scholarly journals Analysis of Plasmodium vivax schizont transcriptomes from field isolates reveals heterogeneity of expression of genes involved in host-parasite interactions

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Sasha V. Siegel ◽  
Lia Chappell ◽  
Jessica B. Hostetler ◽  
Chanaki Amaratunga ◽  
Seila Suon ◽  
...  
Keyword(s):  
Plasmodium Vivax ◽  
Ex Vivo ◽  
Culture Method ◽  
Surface Proteins ◽  
Parasite Development ◽  
Field Isolates ◽  
Host Parasite Interactions ◽  
Short Period ◽  
Host Parasite

Abstract Plasmodium vivax gene regulation remains difficult to study due to the lack of a robust in vitro culture method, low parasite densities in peripheral circulation and asynchronous parasite development. We adapted an RNA-seq protocol “DAFT-seq” to sequence the transcriptome of four P. vivax field isolates that were cultured for a short period ex vivo before using a density gradient for schizont enrichment. Transcription was detected from 78% of the PvP01 reference genome, despite being schizont-enriched samples. This extensive data was used to define thousands of 5′ and 3′ untranslated regions, some of which overlapped with neighbouring transcripts, and to improve the gene models of 352 genes, including identifying 20 novel gene transcripts. This dataset has also significantly increased the known amount of heterogeneity between P. vivax schizont transcriptomes from individual patients. The majority of genes found to be differentially expressed between the isolates lack Plasmodium falciparum homologs and are predicted to be involved in host-parasite interactions, with an enrichment in reticulocyte binding proteins, merozoite surface proteins and exported proteins with unknown function. An improved understanding of the diversity within P. vivax transcriptomes will be essential for the prioritisation of novel vaccine targets.

scholarly journals Analysis of Plasmodium vivax schizont transcriptomes from field isolates reveals heterogeneity of expression of genes involved in host-parasite interactions

2020 ◽  
Author(s):  
Sasha V. Siegel ◽  
Lia Chappell ◽  
Jessica B. Hostetler ◽  
Chanaki Amaratunga ◽  
Seila Suon ◽  
...  
Keyword(s):  
Plasmodium Vivax ◽  
Ex Vivo ◽  
Culture Method ◽  
Surface Proteins ◽  
Parasite Development ◽  
Field Isolates ◽  
Host Parasite Interactions ◽  
Short Period ◽  
Host Parasite

AbstractPlasmodium vivax gene regulation remains difficult to study due to the lack of a robust in vitro culture method, low parasite densities in peripheral circulation and asynchronous parasite development. We adapted an RNA-seq protocol “DAFT-seq” to sequence the transcriptome of four P. vivax field isolates that were cultured for a short period ex vivo before using a density gradient for schizont enrichment. Transcription was detected from 78% of the PvP01 reference genome, despite being schizont-enriched samples. This extensive data was used to define thousands of 5’ and 3’ untranslated regions (UTRs), some of which overlapped with neighbouring transcripts, and to improve the gene models of 352 genes, including identifying 20 novel gene transcripts. This dataset has also significantly increased the known amount of heterogeneity between P. vivax schizont transcriptomes from individual patients. The majority of genes found to be differentially expressed between the isolates lack Plasmodium falciparum homologs and are predicted to be involved in host-parasite interactions, with an enrichment in reticulocyte binding proteins, merozoite surface proteins and exported proteins with unknown function. An improved understanding of the diversity within P. vivax transcriptomes will be essential for the prioritisation of novel vaccine targets.

scholarly journals FLAgellum Member 8 modulates extravascular trypanosome distribution in the mammalian host

2021 ◽  
Author(s):  
Estefanía Calvo Alvarez ◽  
Aline Crouzols ◽  
Brice Rotureau
Keyword(s):  
Blood Circulation ◽  
Parasite Development ◽  
Mammalian Host ◽  
Host Parasite Interactions ◽  
Flagellar Protein ◽  
Host Parasite ◽  
Parasite Populations ◽  
Extravascular Compartment ◽  
Host Tissues

