited to, the following:
Bacillary angiomatosis
Candidiasis, oropharyngeal (thrush)
Candidiasis, vulvovaginal; persistent, frequent, or poorly responsive to therapy
Cervical dysplasia (moderate or severe)/cervical carcinoma in situ
Constitutional symptoms, such as fever (38.5ºC) or diarrhea lasting >1 month
Hairy leukoplakia, oral
Herpes zoster (shingles), involving at least two distinct episodes or more than one dermatome
Idiopathic thrombocytopenic purpura
Listeriosis
Pelvic inflammatory disease, particularly if complicated by tubo-ovarian abscess
Peripheral neuropathy

Category C (AIDS surveillance case definition.)
Candidiasis of bronchi, trachea, or lungs
Candidiasis, esophageal
Cervical cancer, invasive
Coccidioidomycosis, disseminated or extrapulmonary
Cryptococcosis, extrapulmonary
Cryptosporidiosis, chronic intestinal (>1 month’s duration)
Cytomegalovirus disease (other than liver, spleen, or nodes)
Cytomegalovirus retinitis (with loss of vision)

Encephalopathy, HIV-related
Herpes simplex: chronic ulcer(s) (>1 month’s duration); or bronchitis,
pneumonia, or esophagitis
Histoplasmosis, disseminated or extrapulmonary
Isosporiasis, chronic intestinal (>1 month’s duration)
Kaposi’s sarcoma
Lymphoma, Burkitt’s (or equivalent term)
Lymphoma, primary, of brain
Mycobacterium avium complex or M. kansasii, disseminated or
extrapulmonary

Mycobacterium tuberculosis, any site (pulmonary or extrapulmonary)
Mycobacterium, other species or unidentified species, disseminated or
extrapulmonary
Pneumocystis jiroveci pneumonia
Pneumonia, recurrent
Progressive multifocal leukoencephalopathy
Salmonella septicemia, recurrent
Toxoplasmosis of brain
Wasting syndrome due to HIV

ACUTE HIV INFECTION SYNDROME
 Primary HIV infection should be considered in any patient with possible HIV
exposure who presents with fever of unknown cause.
 Develops 2 to 4 weeks after initial exposure to HIV, although incubation
periods of 6 days to 6 weeks.
 The period of illness associated with acute infection is generally 10 days but
might vary from 3 to 25 days.
Fever
Mylagias
Fatigue
Pharyngitis
Weight loss
Headache.

 The Mononucleosis-like illness (fever, pharyngitis, and adenopathy) might not
always be evident.
 Acute HIV infection might be associated with a myriad of dermatologic
manifestations.
 The typical rash is mononucleosis like, nonpruritic, erythematous, and
maculopapular and distributed on the face and trunk but might involve the
palms and soles or be generalized.
Desquamation of the palms and soles,
Generalized urticaria
Vesiculopustular rash
Erythema multiforme or
Alopecia as part of the presenting complex.

 Oral and genital mucocutaneous ulceration is also typically associated with
acute HIV infection.
 Gastrointestinal (GI) manifestations might dominate the acute viral syndrome,
with abdominal pain, vomiting, and diarrhea being the primary manifestations.
 Severe esophageal ulceration.
 Headache and photophobia.
More unusual nervous system manifestations include
 Acute fulminating encephalopathy,
 Brachial neuritis,
 Myelopathy,
 Peripheral neuropathy,

 Facial nerve palsy,
 Cauda equina syndrome, and
 Guillain-Barré syndrome.
Profound depression of the CD4+ T-cell count after acute infection might
be associated with opportunistic infections (OIs), such as
 Oral-esophageal candidiasis,
 Pneumocystis carinii pneumonia (PCP),
 Cytomegalovirus (CMV) colitis or encephalitis,
 Cerebral toxoplasmosis
 Tuberculosis.

METABOLIC MANIFESTATIONS
 Hyperlipidemia
 Lipoatrophy
 Lactic acidosis
 Insulin resistance and Hyperglycemia.
 Hypertriglyceridemia.
 Lipohypertrophy Buffalo hump, Increased torso and abdominal girth &
Breast size.
 Lipodystrophy Characterized by dramatic relocation of fat from the
subcutaneous compartment, which atrophies, into the visceral compartment, where
fat accumulation is noted.
 Highly active antiretroviral therapy (HAART), which contributes to altered lipid
metabolism in addition to impaired mitochondrial function, contributes
significantly to this syndrome.

