The document discusses laboratory diagnosis of HIV. It provides an introduction to HIV and AIDS, including how HIV infects and destroys CD4+ lymphocytes, leading to immunodeficiency. It also discusses the taxonomy of HIV, morphology, epidemiology in Nigeria and globally, reservoirs and transmission routes. Testing methods for HIV include antibody, nucleic acid and antigen testing to diagnose infection.

1. LABORATORY DIAGNOSIS OF HIV
PRESENTED BY
MLS IGWE CHARLES S
MLS EDEH CALEB
MLS ADESUE S.T
MLS FOLAYOWON AFISU
MLS IZIEGBE OSUNDE
MLS ESTHER I. E
MOLECULAR UNIT
FEDERAL MEDICAL CENTRE, KEFFI
24th AUGUST, 2023
1

2. INTRODUCTION
• Human immunodeficiency virus (HIV) infection
results from 1 of 2 similar retroviruses (HIV-1 and
HIV-2) that destroy CD4+ lymphocytes and impair
cell-mediated immunity, increasing risk of certain
infections and cancers.
• Initial infection may cause nonspecific febrile
illness. Risk of subsequent manifestations—related
to immunodeficiency—is proportional to the level
of CD4+ lymphocyte depletion.
2

3. INTRODUCTION
• HIV can directly damage the brain, gonads, kidneys,
and heart, causing cognitive impairment,
hypogonadism, renal insufficiency, and
cardiomyopathy.
• Manifestations range from asymptomatic carriage
to acquired immune deficiency syndrome (AIDS),
which is defined by the presence of an AIDS-
defining illness (serious opportunistic infections or
cancers) or a CD4 count of < 200/mcL. People die of
AIDS because the disease remains undetected for
long periods of time.
3

4. INTRODUCTION
• HIV diagnostic testing has come a long way since it
was introduced in the early 1980s. Early diagnosis is
key to successful treatment of HIV.
• HIV infection can be diagnosed by antibody, nucleic
acid (HIV RNA), or antigen (p24) testing. Screening
should be routinely offered to all adults and
adolescents.
• Treatment aims to suppress HIV replication by using
combinations of ≥ 2 drugs that inhibit HIV enzymes;
treatment can restore immune function in most
patients if suppression of replication is sustained.
4

5. LITERATURE REVIEW
Human immunodeficiency virus (HIV) is the chief contributor
to global burden of disease culminating to morbidity and
mortality (CDC 2017).
In 2010, HIV was the fifth leading cause of disability-adjusted
life years in people of all ages and leading cause for people
aged 30–44 years.
The human immunodeficiency virus (HIV) is a lentivirus
(family of retroviruses) that targets white blood cells vital to
the human immune system including macrophages, dendritic
cells and especially helper T cells (CD4 lymphocytes)
(Cunningham et al., 2022).
.
5

6. LITERATURE REVIEW
• It is classified as a member of the family
Retroviridae and genus Lentivirus based on the
biological, morphological, and genetic properties.
• It infects different cells of the immune system, such
as CD4+ T cells (T-helper cells), dendritic cells, and
macrophages.
• HIV has two subtypes: HIV-1 and HIV-2. Among
these strains, HIV-1 is the most virulent and
pathogenic
6

7. LITERATURE REVIEW
• The human immunodeficiency viruses (HIV) are two
species of Lentivirus (a subgroup of retrovirus) that
infect humans. Over time, they cause acquired
immunodeficiency syndrome (AIDS), ( Douek et al.,
2009)
• condition in which progressive failure of the
immune system allows life-threatening
opportunistic infections and cancers to thrive.
(Powell et al., 2016)Without treatment, average
survival time after infection with HIV is estimated
to be 9 to 11 years, depending on the HIV subtype.
7

8. LITERATURE REVIEW
• AIDS was first recognized in the United States in 1981
following a sudden outbreak of opportunistic
infections, including Pneumocystis carinii pneumonia
and Kaposi’s sarcoma (KS) among homosexual men
(Durack, 1981; Gottlieb et al., 1981; Masur et al., 1981).
• Subsequent epidemiologic studies implicated an
infectious agent that was transmitted during sexual
intercourse, through intravenous drug abuse, by
therapies utilizing blood and blood products, and
vertically from mother to child (Broder et al., 1994).
8

