HIV Testing Throughout the Ages

1
HIV Testing Throughout the Ages

• Ronald D. Wilcox MD FAAP
• Asst. Professor of Medicine and Pediatrics LSU
  Health Sciences Center - New Orleans
• PI/Medical Director Delta AETC

2
www.deltaaetc.org
504-903-0788
LA Coordinator Dana Gray
HIV Trainer Christina Price Nurse Educators
Sheryl Thornton, Edwina McBride
3
Objectives

• To review the appropriate testing for HIV in
  patients based on their age and clinical
  presentation
• To review the acute retroviral syndrome
• To review rapid testing for HIV
• To review the new CDC guidelines for HIV
  screening from 2006

4
Case

• 21 year old woman comes to your clinic for the
  first time. She has no significant past medical
  history except occasionally has candidal
  vulvovaginitis.

5
Antibody Testing

• ELISA-EIA
• Western Blot
• Indirect Immunofluorescence Assay (IFA)

6
Enzyme-Linked ImmunoSorbent Assay (ELISA) HIV
Enzyme Immunoassay (EIA)
Photograph courtesy of http//www.alibaba.com/
7
ELISA - EIA

• Apply a sample of HIV antigens
• Apply patients serum containing antibodies
• Apply second antibody
• (anti-human immunoglobulins)
• Apply a color changing substrate

Positive ELISA
Negative ELISA
8
ELISA - EIA
9
Western Blot (Immunoblot)

• Detects proteins in a given tissue sample
• HIV infected cells are opened and their proteins
  placed into a gel
• Proteins are separated by mass using gel
  electrophoresis
• Separated proteins are transferred from gel to a
  membrane (blotting)

Photograph courtesy of Wikipedia Encyclopedia
10
Structure of HIV
Illustration from SCIENCE TECHNOLOGYAugust
27, 2001 Volume 79, Number 35 CENEAR 79 35 pp.
37-44
11
Western Blot
Controls
Samples

• Procedure continues similar to ELISA
• Results are read
• If no viral band are present, the result is
  negative
• If less than the required number of viral bands
  are detected, the result is indeterminate

-

-
Indeterminate
Picture court4esy of www.msichicago.org/ed/AIDS/hi
vtst5.htm
12
Western Blot

• Positive Western Blot
• 3 bands representative of the three major gene
  products of HIV gp120/160, gp41, p24
• or, 2 of 3 in high risk individuals
  

13
Interpreting HIV Serology

• HIV Positive
  ELISA AND
  Western Blot

14
Interpreting HIV Serology

• HIV Indeterminate Positive
  ELISA
  Only 1 band present on WB

15
Explanations for HIV Indeterminate

• Early infection, seroconverting
• Advanced infection with decreased p24 antibodies
• Cross-reactive antibodies or auto-antibodies from
  CVD, autoimmune process, or malignancy
• HIV-2 infection O clade HIV-1
• Experimental HIV vaccine recipient

16
Interpreting HIV Serology
17
Case

• The patient tests negative by ELISA/Western Blot.
• 1 year later she returns. She reports a sore
  throat recently and 10 weight loss over the past
  2 weeks. She has a new sexual partner for the
  past 2 months.

18
Case

• On physical exam you note
• A faint erythematous diffuse rash
• Diffuse lymphadenopathy
• Splenomegaly
• Mild erythema of her posterior oropharynx

19
HIV Infection
20
Acute Retroviral Syndrome

• Primary HIV infection
• Often overlooked
• 50 - 90 experience some symptoms
• Symptoms occur 1-3 weeks after exposure
• Seroconvert 6-12 weeks after exposure

21
Acute Retroviral Syndrome- Symptoms

• Fever (96)
• Lymphadenopathy (74)
• Pharyngitis (70)
• Rash (70)
• Myalgia/arthralgia (54)

22
Rash of Acute Retroviral Syndrome
23
Acute Retroviral Syndrome- Symptoms

• Headache
• Diarrhea
• Nausea and vomiting
• Hepatosplenomegaly
• Weight loss
• Thrush
• Neurologic symptoms

24
Acute Retroviral Syndrome- Differential Diagnosis

• Mononucleosis
• Cytomegalovirus
• Primary HSV
• Viral hepatitis
• Rubella
• Toxoplasmosis

• Secondary syphilis
• Measles
• Disseminated GC
• Pityriasis rosea
• Acute rheumatologic
• Drug reaction

25
APTIMA HIV-1 RNA Qual Assay

• Used for either acute or chronic infection
• Can be found early in infection before antibodies
  are formed
• Still needs verification

26
Case

• The patients p24 antigen is negative and ELISA
  done 3 months later is also negative.
• The patient is lost-to-follow-up for care. She
  presents to the hospital 2 years later where you
  are moonlighting, in labor, pregnant at 36 weeks.