The African trypanosome flagellum is essential in multiple aspects of the parasite development. In the mammalian infective form of this protist, FLAgellar Member 8 (FLAM8) is a large protein distributed along the entire flagellum that is suspected to be involved in host-parasite interactions. Analyses of knockdown and knockout trypanosomes demonstrated that FLAM8 is not essential in vitro for survival, growth, motility and slender to stumpy differentiation. Functional investigations in experimental infections showed that FLAM8 -deprived trypanosomes are able to establish and maintain the infection in the blood circulation, and to differentiate into transmissible stumpy forms. However, bioluminescence imaging revealed that FLAM8 -null parasites exhibit an impaired dissemination in the extravascular compartment, especially in the skin, that is partially restored by the addition of a single rescue copy of FLAM8 . To our knowledge, FLAM8 is the first example of a flagellar protein that modulates T. brucei parasite distribution in the host tissues, contributing to the maintenance of extravascular parasite populations in mammalian anatomical niches.

scholarly journals A decellularized human corneal scaffold for anterior corneal surface reconstruction

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Naresh Polisetti ◽  
Anke Schmid ◽  
Ursula Schlötzer-Schrehardt ◽  
Philip Maier ◽  
Stefan J. Lang ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Ex Vivo ◽  
Sodium Deoxycholate ◽  
Nuclear Material ◽  
Biological Properties ◽  
Host Cells ◽  
Human Cornea ◽  
Human Donor ◽  
Short Period

AbstractAllogenic transplants of the cornea are prone to rejection, especially in repetitive transplantation and in scarred or highly vascularized recipient sites. Patients with these ailments would particularly benefit from the possibility to use non-immunogenic decellularized tissue scaffolds for transplantation, which may be repopulated by host cells in situ or in vitro. So, the aim of this study was to develop a fast and efficient decellularization method for creating a human corneal extracellular matrix scaffold suitable for repopulation with human cells from the corneal limbus. To decellularize human donor corneas, sodium deoxycholate, deoxyribonuclease I, and dextran were assessed to remove cells and nuclei and to control tissue swelling, respectively. We evaluated the decellularization effects on the ultrastructure, optical, mechanical, and biological properties of the human cornea. Scaffold recellularization was studied using primary human limbal epithelial cells, stromal cells, and melanocytes in vitro and a lamellar transplantation approach ex vivo. Our data strongly suggest that this approach allowed the effective removal of cellular and nuclear material in a very short period of time while preserving extracellular matrix proteins, glycosaminoglycans, tissue structure, and optical transmission properties. In vitro recellularization demonstrated good biocompatibility of the decellularized human cornea and ex vivo transplantation revealed complete epithelialization and stromal repopulation from the host tissue. Thus, the generated decellularized human corneal scaffold could be a promising biological material for anterior corneal reconstruction in the treatment of corneal defects.

scholarly journals Ex VivoMaturation Assay for Testing Antimalarial Sensitivity of Rodent Malaria Parasites

2016 ◽  
Vol 60 (11) ◽  
pp. 6859-6866 ◽  
Author(s):  
Zi Wei Chang ◽  
Benoit Malleret ◽  
Bruce Russell ◽  
Laurent Rénia ◽  
Carla Claser
Keyword(s):  
Ex Vivo ◽  
Single Step ◽  
Parasite Development ◽  
Ring Stage ◽  
Malaria Parasites ◽  
Rodent Malaria ◽  
Content Type ◽  
Parasite Biology

ABSTRACTEx vivoassay systems provide a powerful approach to studying human malaria parasite biology and to testing antimalarials. For rodent malaria parasites, short-termin vitroculture andex vivoantimalarial susceptibility assays are relatively cumbersome, relying onin vivopassage for synchronization, since ring-stage parasites are an essential starting material. Here, we describe a new approach based on the enrichment of ring-stagePlasmodium berghei,P. yoelii, andP. vinckei vinckeiusing a single-step Percoll gradient. Importantly, we demonstrate that the enriched ring-stage parasites develop synchronously regardless of the parasite strain or species used. Using a flow cytometry assay with Hoechst and ethidium or MitoTracker dye, we show that parasite development is easily and rapidly monitored. Finally, we demonstrate that this approach can be used to screen antimalarial drugs.

scholarly journals The three-spined stickleback-Schistocephalus solidussystem: an experimental model for investigating host-parasite interactions in fish