9. LITERATURE REVIEW
• In most cases, HIV is a sexually transmitted infection and
occurs by contact with or transfer of blood, pre-ejaculate,
semen, and vaginal fluids.(Rodger et al., 2019). Non-
sexual transmission can occur from an infected mother to
her infant during pregnancy, during childbirth by
exposure to her blood or vaginal fluid, and through breast
milk (Mabuka et al., 2012).
• Within these bodily fluids, HIV is present as both free
virus particles and virus within infected immune cells.
Research has shown (for both same-sex and opposite-sex
couples) that HIV is untransmittable through condomless
sexual intercourse if the HIV-positive partner has a
consistently undetectable viral load (Eisinger et al., 2019).
9

10. LITERATURE REVIEW
• HIV infects vital cells in the human immune system,
such as helper T cells (specifically CD4+ T cells),
macrophages, and dendritic cells. HIV infection
leads to low levels of CD4+ T cells through a
number of mechanisms, including pyroptosis of
abortively infected T cells (Doitsh et al., 2014),
apoptosis of uninfected bystander cells (Garg et al.,
2012), direct viral killing of infected cells, and killing
of infected CD4+ T cells by CD8+ cytotoxic
lymphocytes that recognize infected cells.
10

11. LITERATURE REVIEW
• When CD4+ T cell numbers decline below a critical
level, cell-mediated immunity is lost, and the body
becomes progressively more susceptible to
opportunistic infections, leading to the development
of AIDS.
• Without antiretroviral treatment the disease
typically advances through several phases:
• First year: rapid and transient decline in the number
of CD4 lymphocytes;
11

12. LITERATURE REVIEW
• Progression over anywhere from a few months to
more than 10 years: CD4+ T cell numbers slowly
decline to below a critical level and the immune
system can no longer fight opportunistic infections
• Final stage: Onset of full-blown AIDS (Acquired
Immune Deficiency Syndrome), marked by infection
with one or more HIV-associated opportunistic
infections or cancers and ultimately death.
12

13. TAXONOMY ( SCIENTIFIC
CLASSIFICATION)
• Groups included
• Human
immunodeficiency virus 1
• Human
immunodeficiency virus 2
• (unranked): Virus
• Virus
• Realm: Riboviria
• Kingdom: Pararnavirae
• Phylum: Artverviricota
• Class: Revtraviricetes
• Order: Ortevirales
• Family: Retroviridae
• Subfamily: Orthoretrovirinae
• Genus: Lentivirus
13

14. MORPHOLOGY
• HIV is a spherical, enveloped virus, which measures
up to 120 nm in diameter (Fig. 68-1). It has a
unique three-layered structure:
• (i) the innermost genome layer,
• (ii) middle cone-shaped nucleocapsid
• (iii) an outer membrane of glycopro-tein
surrounded by lipoprotein envelope.
14

15. • Viral schematic morphology: (slideshares. net.,
2019) 15

16. EPIDEMIOLOGY
• The Global HIV and AIDS Epidemic
• HIV, the virus that causes AIDS, is one of the world’s most
serious public health challenges. But there is a global
commitment to stopping new HIV infections and ensuring
that everyone with HIV has access to HIV treatment.
• The latest statistics on HIV around the world from UNAIDS
include:
• Number of People with HIV—There were approximately
39 million people across the globe with HIV in 2022. Of
these, 37.5 million were adults and 1.5 million were
children (<15 years old). In addition, 53% were women
and girls (USAID 2022).
16

17. • USAID 2022
17

18. EPIDEMIOLOGY
• New HIV Infections—
• An estimated 1.3 million individuals worldwide
acquired HIV in 2022, marking a 38% decline in new
HIV infections since 2010 and 59% since the peak in
1995.
• New HIV infections, or “HIV incidence,” refers to the
estimated number of people who newly acquired HIV
during a given period such as a year, which is
different from the number of people diagnosed with
HIV during a year. (Some people may have HIV but
not know it.) Women and girls accounted for 46% of
all new HIV infections in 2022.
18