27
Rapid Testing

• Currently six rapid tests are available in the US
  
• OraQuick Rapid HIV-1/2 Antibody Test
• Reveal Rapid HIV-1 Antibody Test
• Uni-Gold Recombigen HIV Test
• Multispot HIV-1/HIV-2 Rapid Test
• HIV 1 / 2 Stat Pak
• Sure Check HIV 1 / 2 Assay
• All are interpreted visually

28
OraQuick Rapid HIV-1 Antibody Test

• Easy to use untrained first-time users report
  98.5 of results correctly
• Read test results in 20-60 minutes
• Sensitivity 99.3 - 99.6
• Specificity 99.8 - 100
• CLIA-wavered for oral fluid and whole blood

29
OraQuick Advance

• Nitrocellulose strip
• Test Region (T)
• Synthetic gp41 peptides representing the HIV-1
  envelope
• Synthetic gp36 region of the HIV-2 envelope
• Control Region (C)
• Goat antihuman IgG

Photograph courtesy of the CDC
www.cdc.gov/hiv/rapid_testing
30
Obtain a finger stick specimen
Insert loop into vial and stir
Photographs courtesy of the CDC
www.cdc.gov/hiv/rapid_testing
31
OraQuick Advance
Photographs courtesy of the CDC
www.cdc.gov/hiv/rapid_testing
Collect oral fluid specimens by swabbing gums
with test device.
32
OraQuick Advance

• Insert Device
• Read test in 20-40 minutes

Photograph courtesy of the CDC
www.cdc.gov/hiv/rapid_testing
33
Interpreting the Results

• HIV antibodies bind to peptides causing a red
  line to appear in the T
• location

• Red lines at both T and C locations indicate a
  valid reactive test
• A red line at only the C location indicates a
  valid negative test

Reactive Control
Positive HIV-1/2

• If no red line appears, the
• test is invalid

Photograph courtesy of the CDC
www.cdc.gov/hiv/rapid_testing
34
Uni-Gold Recombigen

• Whole blood or serum/plasma
• CLIA-wavered for whole blood
• Sensitivity 100
• Specificity 99.7 99.8

35
Uni-Gold Recombigen
Photographs courtesy of the CDC
www.cdc.gov/hiv/rapid_testing
36
Uni-Gold Recombigen
Positive
Negative
Photographs courtesy of the CDC
www.cdc.gov/hiv/rapid_testing
37
Reveal G-2 / Multi-Spot

• Serum or Plasma
• NOT CLIA-wavered
• Sensitivity 99.8 - 100
• Specificity 98.6 99.9
• Multi-Spot also detects HIV-2

38
Reveal Rapid HIV-1 Antibody Test

• Takes 3 minutes to run
• Test is usually run in a clinical laboratory
• Utilizes centrifuged blood and plasma samples
• Utilizes several reagents

Photograph courtesy of the CDC
www.cdc.gov/hiv/rapid_testing
39
Reveal Rapid HIV-1 Antibody Test
2
1
Reactive
4
3
Negative
40
Multispot HIV-1/HIV-2 Rapid Test

• Utilizes serum and plasma to distinguish between
  HIV-1 and HIV-2
• Test consists of one test cartridge and five
  reagents
• Reagents must be refrigerated
• Test can be completed in 15 minutes

Photographs courtesy of the CDC
www.cdc.gov/hiv/rapid_testing
41
Multispot HIV-1/HIV-2 Rapid Test
HIV-1
Reactive Control
HIV-1
HIV-2
Negative
HIV-1 HIV-2 Positive
42
Additional Rapid Tests
HIV 1 /2 STAT-PAK
SURE CHECK HIV 1/2 ASSAY
43
Rapid Testing

• Appropriate settings
• CBO Outreach
• Late-Presenting Mothers
• Occupational Exposures
• Emergency Rooms
• Providers Offices

44
Case

• The patients HIV rapid test was positive.
• Confirmation by ELISA and Western Blot were also
  positive
• The infant is the brought to the pediatrician 2
  weeks later.

