Tests and monitoring in HIV
infection
UK standard of care and some other useful
tests
Matthew Williams
UK CAB
Tests and monitoring in HIV
infection
UK standard of care and some other useful
tests
Tests and monitoring in HIV
infection
CD4 count
Viral load
Resistance
Therapeutic drug monitoring
Tests and monitoring in HIV
infection
CD4 count
Blood test
●Used to judge how far HIV disease has
advanced
●Helps predict the risk of opportunistic
infections
●Most useful when it is compared with the
count obtained from an earlier test.
●
Tests and monitoring in HIV
infection
CD4 count
CD4 dips on HIV infection from a normal
count of 500-1,500 cells in a cubic millimeter
(mm3) of blood (a drop, more or less),
recovers somewhat, then falls over time
down to as low as 0.
Tests and monitoring in HIV
infection
CD4 count
CD4 <200 = greater risk of opportunistic
infections (OIs) = “AIDS” = threshold for
prophylaxis eg for PCP = bottom of UK
threshold for starting combination therapy
CD4 <50 = very great risk of OIs
Tests and monitoring in HIV
infection
CD4 count
CD4 over 350: treatment not recommended
CD4 any count: treatment recommended if
“symptomatic”
BHIVA, Treatment of HIV-infected adults with antiretroviral therapy
(2006)
Tests and monitoring in HIV
infection
CD4 count
CD4 count lower in pregnancy – temporary
drop of 50 cells/mm3
Tests and monitoring in HIV
infection
CD4 count
CD4% - CD4 cells as proportion of all
lymphocytes (white blood cells), normally
about 40% in adults
CD4% is used to monitor babies and
children who have higher CD4 counts
Tests and monitoring in HIV
infection
Viral load
Blood test (can be other serum eg CSF)
●Used to judge whether treatment is working
(early infection?)
●Helps predict the risk of disease
progression?
●Most useful when it is compared with the
count obtained from an earlier test.
●
Tests and monitoring in HIV
infection
Viral load
After infection, viral load surges to a very
high for the first weeks or months
Often 1,000,000+ copies in a millilitre (mL) of
blood, when you are very infectious
Viral load falls as the body controls HIV
infection then rises over time as immunity is
damaged
Tests and monitoring in HIV
infection
Viral load
When you are on HIV treatment, your viral
load should be reduced to “undetectable” =
<50 copies/mL = 1.7 log10
Tests and monitoring in HIV
infection
Viral load
Blip = 1 viral load test detectable (over 50
copies) – 2 of these may be a trend and
indicate “virological failure”
Tests and monitoring in HIV
infection
Viral load
Viral load is usually a PCR (polymerase
chain reaction) quantitative (counting) test
for HIV RNA - research tests can measure
viral load below 50 copies
Other tests: bDNA (branched DNA), NASBA
(nucleic acid sequence based amplification)
Tests and monitoring in HIV
infection
Resistance test
Blood test (can be other serum eg CSF)
●Used to judge whether treatment will work
●Involves interpretation
●Two types: genotype and phenotype
●Viral load needs to be over 500 copies/mL
for the test to work
●
Tests and monitoring in HIV
infection
Resistance test
Resistance test
Tests and monitoring in HIV
infection
Resistance test
Genotype looks at genetic make up of HIV
viruses in infection and compares this to
what is known about which mutations lead
to resistance
Phenotype looks at whether the drugs work
in a test tube
Tests and monitoring in HIV
infection
Resistance test
Minority species of resistant virus may be missed
by conventional resistance testing (ie if less than
10% of your virus is resistant the test may not pick
this up).
In patients without evidence of transmitted
resistance, a suboptimal virological response to
first-line therapy (<1 log10 copies/mL drop in viral
load by 4–8 weeks) should prompt resistance
testing at that time.
