Author | Dias, Felipe de Almeida | |
Author | Guerra, Barbara | |
Author | Vieira, Larissa Rezende | |
Author | Perdomo, Hugo Diego | |
Author | Gandara, Ana Caroline Paiva | |
Author | Amaral, Raquel Juliana Vionette do | |
Author | Vollú, Renata Estebanez | |
Author | Gomes, Suzete Araujo Oliveira | |
Author | Lara, Flavio Alves | |
Author | Sorgine, Marcos Henrique Ferreira | |
Author | Medei, Emiliano | |
Author | Oliveira, Pedro Lagerblad de | |
Author | Salmon, Didier | |
Access date | 2015-11-19T11:52:16Z | |
Available date | 2015-11-19T11:52:16Z | |
Document date | 2015 | |
Citation | DIAS, Felipe de ALmeida; et al. Monitoring of the Parasite Load in the Digestive Tract of Rhodnius prolixus by Combined qPCR Analysis and Imaging Techniques Provides New Insights into the Trypanosome Life Cycle. PLoS Negl Trop Dis, v.9, n.10, e0004186, 23p, Oct. 2015. | pt_BR |
ISSN | 1935-2727 | |
URI | https://www.arca.fiocruz.br/handle/icict/12281 | |
Language | eng | pt_BR |
Publisher | Plos One | pt_BR |
Rights | open access | |
Title | Monitoring of the Parasite Load in the Digestive Tract of Rhodnius prolixus by Combined qPCR Analysis and Imaging Techniques Provides New Insights into the Trypanosome Life Cycle | pt_BR |
Type | Article | pt_BR |
DOI | 10.1371/journal.pntd.0004186 | |
Abstract | Background
Here we report the monitoring of the digestive tract colonization of Rhodnius prolixus by Trypanosoma
cruzi using an accurate determination of the parasite load by qPCR coupled with
fluorescence and bioluminescence imaging (BLI). These complementary methods revealed
critical steps necessary for the parasite population to colonize the insect gut and establish
vector infection.
Methodology/Principal Findings
qPCR analysis of the parasite load in the insect gut showed several limitations due mainly
to the presence of digestive-derived products that are thought to degrade DNA and inhibit
further the PCR reaction. We developed a real-time PCR strategy targeting the T. cruzi
repetitive satellite DNA sequence using as internal standard for normalization, an exogenous
heterologous DNA spiked into insect samples extract, to precisely quantify the parasite
load in each segment of the insect gut (anterior midgut, AM, posterior midgut, PM, and
hindgut, H). Using combined fluorescence microscopy and BLI imaging as well as qPCR
analysis, we showed that during their journey through the insect digestive tract, most of the
parasites are lysed in the AM during the first 24 hours independently of the gut microbiota.During this short period, live parasites move through the PM to establish the onset of infection.
At days 3–4 post-infection (p.i.), the parasite population begins to colonize the H to
reach a climax at day 7 p.i., which is maintained during the next two weeks. Remarkably,
the fluctuation of the parasite number in H remains relatively stable over the two weeks after
refeeding, while the populations residing in the AM and PM increases slightly and probably
constitutes the reservoirs of dividing epimastigotes.
Conclusions/Significance
These data show that a tuned dynamic control of the population operates in the insect gut to
maintain an equilibrium between non-dividing infective trypomastigote forms and dividing
epimastigote forms of the parasite, which is crucial for vector competence. | pt_BR |
Affilliation | Universidade Federal do Rio de Janeiro. Instituto de Bioquímica Médica Leopoldo de Meis. Rio de Janeiro, RJ, Brasil. | pt_BR |
Affilliation | Universidade Federal do Rio de Janeiro. Centro Nacional de Biologia Estrutural e Bioimagem—CENABIO. Rio de Janeiro, RJ, Brasil. | pt_BR |
Affilliation | Universidade Federal do Rio de Janeiro. Instituto de Bioquímica Médica Leopoldo de Meis. Rio de Janeiro, RJ, Brasil. | pt_BR |
Affilliation | Universidade Federal do Rio de Janeiro. Instituto de Bioquímica Médica Leopoldo de Meis. Rio de Janeiro, RJ, Brasil. | pt_BR |
Affilliation | Universidade Federal do Rio de Janeiro. Instituto de Bioquímica Médica Leopoldo de Meis. Rio de Janeiro, RJ, Brasil. | pt_BR |
Affilliation | Universidade Federal do Rio de Janeiro. Instituto de Bioquímica Médica Leopoldo de Meis. Rio de Janeiro, RJ, Brasil. | pt_BR |
Affilliation | Universidade Federal do Rio de Janeiro. Instituto de Microbiologia Professor Paulo de Góes. Rio de Janeiro, RJ, Brasil. | pt_BR |
Affilliation | Universidade Federal Fluminense. Departamento de Biologia Geral. Niterói, RJ, Brasil. | pt_BR |
Affilliation | Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, Brasil. | pt_BR |
Affilliation | Universidade Federal do Rio de Janeiro. Instituto de Bioquímica Médica Leopoldo de Meis. Rio de Janeiro, RJ, Brasil / Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular (INCT-EM). Rio de Janeiro, RJ, Brasil. | pt_BR |
Affilliation | Universidade Federal do Rio de Janeiro. Centro Nacional de Biologia Estrutural e Bioimagem—CENABIO. Rio de Janeiro, RJ, Brasil. | pt_BR |
Affilliation | Universidade Federal do Rio de Janeiro. Instituto de Bioquímica Médica Leopoldo de Meis. Rio de Janeiro, RJ, Brasil / Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular (INCT-EM). Rio de Janeiro, RJ, Brasil. | pt_BR |
Affilliation | Universidade Federal do Rio de Janeiro. Instituto de Bioquímica Médica Leopoldo de Meis. Rio de Janeiro, RJ, Brasil. | pt_BR |
Subject | Rhodnius prolixus | pt_BR |
Subject | Trypanosome Life Cycle | pt_BR |
Subject | Parasite Load | pt_BR |
Subject | Digestive Tract | pt_BR |
DeCS | Reação em Cadeia da Polimerase | pt_BR |
DeCS | Carga Parasitária | pt_BR |
DeCS | Trato Gastrointestinal | pt_BR |
DeCS | Trypanosoma cruzi | pt_BR |