HIV Lipodystrophy
Buffalo hump

PULMONARY MANIFESTATIONS
 Pulmonary manifestations of HIV infection can be broadly divided into infectious,
noninfectious, and malignant categories.
Lymphocytic interstitial pneumonitis (LIP) is primarily a disease of HIV-infected
pediatric patients.
 Approximately 30% to 40% of perinatally infected children and 12% of transfusion-
related HIV infected children will have LIP.
Clinically
Insidious onset of non-productive cough
Generalized lymphadenopathy
Digital clubbing.
Increased frequency of bacterial pneumonia and resultant bronchiectasis.
 Radiograph Diffuse, symmetric, reticulonodular or nodular pattern with or
without hilar or mediastinal adenopathy.

Lymphocytic interstitial
pneumonitis (LIP)

PULMONARY MANIFESTATIONS
Nonspecific interstitial pneumonitis (NIP) is more prevalent in the adult
HIV-infected population.
 NIP develops when the CD4+ T cells and total lymphocyte counts are still
preserved.
 Histology Patchy lymphocytic infiltrate composed of B cells in focal
aggregates, with T cells more extensively distributed.
HIV-related pulmonary hypertension (HRPH) occurs with a frequency
of approximately 1 per 200 HIV-infected patients.
 The most dominant histopathologic finding in cases of HRPH is plexiform
arteriopathy.
 An increased prevalence of HLADR6 and HLA-DR52 has been
demonstrated in patients with pulmonary hypertension and HIV disease.

Pathology of the nonspecific interstitial pneumonia (NSIP) pattern
(a) Uniform and diffuse thickening of the alveolar walls is a typical finding in nsip.
(b) Higher magnification shows alveolar septa thickened by the inflammatory cell
infiltrate and mild interstitial fibrosis, without remodeling the basic lung
architecture.

Plexogenic pulmonary arteriopathy
featuring a plexiform lesion (upper-left
centre) consisting of a number of slit-
like vessels lined by cobblestone-like
endothelium and surrounded by loose-
textured fibrous tissue. The lesion is
flanked by distended vessels, the so-
called vein-like branches, which are
presumed to represent widely distended
branches of the feeding axial artery.

CARDIAC MANIFESTATIONS
 HIV infection is associated with cardiac abnormalities in almost 70% of infected
patients.
 Cardiac complications include pericardial effusion, myocarditis, endocarditis,
dilated cardiomyopathy, pulmonary hypertension, and cardiac tumors.
Small pericardial effusions(20% patients) due to Infections, lymphoma, KS,
myocardial infarction, fibrinous exudate, myocarditis, and endocarditis.
Infectious endocarditis is most commonly associated with intravenous drug use.
 Right-sided heart valves are involved primarily, and Staphylococcus aureus and
Streptococcus viridans are the most commonly isolated pathogens.
Lymphocytic myocarditis CMV, Cryptococcus neoformans, Candida species,
Toxoplasma gondii, sarcosporidium, Histoplasma capsulatum, Mycobacterium
tuberculosis, Coxsackie virus, and Aspergillus species.

CARDIAC MANIFESTATIONS
Dilated cardiomyopathy
 Most often noted in the late stages of infection, when CD4+ T-cell counts are low,
it affects up to 40% of HIV-infected patients.
 Cardiac involvement with KS in an HIV-infected patient usually occurs as a part
of disseminated KS.
 There is a predilection to involve the subepicardial tissue adjacent to a major
coronary artery.
 Pericardial and myocardial involvement has also been reported.
Primary NHL affecting cardiac tissue is extremely rare, but involvement as a
result of dissemination occurs in approximately 15% of cases.
Polypoid masses of the endocardium are noted most often.
 Patients might be asymptomatic initially but rapidly progress to pericardial
effusion, tamponade, and heart failure.