19. EPIDEMIOLOGY
• HIV Testing & Knowledge of HIV Status—
• Approximately 86% of people with HIV globally
knew their HIV status in 2022.
• The remaining 14% (about 5.5 million people) did
not know they had HIV and still needed access to
HIV testing services. HIV testing is an essential
gateway to HIV prevention, treatment, care, and
support services.
• The global target for HIV status awareness is 95%
by 2025.
19

20. • UNAID 2022
20

21. EPIDEMIOLOGY
• HIV Treatment Access—
• As of the end of 2022, 29.8 million people with HIV (76% of
all people with HIV) were accessing antiretroviral therapy
(ART) globally. HIV treatment access is key to the global
effort to end AIDS as a public health threat. People with HIV
who are aware of their status, take ART as prescribed, and
get and keep an undetectable viral load can live long and
healthy lives and will not transmit HIV to their HIV-negative
partners through sex. This is sometimes referred to as
“undetectable = untransmittable” or U=U. The global
targetsExit Disclaimer for 2025 include 95% of all people
with diagnosed HIV receive sustained ART and 95% of those
individuals on treatment achieve and maintain HIV viral
suppression.
21

22. EPIDEMIOLOGY
• HIV Care Continuum—
• The term HIV care continuum refers to the sequence of
steps a person with HIV takes from diagnosis through
receiving treatment until his or her viral load is
suppressed to an undetectable level.. The stages are:
• being diagnosed with HIV;
• being linked to medical care;
• starting ART; adhering to the treatment regimen;
• and, finally, having HIV suppressed to undetectable
levels in the blood. UNAIDS reports that in 2022,
among all people with HIV worldwide:
22

23. EPIDEMIOLOGY
• Perinatal Transmission—In 2022, globally, 82% of pregnant
people with HIV had access to ART to prevent transmitting
HIV to their babies during pregnancy and childbirth and to
protect their own health.
• AIDS-related Deaths—AIDS-related deaths have been
reduced by 69% since the peak in 2004. In 2022, around
630,000 people died from AIDS-related illnesses worldwide,
compared to 2 million people in 2004 and 1.3 million in
2010.
• Regional Impact—Certain regions of the globe are
disproportionately affected by HIV. In 2022, there were 20.8
million people with HIV in eastern and southern Africa, 4.8
million in western and central Africa, 6.5 million in Asia and
the Pacific, and 2.3 million in Western and Central Europe
and North America.
23

24. GLOBAL DATA OF PEOPLE LIVING
WITH HIV
• Globally,
• An estimated 39.0 million [33.1–45.7 million] people
were living with HIV at the end of 2022.
• 1.5 million [1.2–2.1 million] children (0–14 years old).
• HIV incidence
• 1.3 million [1.0–1.7 million] people acquired HIV in
2022. Since 2010, the number of people acquiring
HIV has been reduced by 38%, from 2.1 million [1.6–
2.8 million].
• 130 000 [90 000–210 000] children acquired HIV in
2022.
24

25. GLOBAL DATA OF PEOPLE LIVING
WITH HIV
• HIV-related mortality
• In 2022, 630 000 [480 000–880 000] people died from
HIV-related causes globally. Since 2010, HIV-related
deaths have been reduced by 51%, from 1.3 million
[970 000–1.8 million]. The global HIV epidemic claimed
69% fewer lives in 2022 since the peak in 2004.
• 84 000 [56 000–120 000] children died from HIV-
related causes in 2022.
• HIV continues to be a major global public health issue,
claiming 40.4 million [32.9–51.3 million] lives so far
(WHO 2022).
25

26. NIGERIA HIV EPIDEMIOLOGY
• In 2021, 1.9 million people in Nigeria were living with
HIV. Women were the most affected group, counting
1.1 thousand individuals. Also, children up to age 14
who were HIV positive equaled 170 thousand.
• 1.9 million people with HIV
• 1.3% adult HIV prevalence
• 74,000 new HIV infections
• 51,000 AIDS-related deaths
• 1.7 million people on antiretroviral treatment
(National Agency for the Control of AIDS (NACA)
2021).
26