45
(No Transcript)
46
HIV Testing in Infancy

• HIV cDNA PCR
• HIV-1 nucleic acid detection by PCR assay of cDNA
  extracted from peripheral blood mononuclear cells
• Test range 1-10 DNA copies
• Single test (1-36 months of age)
• Sensitivity 95
• Specificity 97
• False negatives non-B subtypes

47
HIV Testing in Infancy

• Recommended testing schedule for cDNA PCR
• Within first 48 hours
• Do NOT use umbilical cord blood
• 1-2 months
• 2-4 months
• (4-6 months)
• Positive test repeat as soon as possible on a
  second blood sample

48
HIV Testing in Infancy

• Other tests
• HIV ELISA / Western blot
• Up to 18 months or longer would be testing for
  maternal antibodies
• p24 Antigen
• Less sensitive
• False positives in first month of life
• HIV culture
• Expensive and not readily available
• HIV RNA PCR
• Negative test NOT exclusionary in patients lt 18
  months of age
• Can have false positives at low levels

49
HIV Testing in the Toddler

• If a child had perinatal testing that was
  negative
• HIV ELISA / Western Blot are then ordered at
  12-18 months of age
• Can also be considered negative if 2 blood
  samples obtained at least one month apart after 6
  months of age are negative

50
Case

• The patients 54 year old mother comes with the
  patient at the next visit. She reports that she
  has only had problems with hypertension and
  occasional bladder infections. She has never had
  any STDs. She states that her husband died about
  5 years ago and she has only had one other sexual
  partner since his death. She asks whether she
  should have an HIV test since her daughter is
  positive.

51
CDC Guidelines for Screening and Testing
52
HIV Serologic Screening Recommended

• Persons between 13 and 64 years of age routinely
  offered at entry to care
• Persons with STDs
• Persons with tuberculosis

53
HIV Serologic Screening Recommended

• Women considering conception and pregnancy
• All pregnant women
• Women in delivery with undocumented HIV status
• Infants born to mothers of undocumented HIV status

54
HIV Diagnostic Testing

• Signs or symptoms of HIV infection
• Patients at high risk for HIV based on risk
  assessment, offered yearly

55
HIV Serologic Testing Recommended

• ANYONE WHO REQUESTS TO BE
  TESTED
• Consider testing anyone who gets admitted to the
  hospital with an infectious process

56
Opt-Out Testing

• Oral or written information given at time of
  testing at an appropriate health literacy level
  for the patient
• Explaining what HIV infection is
• Describing ways to prevent transmission
• Meaning of positive and negative results
• Testing is voluntary and never coerced and prior
  knowledge is still needed. Patient then given
  the opportunity to decline testing
• If refuses, explore reasons. Offer at subsequent
  visits.
• Document refusal in patient record
• General consent for medical care is sufficient
  for HIV testing

57
Giving results

• Negative test results
• Can be conveyed without personal direct contact
• High risk patients should be encouraged to get
  retested in future
• Positive results
• Conveyed in private setting in person
• Assure confidentiality. Do NOT use family
  members as translators
• Discuss partner notification
• Document in patient record
• Refer for care

58
Other considerations

• CDC recommendations do not supercede local laws
• Facilities and institutions may have their own
  requirements
• Rapid results reported in 20-30 minutes and
  substantially decrease amount of patients not
  receiving results but positive results MUST be
  confirmed with ELISA/Western Blot

59
Referral to Specialty Care

• Consider referral to HIV specialist if
• CD4 count 350 or lower
• Viral load gt 100,000 copies/ml
• Pregnant
• In the acute retroviral phase
• Symptomatic
• Has accompanying co-morbidities, such as TB,
  hepatitis B or C
• Has an ADIS-related malignancy, ie NHL, Kaposi
  sarcoma

60
FACES 821-4611

• Program Director Barbara Brown
• Asst Director/Health Ed Supervisor Claudia
  Medina
• Medical Director Ronald Wilcox
• Regional Program Coordinator Jamie Segura
• Hispanic Program Coordinator Oscar Salinas
• Case Managers Adranell Murray, Kennie Emezie,
  Anabella Salazar-Issusi, Karen Wright
• Treatment Adherence Nurses Carrie Galpin, Donna
  Stuart, Tabitha Blann
• Mental Health Specialist Mary Boutte, Natalie
  Griffin
• Decision Support Analyst Amal Elfarra
• Peer Advocate Mimi Alvarado
• Family Advocacy/Community Liaison Gina Brown
• Secretary Jeanne Eddington
• Direct Intake Specialist Amanda Raker

61
Summation

• Infants
• HIV cDNA PCR up to 15 months of age
• 18 months of age or older
• Chronic infection ELISA / Western Blot
• Acute infection p24 antigen, APTIMA testing
• Rapid testing late presenting mothers, office
  setting, emergency rooms
• CDC Guidelines for testing offered to everyone
  13-64 years of age at entry to care, repeat based
  on risk

62
Contact Information

• DAETC www.deltaaetc.org
• Phone [phone]
• Rwilco_at_lsuhsc.edu
• Office 504-903-7301
• Pager 504-363-1692