NEUROLOGIC MANIFESTATIONS
 The most prevalent and important is subacute encephalitis, also
referred to as AIDS encephalopathy or AIDS dementia complex.
Aseptic meningitis
Peripheral neuropathy
Vacuolar myelopathy.
 HIV infection predominantly results in peripheral neuropathy that is
chronic, distal, symmetric, and predominantly sensory(50% of
infected patients).
Inflammatory demyelinating polyneuropathy Weakness, mild
sensory deficits, and areflexia.

NEUROLOGIC MANIFESTATIONS
Guillain-Barré–like syndrome,
Cranial nerve involvement
Mononeuropathy multiplex.
Vacuolar myelopathy Progressive spastic paraparesis that evolves
during a period of weeks to months.
 The lesion typically involves the posterior columns and clinically
presents as gait ataxia, leg weakness, upper motor neuron signs,
incontinence, and posterior column deficits.

GENITOURINARY MANIFESTATIONS
 Sclerosing glomerulopathy
 HIV-associated nephropathy (HIV-AN)
 Clinical presentation varies from nephrotic-range proteinuria to
subnephrotic proteinuria.
 The incidence of hypertension is variable, even in the face of significant
renal insufficiency.
Additional glomerulonephropathies in HIV-infected patients
Immune complex type (mesangioproliferative),
Diffuse proliferative,
Membranous nephropathy,

Lupus-like nephropathy, and
IgA nephropathy.
Immune complex glomerulonephritis.
 HIV-AN in the presence of advanced AIDS is associated with rapid-
onset renal failure and death.
 Infections of the renal tract in HIV-infected patients might be related
to common urinary pathogens, as well as opportunistic pathogens.
 CMV infection involving the kidneys is generally related to systemic
infection and is frequently associated with acute tubular necrosis.

 Prostatitis with an increased frequency of abscess formation in patients with AIDS.
 Symptoms of frequency and dysuria.
 HIV and CMV might cause localized cystitis.
 Toxoplasma gondii has been associated with bladder infection presenting as fever
alone in an HIV-infected patient.
 A wide variety of pathogens have been identified, including HIV, CMV, Toxoplasma
gondii, Histoplasma capsulatum, Candida species, and Mycobacterium avium complex
(MAC).
 Epididymis Candida species, Salmonella species, MAC, and CMV.
 Malignant lesions can affect any part of the urinary tract.
 Penile KS might be the presenting feature of HIV infection in up to 2% of patients.
 An estimated 20% of patients with KS will have genital lesions.

RHEUMATOLOGIC MANIFESTATIONS
 A seronegative arthritis (distinct from spondyloarthropathy or rheumatoid
arthritis) oligoarticular, affecting primarily the lower extremities, self-
limited.
 Homosexual men might have an increased risk for gonococcal arthritis or
postinfectious reactive arthritis associated with genital or GI tract
infections (Reiter’s syndrome) Asymmetric oligoarticular arthritis,
primarily of the large joints of the lower extremities, and sacroiliitis.
 Urethritis, conjunctivitis, keratoderma blennorrhagicum, and circinate
balanitis might also be present.
 Polymyositis is considered the most common of the HIV-associated
myopathies.
 Typically, presents with a subacute, progressive, proximal muscle
weakness associated with an elevated creatine kinase level.

GI MANIFESTATIONS
 Candida species cause dysphagia or odynophagia (or both) and account for
60% to 75% of esophageal symptoms in patients with AIDS.
 Although oropharyngeal candidiasis is most commonly associated with Candida
species esophagitis, it might also be associated with CMV or idiopathic.
 In addition to Candida species, CMV, HIV, and HSV infection result in
esophageal ulceration.
 Esophageal ulcerations caused by HSV are usually multiple, deep, and
erythematous, with overhanging edges.
 HIV infection might be associated with esophageal ulcers, ranging in size from 2
to 15 mm, causing odynophagia, dysphagia, and esophagospasm; symptoms
and ulcers resolve spontaneously.
 Other infections reported to cause esophageal disease in patients with AIDS
include Torulopsis glabrata, Cryptosporidium species, and Mycobacterium
tuberculosis

GI MANIFESTATIONS
Abdominal pain in HIV-infected patients
 Biliary tract disorders Acalculous cholecystitis and AIDS cholangiopathy.
 Acalculous cholecystitis Previous OI, such as CMV or Cryptosporidium species.
AIDS cholangiopathy comprises a spectrum of biliary tract disorders, including
Papillary stenosis,
Sclerosing cholangitis,
A combination of papillary stenosis and sclerosing cholangitis, and
Long extrahepatic bile duct strictures.
 Clinical presentation includes fever, right upper quadrant pain, diarrhea, and weight loss.

