Tsetse Transmission Research Articles

A conserved trypanosomatid differentiation regulator controls substrate attachment and morphological development in Trypanosoma congolense.

Trypanosomatid parasites undergo developmental regulation to adapt to the different environments encountered during their life cycle. In Trypanosoma brucei, a genome wide selectional screen previously identified a regulator of the protein family ESAG9, which is highly expressed in stumpy forms, a morphologically distinct bloodstream stage adapted for tsetse transmission. This regulator, TbREG9.1, has an orthologue in Trypanosoma congolense, despite the absence of a stumpy morphotype in that parasite species, which is an important cause of livestock trypanosomosis. RNAi mediated gene silencing of TcREG9.1 in Trypanosoma congolense caused a loss of attachment of the parasites to a surface substrate in vitro, a key feature of the biology of these parasites that is distinct from T. brucei. This detachment was phenocopied by treatment of the parasites with a phosphodiesterase inhibitor, which also promotes detachment in the insect trypanosomatid Crithidia fasciculata. RNAseq analysis revealed that TcREG9.1 silencing caused the upregulation of mRNAs for several classes of surface molecules, including transferrin receptor-like molecules, immunoreactive proteins in experimental bovine infections, and molecules related to those associated with stumpy development in T. brucei. Depletion of TcREG9.1 in vivo also generated an enhanced level of parasites in the blood circulation consistent with reduced parasite attachment to the microvasculature. The morphological progression to insect forms of the parasite was also perturbed. We propose a model whereby TcREG9.1 acts as a regulator of attachment and development, with detached parasites being adapted for transmission.

Tsetse Invasion as an Emerging Threat to Socioecological Resilience of Pastoral Communities in Karamoja, Uganda

Over 70% of Uganda is infested by the tsetse fly, which has negative effects on human and livestock health. From colonial to post-independent Uganda, the Government of Uganda has worked to eradicate the tsetse menace. Despite these efforts, recent veterinary reports from the Karamoja sub-region have indicated widespread tsetse invasion. This study investigated the potential impact of tsetse invasion on the socioecological resilience of pastoral communities in the Karamoja sub-region. Results indicated that tsetse invasion is spreading from north to south of Karamoja. The tsetse transmission route emerging from southern Karamoja is perceived to be a continuation of the tsetse belt from West Pokot, Kenya. Cases of livestock deaths, livestock abortions, decreased milk yields, restricted access to prime grazing lands, heightened human-wildlife conflicts and disruption on crop cultivation have been reported. A computed socioecological resilience index in the study area was positive but low. Owing to the transboundary characteristics of tsetse invasions and sources and the associated documented effects, an urgent, strategic and system-wide intervention should be undertaken to control the tsetse invasion in this sub-region.

Tsetse Fly Transmission Studies of African Trypanosomes.

African trypanosomes are naturally transmitted by bloodsucking tsetse flies in sub-Saharan Africa and these transmission cycles can be reproduced in the laboratory if clean tsetse flies and suitable trypanosomes are available for experiments. Tsetse transmission gives access to more trypanosome developmental stages than are available from in vitro culture, albeit in very small numbers; for example, the sexual stages of Trypanosoma brucei have been isolated from infected tsetse salivary glands, but have not yet been reported from culture. Tsetse transmission also allows for the natural transition between different developmental stages to be studied.Both wild-type and genetically modified trypanosomes have been successfully fly transmitted, and it is possible to manipulate the trypanosome environment inside the fly to some extent, for example, the induction of expression of genes controlled by the Tet repressor by feeding flies with tetracycline.