27. HIV PREVALENCE IN NIGERIA 2022/2023
• Akwa Ibom 5.6%
• Benue 4.9%
• Rivers 3.8%
• Taraba 2.7%
• Anambra 2.4%
• Enugu 2.1%
• Abia 2.1%
• Delta 1.9%
• Nasarawa 1.9%
• Edo 1.8%
• Bayelsa 1.8%
• Cross River 1.7%
• Imo 1.6%
• Plateau 1.5%
• FCT 1.5%
• Lagos 1.3%
• Borno 1.3%
• Adamawa 1.3%
• Ogun 1.2%
• Gombe 1.2%
• Kaduna 1.0%
• Kogi 1.0%
• Kwara 1.0%
• Ondo 0.9%
• Osun 0.9%
• Oyo 0.9%
• Ebonyi 0.8%
• Niger 0.7%
• Ekiti 0.7%
• Kebbi 0.6%
• Kano 0.5%
• Zamfara 0.5%
• Yobe 0.4%
• Bauchi 0.4%
• Sokoto 0.4%
• Jigawa 0.3%
• Katsina 0.3% 27

28. HIV prevalence rate according to Zones
2022/2023
Zones HIV prevalence (%)
• South-South 3.1
• South-East 1.9
• South-West 1.2
• North-Central 2.1
• North-West 0.6
• North-East 1.1
• HIV NIGERIA - UNAID 2022
28

29. (Institute of Human Virology Nigeria 2019)
29

30. RESERVOIR, SOURCE, AND
TRANSMISSION OF INFECTION
• HIV is primarily a human infection.
• Humans infected with HIV and AIDS are the
reservoir of infection.
• The high titer of HIV is found in the blood, semen,
and vaginal secretions of the infected people;
hence these are important sources of infection.
• The virus is also present in the breast milk of an
infected mother.
30

31. TRANSMISSION
• Transmission of HIV infection: HIV infection occurs either by the transfer
of HIV-infected cells or free HIV not associated with cells. HIV
transmission occurs in following ways.
• HIV is spread through body fluids such as
• blood,
• semen, vaginal fluid,
• pre-ejaculate, or
• breast milk.
• Unprotected sexual contact is the most common means of transmission.
• Other means of transmission include
• blood transfusions
• sharing of needles in IV drug use,
• mother-to-child transmission in pregnancy,
• childbirth
• breastfeeding.
31

32. TRANSMISSION OF HIV
Transmission of HIV USAID 2022
32

33. PATHOGENESIS AND IMMUNITY
RESPONSE
• Pathogenesis and Immunity
• HIV is primarily a sexually transmitted pathogen
transmitted by high-risk behaviors, such as unprotected
intercourse, male homosexual intercourse, and also by
intravenous (IV) drug abuse.
• HIV infects the cells of the immune system and destroys
them or makes them ineffective so that the immune
system deteriorates and can no longer fight infection.
• The tropism of the HIV for CD4-expressing T-cells and
macrophages is the principal determinant of the
pathogenicity of HIV.
33

34. PATHOGENESIS AND IMMUNITY
RESPONSE
• HIV shows tropism for all the cells expressing CD4 anti-gens
on their cell surfaces. The CD4 antigens act as receptors for
HIV. The virus infects helper T cells and kills them, result-ing
in HIV-induced immunosuppression, leading to full-blown
AIDS—a key feature of the pathogenesis of HIV infection.
This makes the patient most susceptible to opportunistic
infections and certain cancerous conditions, such as Kaposi’s
sarcoma and lymphoma. However, the virus does not
directly cause any tumor, because HIV genes are not found
in these tumor cells
• .Without antiretroviral treatment the disease typically
advances through several phases:
• First year: rapid and transient decline in the number of CD4
lymphocytes;
34

35. PATHOGENESIS AND IMMUNITY
RESPONSE
• Progression over anywhere from a few months to
more than 10 years: CD4+ T cell numbers slowly
decline to below a critical level and the immune
system can no longer fight opportunistic infections;
• Final stage: Onset of full-blown AIDS (Acquired
Immune Deficiency Syndrome), marked by infection
with one or more HIV-associated opportunistic
infections or cancers and ultimately death.
35

