Dracunculiasis (Guinea-Worm Disease)

Symposium: Top Papers in Neglected Tropical Diseases April 23th, 2018

Top Papers in NTDs Diagnostic Parasitology

Míriam J. Álvarez-Martínez M.D., Ph.D.

Microbiology Department Hospital Clinic, Barcelona (Spain) ISGlobal (Barcelona Institute for Global Health) School of Medicine-University of Barcelona [email protected] eLibrary © by author Neglected Tropical Diseases, NTDs

WHO recommends five strategies for the prevention, control, elimination and eradication of neglected tropical diseases.

• Preventive chemotherapy • Intensified disease management • Vector and intermediate host control • Veterinary public health at the human– animal interface • Provision of safe water, sanitation and hygiene ESCMID eLibrary Accelerating work to overcome the global © impact of byneglected tropical author diseases – A roadmap for implementation, WHO, 2012 PARASITIC WHO-NTDs

• Buruli ulcer • Onchocerciasis • Chagas disease • Rabies • Dengue and Chikungunya • Scabies and other ectoparasites • Dracunculiasis (guinea-worm disease) • Schistosomiasis • Echinococcosis • Soil-transmitted helminthiases • Foodborne trematodiases • Snakebite envenoming • Human African Trypanosomiasis • Taeniasis/Cysticercosis • Leishmaniasis • Trachoma • Leprosy • Yaws • Lymphatic filariasis • Mycetoma, chromoblastomycosis andESCMID other deep mycoses eLibrary © by author PAPER’S SELECTION

PUBMED: DIAGNOSIS +HUMAN PARASITES +PROTOZOAN+HELMINTHS +DATE (05/01/2017-TODAY)

RESULT: 477 PAPERS ESCMID eLibrary © by author10 PAPERS PARASITIC WHO-NTDs

ESCMID eLibrary © by author Chagas Disease: Molecular Diagnosis & LAMP

o Trypanosoma cruzi, a parasite transmitted to humans from hematophagous insects, causes Chagas disease, a Neglected Tropical Disease with public health impact, affecting 7 million people in Latin America. o Although mainly related to low income populations inhabiting rural environments, migrations have conveyed Chagas Disease to urban areas of endemic and non-endemic countries. o Often presents non-specific symptoms, and direct, low cost Microscopy-based diagnosis only detects acute infections, missing a high proportion of cases. o Serology is the “gold standard” diagnostic technique for chronic stages and needs the concordance of at least two different assaysESCMID to confirm infection. eLibrary Recent advances in Genetics and Molecular Diagnosis of Parasitic Protozoa: Molecular diagnosis of Trypanosoma cruzi. Acta Trop. 2018 Feb 21 (review) © by author Chagas Disease: Molecular Diagnosis & LAMP

o Standardized and validated PCR are now available tools with high sensitivity and specificity.

o Best performing methods selected are based on highly repetitive satellite DNA (SatDNA) or minicircle DNA (kDNA) sequences.

o In qPCR assays, analytical sensitivity of kDNA qPCR was higher than that of SatDNA qPCR, with limits of detection of 0.234 and 0.698 parasite equivalents/mL, respectively.

o High concordance was observed between both methods in proficiencyESCMID panels and clinical eLibrary specimens. Recent advances in Genetics and Molecular Diagnosis of Parasitic Protozoa: Molecular diagnosis of Trypanosoma cruzi. Acta Trop. 2018 Feb 21 (review) © by author Chagas Disease: Molecular Diagnosis & LAMP o To evaluate the analytical sensitivity and specificity of a prototype kit based on Loop mediated isothermal amplification (LAMP), using standardized qPCR as a comparator prototype for the amplification of satellite sequence of T.cruzi . o Peripheral blood specimens belonging to Chagas disease patients, including acute, congenital, chronic and reactivated cases (N = 23), & seronegative controls (N = 10) were evaluated by LAMP in comparison to qPCR. o LAMP was able to amplify DNAs from T. cruzi stocks representative of the six DTUs, it did not amplify DNAs from Leishmania sp, T. brucei sp, T. rangeli, P. falciparum and noninfected human DNA. o Analytical sensitivity was 1x10-2 fg/μL (qPCR detected up to 1x 10−1 fg/μL of CL Brener DNA and 1 fg/μl of Sylvio X10 DNA) ESCMID eLibrary Analytical sensitivity and specificity of a loop-mediated isothermal amplification (LAMP) kit prototype for detection of Trypanosoma cruzi DNA in human blood© samples. by PLoS Negl Tropauthor Dis. 2017 Jul 20;11(7) Chagas Disease: Molecular Diagnosis & LAMP

