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Performance of a real-time PCR approach for diagnosing Schistosoma haematobium infections of different intensity in urine samples from Zanzibar

BACKGROUND: Efforts to control and eliminate schistosomiasis have accelerated over the past decade. As parasite burden, associated morbidity and egg excretion decrease, diagnosis with standard parasitological methods becomes harder. We assessed the robustness and performance of a real-time PCR (qPCR...

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Autores principales: Keller, Dominique, Rothen, Julian, Dangy, Jean-Pierre, Saner, Corina, Daubenberger, Claudia, Allan, Fiona, Ame, Shaali M., Ali, Said M., Kabole, Fatma, Hattendorf, Jan, Rollinson, David, Seyfarth, Ralf, Knopp, Stefanie
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2020
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Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7487541/
https://www.ncbi.nlm.nih.gov/pubmed/32887642
http://dx.doi.org/10.1186/s40249-020-00726-y
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author Keller, Dominique
Rothen, Julian
Dangy, Jean-Pierre
Saner, Corina
Daubenberger, Claudia
Allan, Fiona
Ame, Shaali M.
Ali, Said M.
Kabole, Fatma
Hattendorf, Jan
Rollinson, David
Seyfarth, Ralf
Knopp, Stefanie
author_facet Keller, Dominique
Rothen, Julian
Dangy, Jean-Pierre
Saner, Corina
Daubenberger, Claudia
Allan, Fiona
Ame, Shaali M.
Ali, Said M.
Kabole, Fatma
Hattendorf, Jan
Rollinson, David
Seyfarth, Ralf
Knopp, Stefanie
author_sort Keller, Dominique
collection PubMed
description BACKGROUND: Efforts to control and eliminate schistosomiasis have accelerated over the past decade. As parasite burden, associated morbidity and egg excretion decrease, diagnosis with standard parasitological methods becomes harder. We assessed the robustness and performance of a real-time PCR (qPCR) approach in comparison with urine filtration microscopy and reagent strip testing for the diagnosis of Schistosoma haematobium infections of different intensities. METHODS: The robustness of DNA isolation and qPCR was validated in eight laboratories from Europe and Africa. Subsequently, 792 urine samples collected during cross-sectional surveys of the Zanzibar Elimination of Schistosomiasis Transmission (ZEST) project in 2012–2017 were examined with qPCR in 2018. Diagnostic sensitivity of the qPCR was calculated at different infection intensity categories, using urine filtration microscopy as reference test. Spearman’s rank correlation between Ct-values and S. haematobium egg counts was assessed and Ct-value percentiles for infection intensity categories determined. RESULTS: S. haematobium Dra1 DNA-positive samples were identified correctly in all eight laboratories. Examination of urine samples from Zanzibar revealed Dra1 DNA in 26.8% (212/792) by qPCR, S. haematobium eggs in 13.3% (105/792) by urine filtration, and microhaematuria in 13.8% (109/792) by reagent strips. Sensitivity of the qPCR increased with augmenting egg counts: 80.6% (29/36) for counts between 1 and 4 eggs, 83.3% (15/18) for counts between 5 and 9 eggs, 100% (23/23) for counts between 10 and 49 eggs, and 96.4% (27/28) for counts of 50+ eggs. There was a significant negative correlation between Ct-values and egg counts (Spearman’s rho = − 0.49, P < 0.001). Seventy-five percent of the Ct-values were ≥ 33 in the egg-negative category, < 31 in the light intensity category, and < 24 in the heavy intensity category. CONCLUSIONS: While the sensitivity of the qPCR was ~ 80% for very light intensity infections (egg counts < 10), in general, the Dra1 based qPCR assay detected twice as many S. haematobium infections compared with classical parasitological tests. The qPCR is hence a sensitive, urine-based approach for S. haematobium diagnosis that can be used for impact assessment of schistosomiasis elimination programmes, individual diagnosis, and in improved format also for verification and certification of elimination. TRIAL REGISTRATION: ISRCTN, ISRCTN48837681. Registered 05 September 2012 - Retrospectively registered.