36. 36

37. SIGNS AND SYMPTOMS
37

38. HIV DIAGNOSIS
• Diagnostic testing
• Conventional screening for HIV is usually first done through a blood sample,
sputum or urine test to detect the presence of HIV antibodies.
• Diagnosis can best be achieved by serologic testing but the monitoring of HIV
disease progression is mostly accomplished by the quantitation of CD4 T cells
and viral RNA.
• HIV Tests for Screening and Diagnosis
• HIV tests are very accurate, but no test can detect the virus immediately after
infection. How soon a test can detect HIV depends on the type of test being
used.
• There are three types of HIV tests:
• antibody tests,
• antigen/antibody tests, and
• nucleic acid tests (NAT)
( CDC., 2022).
38

39. LABORATORY DIAGNOSIS
• HIV diagnosis is achieved by utilizing variety of
• immunological and
• molecular methods, like
• ELISA,
• rapid diagnostics,
• Western blotting,
• indirect immunoassays,
• Molecular (NAT, VIRAL LOAD TEST. CD4 COUNT) assays are
currently used
• antiretroviral monitoring and
• drug resistance characterization in developed countries. what
nucleic acid-based tests
39

40. LABORATORY DIAGNOSIS
• Serologic tests
• These tests include demonstration of antigens and antibodies in
the serum.
• The tests have been classified as:
• HIV antigen-antibody laboratory-based tests.
• HIV antigen-antibody point-of-care tests.
• HIV antibody laboratory-based tests.
• HIV antibody point-of-care tests.
• HIV 1 and 2 differentiation tests.
• HIV-1 Western Blot test.
• HIV Nucleic acid diagnostic tests.
• In-home HIV tests.
40

41. LABORATORY DIAGNOSIS
HIV antigen-antibody laboratory-based tests
• These immunoassay tests are the preferred screening tests
which detect HIV-1 p24 (capsid) antigen and antibodies (IgM
and IgG) to HIV-1 and HIV-2. (Figure 1A–C)
• These antigen-antibody test detect HIV infection much earlier
than the antibody-based tests.
• If found positive in these tests, then it may require a
confirmatory test.
• Limitation of these tests are cross-reactivity to HIV-1 p24
antigen seen in HIV-2 infected persons.
• Examples are ADVIA Centaur HIV Ag/Ab Combo (CHIV) Assay,
ARCHITECT HIV Ag/Ab Combo, BioPlex 2200 HIV Ag-Ab Assay,
Elecsys HIV Combi PT, GS HIV Combo Ag/Ab EIA, & VITROS HIV
Combo Test (Delaney et al., 2017).
41

42. 42

43. LABORATORY DIAGNOSIS
HIV antigen-antibody point-of-care tests
• This assay is a single use, rapid test which is a point-of-
care test for the detection of HIV-1 p24 antigen,
antibodies to HIV-1 (group 0), and antibodies to HIV-2.
• This test does not differentiate HIV-1 and HIV-2
antibodies.
• This test is less sensitive for acute or recent HIV
infection when compared to laboratory-based HIV-1/2
antigen-antibody tests. Example: Abbott Determine
HIV-1/2 Ag/Ab Combo (Maciotra et al., 2017)
43

44. LABORATORY DIAGNOSIS
HIV antibody laboratory based tests
• Laboratory-based HIV antibody tests were the first to be
used for screening HIV since 20 years which has been
replace by HIV antigen-antibody tests.
• These tests can detect the IgM/IgG-sensitive assays for HIV
IgM antibodies in 23–25 days after HIV infection. Window
period is around 90 days.
• The positive result in this tests would require an
confirmatory test. Examples are: ADVIA Centaur HIV 1/O/2
Enhanced, Avioq HIV-1 Microelisa System, Genetic Systems
(GS) HIV-1/HIV-2 Plus O EIA, VITROS Anti-HIV 1 + 2 Assay
(Hurt et al., 2017).
44