ESCMID eLibrary Analytical sensitivity and specificity of a loop-mediated isothermal amplification (LAMP) kit prototype for detection of Trypanosoma cruzi DNA in human blood© samples. by PLoS Negl Tropauthor Dis. 2017 Jul 20;11(7) Chagas Disease: Molecular Diagnosis & LAMP

o Molecular diagnostic tests may be employed

• epidemiological surveys of transmission • early diagnosis of • congenital transmission • acute infections due to oral transmission • transfusion or transplantation routes • reactivation due to immunosuppression • monitoring of treatment response in chronically infected patients receiving trypanocidal chemotherapy. ESCMID eLibrary Recent advances in Genetics and Molecular Diagnosis of Parasitic Protozoa: Molecular diagnosis of Trypanosoma cruzi. Acta Trop. 2018 Feb 21 (review) © by author Chagas Disease Standardized and validated PCR and LAMP are now available laboratory tools with high sensitivity and specificity

ESCMID eLibrary © by author ESCMID eLibrary © by authorhttp://www.who.int/chagas/en/ Echinococcosis

ESCMID eLibrary © by author Echinococcosis:

o Alveolar echinococcosis caused by Echinococcus multilocularis is an infrequent zoonosis with a high degree of disability, morbidity, and mortality, especially in disease clusters of the northern hemisphere. o Incubation period of approx.5–15 years, primary organ affected is the liver (>95% of alveolar echinococcosis cases) o Extrahepatic involvement is rare, occurs by continuous growth or through haematogenous spread, metastasising from the liver to the lungs, spleen, CNS, bones, lymph nodes, or muscle. o Osseous manifestation is 0·02–1·00% of alveolar echinococcosis cases. o A case of extrahepatic active alveolar echinococcosis affecting ESCMIDthe lumbar vertebrae, with eLibraryfurther lesions of the liver and brain. Vertebral alveolar echinococcosis-a case report, systematic analysis, and review of the literature. Lancet Infect Dis. 2018 Mar;18(3):e87 -e98. © by author Echinococcosis:

o Feb. 2016, 75-y woman with progressive back pain and substantial weight loss. o Lab: leucocytosis and paraproteinaemia (IgG λ). o CT: osteolytic lesions in the lumbar L3 and L4 multiple myeloma was suspected o Bone marrow biopsy: June, 2016 diagnosed of BCR-ABL-positive chronic myeloid leukaemia started treatment with nilotinib. o Back pain despite high doses of analgesics o Lumbar MRI: diffuse, infiltrative, vertebral and paravertebral, multicystic, slightly ring-enhancing mass protruding intraspinally and compressing the spinal cord at L1–L4 . Suspected extramedullary haemopoiesis. ESCMID eLibrary Vertebral alveolar echinococcosis-a case report, systematic analysis, and review of the literature. Lancet Infect Dis. 2018 Mar;18(3):e87 -e98. © by author Echinococcosis:

o Transpedicular biopsy of this lesion in Sep., 2016. o Intraoperatively, 40 mL of pus-like fluid aspirated for pathological and microbiological tests. o Periodic acid-Schiff (PAS)-positive lamellar structures and granulomatous inflammation were identified microscopically, suggesting a diagnosis of parasitic disease, specifically echinococcosis. o PCR confirmed the diagnosis of alveolar echinococcosis in biopsy tissue. o Serology was positive for E granulosus (64 AU/mL), but also for specific epitopes of E multilocularis (17 AU/mL against Em2 and ESCMID28 AU/mL against Em18)***** eLibrary Vertebral alveolar echinococcosis-a case report, systematic analysis, and review of the literature. Lancet Infect Dis. 2018 Mar;18(3):e87 -e98. © by author ESCMID eLibrary © by author Echinococcosis:

o Head-to pelvic body CT scan: several small hypodense lesions in the liver (one of them calcified) and calcified structures in the left basal ganglia.

o Albendazole (400 mg twice daily, orally) 2 weeks after the diagnostic biopsy. As a result of debilitating pain despite 2 months of albendazole treatment, the patient was offered palliative surgery. o 7 months after the surgery, dramatic improvement, almost pain ESCMIDfree and able to walk with walker. eLibrary Vertebral alveolar echinococcosis-a case report, systematic analysis, and review of the literature. Lancet Infect Dis. 2018 Mar;18(3):e87 -e98. © by author Alveolar Echinococcosis Complexity of extrahepatic alveolar echinococcosis manifestations and the necessity of an interdisciplinary approach.

ESCMID eLibrary © by author Human African Trypanosomiasis

ESCMID eLibrary © by author Human African Trypanosomiasis: Diagnosis review

Serodiagnostic tests only for T. brucei gambiense Card Agglutination Test for Trypanosomiasis (CATT ) blood, plasma/ serum, 5 min screening of at-risk populations by mobile teams

RDTs: HAT SeroK-SeT (Coris BioConcept, Gembloux, Belgium)

SD Bioline HAT 1.0 (Standard Diagnostics, Yongin, South Korea

ASSURED (Affordable, Sensitive, ESCMID eLibrarySpecific, User-friendly, Rapid and robust, passive screening and Human African trypanosomiasis. Lancet. 2017 Nov 25;390(10110):2397-2409. surveillance © by author Human African Trypanosomiasis : Diagnosis review

o CATT and rapid diagnostic tests are not 100% specific. Particularly when disease prevalence is low, their positive predictive value becomes critically low—eg, with a specificity of 98% and a prevalence of 0·1%, the positive predictive value is 4·5%. Currently, in most human African trypanosomiasis foci, prevalence is far below 0·1% and serological screening tests yield about 99 false-positive results for every true positive.

o Immune trypanolysis and ELISAs are applicable in laboratory conditions on serum, plasma, and dried blood spots. High specificity and sensitivity, applicability to dried blood spots, and adaptability to animal specimens make them excellent tools for largescale surveys, post-elimination monitoring, and animal reservoir studies.

o No field-applicable serodiagnostic test exists for T. brucei rhodesiense infection. Efforts to develop second-generation rapid diagnostic tests able to detect both T. brucei gambiense and T. brucei rhodesiense infection are ongoing, but the risk of cross-reaction with antibodies against non-human infective trypanosomes is ESCMIDever-present eLibrary Human African trypanosomiasis . Lancet.©2017 Novby 25;390(10110):2397 author-2409. Human African Trypanosomiasis :

o Parasitological confirmation of T. brucei gambiense infection is achieved by microscopic examination of a lymph node aspirate or by concentration techniques applied on blood or on cerebrospinal fluid. o Microscope should be adjusted for maximum light diffraction o T. brucei rhodesiense infection, presents with higher parasitaemia, stained blood thin film or thick drop, or chancre aspirate can be considered if the more sensitive concentration techniques are not available.

o Stage determination relies on the examination of CSF. o Patients with five or fewer white blood cells per µL and no trypanosomes in CSF are classified as first stage o those with more than five white blood cells per µL or trypanosomes ESCMIDin the cerebrospinal fluid aseLibrary second stage Human African trypanosomiasis . Lancet.©2017 Novby 25;390(10110):2397 author-2409. Human African Trypanosomiasis: Diagnosis review

