Expression of foreign proteins in Trypanosoma congolense

Downey, Nicholas; Donelson, John E.

doi:10.1016/s0166-6851(99)00136-x

Cited by 11 publications

(8 citation statements)

References 23 publications

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“…Most sequences were homologous to CP2 and assigned to SAV1 (21 sequences), SAV2 (10) or SAV3 (7). Sequences of SAV1 predominated and, despite preferentially showing typical triad (CHN), this group included the first variant (SHN) reported in T. congolense (Downey and Donelson, 1999) in the strain TRUM 183.…”

Section: Repertoire and Genealogy Of The Catalytic Domains Of Catl-limentioning

confidence: 98%

Congopain genes diverged to become specific to Savannah, Forest and Kilifi subgroups of Trypanosoma congolense, and are valuable for diagnosis, genotyping and phylogenetic inferences

Rodrigues

Ortíz

Costa-Martins

et al. 2014

Infection, Genetics and Evolution

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Section: Repertoire and Genealogy Of The Catalytic Domains Of Catl-limentioning

confidence: 98%

Congopain genes diverged to become specific to Savannah, Forest and Kilifi subgroups of Trypanosoma congolense, and are valuable for diagnosis, genotyping and phylogenetic inferences

Rodrigues

Ortíz

Costa-Martins

et al. 2014

Infection, Genetics and Evolution

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“…Transgene expression and RNA interference (RNAi) in T. congolense PCF have been reported previously [11],[27]. Specifically, the tetracycline inducible system first used in T. brucei (constitutive expression of RNA polymerase and tetracycline repressor, [5]) has been adapted for T. congolense by Inoue and colleagues in the TRUM183 uncloned serodeme [11].…”

Section: Resultsmentioning

confidence: 99%

Complete In Vitro Life Cycle of Trypanosoma congolense: Development of Genetic Tools

et al. 2010

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BackgroundAnimal African trypanosomosis, a disease mainly caused by the protozoan parasite Trypanosoma congolense, is a major constraint to livestock productivity and has a significant impact in the developing countries of Africa. RNA interference (RNAi) has been used to study gene function and identify drug and vaccine targets in a variety of organisms including trypanosomes. However, trypanosome RNAi studies have mainly been conducted in T. brucei, as a model for human infection, largely ignoring livestock parasites of economical importance such as T. congolense, which displays different pathogenesis profiles. The whole T. congolense life cycle can be completed in vitro, but this attractive model displayed important limitations: (i) genetic tools were currently limited to insect forms and production of modified infectious BSF through differentiation was never achieved, (ii) in vitro differentiation techniques lasted several months, (iii) absence of long-term bloodstream forms (BSF) in vitro culture prevented genomic analyses.Methodology/Principal FindingsWe optimized culture conditions for each developmental stage and secured the differentiation steps. Specifically, we devised a medium adapted for the strenuous development of stable long-term BSF culture. Using Amaxa nucleofection technology, we greatly improved the transfection rate of the insect form and designed an inducible transgene expression system using the IL3000 reference strain. We tested it by expression of reporter genes and through RNAi. Subsequently, we achieved the complete in vitro life cycle with dramatically shortened time requirements for various wild type and transgenic strains. Finally, we established the use of modified strains for experimental infections and underlined a host adaptation phase requirement.Conclusions/SignificanceWe devised an improved T. congolense model, which offers the opportunity to perform functional genomics analyses throughout the whole life cycle. It represents a very useful tool to understand pathogenesis mechanisms and to study potential therapeutic targets either in vitro or in vivo using a mouse model.

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“…Major breakthroughs have recently been made in the study of T. congolense , namely the development and standardization of axenic cultures and the development of transfection techniques [35], [36]. Researchers had showed growing interest in T. vivax adaptation to experimental animals from more than twenty years after the encouraging studies of Leeflang in the 1970s, and the description being made of parasite isolates that were infective to rodents [22], [37], [38].…”

Section: Discussionmentioning

confidence: 99%

“…The development in the 1990s of Trypanosomatid transfection using Gene Pulser systems [42]–[45] was then adapted to other species [36], [46], [47]. Despite a number of similar transfection parameters shared by all Kinetoplastidae , recombination efficiencies and susceptibility to drug selection diverged.…”

Section: Discussionmentioning

confidence: 99%

“…This may suggest that T. vivax is not able to maintain episomal DNA, unlike T. brucei and T. congolense that carry out particular sequences (i.e. parp ) that promote episomal mantainance [36], [48]. Alternatively, T. vivax circular DNA may be integrated but less efficiently than linear plasmids, resulting in a smaller number of live parasites that do not survive culture conditions.…”

Section: Discussionmentioning

confidence: 99%

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Genetic Engineering of Trypanosoma (Dutonella) vivax and In Vitro Differentiation under Axenic Conditions

D’Archivio¹,

Medina²,

Cosson³

et al. 2011

PLoS Negl Trop Dis

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Trypanosoma vivax is one of the most common parasites responsible for animal trypanosomosis, and although this disease is widespread in Africa and Latin America, very few studies have been conducted on the parasite's biology. This is in part due to the fact that no reproducible experimental methods had been developed to maintain the different evolutive forms of this trypanosome under laboratory conditions. Appropriate protocols were developed in the 1990s for the axenic maintenance of three major animal Trypanosoma species: T. b. brucei, T. congolense and T. vivax. These pioneer studies rapidly led to the successful genetic manipulation of T. b. brucei and T. congolense. Advances were made in the understanding of these parasites' biology and virulence, and new drug targets were identified. By contrast, challenging in vitro conditions have been developed for T. vivax in the past, and this per se has contributed to defer both its genetic manipulation and subsequent gene function studies. Here we report on the optimization of non-infective T. vivax epimastigote axenic cultures and on the process of parasite in vitro differentiation into metacyclic infective forms. We have also constructed the first T. vivax specific expression vector that drives constitutive expression of the luciferase reporter gene. This vector was then used to establish and optimize epimastigote transfection. We then developed highly reproducible conditions that can be used to obtain and select stably transfected mutants that continue metacyclogenesis and are infectious in immunocompetent rodents.

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Expression of foreign proteins in Trypanosoma congolense

Cited by 11 publications

References 23 publications

Congopain genes diverged to become specific to Savannah, Forest and Kilifi subgroups of Trypanosoma congolense, and are valuable for diagnosis, genotyping and phylogenetic inferences

Congopain genes diverged to become specific to Savannah, Forest and Kilifi subgroups of Trypanosoma congolense, and are valuable for diagnosis, genotyping and phylogenetic inferences

Complete In Vitro Life Cycle of Trypanosoma congolense: Development of Genetic Tools

Genetic Engineering of Trypanosoma (Dutonella) vivax and In Vitro Differentiation under Axenic Conditions

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