<i>Trypanosoma cruzi</i>
            : single cell live imaging inside infected tissues

Ferreira, Bianca Rodrigues Lima; Orikaza, Cristina Mary; Cordero, Esteban; Mortara, Renato Arruda

doi:10.1111/cmi.12553

Cited by 7 publications

(8 citation statements)

References 20 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…In these forms, the kinetoplast remains discoid and anterior and the nucleus is spherical but the cell body is elongated in comparison to an amastigote (Fig 8C, orange box and inset). Similar forms have been reported once before in a very early stage infection (day 8) [35]. These epimastigote-like forms, which we detected repeatedly in tissue samples, often co-existed with dividing amastigotes and differentiating trypomastigotes in the same infected cell, and could be observed by live cell imaging in vitro (S5 Fig).…”

Section: Multiple Morphological Forms Of T Cruzi Are Present In Deepsupporting

confidence: 87%

Intracellular DNA replication and differentiation of Trypanosoma cruzi is asynchronous within individual host cells in vivo at all stages of infection

et al. 2020

View full text Add to dashboard Cite

Investigations into intracellular replication and differentiation of Trypanosoma cruzi within the mammalian host have been restricted by limitations in our ability to detect parasitized cells throughout the course of infection. We have overcome this problem by generating genetically modified parasites that express a bioluminescent/fluorescent fusion protein. By combining in vivo imaging and confocal microscopy, this has enabled us to routinely visualise murine infections at the level of individual host cells. These studies reveal that intracellular parasite replication is an asynchronous process, irrespective of tissue location or disease stage. Furthermore, using TUNEL assays and EdU labelling, we demonstrate that within individual infected cells, replication of both mitochondrial (kDNA) and nuclear genomes is not coordinated within the parasite population, and that replicating amastigotes and non-replicating trypomastigotes can co-exist in the same cell. Finally, we report the presence of distinct non-canonical morphological forms of T. cruzi in the mammalian host. These appear to represent transitional forms in the amastigote to trypomastigote differentiation process. Therefore, the intracellular life-cycle of T. cruzi in vivo is more complex than previously realised, with potential implications for our understanding of disease pathogenesis, immune evasion and drug development. Dissecting the mechanisms involved will be an important experimental challenge.

show abstract

Section: Multiple Morphological Forms Of T Cruzi Are Present In Deepsupporting

confidence: 87%

Intracellular DNA replication and differentiation of Trypanosoma cruzi is asynchronous within individual host cells in vivo at all stages of infection

et al. 2020

View full text Add to dashboard Cite

show abstract

“…The same procedure were performed in CL-DsRed infected mice, but due to weak fluorescence we could not obtain representative images. Nonetheless, images of CL-DsRed infected tissues (mice) can be found in our previous work ( Ferreira et al, 2016 ).…”

Section: Resultsmentioning

confidence: 99%

“…We used G (TcI) ( Yoshida, 1983 ) and CL (TcVI) ( Brener and Chiari, 1963 ) T. cruzi strains, wild-type or transfected with GFP (G strain) ( Cruz et al, 2012 ) or DsRed (CL strain) ( Ferreira et al, 2016 ) in pTREX plasmids. Tissue-culture derived trypomastigotes (TCTs), analogous to bloodstream trypomastigotes, were obtained from the supernatant of infected Vero cells (Instituto Adolfo Lutz, São Paulo, SP, Brazil) that were grown in RPMI 1640 medium (Vitrocell, Embriolife, Brazil) supplemented with 2% fetal bovine serum (FBS) (Gibco, Thermo Fisher Scientific, United States), 10 μg/mL streptomycin, 100 U/mL penicillin and 40 μg/mL gentamicin (antibiotics from Sigma-Aldrich, United States) at 37°C and 5% CO 2 .…”

Section: Methodsmentioning

confidence: 99%

“…The organs were placed onto a cold paraffin surface, and slices were cut with two parallel GEM single-edge blades, immersed in PBS containing 1 μM Hoechst 33342 (Invitrogen, Carlsbad, CA, United States) to label nuclei and then transferred to glass-bottom dishes (MatTek Corporation, Ashland, MA, United States) in RPMI-1640 medium (Vitrocell, Embriolife, Brazil) supplemented with 10% FBS (Gibco, Thermo Fisher Scientific, United States). Ex vivo specimens were transferred to a stage-top incubator at 37°C and 5% CO 2 with controlled humidity (Tokai Hit, Japan) on a Leica SP5 TS confocal microscope and imaged with a 63 × NA 1.40 oil-immersion objective using the resonant scanner (8000 Hz) mode, as described ( Ferreira et al, 2016 ). Final imaging processing was performed using Imaris software 7.0 (Bitplane).…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

BALB/c and C57BL/6 Mice Cytokine Responses to Trypanosoma cruzi Infection Are Independent of Parasite Strain Infectivity

Ferreira

Brito

et al. 2018

Front. Microbiol.