BHIVA, Treatment of HIV-infected adults with antiretroviral therapy
(2006)
Tests and monitoring in HIV
infection
Resistance test
Tests and monitoring in HIV
infection
Therapeutic drug monitoring (TDM)
Measures drug levels in the blood - requires
blood samples at recorded time intervals after a
drug dose
Practical uses for NNRTIs and PIs
Tests and monitoring in HIV
infection
Therapeutic drug monitoring (TDM)
Freely available at low (£45/drug) or no cost (if
covered by drug company) from University of
Liverpool Department of Pharmacology.
http://www.hiv-druginteractions.org/
http://www.delphicdiagnostics.com/
Tests and monitoring in HIV
infection
Therapeutic drug monitoring (TDM)
Drug levels vary – much evidence of this –
recommended dose is based on averages
University of Liverpool TDM audit – 20-25% of
children on NNRTIs or PIs using drugs below
therapeutic level.
http://www.i-base.info/htb/v7/htb7-6/University.html
Tests and monitoring in HIV
infection
UK standard of care (BHIVA)
i)
ii)
iii)
iv)
All patients should have:
a resistance test at diagnosis,
before starting HAART,
if viral load does not drop by <1log10 after 48 weeks after starting HAART, (genotype)
after virological failure
Tests and monitoring in HIV
infection
UK standard of care
BHIVA – viral load before and 4-8 weeks
after starting treatment (as necessary
thereafter)
Tests and monitoring in HIV
infection
UK standard of care
BHIVA – CD4 count before treatment (as
necessary thereafter)
Tests and monitoring in HIV
infection
UK standard of care
BHIVA – TDM for management of drug
interactions, pregnancy and paediatrics,
highly treatment-experienced patients when
TDM and resistance test results can be
integrated, kidney or liver impairment,
transplant patients, drug toxicity, alternative
dosing where safety and efficacy has not
been established
Tests and monitoring in HIV
infection
UK standard of care and some other useful
tests
Tests and monitoring in HIV
infection
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Liver enzymes
Kidney function
Urine
Albumin
Bone density
Glucose
Platelets
Red blood count
White blood count
Lipids
Tests and monitoring in HIV
infection
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C-reactive protein
DEXA scan
Chest x-ray
Sputum serology
Lactic acid and blood pH
Haemoglobin and iron
Serum urea
Creatine phosphokinase and lactate
dehydrogenase
Electrolytes
Tests and monitoring in HIV
infection
Tests = blood, blood and more blood...
Many tests require a blood sample
Tests and monitoring in HIV
infection
Blood glucose
Blood cholesterols and triglycerides
Kidney function
C-reactive protein
Liver enzymes
Anaemia
Lactic acidosis
Tests and monitoring in HIV
infection
Blood glucose
Normal range 4-8 millimoles per litre
(µmol/L)
High glucose ?= kidney disease,
neuropathy, insulin resistance,
cardiovascular disease
Drugs: PIs, tenofovir, AZT?
Tests and monitoring in HIV
infection
Blood cholesterols and triglycerides
Low-density lipoproteins (LDLs) or "bad"
cholesterol, and high-density lipoproteins
(HDLs) or "good" cholesterol – ratio is key
measure
Insulin resistance, metabolic syndrome,
cardiovascular disease
Drugs: PIs, AZT, efavirenz
Tests and monitoring in HIV
infection
Blood cholesterols and triglycerides
Total cholesterol
Target level under 5.2 µmol/L (4-6.4 µmol/L)
Tests and monitoring in HIV
infection
Blood cholesterols and triglycerides
LDL cholesterol
Target level under 3.4 µmol/L
Tests and monitoring in HIV
infection
Blood cholesterols and triglycerides
HDL cholesterol
Target level over 0.9 µmol/L
Tests and monitoring in HIV
infection
Blood cholesterols and triglycerides
Cholesterol – CV risk cannot be judged on
cholesterol levels alone, HDL:LDL ratio,
lifestyle, BMI, age and other factors are
important
Smoking!
Tests and monitoring in HIV
infection
Blood cholesterols and triglycerides
Triglycerides
Fasted levels <2.2 µmol/L normal, 2.2–4.4
µmol/l borderline, >11 µmol/L very high
Each +1.1 µmol/L increased the risk of a
heart attack by about 25% in men and 60%
in women
Tests and monitoring in HIV
infection
Urine
Dipstick test
Protein – kidney (tenofovir) << mostly albumin
●Glucose – insulin resistance (PIs), kidney
(tenofovir, indinavir?, atazanavir?)