Molecular

o Interpreted with caution in clinical practice, even for exported cases. All formats have poor diagnostic accuracy (even for stage determination and posttreatment follow-up), poor reproducibility, and incompatibility with diagnostic facilities in endemic countries.

o In the context of disease elimination, identification of the subspecies of T. brucei (eg, in tsetse, animals, and human beings) might be useful since atypical infections with animal trypanosomes are possible. o T. brucei gambiense-specific and T. brucei rhodesiense-specific PCR assays exist, but they target single-copy genes; hence, their ESCMIDanalytical sensitivity is poor eLibrary. Human African trypanosomiasis . Lancet.©2017 Novby 25;390(10110):2397 author-2409. Human African Trypanosomiasis: Improving the specificity of RDTs Molecular methods no accuracy

ESCMID eLibrary © by author Leishmaniasis

ESCMID eLibrary © by author© CDC Leishmaniasis: Antigenuria as Predictor o VL/HIV coinfection is now a major problem in some low resource settings. The highest burden globally is found in North-West Ethiopia, around 20% of VL patients are HIV co-infected. o The KAtex urine antigen test detects Leishmania antigen, which is a direct marker of infection. o Presence of urine antigen during follow-up of HIV patients was found predictive of VL relapse in areas where L. infantum is present but has not been explored in L. donovani endemic areas. o Nested within a two-site clinical trial in Ethiopia (2011–2015), to assess whether, 1- levels of Leishmania antigenuria measured at VL diagnosis were associated with initial treatment failure. 2- levels of Leishmania antigenuria at the end of treatment (parasitologically- ESCMIDconﬁrmed cure) were associated eLibrarywith subsequent relapse. Leishmania Antigenuria to Predict Initial Treatment Failure and Relapse in Visceral Leishmaniasis/HIV Coinfected Patients: An Exploratory Study Nested © Within aby Clinical Trial inauthor Ethiopia. Front Cell Infect Microbiol. 2018 Mar 29;8:94 Leishmaniasis:Antigenuria as Predictor

o Positive association between the tissue parasite load at the initial VL diagnosis and both the level of Leishmania antigen in the urine at VL diagnosis and at the end of treatment. o A high tissue parasite load at VL diagnosis was also associated with relapse.

o The level of urine antigen at the time of cure could be used before discharge to identify those at highest risk of relapse, who could be targeted for closer medical follow-up or (prolonged) secondary prophylaxis. ESCMID eLibrary Leishmania Antigenuria to Predict Initial Treatment Failure and Relapse in Visceral Leishmaniasis/HIV Coinfected Patients: An Exploratory Study Nested Within a Clinical Trial in Ethiopia. Front Cell Infect Microbiol. 2018 Mar 29;8:94 © by author Leishmaniasis:Antigenuria as Predictor

ESCMID eLibrary Leishmania Antigenuria to Predict Initial Treatment Failure and Relapse in Visceral Leishmaniasis/HIV Coinfected Patients: An Exploratory Study Nested © Within aby Clinical Trial inauthor Ethiopia. Front Cell Infect Microbiol. 2018 Mar 29;8:94 ESCMID eLibrary © by author © WHO Leishmaniasis: LAMP

o Evaluation LAMP kit for the diagnosis of leishmaniasis at the POC, the Loopamp™ Leishmania Detection Kit uses primers targeting 18S rRNA gene kDNA minicircles, specific for genus Leishmania

