Diagnosis of leptospirosis


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GRF 2nd One Health Summit 2013: Presentation by Rudy Hartskeerl, WHO/FAO/OIE and National Leptospirosis Reference Centre

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Diagnosis of leptospirosis

  1. 1. Diagnosis of leptospirosis Rudy A. Hartskeerl WHO/FAO/OIE and National Leptospirosis Reference Centre
  2. 2. Clinical manifestations • Large variety of signs and symptoms mimicking several other diseases (dengue, hanta virus, yellow fever, hepatitis, salmonellosis, brucellosis, malaria, pneumonia) • Asymptomatic, mild (flu-like) to severe course • Weil’s syndrome (kidney failure, liver impairment (jaundice), hemorrhage: CFR 10-20% • Severe Pulmonary Hemorrhage Syndrome (SPHS): globally emerging form with CFR 50-80% • Diagnosis on clinical grounds is challenging Amsterdam, The Netherlands www.kit.nl
  3. 3. Conventional tests Conventional tests: DFM, MAT, ELISA, IFAT, IHA, MSAT etc. Many drawbacks: slow, unreliable, low detection threshold, difficult to standardize, require well-trained personnel and/or expensive media/equipment Isolation confirms leptospirosis but is too slow for individual diagnosis and has mainly epidemiological value Amsterdam, The Netherlands www.kit.nl
  4. 4. Laboratory diagnosis Serological tests MAT Gold or reference standard Requires panels of ‘locally’ representative serovars May provide indication for infecting serogroup ELISA IgM antibodies against leptospires Genus specific In-house ELISA: dilution titers Both tests are restricted to well equipped expert labs Amsterdam, The Netherlands www.kit.nl
  5. 5. Sensitivity of MAT and IgM ELISA according to stage of disease Main findings • Serology does not contribute to confirmation at early acute phase • ‘Gold standard’ MAT has sensitivity of 80% • Use of paired samples greatly improves case detection Amsterdam, The Netherlands www.kit.nl
  6. 6. Rapid Diagnostic Tests for use at ‘peripheral’ level Test/kit Manufacturer Technology comments IHA Mayo Medical Lab (USA) Indirect hemagglutination Leptospira-MC test Japan Lyophilization Lab Microcapsule agglutination test LeptoTek LFA Organon Teknika (Netherlands) Lateral flow assay Expected soon by IMACCESS Test-it Life Assays (South Africa) Lateral flow assay Human and veterinary Leptocheck-WB Zephyr (India) Lateral flow assay SD Leptospira LF Standard Diagnostics (Korea) Lateral flow assay IgM LFA Omega Diagnostics (UK) Lateral flow assay LeptoTek DriDot BioMerieux (France) Latex cardagglutination test Leptorapide Linnodee (Northern Ireland) Latex cardagglutination test Not available GLEAN paper: Picardeau et al., Rapid tests for diagnosis of leptospirosis: current tools and emerging technologies. Diag Microbiol Infect Dis (2013); Table 1 Amsterdam, The Netherlands www.kit.nl
  7. 7. Leptocheck WB (Zephyr); sensitivity 2005-2011 100 90 80 70 60 % 50 40 30 20 10 0 "2005" "2006" "2009" "2010" "2011"
  8. 8. Drawbacks rapid tests • Confirmation by conventional tests is required • Relatively low diagnotic accuracy • Varying diagnostic accuracy => repeated local evaluations • Need for improved QA at production and/or test performer (easy does not exclude experience) • Based on serology: too late for early diagnosis and start of effective antibiotic treatment • Rapid diagnosis is different from early diagnosis! Amsterdam, The Netherlands www.kit.nl
  9. 9. Importance of high quality laboratories Adequate laboratory capacity is indispensible for: • Early detection of diseases • Disease surveillance • Evidence-based diagnostic decision making Amsterdam, The Netherlands www.kit.nl
  10. 10. Quality Management System Implementation of a quality management system (QMS) is a key intervention to break the negative spiral • • www.kit.nl The internationally accepted quality standard for medical laboratories is ISO 15189 • Amsterdam, The Netherlands The requirements for a wellfunctioning QMS are formulated in quality standards A laboratory is accredited when it complies with the complete standard