GI MANIFESTATIONS
 The CD4+ T-cell count is frequently less than 200/mm3.
 The cause of this condition includes CMV, Cryptosporidium species,
Microsporida species, MAC, and chronic inflammation.
 Pancreatitis was not a significant problem in HIV-infected patients until the
introduction of dideoxyinosine.
 Clinical presentation is similar to that seen in HIV-negative patients with
pancreatitis
 Causes include infection (CMV, MAC, and Cryptosporidium, Cryptococcus,
and Toxoplasma species)
 Medication (pentamidine, dideoxyinosine, dideoxycytidine, lamivudine, and
trimethoprimsulfamethoxazole), and malignancy.
 Hypertriglyceridemia, common in patients with AIDS, has also been associated
with pancreatitis.

Laboratory DiagnosisCollection, Storage, and Transport
of Specimens for HIV Testing
 Currently, the diagnosis and monitoring of HIV infection is performed on blood
specimens.
 Generally for serological tests (antigen and antibody detection) serum/plasma/whole
blood is used, whereas
 For CD4 enumeration tests only whole blood collected in K2/K3 ethylene diamine tetra
acetic acid (EDTA) evacuated tubes is used.
 For DNA/RNA PCR, Dried Blood Spots (DBS) or whole blood collected in K2/K3
EDTA is used.
Blood Collection
 Ensure pre-test counselling is done and informed consent has been obtained.
 Identify the person using at least 2 identifiers. (e.g., name, identification number (ID),age,
gender).

Laboratory DiagnosisCollection, Storage, and Transport
of Specimens for HIV Testing
Separation of Sera
 The blood collected is allowed to clot for 30 minutes.
 The serum should be separated as soon as possible and should be refrigerated.
 The test tube/red top evacuated tube, containing collected blood, should be centrifuged at
2000 to 3000 revolutions per minute (rpm) for ten minutes to separate the serum.
 The serum should then be aliquoted in pre-labelled screw-capped, sterile, O-ring storage
vials using micropipette tips for testing/storage.
 No preservatives should be added as they interfere with the testing.
Storage of sera
 The sera can be stored at 2 to 8 C in the refrigerator for only up to a week.
 For longer storage, specimens need to be kept frozen at -200 C.
 Repeated freeze-thawing should be avoided.

 Shipment of infectious agents is permitted as per
the International Air Transport Association’s
(IATA) Regulations.
 Packaging requires a 3-layer system.
 The specimen tube, in which serum is to be
transported, should not have cracks/leaks.
 Preferably, it should be made of plastic and be
screw capped.
 The outside of the container should be checked
for any visible contamination with blood that
should be disinfected.
 Place the tube containing the specimen in a leak-
proof container (e.g., a sealed plastic bag with a
zip-lock or, alternatively, the bag may be stapled
and taped).
 A request slip should have all of the patient’s
details (i.e., name, age, sex, risk factors history of
previous testing, etc.) and should accompany the
specimen.
 For mailing, another box is used containing the
mailing label and a biohazard sign.

Serological Diagnosis of HIV Infection
 Laboratory diagnosis by HIV testing is the only method of determining the HIV
status of an infected individual’s infected blood, blood products, organs, and
tissues.
 HIV diagnosis at ICTCs and other laboratories is based on the demonstration
of antibodies.
 Antibody detection can be done using an ELISA test, rapid test, and western
blot test.
 These tests are used as screening tests and/or confirmatory tests.
 All tests should be performed and interpreted as per test instruction manuals that
are supplied with the kit.
 HIV testing should be based on testing strategy and algorithm.
 Testing laboratories should ensure pre and post-test counselling for every
individual and confidentiality to be maintained.

