Validation of a novel multiplex real-time PCR assay for Trypanosoma cruzi detection and quantification in açai pulp.

Autor: Finamore-Araujo P; Plataforma Fiocruz de PCR em Tempo Real RPT09A -Laboratório de Biologia Molecular e Doenças Endêmicas, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, Brazil., Faier-Pereira A; Plataforma Fiocruz de PCR em Tempo Real RPT09A -Laboratório de Biologia Molecular e Doenças Endêmicas, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, Brazil., Ramon do Nascimento Brito C; Departamento de Análises Clínicas e Toxicológicas, Centro de Ciências da Saúde, Universidade Federal do Rio Grande do Norte, Natal, RN, Brazil., Gomes Peres E; Departamento de Química, Universidade Federal do Amazonas, Manaus, AM, Brazil., Kazumy de Lima Yamaguchi K; Instituto de Saúde e Biotecnologia, Universidade Federal do Amazonas-Campus Coari, Amazonas, Brazil., Trotta Barroso Ferreira R; Instituto Nacional de Controle de Qualidade em Saúde, Fundação Oswaldo Cruz, Rio de Janeiro, Brazil., Moreira OC; Plataforma Fiocruz de PCR em Tempo Real RPT09A -Laboratório de Biologia Molecular e Doenças Endêmicas, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, Brazil.
Jazyk: angličtina
Zdroj: PloS one [PLoS One] 2021 Feb 02; Vol. 16 (2), pp. e0246435. Date of Electronic Publication: 2021 Feb 02 (Print Publication: 2021).
DOI: 10.1371/journal.pone.0246435
Abstrakt: 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.
Competing Interests: The authors have declared that no competing interests exist.
Databáze: MEDLINE
Externí odkaz:
Nepřihlášeným uživatelům se plný text nezobrazuje