English

Ethiopia is one of African country with nine regions and two city administration of which five regions were infected with more than one species of tsetse flies. Known species in Ethiopia are five in number namely Glossina pallidipes, G .morsitans, G. fuscipes, G. tachinoides and G. longipennis. Most tsetse transmission is cyclic and begins when blood from a trypanosome-infected animal is ingested by the fly. The clinical feature of the disease follows the level or burdens of tsetse challenge species. The main feature is anemia results in a progressive drop in packed cell volume, a non-specific but useful indicator in endemic areas. The most sensitive rapid method is examining a wet mount of the buffy coat area of a PCV tube after centrifugation, looking for motile parasites. The prevalence of trypanosomosis in enzootic area can be reduced by parasite control, vector control, host resistance protection prophylactic treatment and good husbandry management system. The methods of tsetse fly control involved bush clearing, elimination of game animals on which tsetse feed, and the sterile male technique (sterile insect techniques). Since female tsetse only mates once in a lifetime, this technique is theoretically able to eradicate a targeted tsetse species. Trypanotolerant animals are very important in tsetse fly challenging areas, but most countries did not accept them due to their low production of milk than indigenous breed. In conclusion, prevalence of trypanosomosis is devastating diseases of cattle in Ethiopia with both direct and indirect economic losses. Key words: Bovine, trypanosomosis, nagana, tsetse fly, protozoa.

First PATTEC consultative workshop on strategies, technical advances and partnerships in tsetse and trypanosomosis management

Tsetse and trypanosomiasis (T&T) remain endemic in many countries, although many efforts have been made in recent decades within the framework of the Pan-African Tsetse and Trypanosomiasis Eradication Campaign (PATTEC) Initiative, and despite progress observed both in research and development, and in the commitment of many partners to support control efforts. In order to create a functional multipartite platform to control T&T, a consultative workshop involving all stakeholders was held in 2014. In conclusion of the workshop, the competence and multidisciplinary of the various partners was acknowledged, as well as the need for progress in areas such as the revision of chemoprophylaxis and treatment strategies with regard animal trypanosomoses, the application of human African trypanosomiasis surveillance strategies, and the need to understand better the mechanism of resistance to trypanocides and the factors that maintain trypanosomiasis without tsetse transmission. The need to centralize the results of operational research to improve policy formulation and management techniques was also discussed. Geographic information systems were acknowledged as planning and management tools to control T&T. This first consultation workshop ended with a plea for better communication and a clear definition of roles as a guarantee of success of an effective partnership.

Trypanosome genetics: Populations, phenotypes and diversity

In the last decade, there has been a wide range of studies using a series of molecular markers to investigate the genotypic diversity of some of the important species of African trypanosomes. Here, we review this work and provide an update of our current understanding of the mechanisms that generate this diversity based on population genetic analysis. In parallel with field based studies, our knowledge of the key features of the system of genetic exchange in Trypanosoma brucei, based on laboratory analysis, has reached the point at which this system can be used as a tool to determine the genetic basis of a phenotype. In this context, we have outlined our current knowledge of the basis for phenotypic variation among strains of trypanosomes, and highlight that this is a relatively under researched area, except for work on drug resistance. There is clear evidence for ‘strain’-specific variation in tsetse transmission, a range of virulence/pathogenesis phenotypes and the ability to cross the blood brain barrier. The potential for using genetic analysis to dissect these phenotypes is illustrated by the recent work defining a locus determining organomegaly for T. brucei. When these results are considered in relation to the body of research on the variability of the host response to infection, it is clear that there is a need to integrate the study of host and parasite diversity in relation to understanding infection outcome.

Developmental regulation and extracellular release of a VSG expression-site-associated gene product from Trypanosoma brucei bloodstream forms

Trypanosomes evade host immunity by exchanging variant surface glycoprotein (VSG) coats. VSG genes are transcribed from telomeric expression sites, which contain a diverse family of expression-site-associated genes (ESAGs). We have discovered that the mRNAs for one ESAG family, ESAG9, are strongly developmentally regulated, being enriched in stumpy forms, a life-cycle stage in the mammalian bloodstream that is important for the maintenance of chronic parasite infections and for tsetse transmission. ESAG9 gene sequences are highly diverse in the genome and encode proteins with weak similarity to the massively diverse MASP proteins in Trypanosoma cruzi. We demonstrate that ESAG9 proteins are modified by N-glycosylation and can be shed to the external milieu, this being dependent upon coexpression with at least one other family member. The expression profile and extracellular release of ESAG9 proteins represents a novel and unexpected aspect of the transmission biology of trypanosomes in their mammalian host. We suggest that these molecules might interact with the external environment, with possible implications for infection chronicity or parasite transmission.