Serological Diagnosis of HIV Infection
Objectives of Testing
Transfusion and transplant safety
Diagnosis of HIV infection in symptomatic and asymptomatic individuals
Prevention of parent to child transmission
For Post-Exposure Prophylaxis (PEP)
Epidemiological surveillance using unlinked anonymous HIV testing
Research
Detection of Anti-HIV Antibodies
 The central component in the diagnosis of HIV infection is the detection of anti-HIV antibodies in
serum, plasma, or whole blood.
 Urine and saliva may be tested using specific kits.
 Some of these assays can differentiate between HIV-1 and HIV-2 infections.
 Differentiation between HIV-1 and HIV-2 is required since the treatment varies for the two types.

Serological Diagnosis of HIV Infection
Screening Tests
 Serological tests for the detection of HIV are classified as first to fourth generation
tests based on the type of antigens used and principle of the assays.
 NACO recommends the use of rapid test kits, which detect >99.5% of all HIV-infected
individuals and have false-positive results in <2% of all those who are tested.
Commonly used screening tests are
Enzyme Linked Immunosorbent Assay (ELISA)
Rapid tests
Immunoconcentration/Dot Blot assay (vertical flow)
Agglutination assay
Immunochromatographic assay (lateral flow)
Dipstick and comb assay based on Enzyme Immune Assay (EIA)

Screening Tests
On the basis of the principle of the test, ELISA can be divided into
Indirect
Competitive
Sandwich
Capture
Factors which may affect the test results
Pre-analytical:
Haemolysed sample
Grossly lipaemic samples
Repeated freezing and thawing
Contaminated samples and reagents
Improperly stored, expired and deteriorated reagents

Screening Tests
Analytical:
Pipetting errors
Improper incubation time and temperature
Improper washing procedure
Carry over from the adjacent specimen
Equipment malfunction
Glove -powder aerosol
Calculation errors
Post-analytical:
Transcription errors

Screening Tests
False Positive ELISA Results
Autoimmune diseases
Alcoholic hepatitis
Primary biliary cirrhosis
Leprosy
Multiple pregnancies
Common causes of a false negative result
Technical errors
The test may be negative during the window period and during the end stage of the
disease
Rapid Anti-HIV Tests
 Several rapid tests have been developed using recombinant and/or synthetic antigens.

Screening Tests
Commonly employed rapid anti-HIV tests are based on the principle of
Immunoconcentration/dot blot immunoassay (vertical flow),
Immunochromatographic (lateral flow),
Particle agglutination (e.g., gelatine or latex),
Dipstick and Comb assay based on EIA.
Western Blot Test
 In the western blot, the various HIV specific recombinant or synthetic antigens are adsorbed onto
nitrocellulose paper.
 The antibody, when present, attaches to the antigen on the strip and the antigen antibody complex
is then detected using enzyme conjugate and substrate.
 The test procedure should be carried out as per the kit insert.
 WB tests detect the presence of antibodies against specific HIV proteins, which are seen as bands
on the test strip.
 The test results are interpreted as per kit instructions.

Sequence of the
appearance of
laboratory
markers
following HIV-1
infection

Screening Tests
Limitations of Antibody Assays
 Antibodies are not detectable in the window period.
 Therefore, antibody detection tests are of no use during this period.
 Diagnostic tests based on antibody detection are also not useful in the diagnosis of
infection in children below 18 months of age.
 Babies born to HIV positive mothers may have passively acquired maternal antibodies.
 In this situation, tests that detect the viral genome may be done for early diagnosis.
 NACO is now promoting the use of the DBS technique for early infant diagnosis, based
on the detection of HIV-1 DNA viral nucleic acid.
Molecular tests are of particular importance in detection of
 Infection< 18 months children
 Acute HIV infection during window period.

Molecular & Other Assays
Tests utilized
Qualitative Polymerase Chain Reaction for HIV DNA [qualitative AMPLICOR HIV-1 DNA
PCR Test, ver. 1.5 (Roche)]
 NACO’s first choice for the diagnosis of HIV-1 infection in infants and children less than 18
months of age (starting at 6 weeks of age or at the earliest opportunity thereafter) is the HIV-1
DNA PCR test.
Qualitative Transcription-mediated Amplification Assay for HIV RNA[APTIMA HIV-1
qualitative RNA assay (Gen-Probe)]
Quantitative HIV-1 RNAAssays
Other Tests Virus Isolation
 HIV isolation requires co-cultivation of peripheral blood mononuclear cells (PBMCs), from an
infected individual and mitogen stimulated PBMCs from an HIV-uninfected individual and takes
up to 6 weeks in a medium containing interleukin- 2.
 The replication of HIV can be detected by measuring the p24 antigen by ELISA, or reverse
transcriptase activity in culture supernatant.