View full text Add to dashboard Cite

Trypanosoma cruzi is the etiologic agent of Chagas’ disease, which affects 6–7 million people worldwide. Different strains of T. cruzi present specific genotypic and phenotypic characteristics that affect the host–pathogen interactions, and thus, the parasite has been classified into six groups (TcI to TcVI). T. cruzi infection presents two clinical phases, acute and chronic, both with distinct characteristics and important participation by the immune system. However, the specific contributions of parasite and host factors in the disease phases are not yet fully understood. The murine model for Chagas’ disease is well-established and reproduces important features of the human infection, providing an experimental basis for the study of host lineages and parasite strains. Thus, we evaluated acute and chronic infection by the G (TcI) and CL (TcVI) strains of T. cruzi, which have distinct tropisms and infectivity, in two inbred mice lineages (C57BL/6 and BALB/c) that display variable degrees of susceptibility to different T. cruzi strains. Analysis of the parasite loads in host tissues by qPCR showed that CL strain established an infection faster than the G strain; at the same time, the response in BALB/c mice, although diverse in terms of cytokine secretion, was initiated earlier than that in C57BL/6 mice. At the parasitemia peak in the acute phase, we observed, either by confocal microscopy or by qPCR, that the infection was disseminated in all groups analyzed, with some differences concerning parasite tropism; at this point, all animals responded to infection by increasing the serum concentrations of cytokines. However, BALB/c mice seemed to better regulate the immune response than C57BL/6 mice. Indeed, in the chronic phase, C57BL/6 mice still presented exacerbated cytokine and chemokine responses. In summary, our results indicate that in these experimental models, the deregulation of immune response that is typical of chronic Chagas’ disease may be due to control loss over pro- and anti-inflammatory cytokines early in the acute phase of the disease, depending primarily on the host background rather than the parasite strain.

show abstract

“…The parasites used in proliferation and flow cytometry assays were genetically modified to express the enzyme β -galactosidase ( T. cruzi -LacZ [9]). To illustrate the inhibition of parasite proliferation, a T. cruzi expressing GFP (G-GFP strain [10]) was used.…”

Section: Methodsmentioning

confidence: 99%

Phenothiazinium Dyes Are Active againstTrypanosoma cruziIn Vitro

Portapilla

Pereira

Costa

et al. 2019

BioMed Research International

View full text Add to dashboard Cite

Chagas disease is a tropical illness caused by the protozoan Trypanosoma cruzi. The disease affects populations of the Americas and has been spread to other continents due to the migration process. The disease is partially controlled by two drugs, Benznidazole and Nifurtimox. These molecules are active in the acute phase of the infection but are usually ineffective during the symptomatic chronic phase. Several research groups have developed novel candidates to control Chagas disease; however, no novel commercial formulation is available. In this article, we described the anti-T. cruzi effects of phenothiazinium dyes in amastigote and trypomastigote forms of the parasite. Methylene Blue, New Methylene Blue, Toluidine Blue O, and 1,9-Dimethyl Methylene Blue inhibited the parasite proliferation at nanomolar concentrations and also demonstrated low toxicity in host cells. Moreover, combinations of phenothiazinium dyes indicated a synergic pattern against amastigotes compared to the Benznidazole counterparts. Phenothiazinium dyes levels of reactive oxygen species (ROS) and decreased the mitochondrial potential in trypomastigotes, indicating the mechanism of action of the dyes in T. cruzi. Our article offers a basis for future strategies for the control of Chagas disease using low-cost formulations, an important point for endemic underdeveloped regions.

show abstract

Trypanosoma cruzi : single cell live imaging inside infected tissues

Cited by 7 publications

References 20 publications

Intracellular DNA replication and differentiation of Trypanosoma cruzi is asynchronous within individual host cells in vivo at all stages of infection

Intracellular DNA replication and differentiation of Trypanosoma cruzi is asynchronous within individual host cells in vivo at all stages of infection

BALB/c and C57BL/6 Mice Cytokine Responses to Trypanosoma cruzi Infection Are Independent of Parasite Strain Infectivity

Phenothiazinium Dyes Are Active againstTrypanosoma cruziIn Vitro

Contact Info

Product

Resources

About