  11. 11. Observations The problem is that quality standards indicate what shall be implemented, not how it must be implemented. Observations: • Interest of laboratories to become accredited is increasing • It is difficult to translate the international quality standard into practice Amsterdam, The Netherlands www.kit.nl
  12. 12. Electronic guidance tools • • • • GLI tool: Global Laboratory Initiative Stepwise Process towards TB Laboratory Accreditation Tool translate the ISO 15189 quality standard into: • Sequence of practical steps for implementation • Links to support materials (trainings, templates, …) where appropriate Available online-www.GLIquality.org Free of charge Amsterdam, The Netherlands www.kit.nl
  13. 13. Electronic guidance tools WHO LQSI tool: World Health Organization Laboratory Quality Stepwise Implementation tool – last stages of review. Amsterdam, The Netherlands www.kit.nl
  14. 14. In the context of OneHealth LQSI tool and GLI tool focus on medical laboratories. To prevent problems related with global urbanization other laboratories need to be of adequate capacity as well: • • • • • Food security laboratories Industrial laboratories (pharmaceutics/food production/etc.) Veterinary laboratories Forensic laboratories Etc. Amsterdam, The Netherlands www.kit.nl ISO 15189 Similar tools needed for ISO 17025: Test and Calibration laboratories
  15. 15. Early diagnosis Application of current rapid diagnostic tests (RDT) does not provide early diagnosis! Amsterdam, The Netherlands www.kit.nl
  16. 16. Early diagnosis •Early diagnostic gap with serology •Leptospires circulate in the blood notably first 4-7 days after onset of disease: Several NA amplification techniques on blood samples like PCR can fill in this gap Amsterdam, The Netherlands www.kit.nl
  17. 17. Hand held PCR equipment • • • • • • • Battery-powered stand-alone device: More than 4 hours of operation on a single charge High amplification speed: Extreme high efficiency: Near single-copy amplification sensitivity Dynamic range sufficient for genetic identification: Unmatched well-to-well uniformity: Simple and adaptable: Easy to program and compatible with standard tools Affordable costs Amsterdam, The Netherlands www.kit.nl T-COR 4 qPCR Palm PCR F1-12
  18. 18. Future (tests) • Early, easy and rapid – antigen detection based? • Same test for humans and animals • Discriminative (multiplex testing/differential diagnosis) • Traceable (online data transfer to e.g. MoH and MoA) - early outbreak warning (RDT with QR/barcodes or chip; data and location recording by smart phone?) Amsterdam, The Netherlands www.kit.nl
  19. 19. Future tests: Antigen detection Antigen test sens 102-103/ml – improve by use of nanobiosensors? Isothermal Recombinase Polymerase Amplification (RPA): multiplex, easy design (37-42oC, runtime 30 min, PCR-like primers/probes) 10 -2 10 -3 10 -4 Piepenburg et al. (2006). DNA detection using recombination proteins. PLoS Biol 4:e204 Amsterdam, The Netherlands www.kit.nl 10 10 10 -5 -6 -7 10 -8 H2 O Pos. C.
  20. 20. Quantification market for RDTs in Indonesia Incidence 50 per 100,000 10-20% of patients with compatible symptoms have lepto 120.000 people acquire leptospirosis/ yr 7 x 120.000 = 840.000 patients/yr require leptotest 2 tests per patient (paired samples) = 1.680.000 tests required population 240.000.000 Market might be increased in case of multiplex testing with more common infectious diseases in differential diagnosis!
  21. 21. Summary • • • • • • • Leptospirosis is difficult to diagnose in the clinic: laboratory support is indispensable Conventional diagnostic tests are inadequate RDT are helpful but ‘are late’, lack diagnostic accuracy and consistency Early and rapid diagnosis is possible with molecular antigenbased tests: applicable on both humans and animals Test results should be digitally transmittable and associated with data, important for surveillance and early outbreak warning Implementation of QA at several stages is badly needed There is a potential market for leptospirosis RDT notably when associated with multiplex testing for differential diagnosis Amsterdam, The Netherlands www.kit.nl
  22. 22. Thanks