Nye diagnostiske metoder for TB

Transkript

1 U N I V E R S I T Y O F B E R G E N Nye diagnostiske metoder for TB Tehmina Mustafa Professor, Senter for internasjonal helse, UiB Overlege, Lungeavdelingen, HUS

2 Agenda Diagnostiske metoder og ny utvikling Ny diagnostiske metode- EPTB basert på påvisning av mykobakteriell antigen

3 Laboratorieundersøkelser Direkt: Mykobakteria eller antigener Mikroskopi Bakteriologisk dyrkning NAAT (PCR) Antigen påvisning Indirekt: Basert på immunrespons TST/Mantoux Interferon-gamma release assays (IGRAs) Histology/cytology Hematologiske og biokjemiske prøver Serologi- ikke anbefalt

4 Mikroscopi- AFB Sentralt i global TB kontroll Billig USD Rask Behandlingsresponse Fanger opp smitteførende 1 smitteførende nye smitte 1 ny smitteførende Begrensninger: Lav sensitivitet - krever 10,000 bacilli per ml Lav spesifisitet: Atypiske mykobakteria er AFB Utvikling: Automatisering Analysert-datamakin

5 Dyrkning Gull standard Fast eller flytende medium Flytende-MGIT-Raskere -WHO anbefaler SN 80-98% ( bakterier/ml), SP % Fenotypiske resistens, behandlingsresponsen Begrensening: Tar tid: 4-8 uker Rask identifisering - culture isolate Strip speciation test TB-specific antigen (MPT 64)-15 min

6 Nucleic-Acid Amplification Tests (NAAT)- PCR basert metoder Molekylær diagnose i TB- veien fremover Høy SN %, Høy SP % Nyttig- mikroskopi-negativ; Barn, HIV-infiserte, EPTB Rask Mulighet for samtidig påvisning av genotypiske resistens -spesifikke mutasjoner (INH, Rif) In-house eller kommersiell 3 metoder Kassett-basert (Xpert MTB/RIF) Line probe assay (LPA) Loop-mediated amplification (LAMP)

7 Xpert MTB/RIF assay Stor fremskritt Kassett-basert lukket sytem- ikke kontaminering Rask (2 t) Samtidig påvisning av MTB & RIF resistens (MDR-TB) Mikroskopi-negativ ekspektorat Sensitivitet: 57% -83% WHO anbefaler

8 Center for International Health, Department of Global Public HEalth & Primary Care De nyeste versjonene av Xpert «Xpert-Ultra»-kassetter- høyere SN «Expert Omni»- bærbar- point-of-care «Xpert Xpand»- XDR-TB- mutasjoner resistanse mot isoniazid, fluorokinoloner og 2.linje sprøytemidler Flere i «pipe-line»

9 Center for International Health, Department of Global Public HEalth & Primary Care Utfordringer-implementering i høy TB-endemiske land

10 NAAT- Line probe assay (LPA) Linje probe assays: samtidig påvisning av M.TB & genotypiske resistens -spesifikke mutasjoner (INH, Rif) Anvendes -mikroskopi-positive ekspektorat 5 timer Begrensning: Ikke egnet for mikroskopi-negativ prøver

11 Loop-mediated isothermal amplification (LAMP) simple visual colorimetric read-out No repeated heating and cooling cycles low power requirement WHO- not yet recommended

12 TB i Norge-organfordeling Kilde: FHI

13 Ekstrapulmonal tuberkulose Diagnostiske utfordringer Paucibacillary Mikrobiologiske metoder- lav sensitivitet direkte mikroskopi (0-10%) dyrkning(0-22%) NAAT baserte metoder: Bedre men varierende sensitivitet, Kontaminering, kostbar Histologi: ikke spesifikk Immunesvekkede- atypisk histologi

14 Detection of MPT64-antigen from the biopsies, aspirates, cell smears by immunohistochemistry Robust, rapid, sensitivity/specificity comparable to sensitive PCR-based tests

15 Center for International Health, Department of Global Public HEalth & Primary Care MPT64 in HIV-TB coinfected pleural TB lesions

16 MPT64 antigen in HIV-TB coinfected pleural TB lesions- atypical histology H&E stain Acid-fast stain IHC-MPT64 IHC- MPT64

17 Immunocytochemical staining of mycobacterial antigen MPT64 Lymfeknute Aspirat Pleuravæske

18 MPT64 antigen detection test- previous studies Country Site of disease Reference standard India South Africa HIV-coinfected Lymph node, Pleural fluids, Abdominal TB (peritoneum, intestinal wall, lymph nodes, ascitic fluid), CNS (CSF) Pleural biopsies ZN sensitivity < 10%, Culture sensitivity < 28% SN SP N-PCR 88-93% Response treatment 80% 100% Tanzania LN biopsies N-PCR 90% 83% Norway LN biopsies N-PCR 95% 62% Ethiopia Pleural fluid aspirates & Lymph node aspirates PCR 88 % 90% 5 Publications: Diagnostic Pathology 2007, 2:36, Appl Immunohistochem Mol Morphol :554, Diagn Cytopathol :782, BMC Infectious Diseases 2014, 14:585, Appl Immunohistochem Mol Morphol 2017;25:

19 U N I V E R S I T Y O F B E R G E N Center for International Health, Department of Global Public Health & Primary Care Project Title: Improving diagnosis of extrapulmonary tuberculosis by implementation of a sensitive and specific antigen detection test in routine diagnostic settings Support: Research Council of Norway through the Global Health and Vaccination Programme (GLOBVAC), project number Study is part of the EDCTP2 programme supported by the European Union Helse Vests forskningsmidler

20 Primary objective: To improve the diagnosis of extrapulmonary TB by introducing a sensitive and specific test in the selected tertiary care hospitals in high TB and high-low HIV burden countries using a routine TB control programme.

21 Center for International Health, Department of Global Public HEalth & Primary Care Partners: Pakistan, India, Tanzania, Zanzibar, Norway Zanzibar

22 Study design Center for International Health, Department of Global Public HEalth & Primary Care Biobank

23 Center for International Health, Department of Global Public HEalth & Primary Care Implementaion of test in zanzibar Melissa Davidsen Jørstad-PhDcandidate Mariam Abdallah doctor at MMH

24 Performance of test in zanzibar Center for International Health, Department of Global Public HEalth & Primary Care Number specimens Number of specimens (%) positive by ZN LJ culture GenXpert MPT64 test TB, all specimens FNAC TB-LN Non-TB, all specimens FNAC non-tb LN All case: Sensitivity (SN) 69%, Specificity (SP) 95% Lymphadenitis: SN 79%, SP 97% Pediatric Lymphadenitis: SN 100%, SP 96%

25 Acknowledgement Center for International Health, Department of Global Public HEalth & Primary Care Professor Lisbet Sviland, Department of Pathology, Haukeland University Hospital Professor Harald Wiker, Department of Clinical Science, UiB Professor Anne Ma Dyrhol-Riise, Department of Infectious Diseases, UiO/ OUS Melissa D Jørstad, UiB, PhD scholar Masafiri Marijani, Pathologist, Mnazi Mmoja Hospital, Zanzibar