Transcriptome analysis of differentiating trypanosomes reveals the existence of multiple post-transcriptional regulons

BackgroundTrypanosome gene expression is regulated almost exclusively at the post-transcriptional level, with mRNA degradation playing a decisive role. When trypanosomes are transferred from the blood of a mammal to the midgut of a Tsetse fly, they transform to procyclic forms: gene expression is reprogrammed, changing the cell surface and switching the mode of energy metabolism. Within the blood, trypanosomes can pre-adapt for Tsetse transmission, becoming growth-arrested stumpy forms. We describe here the transitions in gene expression that occur during differentiation of in-vitro cultured bloodstream forms to procyclic forms.ResultsSome mRNAs showed changes within 30 min of cis-aconitate addition, whereas others responded 12-24 hours later. For the first 12 h after addition of cis-aconitate, cells accumulated at the G1 phase of the cell cycle, and showed decreases in mRNAs required for proliferation, mimicking the changes seen in stumpy forms: many mRNAs needed for ribosomal and flagellar biogenesis showed striking co-regulation. Other mRNAs encoding components of signal transduction pathways and potential regulators were specifically induced only during differentiation. Messenger RNAs encoding proteins required for individual metabolic pathways were often co-regulated.ConclusionTrypanosome genes form post-transcriptional regulons in which mRNAs with functions in particular pathways, or encoding components of protein complexes, show almost identical patterns of regulation.

Intraclonal mating occurs during tsetse transmission of Trypanosoma brucei

BackgroundMating in Trypanosoma brucei is a non-obligatory event, triggered by the co-occurrence of different strains in the salivary glands of the vector. Recombinants that result from intra- rather than interclonal mating have been detected, but only in crosses of two different trypanosome strains. This has led to the hypothesis that when trypanosomes recognize a different strain, they release a diffusible factor or pheromone that triggers mating in any cell in the vicinity whether it is of the same or a different strain. This idea assumes that the trypanosome can recognize self and non-self, although there is as yet no evidence for the existence of mating types in T. brucei.ResultsWe investigated intraclonal mating in T. b. brucei by crossing red and green fluorescent lines of a single strain, so that recombinant progeny can be detected in the fly by yellow fluorescence. For strain 1738, seven flies had both red and green trypanosomes in the salivary glands and, in three, yellow trypanosomes were also observed, although they could not be recovered for subsequent analysis. Nonetheless, both red and non-fluorescent clones from these flies had recombinant genotypes as judged by microsatellite and karyotype analyses, and some also had raised DNA contents, suggesting recombination or genome duplication. Strain J10 produced similar results indicative of intraclonal mating. In contrast, trypanosome clones recovered from other flies showed that genotypes can be transmitted with fidelity. When a yellow hybrid clone expressing both red and green fluorescent protein genes was transmitted, the salivary glands contained a mixture of fluorescent-coloured trypanosomes, but only yellow and red clones were recovered. While loss of the GFP gene in the red clones could have resulted from gene conversion, some of these clones showed loss of heterozygosity and raised DNA contents as in the other single strain transmissions. Our observations suggest that many recombinants are non-viable after intraclonal mating.ConclusionWe have demonstrated intraclonal mating during fly transmission of T. b. brucei, contrary to previous findings that recombination occurs only when another strain is present. It is thus no longer possible to assume that T. b. brucei remains genetically unaltered after fly transmission.

Unravelling the phylogenetic relationships of African trypanosomes of suids.

African trypanosomes of the subgenera Nannomonas and Pycnomonas have been recorded from both wild and domestic suids. However, complete descriptions of some of these trypanosomes with regard to host range, pathogenicity, transmission and distribution are still lacking. Neither the recently described Trypanosoma (Nannomonas) godfreyi nor Trypanosoma (Nannomonas) congolense Tsavo have been isolated from mammalian hosts, while Trypanosoma (Pycnomonas) suis remains the rarest of the Salivarian trypanosomes. The only isolate presumed to be of the latter species is maintained at the Kenya Trypanosomiasis Research Institute, Nairobi. We present here the results of characterization of this isolate by morphology, tsetse transmission, the use of species-specific DNA probes and DNA sequence analysis. Morphology in stained blood smears revealed a small trypanosome with a free flagellum. Experimental transmission through Glossina morsitans morsitans showed a developmental cycle typical of subgenus Nannomonas A positive identification was obtained with species-specific PCR primers for T. congolense Tsavo; moreover, the sequence of the SSU rRNA gene was almost identical to that of T. congolense Tsavo on database. In phylogenetic analysis of the SSU rRNA genes of Salivarian trypanosomes, T. congolense Tsavo grouped with T. simiae rather than T. congolense, suggesting that the name T. simiae Tsavo is more appropriate.

