Laboratory diagnostics CH/HIV/0052/17/10/2017
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1 Laboratory diagnostics CH/HIV/0052/17/10/2017
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3 Contents Primary diagnostics Therapy monitoring Therapy optimisation Co-infections and co-morbidities 3
4 HIV in the laboratory Primary diagnostics Antibody screening test screening, preferably 4th generation (antibody + antigen) Confirmation test immunoblot or also via an NAT (nucleic acid amplification test), only done by 11 regional registered laboratories The regional registered laboratories are selected by the Federal Office of Public Health (FOPH) and are under the control of Swiss National Center for Retroviruses (SNCR). Therapy monitoring using 'surrogate markers HIV-1 or HIV-2 viral load HIV-2 only done by regional registered laboratories if diagnosis is confirmed Establishment of immune cell count particularly CD4+ T lymphocytes 4 HIV diagnostics guidelines 2015 DOI /s x Bundesamt fur Gesundheit. Das schweizerische HIV Testkonzept eine aktualisierte Ubersicht. BAG Bulletin, :
5 HIV in the laboratory Therapy optimisation Resistance tests genotype-based at baseline stage and every time therapy fails, only done by 4 of the regional registered laboratories: Bern, Genf, Lausanne and NZR Zurich Tropism test before any proposed use of a co-receptor antagonist HLA B*5701 status before any proposed use of ART containing abacavir Plasma level particularly in the event of non-compliance, absorption problems, or interactions Routine laboratory testing Normal patient management 5 HIV diagnostics guidelines 2015 DOI /s x Bundesamt fur Gesundheit. Das schweizerische HIV Testkonzept eine aktualisierte Ubersicht. BAG Bulletin, :
6 Primary diagnostics CH/HIV/0052/17/10/2017
7 HIV testing 1st step: screening test ELISA test: combined antibody and antigen test (p24) Specificity: >99.5% confirmation test required False reactive: e.g. pregnancy, vaccinations, viral infections Fast test: for emergency scenarios (surgery, needlestick injurie or childbirth) 2nd step: confirmation test Evidence of antibodies in the immunoblot: Result: Reactive for antibodies positive No antibodies: false reactive signal in screening test negative Alternatively: evidence of HIV RNA (also diagnosis during seroconversion possible) Caution: diagnostic window Evidence of HIV RNA (PCR): 11 days on average following infection (median) 3rd generation ELISA or fast test: up to 12 weeks following infection (median) 4th generation ELISA: up to 6 weeks following infection (median) Home tests not recommended by the Swiss Federal Commission for Sexual Health No antigen component (more false negative results) Lack of support if positive 7 HIV diagnostics guidelines 2015 DOI /s x Hoffmann C. and Rockstroh J. K., HIV 2015/2016 ( Bundesamt fur Gesundheit. Das schweizerische HIV Testkonzept eine aktualisierte Ubersicht. BAG Bulletin, : Artikel «Wer es nicht wissen will, macht auch keinen Heimtest», von Prof. Dr. Pietro Vernazza Swiss, Aids News 2.Juli 2014
8 HIV testing Screening test ELISA test: combined antibody and antigen test (p24) Specificity: >99.5% confirmation test required False reactive: e.g. pregnancy, vaccinations, viral infections Fast test: for emergency scenarios (surgery, needlestick injuries or childbirth) Negative Reactive Diagnostic window HIV RNA (PCR): approx. 11 days following infection (median) 3rd generation ELISA or fast test: up to 12 weeks following infection (median) 4th generation ELISA: up to 6 weeks following infection (median) Negative Confirmation test Evidence of antibodies in the immunoblot* Positive Reactive for antibodies *Alternatively, evidence of HIV RNA (diagnosis during seroconversion possible) 8 HIV diagnostics guidelines 2015 DOI /s x Hoffmann C. and Rockstroh J. K., HIV 2015/2016 ( Bundesamt fur Gesundheit. Das schweizerische HIV Testkonzept eine aktualisierte Ubersicht. BAG Bulletin, :
9 HIV testing the technical concept The following requirements must be met for diagnosis of HIV-positive in accordance with the criteria of the FOPH HIV test concept : 1. Reactivity in at least two different types of HIV tests 2. Reactivity in at least two different test materials (to exclude mix-ups) 3. Identification of the HIV type (HIV-1 and/or HIV-2) 4. In the case of HIV-1, identification of the virus group (M or possibly O, N, P) - this requires genetic resistance testing 5. Quantification of plasma HIV-1 RNA (viral load 6. Classification as new (up to 12 months) or older infection 7. HIV genetic resistance test with identification of the HIV subtype 9 Modified from: Landesweite standartisierter Bestätigungsprozess für HIV-Infektionen,
10 HIV testing in practice Clarification/Consent Document consent Signature not required Counselling/Clarification Step-by-step diagnostic approach Limitations due to diagnostic windows Notification of findings Negative: also possible by phone Positive: Only face-to-face with a doctor Help with finding a practice specialising in HIV Information regarding the advice available from AIDS support agencies Anonymous reporting only for the gathering of epidemiological data (FOPH) HIV testing can be performed by any doctor. Support and treatment for HIV infection should be provided by experts! 10 Hoffmann C. and Rockstroh J. K., HIV 2015/2016 ( Bundesamt fur Gesundheit. Das schweizerische HIV Testkonzept eine aktualisierte Ubersicht. BAG Bulletin, :
