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Improved DNA extraction technique from clot for the diagnosis of Chagas disease

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dc.contributor.author Mayta, Holger
dc.contributor.author Romero, Yomara K.
dc.contributor.author Pando, Alejandra
dc.contributor.author Verastegui Pimentel, Manuela Renee
dc.contributor.author Tinajeros, Freddy
dc.contributor.author Bozo, Ricardo
dc.contributor.author Henderson-Frost, Josephine
dc.contributor.author Colanzi, Rony
dc.contributor.author Flores, Jorge
dc.contributor.author Lerner, Richard
dc.contributor.author Bern, Caryn
dc.contributor.author Gilman, Robert Hugh
dc.date.accessioned 2019-07-04T17:00:16Z
dc.date.available 2019-07-04T17:00:16Z
dc.date.issued 2019
dc.identifier.uri https://hdl.handle.net/20.500.12866/6829
dc.description.abstract BACKGROUND: The detection of Trypanosoma cruzi genetic material in clinical samples is considered an important diagnostic tool for Chagas disease. We have previously demonstrated that PCR using clot samples yields greater sensitivity than either buffy coat or whole blood samples. However, phenol-chloroform DNA extraction from clot samples is difficult and toxic. The objective of the present study was to improve and develop a more sensitive method to recover parasite DNA from clot samples for the diagnosis of Chagas disease. METHODOLOGY/PRINCIPAL FINDINGS: A total of 265 match pair samples of whole blood-guanidine (GEB) and clot samples were analyzed; 150 were from Chagas seropositive subjects. DNA was extracted from both whole blood-guanidine samples, using a previously standardized methodology, and from clot samples, using a newly developed methodology based on a combination of the FastPrep technique and the standard method for GEB extraction. A qPCR targeting the nuclear satellite sequences was used to compare the sample source and the extraction method. Of the 150 samples from Chagas positive individuals by serology, 47 samples tested positive by qPCR with DNA extracted by both GEB and clot, but an additional 13 samples tested positive only in DNA extracted from clot. No serology-negative samples resulted positive when tested by qPCR. CONCLUSIONS: The new methodology for DNA extraction from clot samples improves the molecular diagnosis of Chagas disease. en_US
dc.language.iso eng
dc.publisher Public Library of Science
dc.relation.ispartofseries PLoS Neglected Tropical Diseases
dc.rights info:eu-repo/semantics/restrictedAccess
dc.rights.uri https://creativecommons.org/licenses/by-nc-nd/4.0/deed.es
dc.subject Article en_US
dc.subject blood en_US
dc.subject blood clot en_US
dc.subject blood clot lysis en_US
dc.subject blood sampling en_US
dc.subject Chagas disease en_US
dc.subject Chagas Disease en_US
dc.subject controlled study en_US
dc.subject diagnostic test en_US
dc.subject Diagnostic Tests, Routine en_US
dc.subject DNA extraction en_US
dc.subject DNA, Protozoan en_US
dc.subject enzyme linked immunosorbent assay en_US
dc.subject female en_US
dc.subject genetics en_US
dc.subject guanidine en_US
dc.subject hemagglutination test en_US
dc.subject hemagglutination test kit en_US
dc.subject human en_US
dc.subject Humans en_US
dc.subject isolation and purification en_US
dc.subject major clinical study en_US
dc.subject molecular diagnosis en_US
dc.subject Molecular Diagnostic Techniques en_US
dc.subject parasitology en_US
dc.subject procedures en_US
dc.subject protozoal DNA en_US
dc.subject real time polymerase chain reaction en_US
dc.subject Real-Time Polymerase Chain Reaction en_US
dc.subject satellite DNA en_US
dc.subject sensitivity and specificity en_US
dc.subject Sensitivity and Specificity en_US
dc.subject Serologic Tests en_US
dc.subject serology en_US
dc.subject Trypanosoma cruzi en_US
dc.title Improved DNA extraction technique from clot for the diagnosis of Chagas disease en_US
dc.type info:eu-repo/semantics/article
dc.identifier.doi https://doi.org/10.1371/journal.pntd.0007024
dc.subject.ocde https://purl.org/pe-repo/ocde/ford#3.03.06
dc.relation.issn 1935-2735

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