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Droplet Digital PCR for the Detection of Plasmodium falciparum DNA in Whole Blood and Serum: A Comparative Analysis with Other Molecular Methods

Background: The estimation of Plasmodium falciparum parasitaemia can vary according to the method used. Recently, droplet digital PCR (ddPCR) has been proposed as a promising approach in the molecular quantitation of Plasmodium, but its ability to predict the actual parasitaemia on clinical samples...

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Autores principales: Pomari, Elena, Silva, Ronaldo, Moro, Lucia, La Marca, Giulia, Perandin, Francesca, Verra, Federica, Bisoffi, Zeno, Piubelli, Chiara
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7350319/
https://www.ncbi.nlm.nih.gov/pubmed/32560386
http://dx.doi.org/10.3390/pathogens9060478
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author Pomari, Elena
Silva, Ronaldo
Moro, Lucia
La Marca, Giulia
Perandin, Francesca
Verra, Federica
Bisoffi, Zeno
Piubelli, Chiara
author_facet Pomari, Elena
Silva, Ronaldo
Moro, Lucia
La Marca, Giulia
Perandin, Francesca
Verra, Federica
Bisoffi, Zeno
Piubelli, Chiara
author_sort Pomari, Elena
collection PubMed
description Background: The estimation of Plasmodium falciparum parasitaemia can vary according to the method used. Recently, droplet digital PCR (ddPCR) has been proposed as a promising approach in the molecular quantitation of Plasmodium, but its ability to predict the actual parasitaemia on clinical samples has not been largely investigated. Moreover, the possibility of applying the ddPCR-sensitive method to serum samples has never been explored. Methods: We used, for the first time, ddPCR on both blood and serum to detect the DNA of P. falciparum in 52 paired samples from 26 patients. ddPCR was compared with loop-mediated isothermal amplification (LAMP) and rtPCR. The correlation between the ddPCR results, microscopy, and clinical parameters was examined. Results: ddPCR and microscopy were found to be strongly correlated (ρ(26) = 0.83111, p < 0.0001) in blood. Samples deviating from the correlation were partially explained by clinical parameters. In serum samples, ddPCR revealed the best performance in detecting P. falciparum DNA, with 77% positive samples among malaria subjects. Conclusion: Absolute quantitation by ddPCR can be a flexible technique for Plasmodium detection, with potential application in the diagnosis of malaria. In particular, ddPCR is a powerful approach for Plasmodium DNA analysis on serum when blood samples are unavailable.
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spelling pubmed-73503192020-07-15 Droplet Digital PCR for the Detection of Plasmodium falciparum DNA in Whole Blood and Serum: A Comparative Analysis with Other Molecular Methods Pomari, Elena Silva, Ronaldo Moro, Lucia La Marca, Giulia Perandin, Francesca Verra, Federica Bisoffi, Zeno Piubelli, Chiara Pathogens Article Background: The estimation of Plasmodium falciparum parasitaemia can vary according to the method used. Recently, droplet digital PCR (ddPCR) has been proposed as a promising approach in the molecular quantitation of Plasmodium, but its ability to predict the actual parasitaemia on clinical samples has not been largely investigated. Moreover, the possibility of applying the ddPCR-sensitive method to serum samples has never been explored. Methods: We used, for the first time, ddPCR on both blood and serum to detect the DNA of P. falciparum in 52 paired samples from 26 patients. ddPCR was compared with loop-mediated isothermal amplification (LAMP) and rtPCR. The correlation between the ddPCR results, microscopy, and clinical parameters was examined. Results: ddPCR and microscopy were found to be strongly correlated (ρ(26) = 0.83111, p < 0.0001) in blood. Samples deviating from the correlation were partially explained by clinical parameters. In serum samples, ddPCR revealed the best performance in detecting P. falciparum DNA, with 77% positive samples among malaria subjects. Conclusion: Absolute quantitation by ddPCR can be a flexible technique for Plasmodium detection, with potential application in the diagnosis of malaria. In particular, ddPCR is a powerful approach for Plasmodium DNA analysis on serum when blood samples are unavailable. MDPI 2020-06-17 /pmc/articles/PMC7350319/ /pubmed/32560386 http://dx.doi.org/10.3390/pathogens9060478 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Pomari, Elena
Silva, Ronaldo
Moro, Lucia
La Marca, Giulia
Perandin, Francesca
Verra, Federica
Bisoffi, Zeno
Piubelli, Chiara
Droplet Digital PCR for the Detection of Plasmodium falciparum DNA in Whole Blood and Serum: A Comparative Analysis with Other Molecular Methods
title Droplet Digital PCR for the Detection of Plasmodium falciparum DNA in Whole Blood and Serum: A Comparative Analysis with Other Molecular Methods
title_full Droplet Digital PCR for the Detection of Plasmodium falciparum DNA in Whole Blood and Serum: A Comparative Analysis with Other Molecular Methods
title_fullStr Droplet Digital PCR for the Detection of Plasmodium falciparum DNA in Whole Blood and Serum: A Comparative Analysis with Other Molecular Methods
title_full_unstemmed Droplet Digital PCR for the Detection of Plasmodium falciparum DNA in Whole Blood and Serum: A Comparative Analysis with Other Molecular Methods
title_short Droplet Digital PCR for the Detection of Plasmodium falciparum DNA in Whole Blood and Serum: A Comparative Analysis with Other Molecular Methods
title_sort droplet digital pcr for the detection of plasmodium falciparum dna in whole blood and serum: a comparative analysis with other molecular methods
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7350319/
https://www.ncbi.nlm.nih.gov/pubmed/32560386
http://dx.doi.org/10.3390/pathogens9060478
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