Parasitology ◽  
2009 ◽  
Vol 137 (3) ◽  
pp. 411-424 ◽  
Author(s):  
I. BARBER ◽  
J. P. SCHARSACK
Keyword(s):  
Immune Responses ◽  
Gasterosteus Aculeatus ◽  
Laboratory Model ◽  
Host Fish ◽  
Schistocephalus Solidus ◽  
Host Parasite Interactions ◽  
Experimental Laboratory ◽  
Host Parasite ◽  
Host Behaviour

SUMMARYPlerocercoids of the pseudophyllidean cestodeSchistocephalus solidusinfect the three-spined sticklebackGasterosteus aculeatus, with important consequences for the biology of host fish. Techniques for culturing the parasitein vitroand generating infective stages that can be used to infect sticklebacks experimentally have been developed, and the system is increasingly used as a laboratory model for investigating aspects of host-parasite interactions. Recent experimental laboratory studies have focused on the immune responses of hosts to infection, the consequences of infection for the growth and reproductive development of host fish and the effects of infection on host behaviour. Here we introduce the host and the parasite, review the major findings of these recent experimental infection studies and identify further aspects of host parasite interactions that might be investigated using the system.

FURTHER STUDIES ON THE PHYSIOLOGY OF CLADOSPORIUM FULVUM CKE. AND RESISTANCE IN TOMATO

1964 ◽  
Vol 42 (10) ◽  
pp. 1365-1386 ◽  
Author(s):  
Ruth L. Lowther
Keyword(s):  
Culture Media ◽  
Carbon Sources ◽  
Amino Nitrogen ◽  
Cladosporium Fulvum ◽  
Host Parasite Interactions ◽  
Race 1 ◽  
Species Characteristics ◽  
Pathogenic Races ◽  
Host Parasite

Some host–parasite interactions of the C. fulvum – tomato leaf complex have been correlated with degrees of utilization in vitro by races of the pathogen of metabolites which have been shown to occur in different amounts in some tomato hosts which react differentially to them. Similarities and differences in the behavior of three pathogenic races were noted when the plants were grown in a number of amino nitrogen and sugar carbon sources. Similarities are interpreted as representing species characteristics; on the other hand, differences, including colony colors in glutamine and γ-aminobutyric acid, appear to be racial characteristics. Further evidence suggesting that certain host metabolites modify or condition the pathogenic expression was obtained from studies of the effect on pathogenicity of culture media differing in amino nitrogen content. From a comparison of the metabolites of host–pathogen complexes of differing reaction types, some insight was gained into probable nutritional requirements for a race 1 susceptible response. Conversely, where these differed in other reaction-type complexes, clues were obtained regarding possible reasons for resistance to this race.

scholarly journals Transmigration of Trypanosoma cruzi trypomastigotes through 3D cultures resembling a physiological environment

2019 ◽  
Author(s):  
Matías Exequiel Rodríguez ◽  
Mariana Rizzi ◽  
Lucas D. Caeiro ◽  
Yamil E. Masip ◽  
Alina Perrone ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Three Dimensional ◽  
3D Cultures ◽  
Monolayer Cultures ◽  
Host Parasite Interactions ◽  
Virulent Strains ◽  
Host Parasite ◽  
3D Spheroids