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spelling pubmed-74875412020-09-15 Performance of a real-time PCR approach for diagnosing Schistosoma haematobium infections of different intensity in urine samples from Zanzibar Keller, Dominique Rothen, Julian Dangy, Jean-Pierre Saner, Corina Daubenberger, Claudia Allan, Fiona Ame, Shaali M. Ali, Said M. Kabole, Fatma Hattendorf, Jan Rollinson, David Seyfarth, Ralf Knopp, Stefanie Infect Dis Poverty Research Article BACKGROUND: Efforts to control and eliminate schistosomiasis have accelerated over the past decade. As parasite burden, associated morbidity and egg excretion decrease, diagnosis with standard parasitological methods becomes harder. We assessed the robustness and performance of a real-time PCR (qPCR) approach in comparison with urine filtration microscopy and reagent strip testing for the diagnosis of Schistosoma haematobium infections of different intensities. METHODS: The robustness of DNA isolation and qPCR was validated in eight laboratories from Europe and Africa. Subsequently, 792 urine samples collected during cross-sectional surveys of the Zanzibar Elimination of Schistosomiasis Transmission (ZEST) project in 2012–2017 were examined with qPCR in 2018. Diagnostic sensitivity of the qPCR was calculated at different infection intensity categories, using urine filtration microscopy as reference test. Spearman’s rank correlation between Ct-values and S. haematobium egg counts was assessed and Ct-value percentiles for infection intensity categories determined. RESULTS: S. haematobium Dra1 DNA-positive samples were identified correctly in all eight laboratories. Examination of urine samples from Zanzibar revealed Dra1 DNA in 26.8% (212/792) by qPCR, S. haematobium eggs in 13.3% (105/792) by urine filtration, and microhaematuria in 13.8% (109/792) by reagent strips. Sensitivity of the qPCR increased with augmenting egg counts: 80.6% (29/36) for counts between 1 and 4 eggs, 83.3% (15/18) for counts between 5 and 9 eggs, 100% (23/23) for counts between 10 and 49 eggs, and 96.4% (27/28) for counts of 50+ eggs. There was a significant negative correlation between Ct-values and egg counts (Spearman’s rho = − 0.49, P < 0.001). Seventy-five percent of the Ct-values were ≥ 33 in the egg-negative category, < 31 in the light intensity category, and < 24 in the heavy intensity category. CONCLUSIONS: While the sensitivity of the qPCR was ~ 80% for very light intensity infections (egg counts < 10), in general, the Dra1 based qPCR assay detected twice as many S. haematobium infections compared with classical parasitological tests. The qPCR is hence a sensitive, urine-based approach for S. haematobium diagnosis that can be used for impact assessment of schistosomiasis elimination programmes, individual diagnosis, and in improved format also for verification and certification of elimination. TRIAL REGISTRATION: ISRCTN, ISRCTN48837681. Registered 05 September 2012 - Retrospectively registered. BioMed Central 2020-09-04 /pmc/articles/PMC7487541/ /pubmed/32887642 http://dx.doi.org/10.1186/s40249-020-00726-y Text en © The Author(s) 2020 Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.
spellingShingle Research Article
Keller, Dominique
Rothen, Julian
Dangy, Jean-Pierre
Saner, Corina
Daubenberger, Claudia
Allan, Fiona
Ame, Shaali M.
Ali, Said M.
Kabole, Fatma
Hattendorf, Jan
Rollinson, David
Seyfarth, Ralf
Knopp, Stefanie
Performance of a real-time PCR approach for diagnosing Schistosoma haematobium infections of different intensity in urine samples from Zanzibar
title Performance of a real-time PCR approach for diagnosing Schistosoma haematobium infections of different intensity in urine samples from Zanzibar
title_full Performance of a real-time PCR approach for diagnosing Schistosoma haematobium infections of different intensity in urine samples from Zanzibar
title_fullStr Performance of a real-time PCR approach for diagnosing Schistosoma haematobium infections of different intensity in urine samples from Zanzibar
title_full_unstemmed Performance of a real-time PCR approach for diagnosing Schistosoma haematobium infections of different intensity in urine samples from Zanzibar
title_short Performance of a real-time PCR approach for diagnosing Schistosoma haematobium infections of different intensity in urine samples from Zanzibar
title_sort performance of a real-time pcr approach for diagnosing schistosoma haematobium infections of different intensity in urine samples from zanzibar
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7487541/
https://www.ncbi.nlm.nih.gov/pubmed/32887642
http://dx.doi.org/10.1186/s40249-020-00726-y
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