45. LABORATORY DIAGNOSIS
HIV antibody point-of-care tests
• Single-use, point-of-care tests can yield result in 40 min.
• These tests can detect antibodies to HIV-1 or HIV-2 or both but
they will not be able to differentiate between HIV-1 and HIV-2.
• These tests are primarily used for testing
• (1) emergency situations
• (2) pregnant women whose HIV status in not known
• (3) occupational, and (
• 4) in patients for whom follow-up for HIV result will not be
possible. Examples are: Chembio DPP HIV 1/2 Assay, Chembio HIV
1/2 STAT-PAK Assay, Chembio SURE CHECK HIV 1/2 Assay, INSTI
HIV-1/HIV-2 Antibody Test, OraQuick ADVANCE Rapid HIV-1/2
Antibody Test, Reveal G4 Rapid HIV-1 Antibody Test (Reveal G4),
Uni-Gold Recombigen HIV-1/2 (Kuhar et al., 2013).
45

46. LABORATORY DIAGNOSIS
HIV 1 and 2 differentiation tests
• These tests will be able to differentiate between HIV-1 and
HIV-2.
• These tests utilize multiple recombinant or synthetic
peptides to detect HIV-1 antibodies and HIV-2 antibodies.
• These immunochromatographic tests will contain 7 lines
which consists of 6 HIV peptides and one control.
• A minimum of 2 envelope peptides (gp160 and gp41) or 1
envelope peptide plus either the p24 or the polymerase
peptide p31 for HIV-1 reactive or HIV-2 envelope peptides
gp36 and gp140 should be present for HIV-2 reactive test.
Example: Geenius HIV 1/2 Supplemental Assay.
46

47. 47

48. LABORATORY DIAGNOSIS
• The Geenius HIV 1/2 Supplemental Assay is a
single-use immunochromatographic test that
utilizes multiple recombinant or synthetic peptides
to detect HIV-1 antibodies (p31, gp160, p24, and
gp41) and HIV-2 antibodies (gp36 and gp140).
• The test cassette as shown here contains seven
test lines, including the six HIV peptides and one
control.
48

49. LABORATORY DIAGNOSIS
• HIV-1 Western blot test
• Western blot test is used as supplemental tests for
those tests which are reactive by rapid tests.
• It can detect the human antibodies for three HIV-1
gene regions: env (gp41, gp120/160), pol (p31, p51,
p66), and gag (p15, p17, p24, p55) (Figure 3A-D).
49

50. • Figure 3.
• (a) Components used in the HIV-1 Western blot; (b) HIV-1 antibodies
bound to HIV-1 antigens on Western blot test strip. (c) Addition of
secondary anti-human antibody linked to enzyme signal. (d) HIV-1
Western blot genes. Source: Illustration: David H. Spach, MD. 50

51. LABORATORY DIAGNOSIS
• This graphic shows the relationship of the HIV-1 genes and
products with the corresponding band on the HIV-1 Western blot.
• CDC and the Association of State and Territorial Public Health
Laboratory Directors (ASTPHLD) have published the criteria for
interpretation of Western blot tests (CDC 1989).
• Positive: A positive Western blot indicates the presence of at least
two of the following bands: p24, gp41, and gp120/160.
• Negative: A negative Western blot is defined by the absence of
any bands.
• Indeterminate: An indeterminate Western blot results from the
presence of any bands, but not meeting positive criteria. Possible
causes of an indeterminate Western blot include early HIV
infection, HIV-2, pregnancy, or cross-reactivity with other
antibodies, such as in persons who have recently received an
influenza immunization or who have autoimmune disorder.
51

52. LABORATORY DIAGNOSIS
• HIV nucleic acid diagnostic tests
• HIV RNA nucleic acid test (NAT) is used in case of
• 1. Reactive HIV-1/2 antigen-antibody immunoassay but a
nonreactive or indeterminate HIV-1/HIV-2 differentiation
test,
• 2. HIV-1/2 antigen-antibody immunoassay is negative but
there is high suspicion of acute HIV, and 3. Confirmative test
for chronic HIV-1 infection.
• The limitation of these tests are cost, time taken to perform
the test is 3 hours and the expert is required to perform the
test. Example: APTIMA HIV-1 RNA Qualitative Assay (Pierce
et al., 2011).
52