●Bilirubin – liver ( X atazanavir)
●
Sensitive but not specific
Tests and monitoring in HIV
infection
Kidney function
Urine dipstick test or blood test = first test
and is indicative but inconclusive
Tests and monitoring in HIV
infection
Kidney function
Serum creatinine (blood, not very specific
or sensitive)
●Creatinine clearance (blood and/or urine
samples over 24 hours)
●Calculated creatinine clearance (1 blood
sample) – also called estimated glomular
filtration rate (eGFR)
●
Creatinine clearance can increase in
pregnancy
Tests and monitoring in HIV
infection
Kidney function
Different formulas to calculate GFR
http://en.wikipedia.org/wiki/Glomerular_filtration_rate
Tests and monitoring in HIV
infection
C-reactive protein
Risk of diabetes, hypertension and CV disease
Low risk: <1mg/L
High risk: >3mg/L
Drugs: PIs?, AZT?
C-reactive protein is a plasma protein
produced by the liver (so, another blood test...)
Tests and monitoring in HIV
infection
C-reactive protein
C-reactive protein is also marker of
inflammation from infection but seems to be a
reliable marker for CV disease in HIV
regardless of HAART
http://gateway.nlm.nih.gov/MeetingAbstracts/102261383.html
Tests and monitoring in HIV
infection
Liver enzymes
AST, ALT, ALP, GGT and bilirubin
Many conditions, some specific indications,
complicated by hepatitis B and C coinfection,
pharmacological agents (all kinds) and food
Drugs: ritonavir, nevirapine, efavirenz,
tipranavir, atazanavir, indinavir, d4T - most
ARVS and many other drugs may affect the
liver
Tests and monitoring in HIV
infection
Liver enzymes
ALT (alanine aminotransferase)
Normal range 7-30 units/L W, 10-55 units/L M
May be more reliable sign of liver damage
Tests and monitoring in HIV
infection
Liver enzymes
AST (aspartate aminotransferase)
Normal range 9-25 units/L W, 10-40 units/L M
Unreliable sign of liver damage
Pregnancy may decrease AST
Tests and monitoring in HIV
infection
Liver enzymes
ALP (alkaline phosphatase)
Normal range 30-100 units/L W, 45-115 units/L M
Non-specific sign of liver damage
Atazanavir and indinavir can raise ALP
Tests and monitoring in HIV
infection
Liver enzymes
GGT (gamma glutamyl transferase)
Normal range >50 units/L W, >65 units/L M
Can be specific sign of liver damage
Tests and monitoring in HIV
infection
Liver enzymes
Interpretation requires experience and the
whole picture
ALP+ GGT normal = bone disease?
●ALP+ GGT+ = bile ducts? liver damage?
●10 x ALT/AST = viral hepatitis? ARVs?
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Tests and monitoring in HIV
infection
Liver enzymes
Bilirubin
Direct (unconjugated) 0-7 µmol/L
Total 0-17 µmol/L
Bilirubin levels slightly higher in males than
females, black Africans.
Drugs: atazanavir, indinavir
Tests and monitoring in HIV
infection
Liver enzymes
Bilirubin
Jaundice clinically detectable at levels above
40 µmol/l.
Exception: with atazanavir (or ritonavir) if
bilirubin levels around 60-70 µmol/l
Tests and monitoring in HIV
infection
Liver enzymes
Bilirubin ++ jaundice
Other enzymes ++ may show no outward sign
Tests and monitoring in HIV
infection
Liver enzymes
Q: what is the most liver-damaging over-thecounter (OTC) medicine?
Tests and monitoring in HIV
infection
Liver enzymes
Q: what is the most liver-damaging over-thecounter (OTC) medicine?
A: Paracetamol
Tests and monitoring in HIV
infection
Liver enzymes
WHO's top 10 liver-damaging medicines
Paracetamol, troglitazone, valproic acid, d4T,
halothene, 3TC, ddI, amiodarone, nevirapine,
cotrimoxazole
The ABCs of liver disease, Edwin J Bernard, NAM
http://www.aidsmap.com/files/file1000630.pdf
Tests and monitoring in HIV
infection
Liver
PT time – Prothrombin Time
Also called INR - International Normalized
Ratio
Evaluate the ability of blood to clot properly
Not an enzyme test
Tests and monitoring in HIV
infection
Liver
PT time – Prothrombin Time
Monitor anti-coagulants?, bleeding disorders,
before surgery
Normal range 11-13.5 seconds
1.5-2 times normal = too slow but no
consensus on calibration of test as marker of
over-fast clotting
Tests and monitoring in HIV
infection
Liver
PT time – Prothrombin Time
INR = (Pt test / PT normal) ISI
ISI = International Sensitivity Index for tissue factor (1-1.4)
Tests and monitoring in HIV
infection
Anaemia
Iron, B12, B6, folic acid, red blood count,
heamoglobin (HGB), mean corpuscular
haemoglobin (MCH), heamocrit (HCT), mean
corpuscular volume (MCV)
Drugs: AZT (pregnant?)
