HIV – CLINICAL MANIFESTATIONS,  DIAGNOSTIC CRITERIA & SAMPLING.pdf

Microbiology, medicine

1. HIV – CLINICAL MANIFESTATIONS,
DIAGNOSTIC CRITERIA & SAMPLING


DR PRIYADHARSHINI S,
POSTGRADUATE,
DEPARTMENT OF MICROBIOLOGY,
CHRI,

2. CLINICAL MANIFESTATIONS
Development from HIV infection to end-stage AIDS through
several phases.

●HIV infection.
●HIV disease
●AIDS.
●Acute Retroviral Syndrome
(ARS)
●clinical latency
●HIV (viral) set point.

3. Physical Examination

5. HIV INFECTION
●When a person first contracts HIV, he or she has an HIV
infection. A person can be infected with HIV but may not be ill.

6. HIV DISEASE
●This develops 6 days to 6 weeks after contracting HIV.
●Flu-like signs and symptoms, including fever, head and muscle
aches, sore throat, and enlarged lymph nodes.

7. Signs and symptoms that indicate this early stage of HIV disease
coincide with high levels of virus replication and are called the
Acute retroviral syndrome (ARS); mild and resolve by themselves
in a month.

However, many people with HIV disease are asymptomatic.

10. Reduction in Viremia :
- Marked by a significant reduction in viremia after the acute phase of HIV
infection.
- Followed by a cytotoxic T-lymphocyte response and humoral antibody
response.
Latent Period:
- Clinically asymptomatic or "latent" period lasts from months to years.
- 90% of HIV proviruses are transcriptionally silent.



CLINICAL LATENCY

11. Continuous CD4+ Cell Loss and Replacement :

- Continuous loss of CD4+ cells in which HIV is replicating.
- Active replacement through stem cell multiplication.
- CD4+ count declines slowly over time.

12. Stable Low-Level Virus Production :
- Host immune response maintains a relatively stable, low level of virus
production referred to as the HIV (viral) set point.
- Estimated production of 10
11
virions and 10
9
CD4 T cells.
Less Cytopathic Virus :
- Virus isolated during this period is less cytopathic for CD4+ cells.
- Virus replicates more slowly than during symptomatic AIDS.
Effective Host Immune Response :
- Immune response is effective enough to maintain a relatively stable, low
level of virus production.

14. PROGRESSION TO AIDS:
●Influenced by virologic and immunologic changes.
●Any Immune response stimulation activates resting T cells, increasing the number of
infected CD4+ cells.
●Highly cytocidal, rapidly multiplying variant appears.
●T-cell precursors in lymphoid organs are infected and killed, so ,reducing the capacity to
generate new CD4+ cells.
●The appearance of HIV mutants with altered antigenic specificity are not recognized by
the existing humoral antibody or cytotoxic T lymphocytes.
●This results in a rapid decline in CD4+ count and loss of immune capacity.

17. End-stage AIDS:
a. Spread of HIV to additional body sites
b. Opportunistic infections in AIDS
c. Malignancies associated with AIDS

18. Acquired Immune Deficiency Syndrome:
●Clinical AIDS is defined as a CD4 cell count of less than 200/cmm
and/or the appearance of AIDS defining illnesses.
●Most people with AIDS also suffer from weight loss, fever, fatigue,
and diarrhea—a set of problems referred to as the AIDS-related
complex (ARC).
●This stage may last for one to three years and results in death
without the intervention of ART.

19. Spread of HIV to additional body sites
●HIV infects cells other than CD4+ lymphocytes, causing additional manifestations of
end-stage disease.

●Monocyte-macrophage lineage infected cells can transport the virus into other organs.

20. Opportunistic infections in AIDS

22. Signs of serious illness
●Temperature ≥ 39°C with headache
●Respiratory rate ≥ 30/min
●Heart rate ≥ 120/min
●SpO2 (pulse oximeter) < 90%
●Altered mental status (e.g., confusion, strange behaviour, reduced consciousness)
●Other neurological problem (persistent severe headache, seizure, paralysis, difficulty
in talking, rapid deterioration of vision) , Unable to walk unaided.

23. Advanced HIV Disease
PLHIV with advanced HIV disease are defined as presenting with
1.CD4 count <200 cells/mm3 or
2.WHO clinical stage 3 or 4 or
3.children aged less than 5 years.

28. Blood Collection, Storage, and Transport

29. Currently, the diagnosis and monitoring of HIV infection is performed on blood specimens
●For serological tests (antigen and antibody detection)
Serum/plasma/whole blood is used,
●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.

30. 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).
●Label the tube for blood collection with at least two patient identifiers.
●Wash hands or disinfect with an alcohol based hand sanitizer.
●Put on gloves to comply with standard precautions.
●Place the individual’s in a supine or sitting position with arm supported under good light.
●For the avoidance of soiling, place absorbent material below the forearm before commencing
venepuncture.

31. ●Explain the procedure briefly to the person.
●Assess the individual’s veins for venepuncture.
●Apply the tourniquet
●Clean the venepuncture site with 70 percent alcohol or povidone-iodine
using a circular motion,
●Immobilize the vein by pressing 1 inch to 2 inches below the
venepuncture site, drawing the skin taut.

