Validation of a novel multiplex real-time PCR assay for Trypanosoma cruzi detection and quantification in açai pulp.
In Brazil, orally acquired T. cruzi infection has become the most relevant transmission mechanisms from public health perspective. Around 70% of new Chagas disease cases have been associated with consumption of contaminated food or beverages. Açai (Euterpe oleracea and Euterpe precatoria) is current...
| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2021-01-01
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| Series: | PLoS ONE |
| Online Access: | https://doi.org/10.1371/journal.pone.0246435 |
| _version_ | 1819012502527148032 |
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| author | Paula Finamore-Araujo Amanda Faier-Pereira Carlos Ramon do Nascimento Brito Eldrinei Gomes Peres Klenicy Kazumy de Lima Yamaguchi Renata Trotta Barroso Ferreira Otacilio Cruz Moreira |
| author_facet | Paula Finamore-Araujo Amanda Faier-Pereira Carlos Ramon do Nascimento Brito Eldrinei Gomes Peres Klenicy Kazumy de Lima Yamaguchi Renata Trotta Barroso Ferreira Otacilio Cruz Moreira |
| author_sort | Paula Finamore-Araujo |
| collection | DOAJ |
| description | In Brazil, orally acquired T. cruzi infection has become the most relevant transmission mechanisms from public health perspective. Around 70% of new Chagas disease cases have been associated with consumption of contaminated food or beverages. Açai (Euterpe oleracea and Euterpe precatoria) is currently one of the most commercialized Amazonian fruits in the Brazilian and international markets. Therefore, it has become important to incorporate in the production process some procedures to measure out effective hygiene and product quality control required by global market. Molecular methods have been developed for rapid detection and quantification of T. cruzi DNA in several biological samples, including food matrices, for epidemiological investigation of Chagas disease and food quality control. However, a high-performance molecular methodology since DNA extraction until detection and quantification of T. cruzi DNA in açai berry pulp is still needed. Herein, a simple DNA extraction methodology was standardized from the supernatant of açai berry pulp stabilized in a 6M Guanidine-HCl/0.2M EDTA buffer. In addition, a multiplex real time qPCR assay, targeting T. cruzi DNA and an Exogenous Internal Positive Control was developed and validated, using reference from all T. cruzi DTUs and commercial samples of açai pulp, from an endemic municipality with previous history of oral Chagas disease outbreak. Thus, a high-sensitivity qPCR assay, that could detect up to 0.01 parasite equivalents/mL in açai, was reached. As of the 45 commercial samples analyzed, 9 (20%) were positive for T. cruzi. This high-sensitive, fast, and easy-to-use molecular assay is compatible with most of the laboratories involved in the investigations of oral Chagas disease outbreaks, representing an important tool to the epidemiology, control, and surveillance of Chagas disease. |
| first_indexed | 2024-12-21T01:45:04Z |
| format | Article |
| id | doaj.art-0a1cf52e80cb49dd8d96a5a6dcb10f09 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-21T01:45:04Z |
| publishDate | 2021-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-0a1cf52e80cb49dd8d96a5a6dcb10f092022-12-21T19:20:02ZengPublic Library of Science (PLoS)PLoS ONE1932-62032021-01-01162e024643510.1371/journal.pone.0246435Validation of a novel multiplex real-time PCR assay for Trypanosoma cruzi detection and quantification in açai pulp.Paula Finamore-AraujoAmanda Faier-PereiraCarlos Ramon do Nascimento BritoEldrinei Gomes PeresKlenicy Kazumy de Lima YamaguchiRenata Trotta Barroso FerreiraOtacilio Cruz MoreiraIn Brazil, orally acquired T. cruzi infection has become the most relevant transmission mechanisms from public health perspective. Around 70% of new Chagas disease cases have been associated with consumption of contaminated food or beverages. Açai (Euterpe oleracea and Euterpe precatoria) is currently one of the most commercialized Amazonian fruits in the Brazilian and international markets. Therefore, it has become important to incorporate in the production process some procedures to measure out effective hygiene and product quality control required by global market. Molecular methods have been developed for rapid detection and quantification of T. cruzi DNA in several biological samples, including food matrices, for epidemiological investigation of Chagas disease and food quality control. However, a high-performance molecular methodology since DNA extraction until detection and quantification of T. cruzi DNA in açai berry pulp is still needed. Herein, a simple DNA extraction methodology was standardized from the supernatant of açai berry pulp stabilized in a 6M Guanidine-HCl/0.2M EDTA buffer. In addition, a multiplex real time qPCR assay, targeting T. cruzi DNA and an Exogenous Internal Positive Control was developed and validated, using reference from all T. cruzi DTUs and commercial samples of açai pulp, from an endemic municipality with previous history of oral Chagas disease outbreak. Thus, a high-sensitivity qPCR assay, that could detect up to 0.01 parasite equivalents/mL in açai, was reached. As of the 45 commercial samples analyzed, 9 (20%) were positive for T. cruzi. This high-sensitive, fast, and easy-to-use molecular assay is compatible with most of the laboratories involved in the investigations of oral Chagas disease outbreaks, representing an important tool to the epidemiology, control, and surveillance of Chagas disease.https://doi.org/10.1371/journal.pone.0246435 |
| spellingShingle | Paula Finamore-Araujo Amanda Faier-Pereira Carlos Ramon do Nascimento Brito Eldrinei Gomes Peres Klenicy Kazumy de Lima Yamaguchi Renata Trotta Barroso Ferreira Otacilio Cruz Moreira Validation of a novel multiplex real-time PCR assay for Trypanosoma cruzi detection and quantification in açai pulp. PLoS ONE |
| title | Validation of a novel multiplex real-time PCR assay for Trypanosoma cruzi detection and quantification in açai pulp. |
| title_full | Validation of a novel multiplex real-time PCR assay for Trypanosoma cruzi detection and quantification in açai pulp. |
| title_fullStr | Validation of a novel multiplex real-time PCR assay for Trypanosoma cruzi detection and quantification in açai pulp. |
| title_full_unstemmed | Validation of a novel multiplex real-time PCR assay for Trypanosoma cruzi detection and quantification in açai pulp. |
| title_short | Validation of a novel multiplex real-time PCR assay for Trypanosoma cruzi detection and quantification in açai pulp. |
| title_sort | validation of a novel multiplex real time pcr assay for trypanosoma cruzi detection and quantification in acai pulp |
| url | https://doi.org/10.1371/journal.pone.0246435 |
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