AbstractChaga’ disease, caused by the kinetoplastid parasite Trypanosoma cruzi, presents a variety of chronic clinical manifestations whose determinants are still unknown but probably influenced by the host-parasite interplay established during the first stages of the infection, when bloodstream circulating trypomastigotes disseminate to different organs and tissues. After leaving the blood, trypomastigotes must migrate through tissues to invade cells and establish a chronic infection. How this process occurs remains unexplored. Three-dimensional (3D) cultures are physiologically relevant because mimic the microarchitecture of tissues and provide an environment similar to the encountered in natural infections. In this work, we combined the 3D culture technology with host-pathogen interaction, by studying transmigration of trypomastigotes into 3D spheroids. T. cruzi strains with similar infection dynamics in 2D monolayer cultures but with different in vivo behavior (CL Brener, virulent; SylvioX10 no virulent) presented different infection rates in spheroids (CL Brener ∼40%, SylvioX10 <10%). Confocal microscopy images evidenced that trypomastigotes from CL Brener and other highly virulent strains presented a great ability to transmigrate inside 3D spheroids: as soon as 4 hours post infection parasites were found at 50 µm in depth inside the spheroids. CL Brener trypomastigotes were evenly distributed and systematically observed in the space between cells, suggesting a paracellular route of transmigration to deepen into the spheroids. On the other hand, poor virulent strains presented a weak migratory capacity and remained in the external layers of spheroids (<10µm) with a patch-like distribution pattern. The invasiveness -understood as the ability to transmigrate deep into spheroids- was not a transferable feature between strains, neither by soluble or secreted factors nor by co-cultivation of trypomastigotes from invasive and non-invasive strains. We also studied the transmigration of recent T. cruzi isolates from children that were born congenitally infected, which showed a high migrant phenotype while an isolate form an infected mother (that never transmitted the infection to any of her 3 children) was significantly less migratory. Altogether, our results demonstrate that in a 3D microenvironment each strain presents a characteristic migration pattern and distribution of parasites in the spheroids that can be associated to their in vivo behavior. Certainly, the findings presented here could not have been studied with traditional 2D monolayer cultures.Author SummaryTrypanosoma cruzi is the protozoan parasite that causes Chaga’ disease, also known as American trypanosomiasis. Experimental models of the infection evidence that different strains of the parasite present different virulence in the host, which cannot be always reproduced in 2D monolayer cultures. Three dimensional (3D) cultures can be useful models to study complex host-parasite interactions because they mimic in vitro the microarchitecture of tissues and provide an environment similar to the encountered in natural infections. In particular, spheroids are small 3D aggregates of cells that interact with each other and with the extracellular matrix that they secrete resembling the original microenvironment both functionally and structurally. Spheroids have rarely been employed to explore infectious diseases and host-parasite interactions. In this work we studied how bloodstream trypomastigotes transmigrate through 3D spheroids mimicking the picture encountered by parasites in tissues soon after leaving circulation. We showed that the behavior of T. cruzi trypomastigotes in 3D cultures reflects their in vivo virulence: virulent strains transmigrate deeply into spheroids while non-virulent strains remain in the external layers of spheroids. Besides, this work demonstrates the usefulness of 3D cultures as an accurate in vitro model for the study of host-pathogen interactions that could not be addressed with conventional monolayer cultures.

scholarly journals A Decellularized Human Corneal Scaffold for Anterior Corneal Surface Reconstruction

2020 ◽  
Author(s):  
Naresh Polisetti ◽  
Anke Schmid ◽  
Ursula Schlötzer-Schrehardt ◽  
Philip Maier ◽  
Stefan Lang ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Ex Vivo ◽  
Sodium Deoxycholate ◽  
Nuclear Material ◽  
Biological Properties ◽  
Host Cells ◽  
Human Cornea ◽  
Human Donor ◽  
Short Period

Abstract Allogenic transplants of the cornea are prone to rejection, especially in repetitive transplantation and in scarred or highly vascularized recipient sites. Patients with these ailments would particularly benefit from the possibility to use non-immunogenic decellularized tissue scaffolds for transplantation, which may be repopulated by host cells in situ or in vitro. So, the aim of this study was to develop a fast and efficient decellularization method for creating a human corneal extracellular matrix scaffold suitable for repopulation with human cells from the corneal limbus. To decellularize human donor corneas, sodium deoxycholate, deoxyribonuclease I, and dextran were assessed to remove cells and nuclei and to control tissue swelling, respectively. We evaluated the decellularization effects on the ultrastructure, optical, mechanical, and biological properties of the human cornea. Scaffold recellularization was studied using primary human limbal epithelial cells, stromal cells, and melanocytes in vitro and a lamellar transplantation approach ex vivo. Our data strongly suggest that this approach allowed the effective removal of cellular and nuclear material in a very short period of time while preserving extracellular matrix proteins, glycosaminoglycans, tissue structure, and optical transmission properties. In vitro recellularization demonstrated good biocompatibility of the decellularized human cornea and ex vivo transplantation revealed complete epithelialization and stromal repopulation from the host tissue. Thus, the generated decellularized human corneal scaffold could be a promising biological material for anterior corneal reconstruction in the treatment of corneal defects.

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