53. LABORATORY DIAGNOSIS
• In-home HIV tests
• This test can be performed at home by the client
itself within 40 minutes by simply collecting an oral
sample and performing the test as per kit literature.
• A confirmatory test will be required if this test is
reactive. Example OraQuick In-Home HIV test (U.S.
FDA 2023)
53

54. HIV laboratory testing algorithms
• HIV laboratory testing algorithms
• There are several algorithms published for HIV
laboratory testing among which CDC, NACO
(National AIDS Control Organization) and APHL are
some of them (Branson et al., 2012).
54

55. 55

56. Monitoring status of HIV infection
• Monitoring status of HIV infection
• The laboratory monitoring of the status of HIV
infection can be carried out by analysis of
• (i) T-cell subset,
• (ii) measurement of HIV RNA, and
• (iii) measurement of B2 microglobulin and
neopterin.
56

57. CD4 MEASUREMENT
• Other tests used in HIV treatment
• The CD4 T-cell count is not a HIV test, but rather a procedure where the number of
CD4 T-cells in the blood is determined.
• A CD4 count does not check for the presence of HIV. It is used to monitor immune
system function in HIV-positive people.
• Declining CD4 T-cell counts are considered to be a marker of progression of HIV
infection. A normal CD4 count can range from 500 cells/mm3 to 1000 cells/mm3.
• The absolute number of CD4 lymphocytes and the ratio of helper T inducer
lymphocytes (CD4:CD8 ratio) are very low in HIV-infected people. The CD4:CD8 T-cell
ratio is reversed to 0.5:1 from a normal level of 2:1
• . If the CD4 count is below 500/ L, it indicates progression of the disease and hence
requires specific therapy against HIV.
• In HIV-positive people, AIDS is officially diagnosed when the count drops below 200
cells/μL or when certain opportunistic infections occur. This use of a CD4 count as an
AIDS criterion was introduced in 1992; the value of 200 was chosen because it
corresponded with a greatly increased likelihood of opportunistic infection. Lower
CD4 counts in people with AIDS are indicators that prophylaxis against certain types
of opportunistic infections should be instituted (Pierce et al., 2011).
57

58. NUCLEIC ACID TEST NATs (VIRAL
LOAD TEST)
• NATs look for the actual virus in the blood. This test
should be considered for people who have had a
recent exposure or a possible exposure with early
symptoms of HIV and have tested negative with an
antibody or antigen/antibody test. A NAT can usually
detect HIV 10 to 33 days after exposure.
• Nucleic-acid-based tests amplify and detect one or
more of several target sequences located in specific
HIV genes, such as HIV-I GAG, HIV-II GAG, HIV-env, or
the HIV-pol.
58

59. NUCLEIC ACID TEST NATs (VIRAL
LOAD TEST)
• Nucleic-acid-based tests amplify and detect one or
more of several target sequences located in specific
HIV genes, such as HIV-I GAG, HIV-II GAG, HIV-env, or
the HIV-pol.
• Measurement of HIV RNA: HIV RNA level in serum is
an important predictive marker of disease
progression and are used as prognostic marker to
monitor the effectiveness of anti-HIV therapies. The
test is also useful for early diagnosis of HIV infection
in infants born to infected mothers.
59

60. NUCLEIC ACID TEST NATs (VIRAL
LOAD TEST)
• In the RT-PCR test, viral RNA is extracted from the
patient's plasma and is treated with reverse
transcriptase (RT) to convert the viral RNA into cDNA.
The polymerase chain reaction (PCR) process is then
applied, using two primers unique to the virus's
genome. After PCR amplification is complete, the
resulting DNA products are hybridized to specific
oligonucleotides bound to the vessel wall, and are then
made visible with a probe bound to an enzyme. The
amount of virus in the sample can be quantified with
sufficient accuracy to detect threefold changes.
60