ESCMID eLibrary Evaluation of fluorimetry and direct visualization to interpret results of a loop-mediated isothermal amplification kit to detect Leishmania DNA. Parasit Vectors. 2018 Apr 17;11(1):250 © by author Leishmaniasis: LAMP

o Support diagnosis of VL when serological diagnosis is useless o VL relapses and test-of-cure o VL/HIV co-infection o Diagnosis of CL cases that require confirmatory diagnosis to initiate systemic treatment, which often presents high toxicity

o Promising approach to be explored in veterinary public health for the diagnosis of canine leishmaniasis. ESCMID eLibrary Evaluation of fluorimetry and direct visualization to interpret results of a loop-mediated isothermal amplification kit to detect Leishmania DNA. Parasit©Vectors. by 2018 Apr 17;11(1):250 author Leishmaniasis: VL-HIV diagnostic problem Antigenuria & LAMP solutions

ESCMID eLibrary © by author ESCMID eLibrary http://www.who.int/neglected_diseases/news/NTD_course_now_accessible_via_eLearning/en/ © by author ESCMID eLibrary ©Postigo R.J./WHO. Child affected by kala-azar in Marsabit county hospital, June 2017 © by author Onchocerciasis

ESCMID eLibrary © by author © CDC Onchocerciasis: Distribution & species identification

o Onchocerciasis elimination progress in Bioko Island (Equatorial Guinea) after 18 years of mass ivermectin intervention.

o General ﬁlariasis situation through a rapid and accurate molecular method.

o Cross-sectional study was conducted in Bioko Island from mid-January to mid-February 2014. ESCMID eLibrary Geographical distribution and species identification of human filariasis and onchocerciasis in Bioko Island, Equatorial Guinea. Acta Trop. 2018 ©Apr;180:12 -by17. author Onchocerciasis: Distribution & species identification o 543 subjects included . Whole blood and one skin snip (lumbar regions) analysed by real time PCR. Two other skin biopsies by an expert microscopist. All positive samples confirmed by sequencing. o Traditional microscopic examination of the skin biopsies failed to detect any microfilariae. but 11 (2.03%) infections were detected in skin using PCR assay O. volvulus-1, Mansonella streptocerca-2 , Mansonella perstans-7 and Loa loa-1. o PCR assays in blood detected 52 filariae-positive individuals (9.6%) with M. perstans or Loa loa. o The low prevalence of O. volvulus confirms the success of the Onchocerciasis Control Programme and suggests that Mass Drug Administration in Bioko Island can be interrupted in the near future. o The very high prevalence of M. perstans found in skin snips assays raises doubts about the reliabilityESCMID of microscope-based diagnosis eLibrary of O. volvulus infections. Geographical distribution and species identification of human filariasis and onchocerciasis in Bioko Island, Equatorial Guinea. Acta Trop. 2018 ©Apr;180:12 -by17. author Onchocerciasis: Distribution & species identification

ESCMID eLibrary Geographical distribution and species identification of human filariasis and onchocerciasis in Bioko Island, Equatorial Guinea. Acta Trop. 2018 ©Apr;180:12 -by17. author Onchocerciasis Molecular diagnosis in ﬁlariasis control programmes

ESCMID eLibrary © by author © P. DiCampo/ The Carter Center WHOESCMID verifies Mexico free of onchocerciasis eLibrary Decades-long mass treatment by ivermectin of affected populations accelerated interruption of transmission © by author Soil-Transmitted Helminthiases Strongyloidiasis

ESCMID eLibrary © by author © CDC Strongyloidiasis: Evaluation of PCR accuracy o Strongyloides stercoralis infection diagnosis lack of a gold standard, sensitivity of traditional parasitological tests (microscopic examination of stool samples and coproculture) is low. o Systematic review with meta-analysis to evaluate the accuracy of PCR for the diagnosis of S. stercoralis infection. Fourteen studies (12 real-time PCR) o Specificity of the techniques resulted high (ranging from 93 to 95% according to the reference test(s) used. o Molecular techniques were compared to parasitological methods with Sensitivity of PCR 71.8% (95% CI 52.2–85.5), Sensitivity decreased to 61.8% (95% CI 42.0– 78.4) when serology was added among the reference tests. o PCR might not be suitable for screening purpose, whereas it might have a role as aESCMID confirmatory test eLibrary Accuracy of molecular biology techniques for the diagnosis of Strongyloides stercoralis infection—A systematic review and meta -analysis. PLoS Negl©Trop Dis 12(2),by 2018 author Strongyloidiasis: Evaluation of PCR accuracy