Tests and monitoring in HIV
infection
Anaemia
Haemocrit
Normal ranges 40-52% M 35-35% W
Low haematocrit = anaemia?
Tests and monitoring in HIV
infection
Anaemia
Haemoglobin
Normal ranges 11.5- to 16.5g W 13-18g M per
100mL blood
Low haemoglobin = anaemia?
Tests and monitoring in HIV
infection
Anaemia
Mean corpuscular volume
Larger = anaemic, B6, B12, folic acid
deficiency?
Smaller = anaemic, iron deficiency?
AZT (and smoking) can increase MCV without
causing anaemia
Tests and monitoring in HIV
infection
Anaemia
Red blood count (total erythrocytes)
Normal range 3.8-5 W 4.5-6.5 M billion per litre
or million per cubic millimitre of blood (1012/L).
Low count = anaemia? but not sensitive or
specific = probably a first test
Tests and monitoring in HIV
infection
The rest...
Blood chemistry
http://www.aidsmeds.com/articles/CSTest_4730.shtml
http://www.aidsmeds.com/articles/CBCTest_4729.shtml
A-Z tests
http://www.aidsmap.com/cms1031936.asp
http://www.labtestsonline.org.uk/
Hepatitis C coinfection – liver and diagnosis
http://www.i-base.info/guides/hepc/livertests.html
http://www.i-base.info/guides/hepc/hcvtesting.html
Tests and monitoring in HIV
infection
Tests to avoid during pregnancy
Amniocentesis
● Chorionic villus sampling
● Fetal scalp sampling
● Cordocentis
● Percutaneous umbilical cord sampling
● Internal fetal labour monitoring (external
ultrasound and fetal monitoring OK)
●
Tests and monitoring in HIV
infection
Plebotomy (having blood taken)
A cruel and unusual punishment - or nice chat
with the nurse?
Tests and monitoring in HIV
infection
Blood count reference ranges
Red blood count (RBC) 3.8 to 5 W 4.5-6.5 M
million per mm3
3
●White blood count (WBC) 4-11 per mm
●Haemoglobin (HGB, Hg) 11.5-16.5 W 13-18 M g
per 100mL
3
●Neutropils 2-7.5 per mm
3
●Lymphocytes 1.3-4 per mm
3
●Platelets 150-440 per per mm
●Mean corpuscular volume (MCV) 80-97
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Tests and monitoring in HIV
infection
Blood count reference ranges
CD4
400-1,600 per mm3
CD4%
32-68%
CD8
140-1000 per mm3
CD4:CD8 ratio 0.9-6
Tests and monitoring in HIV
infection
Sampling
Biggest causes of odd results are ‘sampling
error’, ‘processing error’ or ‘sample
contamination’
wrong tube, wrong person's sample, crosscontamination, sample too hot/cold/old, wrong
reagent, wrongly set up equipment, not
reading instructions, misreading output...
Tests and monitoring in HIV
infection
Results
One result is rarely conclusive
ANY unusual or unexpected results should
ALWAYS be retested before making a
treatment decision
“Normal” is a difficult word – tests refer to
reference ranges, can mean doctor/nurse is
happy with results even though high/low
Tests and monitoring in HIV
infection
Sensitive v specific
Sensitivity = reacts positively
Specificity = reacts positively in right
circumstances
Tests and monitoring in HIV
infection
Sensitive v specific
Sensitivity = rule in
Sensitivity refers to the proportion of people with disease who
have a positive test result
Specificity = rule out
Specificity refers to the proportion of people without disease
who have a negative test result.
Tests and monitoring in HIV
infection
Sensitive v specific
SnNout: when a sign, test or symptom has a high
Sensitivity, a Negative result rules out the diagnosis.
SpPin: when a sign, test or symptom has a high
Specificity, a Positive result rules in the diagnosis.
Tests and monitoring in HIV
infection
Sensitive v specific
Many “rapid” tests are highly sensitive but not
specific enough to be definitive – prone to
sample contamination
Urine dipstick tests
●Fingerprick tests
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