Please use this identifier to cite or link to this item: https://www.arca.fiocruz.br/handle/icict/36232
Title: Ribosomal and non-ribosomal PCR targets for the detection of low-density and mixed malaria infections
Authors: Amaral, Lara Cotta
Robortella, Daniela Rocha
Guimarães, Luiz Felipe Ferreira
Limongi, Jean Ezequiel
Fontes, Cor Jesus Fernandes
Pereira, Dhelio Batista
Brito, Cristiana Ferreira Alves de
Kano, Flora Satiko
Sousa, Taís Nóbrega de
Carvalho, Luzia Helena de
Affilliation: Fundação Oswaldo Cruz. Instituto Rene Rachou. Belo Horizonte, MG, Brasil/Universidade Federal de Minas Gerais. Belo Horizonte, MG, Brasil.
Fundação Oswaldo Cruz. Instituto Rene Rachou. Belo Horizonte, MG, Brasil/Universidade Federal de Minas Gerais. Belo Horizonte, MG, Brasil.
Fundação Oswaldo Cruz. Instituto Rene Rachou. Belo Horizonte, MG, Brasil.
Universidade Federal de Uberlândia. Uberlândia, MG, Brasil.
Universidade Federal de Mato Grosso.Hospital Julio Mueller. Cuiabá. MT, Brasil.
Centro de Pesquisas em Medicina Tropical de Rondônia. Porto Velho, RO, Brasil.
Fundação Oswaldo Cruz. Instituto Rene Rachou. Belo Horizonte, MG, Brasil.
Fundação Oswaldo Cruz. Instituto Rene Rachou. Belo Horizonte, MG, Brasil.
Fundação Oswaldo Cruz. Instituto Rene Rachou. Belo Horizonte, MG, Brasil.
Fundação Oswaldo Cruz. Instituto Rene Rachou. Belo Horizonte, MG, Brasil / Universidade Federal de Minas Gerais. Belo Horizonte, MG, Brasil.
Abstract: Background The unexpected high proportion of submicroscopic malaria infections in areas with low transmission intensity challenges the control and elimination of malaria in the Americas. The current PCR-based assays present limitations as most protocols still rely on amplification of few-copies target gene. Here, the hypothesis was that amplification of different plasmodial targets—ribosomal (18S rRNA) and non-ribosomal multi-copy sequences (Pvr47 for Plasmodium vivax and Pfr364 for Plasmodium falciparum)—could increase the chances of detecting submicroscopic malaria infection. Methods A non-ribosomal real-time PCR assay targeting Pvr47/Pfr364 (NR-qPCR) was established and compared with three additional PCR protocols, two of them based on 18S rRNA gene amplification (Nested-PCR and R-qPCR) and one based on Pvr47/Pfr364 targets (NR-cPCR). The limit of detection of each PCR protocol, at single and artificial mixed P. vivax/P. falciparum infections, was determined by end-point titration curves. Field samples from clinical (n = 110) and subclinical (n = 324) malaria infections were used to evaluate the impact of using multiple molecular targets to detect malaria infections. Results The results demonstrated that an association of ribosomal and non-ribosomal targets did not increase sensitivity to detect submicroscopic malaria infections. Despite of that, artificial mixed-malaria infections demonstrated that the NR-qPCR was the most sensitive protocol to detect low-levels of P. vivax/P. falciparum co-infections. Field studies confirmed that submicroscopic malaria represented a large proportion (up to 77%) of infections among asymptomatic Amazonian residents, with a high proportion of infections (~ 20%) identified only by the NR-qPCR. Conclusions This study presents a new species-specific non-ribosomal PCR assay with potential to identify low-density P. vivax and P. falciparum infections. As the majority of subclinical infections was caused by P. vivax, the commonest form of malaria in the Amazon area, future studies should investigate the potential of Pvr47/Pfr364 to detect mixed-malaria infections in the field. Electronic supplementary material The online version of this article (10.1186/s12936-019-2781-3) contains supplementary material, which is available to authorized users.
Keywords: Malaria
Molecular diagnosis
PCR
Submicroscopic
Mixed-malaria infections
keywords: Malaria
PCR
Issue Date: 2019
Publisher: BMC
Citation: AMARAL, Lara Cotta et al. Ribosomal and non-ribosomal PCR targets for the detection of low-density and mixed malaria infections. Malaria Journal, v. 18, n. 1, p. 1-14, 2019.
DOI: 10.1186/s12936-019-2781-3
ISSN: 1475-2875
Copyright: restricted access
Appears in Collections:MG - IRR - Artigos de Periódicos

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