Chagas disease in Spain: Experience from a reference laboratory

1. Second WHO consultation: Development of a WHO reference panel for the control of Chagas diagnostic tests Geneva, 27 – 28 January 2009 Chagas disease in Spain: Experience from a reference laboratory Teresa Gárate Servicio de Parasitología Centro Nacional de Microbiología

2. Experience from a Parasitology Reference Laboratory • General Role of the Parasitology Lab • Parasitology Lab and Chagas’ disease in Spain

3. Parasitic diseases • AUTOCTHONOUS • Leishmaniasis • Toxoplasmosis • Cryptosporidiasis • Giardiasis • Amebiasis • Fasciolosis • Hydatidosis • Anisakiasis • Toxocariosis • Trichinellosis • IMPORTED • Malaria • Other Leishmaniasis • Chagas disease • Sleeping sickness • Cysticercosis • Schistosomiasis • Filariasis/Oncho • Intestinal parasites National Microbiology Centre

4. The Parasitology Lab works with Spanish hospitals and blood donor centers on Chagas disease diagnosis

5. Other Activities related with T. cruzidiagnosis • Chagas Test evaluation • In house tests • Commercial tests • Control sample preparation • Serum of characterized patients • DNA from parasite strains • Blood spiked with parasites • Training microbiologists in • Microhaemotocrit • PCR • Serological result interpretation

6. PCR ELISA Commercial assays (rapid tests) Implementation of T. cruziDiagnostic Tools in Parasitology Lab Microscopic examination Xenodiagnosis IFAT Culture Real time PCR

7. Evaluation of serological tests: In house ELISAand 3 serum panels

8. Evaluation of serological tests: In house ELISAand 3 serum panels

9. Characteristics of serum samples of the 3 panels used in the evaluation of the In house ELISA Panel 1: Sera from Bolivian individuals with positive or negative xenodiagnosis Panel 2: Sera from population with positive or negative T. cruzi IFAT. Healthy individuals from endemic and non-endemic areas Panel 3: Sera of anti-T. cruzi Specific Panel and healthy individuals from endemic and non-endemic areas Anti-T. cruzi Specific Sera Panel (QPanel, Sao Paulo-Brazil, http://www.panel.com.br) was kindly provided by Dra. Elena Franco

10. Evaluation of serological tests: Commercial ELISAs

11. Characteristics of serum samples used in the evaluation of commercial tests

12. Evaluation of Commercial ELISAs: Sensitivity

13. Evaluation of Commercial ELISAs: Specificity a The specificity was calculated into account results of healthy control b The specificity was calculated into account results of healthy control and other pathologies.

14. Molecular diagnosis of T. cruzi Control assay of limit detection Samples of population at-risk kDNA-PCRmodified (121-122 / HUF-REV) Brittoet al 1993; Cruz et al 2002; Walsh et al 1991; Dorn et al 1997; Gomes et al 1998; Wincker et al 1994; Rubio et al 2002

15. Molecular diagnosis of T. cruzi Diagnostic sensitivity and specificity

16. Chagas disease cases in Spain AUTOCTHONOUS IMPORTED • Blood transfusion • Congenital transmission • Organ transplant • Laboratory accidents • Immigration from endemic area • Long stays in endemic area

17. Latin American population in Spain On December 2008: > 1.8 million

18. Reports of TransfusionalChagas cases in Spain National Haemovigilance System

19. TransfusionalChagas Cases

20. País Vasco A Coruña Madrid Characteristics of Infected donors: Malaga

21. Congenital Chagas disease Newborns examined during 2002 - 2008 Transmission index: ~ 4,5% * Post mortem diagnosis In Spain ≈ 990 000 women of childbearing age Bolivia: 132 421 women Argentina: 94 794 women

22. Laboratory Accidents and Chagas disease Alvar J. 1983 Laboratorio 76(456): 645-648

23. Chagas disease cases in Spain AUTOCTHONOUS IMPORTED • Blood transfusion • Congenital transmission • Organ transplant • Laboratory accidents • Immigration from endemic area • Long stays in endemic area

24. Chagas disease in blood donors at- risk Asturias Madrid

25. Profile of immigrant population Asturias 2006: ≈ 17 000 immigrant 2008: ≈ 20 000 immigrant Madrid 2006: ≈ 400 000 immigrant 2008: ≈ 500 000 immigrant

26. Prevalence in blood donors at-risk (2002-2006) ? P = Seropositive N = Seronegative

27. Seroprevalence in population at-risk: 24% ≈ 1144 Chagasic patients

28. Distribution of cases in Spain

29. Origin of seropositive population

30. Conclusions • Panel of Reference • - Positive samples by different serological tests, and • - Positive samples from chagasic individuals with parasitological, epidemiological and clinical evaluation. • Number of positive sera/panel • - 3 serum samples for kit manufacturers • - 100 serum samples for evaluation of tests • - 3 serum samples for quality control. • Chagas in Spain • - Imported and autochonous types. • - Bolivian group, mainly from Santa Cruz region, shows the highest seropositivity rates and represents the highest risk for T. cruzi infection transmission.

31. Thank you UNIDAD LEISHMANIA Y CHAGAS Carmen Cañavate Javier Nieto Mercedes Rodriguez Israel Cruz Elena Bodas Marta Hernández Rubén González Emilia García Carmen Chicharro María Flores

32. Chagas disease in Spain AUTOCTHONOUS IMPORTED Transfusional and congenital Chagas Immigration and long stays in endemic area Organ transplant and lab incident ? Months Years Chronic phase Acute

33. Level of anti-T. cruziIgG in samples of patients with positive PCR Acute case 1 (Madrid recipient) Acute case 2 (Málaga recipient)