Trypanosoma brucei is a flagellated parasite responsible for sleeping sickness in central Africa. It is transmitted from one host to another via the bite of the tsetse fly (Glossina genus), which is the major mode of contamination. The tsetse fly (Fig. 1), both male and female, feeds exclusively on blood and exhibits unique features such as viviparous reproduction and lactation. It should be considered as a real host for trypanosomes, since the parasites differentiate and proliferate in several fly tissues such as the gut, the proventriculus, and the salivary glands. Trypanosomes adapt to these various environments by modifications in their cell surface composition, metabolism, and morphogenesis. One of the most prominent characteristics of trypanosomes is the presence of a flagellum that is attached to the cell body and drives parasite movement, but also dictates cell morphology. Its length varies according to the life cycle stage, ranging from 3 to 30μm. This organelle is essential for parasite survival. Here, we will discuss the different aspects of trypanosome development in the tsetse fly, with focus on flagellum function.

Human African Trypanosomiasis is a disease characterised by two phases, one with rather unspecific symptoms such as bouts of fever, headaches, weakness or lymphadenopathy, and a second one with severe neurological symptoms including disruption of the sleeping pattern, hence the historical name of the disease, sleeping sickness. It is due to a parasitic protist called Trypanosoma brucei, with only two subspecies being responsible for human infections: T. b. gambiense (98% of the cases) and T. b. rhodesiense. Under the microscope, this extracellular parasite exhibits a long motile flagellum that is attached to the cell body. It performs multiple functions such as motility, morphogenesis and adhesion to specific tissues. The parasite possesses a typical eukaryotic nucleus (35 Mo) and a single mitochondrion whose large genome is condensed in a structure called the kinetoplast. The parasite escapes the host immune response thanks to a sophisticated process of antigenic variation. The disease is exclusively encountered in sub-Saharan Africa because the parasite is transmitted by the bite of the tsetse fly (Fig. 1), an insect that is not found outside this region.

Glossina flies have unique biological peculiarities. In contrast to most insects, they are not oviparous. Mating occurs from five days after birth and larval development takes place into the female uterus. After 14 days, the female lays a single pupa that is lightly buried in the ground. Development continues for 28 days before hatching and emergence, which is completed in only a few minutes. Like other insects, the tsetse fly is often termed a vector, but it should be considered as real host for at least two reasons. First, trypanosomes undergo a long trip in the tsetse fly, developing in a very precise order in different tissues with a lot of specific morphological, genetic, and biochemical adaptations. Second, trypanosomes are likely to have been parasites of insects well before affecting other organisms. The term of vector is probably more appropriate in the case of mechanical transfer of pathogens from one host to the other, as observed with the bite of tabanids and the transmission of Trypanosoma evansi, for example. This parasite is lacking several mitochondrial genes normally encoded on kinetoplast DNA and is unable to develop in the tsetse fly, yet can be passively transferred from one host to the other. Finally, it should be pointed out here that, although the presence of trypanosomes can affect the tsetse fly, the insect itself does not suffer from sleeping sickness. Therefore, it is preferable to say that the tsetse fly transmits trypanosomes, and not the disease. In the laboratory, tsetse flies are bred in netted cages and fed with heated sheep blood using a membrane to mimic the presence of the skin. Flies do not feed directly on blood.

Trypanosomes are ingested with blood when the fly feeds on an infected host and reach the crop where they have to resist dehydration. Next, the “paste” containing blood and trypanosomes is moved to the gut. The parasites differentiate to adapt to this new environment, where they have to face digestive enzymes and exhibit new molecular, biochemical, and metabolic features. They proliferate in the posterior midgut and then need to migrate to the proventriculus. This is a challenging trip for a trypanosome. If we compare with humans, it would be equivalent to swimming seven kilometres in a highly viscous environment like jelly… This is achieved by active forward motility, with the beating flagellum driving migration [1Rotureau B., Ooi C.P., Huet D., Perrot S., Bastin P. Forward motility is essential for trypanosome infection in the tsetse fly Cell. Microbiol. 2014 ;  16 : 425-433 [cross-ref]