61. Schematic PCR procedure slideshares.com 2019
61

62. Staging of HIV and tests
recommended
• Window Period: The time between HIV infection and the accurate detection
of HIV infection by any laboratory test. This period can vary depending the
type of test done to detect HIV infection. CDC has recommended around 45
days window period for the HIV 1/2 antigen–antibody tests and 90 days for
all HIV antibody tests and all HIV point-of-care tests.
• Seroconversion Window Period: The time interval between HIV infection
and the detection of anti-HIV antibodies by any laboratory test. This period
also can vary depending on the type of HIV test used.
• Acute HIV infection: The time interval between the detection of HIV RNA
and anti-HIV antibodies.
• Recent Infection: The time interval from the HIV infection to 6 months of
infection when anti-HIV antibodies are rising.
• Early Infection: The time interval which includes both acute HIV infection
and recent HIV infection.
• Established HIV Infection: The full-blown HIV infection when the anti-HIV
IgG antibody response is fully detectable.
62

63. Staging of HIV and tests
recommended
• This graphic shows the HIV testing algorithm as
recommended in 2014 and 2018 by the Centers for
Disease Control and Prevention (CDC) and Association
of Public Health Laboratories (APHL).
• The stages of HIV infection and the tests that are
recommended are (PAGAA 2019).
• Eclipse Phase: This is the first phase of HIV infection
during which no diagnostic test will be able to detect
HIV infection. HIV nucleic acid test (NAT) is the test
which can detect HIV infection at the earliest.
63

64. Staging of HIV and tests
recommended
• Days following HIV acquisition which of the HIV
diagnostic tests can show positivity for infection are
shown in Figure 5 (Maciotra et al.,2011).
64

65. • Figure 5.
• Timing of positivity for HIV diagnostic tests.This figure shows
estimates for the mean number of days for HIV diagnostic tests to
become positive after acquisition of HIV. Abbreviation: POC = point-
of-careSource: modified from Centers for Disease Control and
Prevention and Association of Public Health Laboratories [5].
65

66. Interpretation of HIV test results
• Interpretation of HIV test results
• If any HIV-1/2 antigen-antibody immunoassay test is
NONREACTIVE, then the test result should be interpreted as
not infected with HIV-1 or HIV-2. If acute HIV is suspected,
then there will be a need to perform HIV-1 RNA test.
• If any HIV-1/2 antigen-antibody immunoassay test is
REACTIVE, then the test should be checked with HIV-1/HIV-
2 differentiation assay result to check for whether the
person is reactive to HIV-1 or HIV-2.
• If any HIV-1/2 antigen-antibody immunoassay is reactive
and HIV-1/ HIV-2 differentiation test is indeterminate for
HIV-1 and nonreactive for HIV-2, then it is indeterminate
result. HIV-1 NAT should be done in this case.
66

67. INTERPRETATION OF VARIOUS LABORATORY
TEST USED IN THE DIAGNOSIS OF HIV
INFECTION SUMMARIZED
67

68. LABORATORY DIAGNOSIS
• Antigen/antibody tests look for both HIV antibodies and
antigens. Antibodies are produced by a person’s immune system
when they’re exposed to viruses like HIV. Antigens are foreign
substances that cause a person’s immune system to activate.
• If a person has HIV, an antigen called p24 is produced before
antibodies develop.
• Antigen/antibody tests are recommended for testing done in labs
and are common in the United States.
• An antigen/antibody test performed by a lab on blood from a vein
can usually detect HIV 18 to 45 days after exposure.
• There is also a rapid antigen/antibody test available that is done
with a finger stick. Antigen/antibody tests done with blood from a
finger stick can take 18 to 90 days after exposure.
68

69. PREVENTION OF HIV
• Ensuring protective intercourse via use of internal or
external condoms
• Using pre-exposure prophylaxis (PrEP) if you don't have
HIV
• Maintaining an undetectable viral load if you have HIV
• Starting HIV therapy if you are pregnant
• Avoiding breastfeeding if you have HIV
• Avoiding shared needles or syringes
• Using post-exposure prophylaxis (PEP).
69

70. TREATMENT
• HIV treatment (called antiretroviral therapy or ART)
• reduces the amount of HIV in the blood (called viral
load),
• reduces HIV-related illness, and
• prevents transmission to others.
• People with HIV who take HIV treatment as
prescribed and get and keep an undetectable viral
load (or stay virally suppressed) will not transmit HIV
to their sex partners.
70

71. REFERENCES
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