Molecular & Other Assays
Antigen Detection
 The HIV-1 p24 antigen is present as either an immune complex, with anti-p24
antibodies, or as a free p24 antigen in the blood of infected individuals.
 The positive p24 test confirms diagnosis of HIV infection; however, a negative
test does not rule out HIV infection.
 The test is based on the ELISA.
 The sensitivity of the test increases with the use of techniques to dissociate the p24
antigen from its antibody, as in immune complex-dissociated (ICD) tests.
 However, despite this advance, the diagnostic usage of p24 antigen assays is much
less frequent than that of NAT.
 This is due to their relatively lower sensitivity.
 HIV p24 antigen assays, with increased sensitivity, are now commercially
available and under evaluation, e.g., the ultrasensitive p24 assay (Perkin- Elmer).

National strategies for HIV Testing
Strategy 1 (for blood transfusion/transplant safety)
 The specimen is subjected to one test for HIV reactivity.
 The test used in strategy 1 must have high sensitivity.
 If non reactive, the specimen is to be considered free of HIV (negative) and
if reactive, the specimen is considered as HIV positive.
 This strategy is used for ensuring donation safety (e.g., blood, blood
products, organs, tissues, sperms etc.).
 The unit of blood that tests reactive (positive) is discarded.
 If the donor is to be notified of his result, based on his prior consent, it
becomes a matter of diagnosis (in which case strategies II & III must be
used after proper counselling) and the donor should be referred to an ICTC
for the confirmation.

Strategy 2 A (used in sentinel surveillance)
 A specimen is tested as above and if it is reactive it is tested further.
 A specimen is considered negative for HIV if the first ELISA or rapid test
reports it so.
 In case it is reactive, it is subjected to a second ELISA or rapid test, which
utilizes a system different from the first one (i.e., the principle of the test
and/or the antigen used is different).
 It is reported positive only if the second ELISA/rapid test also gives a
reactive report like the first test.
 In case the second E/R is non reactive, the result is taken as negative for
sentinel surveillance purposes.
 This type of HIV testing is anonymous and unlinked.

Strategy 2 B (used for diagnosis in symptomatic patients)
 This strategy is used to determine the HIV status of a clinically symptomatic suspected
AIDS cases in which blood/serum/plasma is tested with a highly sensitive screening test.
 The specimen is considered negative if the test gives a non-reactive result.
 In case the test result is reactive the specimen is tested with another test kit (based on a
different principle of test or having a different antigen as compared to the first test).
 If the result is also reactive with the second test kit, the specimen is considered to be
positive for HIV in a symptomatic AIDS case.
 In case a specimen is reactive with the first test kit and non reactive with the second test
kit, the specimen is subjected to a third tiebreaker test.
 If the third test is reactive, the specimen is reported as indeterminate and follow up testing
is undertaken after 2 to 4 weeks.
 In case the third tiebreaker test is non-reactive, the specimen is reported negative.
 Counselling, informed consent, and confidentiality are a must in all these cases.

Strategy 3 (used for diagnosis in asymptomatic patients)
 In strategy 3 the HIV testing done is similar to strategy 2, with the added testing of a third test for a
positive result.
 Positive confirmation of a third reactive E/R test is required for a specimen to be reported HIV
positive.
 If the specimen gives a reactive result with two E/R and non-reactive result with the third assay, it
is reported as “indeterminate” and the patient is called again for repeat testing after 2-4 weeks.
 The test utilized for the first screening should be the one with the highest sensitivity.
 Those used for the second and third tests are those with the highest specificity (to minimize false
positive reactions).
 This strategy is used for the diagnosis of HIV infection in asymptomatic individuals at ICTCs and
PPTCT centres.
 Counselling, informed consent, and confidentiality are a must in these cases.
 Three different kits, with different antigen systems, and/or different principles of testing are
required to follow this strategy.

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