Self-fertilisation in Trypanosoma brucei

We have investigated whether Trypanosoma brucei can undergo self-fertilisation. A group of 27 metacyclic clones derived from the tsetse transmission of a mixture of two genetically marked stocks was analysed and 22 clones were observed to be of non-hybrid phenotype. A group of 10 clones from this non-hybrid subset were then analysed for one isoenzyme, one restriction fragment length polymorphism and three karyotype markers potentially informative for the detection of self-fertilisation. Five of the 10 clones were found to be recombinant for at least one marker and we interpret these recombination events as indicating the clones to be products of self-fertilisation. We have also analysed a limited number of metacyclic clones from stocks of T. brucei each singly transmitted through tsetse flies but, so far, no evidence of recombination has been detected. We conclude that T. brucei is able to self-fertilise but there may be a requirement for the presence of dissimilar stocks to initiate such an event.

Interference in the establishment of tsetsetransmitted Trypanosoma congolense, T. brucei or T. vivax superinfections in goats already infected with T. congolense or T. vivax

An interference phenomenon that delays superinfection with a trypanosome species different from that used for the initial infection has been found to occur in goats. Following tsetse transmission of Trypanosoma brucei to goats already infected with T. congolense, there was a delay in chancre development, as well as in the appearance of T. brucei and anti- T. brucei antibodies in the blood when compared to previously uninfected goats. However, there was no delay in the establishment of a tsetse-transmitted superinfection with T. vivax in goats already infected with either T. congolense or in animals already infected with a different serodeme of T. vivax.

Instability of the Trypanosoma brucei rhodesiense metacyclic variable antigen repertoire.

Trypanosoma brucei rhodesiense undergoes antigenic variation in its mammalian host by changing the glycoprotein composing its surface coat. Trypanosome clones which have the same repertoire of variable antigen types (VATs) are said to belong to the same serodeme. Tsetse flies infected with a particular serodeme extrude infective metacyclic trypanosomes which express only a restricted part of this repertoire. As the only known acquired immunity in African trypanosomiasis is VAT-specific this limitation of metacyclic VAT (M-VAT) repertoire could be important in devising a vaccine. This possibility of immunoprophylaxis could depend, however, on whether or not the M-VAT repertoire is conserved over long periods of repeated cyclical transmission and between epidemics. Studies reported here on isolates made from an East African focus of sleeping sickness over a 20-yr period suggest substantial changes in the M-VATs expressed during this time. Furthermore, we have detected change in expression of 3 M-VATs during sequential tsetse transmission of a clone in the laboratory indicating a possible instability in the organization of M-VAT genes.

Trypanosoma vivax: Sequence of antigenic variants in mice and goats

Three populations composed of different variable antigen types were derived from a clone of the recently isolated mouse-infective Trypanosoma vivax stock Zaria Y486. The antigenic composition of the trypanosome populations which appeared following infection of mice and goats with these populations was compared using the immune lysis test. Significant differences were observed in the antigenic composition of both the initial and relapse populations obtained from goats and mice. These observations suggest that there may be selective growth of different variable antigen types in different hosts with the result that the antigenic composition of a goat-derived population may be greatly altered following subinoculation into normal mice. This can be somewhat reduced however if goat derived populations are subinoculated into lethally irradiated mice. Two antigenically different T. vivax populations were also cyclically transmitted to individual goats in a preliminary study of the effect of tsetse transmission on the antigenic composition of the ingested populations. It was observed that the composition of the first detectable populations in the infected goats was similar but different from that ingested by the tsetse flies.