Cliquez ici pour aller à la section Références]. The parasite next moves to the salivary glands, a process also likely to be governed by the flagellum. However, this could not be experimentally proven given the failure of motility mutants to reach the proventriculus. None of these parasite stages are able to infect mammals. Once trypanosomes arrive in the salivary glands, they therefore need to differentiate in an infective stage. For this, they attach to the epithelium of the salivary glands before starting the differentiation process. This prevents them from being released prematurely with the saliva. This adhesion takes place via the flagellum, which develops unique and complex membrane extensions that wrap around microvilli of the fly epithelial cells. The parasite then undergoes an asymmetric division, with the daughter cell inheriting the old flagellum, which remains attached to the epithelium and the other daughter inheriting the new flagellum, which adopts different morphology and biochemical composition [2Rotureau B., Subota I., Buisson J., Bastin P. A new asymmetric division contributes to the continuous production of infective trypanosomes in the tsetse fly Development 2012 ;  139 : 1842-1850 [cross-ref]

Cliquez ici pour aller à la section Références]. This latter cell differentiates to the infectious stage and is released in the saliva. It is now ready to infect a new host.

After landing on skin, the tsetse fly does not find immediately blood vessels for feeding. Rather, it probes by scratching and dilacerating tissues. During this process, the fly injects saliva and, if it contains trypanosomes, these are delivered in the skin. It was long accepted that trypanosomes rapidly move to the lymph, then to the blood, where they would spend most of their time before reaching the nervous system. However, recent development in imaging techniques allowed researchers to visualise parasite progression in mouse models. Strikingly, trypanosomes are mostly present in the skin, including in cases where they were not detected in the blood. They are highly motile, possibly to escape phagocytic cells. Moreover, skin trypanosomes have been shown to infect naïve tsetse flies, demonstrating their role in parasite transmission [3Capewell P., Cren-Travaille C., Marchesi F., Johnston P., Clucas C., Benson R.A., et al. The skin is a significant but overlooked anatomical reservoir for vector-borne African trypanosomes Elife 2016 ;  5 :

Cliquez ici pour aller à la section Références, 4Caljon G., Van Reet N., De Trez C., Vermeersch M., Perez-Morga D., Van Den Abbeele J. The Dermis as a Delivery Site of Trypanosoma brucei for Tsetse Flies PLoS. Pathog. 2016 ;  12 : e1005744

Cliquez ici pour aller à la section Références]. Analysis of skin biopsies from human patients coming from endemic areas for human African trypanosomiasis revealed the presence of trypanosomes, suggesting that the same phenomenon could occur in humans [3Capewell P., Cren-Travaille C., Marchesi F., Johnston P., Clucas C., Benson R.A., et al. The skin is a significant but overlooked anatomical reservoir for vector-borne African trypanosomes Elife 2016 ;  5 :

Cliquez ici pour aller à la section Références]. This is an amazing finding that could change the paradigm of African trypanosomiasis given the fact that diagnosis is based on the detection of parasites in the blood. This means that trypanosome carriers might go undetected and could contribute to parasite transmission. This could be highly significant in the current context of HAT elimination [5Informal Expert Group on Gambiense H.A.T.R., Buscher P., Bart J.M., Boelaert M., Bucheton B., Cecchi G., et al. Do Cryptic Reservoirs Threaten Gambiense-Sleeping Sickness Elimination? Trends. Parasitol. 2018 ;  34 : 197-207

Cliquez ici pour aller à la section Références].

Trypanosomes show remarkable adaptations to live within different tissues of the tsetse fly with the final aim to ensure their transmission to mammalian hosts. The flagellum is a key organelle to this success, as it drives motility but also controls cell morphogenesis, which is suited to each particular tissue where the parasite develops. Cilia and flagella are known to function as sensory organelles in other organisms and it would not be surprising to see it exploited by trypanosomes to detect their environment [6Shaw S., DeMarco S.F., Rehmann R., Wenzler T., Florini F., Roditi I., et al. Flagellar cAMP signaling controls trypanosome progression through host tissues Nat. Commun. 2019 ;  10 : 803

Cliquez ici pour aller à la section Références].