32. ●Do not recap the needle. Burn/cut the used needle in a needle destroyer.
●Dispose-off the cut needle and/or syringe into a puncture proof sharps
container.
●Discard cotton balls and spirit swabs into an infectious waste container.
●Discard wrapper/cover/cap of the needle into a non-infectious waste
container.
●Discard gloves into an infectious waste container and wash hands.

33. Specimen Transport
●Shipment of infectious agents is permitted as per the International
Air Transport Association’s (IATA) Regulations.
●HIV infected specimens are classified as infectious class 6.2
substances under the United Nations’ (UN) no. 2814.
●The packaging must adhere to UN class 6.2 specifications.
●Packaging requires a 3-layer system

34. ●The specimen tube, in which serum is to be transported, should
not have cracks/leaks.
●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).

35. ●Pack this container inside a cardboard box containing sufficient
material (cotton gauze) to absorb the blood in case the tube
breaks or leaks.

●Cap the box tightly.

36. ●Fasten the request slip securely to the outside of this box.
●This request slip should have all of the patient’s details (i.e., name,
age,Gender, risk factors, history of previous testing, etc.) and should
accompany the specimen.
●The request slip should be placed in a plastic zip lock bag to prevent
smudging on account of spillage.
●For mailing, this box should be placed inside another box containing
the mailing label and a biohazard sign.

38. HIV SCREENING TEST
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.

39. PRINCIPLE
Retroquic-HIV test comprises of a test device striped with distinct bands of
purified gp 120 and gp 41 synthetic peptide specific to HIV 1 at test region '1' and
gp 36 synthetic peptide specific to HIV 2 at test region '2'.
The third band striped at region 'C' corresponds to the assay performance control.

40. First the membrane assembly is hydrated with wash buffer and then the specimen is added.
Antibodies to HIV 1 and/or 2 if present, are captured by the respective antigens.
After washing with wash buffer, Protein A conjugated gold sol reagent is added to reveal the
presence/ absence of bound antibodies.
Post final wash a positive reaction is visualized by the appearance of purple coloured bands at the
test region '1' and/or '2'.
The absence of bands at test region '1' & '2' is a negative test result.
The appearance of control band serves to validate sample addition, reagent and assay
performance.

42. Kit Components:

Retroquic-HIV immunoconcentration test kit for HIV1 and HIV 2 antibodies comprises of
the following components:
1. Ready to use individually pouched, flow through test devices striped with HIV 1
specific purified synthetic peptides at test region '1' and HIV 2 specific purified synthetic
peptides at test region '2' and a blue dyed protein A based control band at region 'C' along
with a specimen dropper and desiccant pouch.
2. Dropper bottle with ready to use wash buffer solution.
3. Dropper bottle with ready to use protein A conjugated gold sol solution.

43. STORAGE AND STABILITY
The unopened Retroquic-HIV kit, as well as kit components upon opening, must be stored
at 2-8°C, till the duration of the shelf life as indicated on the kit / kit component labels.

44. SPECIMEN COLLECTION AND PREPARATION
1. No prior preparation of the patient is required before sample collection by approved techniques.
2. Fresh serum / plasma is preferable. Serum / plasma may be stored at 2-8°C upto 24 hours in case of delay in
testing. For long term storage, freeze the specimen at -20°C.
3. Repeated freezing and thawing of the specimen should be avoided.
4. Do not use haemolysed, clotted, contaminated, viscous / turbid specimen.
5. Specimen containing precipitates or particulate matter must be centrifuged and the clear supernatant only used for
testing.
6. Do not heat -inactivate the specimen.
7. Frozen samples for retrospective studies must be centrifuged at 3000 rpm for 15 minutes and the clear supernatant
must be used for tests.

45. TEST PROCEDURE
1. Bring all reagents and specimen to room temperature (25 - 30°C) before use. Tighten the Wash Buffer solution and Protein A Gold
Conjugate dropper bottle caps in a clockwise direction to pierce the respective dropper bottle nozzles. The addition of specimen / reagents
must be done at the centre of the reaction port, holding the sample dropper / dropper bottles in a vertical position. Ensure the drops are free
falling. Use a new sample dropper for each specimen to avoid cross contamination.
2. Tear open the foil pouches and retrieve the required number of Retroquic-HIV membrane test devices and label appropriately.
3. Add two drops of wash buffer into the reaction port of the device and allow to soak through completely.
4. Using the sample dropper provided, add one drop of the serum / plasma specimen into the reaction port. Allow to soak through completely.
5. Add three drops of wash buffer to the reaction port and allow to soak through completely.
6. Add two drops of protein A gold conjugate to the reaction port and allow to soak through completely.
7. Add two drops of wash buffer and allow the wash buffer to soak through completely.
8. Read and record the results immediately.

47. REFERENCES
National_Guidelines_for_HIV_Care_and_Treatment_2021
National_Guidelines_for_HIV_Testing
Lippincott illustrated review of microbiology
RETROQUIC-HIV-(Device)-IFU