11 Therapy monitoring CH/HIV/0052/17/10/2017
12 Recommendations from EACS Guidelines: monitoring of HIV infection Specifications Upon HIV diagnosis Before therapy starts Follow-up frequency Confirmation of positive HIV antibody test + Virology Immunology HIV viral load in plasma + + Every 3-6 months Genotype-based resistance test + +/- and in the and identification of subtypes event of R5 tropism (if available) + +/- virological failure Absolute and relative CD4 count HLA B*5701 (if available) + + Every 3-6 months + +/- 12 Modified from: EACS Treatment Guidelines; Version 8.0. October 2015
13 Measurement of therapy success using the surrogate markers viral load and CD4 cell count Maximum and lasting suppression of viral load Viral load permanently below 50 copies/ml Restoration and preservation of immune function Target: more than 200 CD4 cells/mm 3 * Antiretroviral therapy Antiretroviral therapy 13 * The risk of AIDS-defining conditions increases significantly at <200 CD4 cells/mm 3 Hoffmann C. and Rockstroh J. K., HIV 2015/2016 ( Image: Gölz J., Understanding and Living with HIV. 2014
14 Measurement of viral load Antiretroviral therapy Commercial systems: RT-PCR, b-dna, and NASBA Thresholds for detection between 20 and 250 copies/ml Results may differ significantly depending on the system used Best to stick with one system if possible The clinically relevant threshold for detection is 50 copies/ml Important to take into account any deviation associated with the method used Viral load may increase temporarily following vaccinations or infections RT-PCR: reverse transcriptase; PCR: polymerase chain reaction; b-dna: branched DNA; NASBA: nucleic acid sequence-based amplification assay 14 Hoffmann C. and Rockstroh J. K., HIV 2015/2016 ( Image: Gölz J., Understanding and Living with HIV. 2014
15 Monitoring of the CD4 cell count Normal ranges: 500-1,000 CD4 cells/mm 3 Proportion of CD4 cells >30% Ratio CD4/CD8 >1.2 Laboratory-related deviations (in the order of CD4/mm 3 ) associated with measurements, particularly at higher values Repeat measurement if values appear implausible Antiretroviral therapy 15 Hoffmann C. and Rockstroh J. K., HIV 2015/2016 ( Image: Gölz J., Understanding and Living with HIV. 2014
16 Therapy optimisation CH/HIV/0052/17/10/2017
17 Genotype-based resistance test and tropism test 1. Gene sequencing 2. Determining the sequence 3. Interpretation using algorithms vpu LTR gag pol vif 9.7 kb vpr env rev tat nef LTR pol sequencing Protease protease inhibitors Reverse transcriptase NRTIs, NNRTIs Integrase integrase inhibitors env sequencing gp41 fusion inhibitors V3 loop CCR5 antagonists 17 Low A. J. et al.,aids Rev. 2008;10:
18 HLA test The hypersensitivity reaction (HSR) to abacavir is associated with the HLA-B*5701 allele The risk for abacavir HSR to occur is significantly increased for patients who test positive for the HLA-B*5701 allele. However, abacavir HSRs have been reported in patients who do not carry this allele A pharmacogenetic test can be used to determine the HLA-B*5701 status (to identify patients at high risk for abacavir HSR) 2 ml of EDTA blood is required for analysis Before initiating treatment with abacavir, screening for HLA-B*5701 should be performed. Abacavir is contraindicated in patients known to carry the HLA-B*5701 allele The patient must provide consent* 18 * According to "Bundesgesetz über genetische Untersuchungen beim Menschen" Mallal S. et al., N Engl J Med 2008; Ma JD et al. PLoS Curr. 2010, 2: RRN1203
19 Monitoring of co-infections and co-morbidities
20 Recommendations from EACS Guidelines: monitoring of co-infections Specifications Upon HIV diagnosis Before therapy starts Follow-up frequency STIs Viral hepatitis Tuberculosis Others Syphilis + Annually/As STI screening + required HAV + HCV + HBV + + X-ray examination + PPD if CD4 cell count >400 + IGRA in selected high-risk populations (if available) Varicella zoster virus serology + Measles/rubella serology + CMV serology + Screening for tropical diseases (e.g. Schistosoma serology) + Leishmania serology +/- +/- Annually/As required Repeat screening following exposure 20 PPD: purified protein derivative IGRA: interferon-gamma release assay Modified from: EACS Treatment Guidelines; Version 8.0. October 2015
21 Recommendations from EACS Guidelines: monitoring of co-morbidities Specifications Upon HIV diagnosis* Before therapy starts Follow-up frequency Haematology Full blood count + + Every 3-12 months Body Body mass index + + Annually Cardiovascular Risk assessment (Framingham Risk Score) + + Every 2 years ECG + +/- As required 21 Blood pressure Blood pressure measurement + + Annually Lipids TC, HDL-c, LDL-c, TG + + Annually Glucose Serum glucose + + Every 6-12 months Lungs X-ray +/- As required Liver Kidneys Bone Risk assessment + + Annually ALT/AST, ALP, bilirubin + + Every 3-12 months Risk assessment + + Annually egfr (CKD-EPI) + + Every 3-12 months Dipstick + + Annually Bone profile: calcium, phosphate, ALP + + Every 6-12 months Risk assessment (FRAX >40 years of age) + + Every 2 years Vitamin D 25(OH) vitamin D + As required Neurocognitive deficits Screening questionnaire + + As required Depression Questionnaire + + As required *In addition, upon HIV diagnosis: haemoglobinopathies, G6PD, COPD screening Modified from: EACS Treatment Guidelines; Version 8.0. October 2015
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