ESCMID eLibrary Accuracy of molecular biology techniques for the diagnosis of Strongyloides stercoralis infection—A systematic review and meta -analysis. PLoS Negl©Trop Dis 12(2),by 2018 author Strongyloidiasis: Evaluation of PCR accuracy

ESCMID eLibrary Accuracy of molecular biology techniques for the diagnosis of Strongyloides stercoralis infection—A systematic review and meta-analysis. PLoS Negl Trop Dis 12(2), 2018 © by author STH: Strongyloidiasis PCR as confirmatory test

ESCMID eLibrary © by author ESCMID eLibrary http://www.who.int/intestinal_worms/resources/en/english_calendarpages.pdf © by author Taeniasis/Cysticercosis

o To know the diagnostic tests used in European laboratories for human taeniosis/cysticercosis.

o To determine potential gaps in their detection.

o To obtain preliminary data on the number of diagnosed taeniosis/CC cases. ESCMID eLibrary Present status of laboratory diagnosis of human taeniosis/cysticercosis in Europe. Eur J Clin Microbiol Infect Dis. 2017 Nov;36(11):2029 -2040 © by author Taeniasis/Cysteicercosis: Diagnosis in Europe

ESCMID eLibrary Present status of laboratory diagnosis of human taeniosis/cysticercosis in Europe. Eur J Clin Microbiol Infect Dis. 2017 Nov;36(11):2029-2040 © by author Taeniasis/Cysteicercosis: Diagnosis in Europe

ESCMID eLibrary Present status of laboratory diagnosis of human taeniosis/cysticercosis in Europe. Eur J Clin Microbiol Infect Dis. 2017 Nov;36(11):2029 -2040 © by author Taeniasis/Cysticercosis:

Diagnosis of T. solium (neuro)cisticercosis T. solium taeniosis in immigrants and travelers from endemic region and sporadic autochthonouscases,continue to be a problem in the EU , Increase knowledge on T. solium taeniosis/NCC infections is needed prevalence seems rather low in Europe despite the fact there is some evidence that NCC may be on the rise ESCMID eLibrary © by author ESCMID eLibrary © WHO © by author Neglected Plasmodium species

o Diagnostic accuracy of malaria RDTs in detecting Pk, Pm, and Po monoinfections. Recently, Pk was reported to be the most common cause of malaria in Malaysia o Microscopy and rapid diagnostic tests (RDTs) are the WHO recommended tools to confirm the diagnosis of all suspected malaria cases targeted malaria antigens used in malaria RDTs

o Histidine-rich protein-2 (HRP2) a Pf-specific antigen o Lactatedehydrogenase (LDH) Plasmodium falciparum–specific, pan-specific and Pv-specific LDH antibodies o Aldolase common to all Plasmodium species (pan-specific) ESCMID eLibrary Systematic review: Performance of RDTs for the detection of Plasmodium knowlesi, Plasmodium malariae, and Plasmodium ovale mono-infections in human blood. J Infect Dis. 2018 Mar 15. © by author Neglected Plasmodium species:

ESCMID eLibrary Asystematic review: Performance of RDTs for the detection of Plasmodium knowlesi, Plasmodium malariae, and Plasmodium ovale mono-infections in human blood. J Infect Dis. 2018 Mar 15. © by author Neglected Plasmodium species:

THE WORLD HEALTH ORGANIZATION believes that despite the complexity of neglected tropical diseases, the targets are achievable. ESCMID eLibrary © by author In Memoriam

Dr. TeresaESCMID Gárate eLibrary Head of Parasitology Department, Nacional Center of Microbiology, Instituto Salud Carlos III (CNM©-ISCIII), by Madrid author THANK YOU FOR YOUR ATTENTION