Trypanosomes are one-celled parasites that cause the disease trypanosomiasis, which is also known as sleeping sickness. Trypanosomiasis is transmitted to humans and animals by a type of fly, known as tse-tse, which is commonly found in sub-Saharan Africa. A bite from the tse-tse fly transfers the trypanosome cells into the host’s bloodstream, where they spread from the blood to the internal organs and brain. This leads to a long-term illness, which can sometimes result in a coma and eventually death.

Once in the blood trypanosomes move around using a structure similar to an underwater propeller called the flagellum. How the trypanosomes move and behave in the blood determines how the infection will progress. Until now it has only been possible to observe trypanosomes in plastic dishes or in blood drawn from infected patients. However, neither of these settings mimic the conditions of the bloodstream, and it is currently impossible to look inside human hosts to watch how trypanosomes move.

To overcome this hurdle, Doro et al. infected zebrafish with Trypanosoma carassii, a close relative of the sub-Saharan trypanosomes that specifically infects fish. Zebrafish are transparent when young, making it possible to observe the parasite in the blood and tissues of live fish using a microscope.

Doro et al. noticed that Trypanosoma carassii cells adapt to different environments in the host by using different swimming techniques. For example, in small capillaries trypanosomes were dragged along with the blood flow, whilst in larger vessels, when blood flow was slow or there were fewer red blood cells, trypanosomes actively swam against the current. The parasites were also able to change direction by using their flagella in a ‘whip-like’ motion. Lastly, it was discovered that Trypanosoma carassii could rapidly attach to blood vessel walls using one end of its cell body, even when blood flow was strong. This behaviour may help the parasites escape from the bloodstream into the surrounding tissues, making the infection more dangerous.

Studying how trypanosomes infect zebrafish at this high level of detail provides new insights into how these parasites move and behave inside a host. An important question that remains to be answered, is how exactly the trypanosomes leave the bloodstream. A better understanding of the whole infection process may hint at new ways of fighting these deadly infections in future.

Trypanosoma is a genus of flagellated protozoa, and most species proliferate in the blood and tissue fluids of their host. The best-known and most commonly studied trypanosomes are those that cause human diseases: Human African Trypanosomiasis caused by Trypanosoma brucei and American Trypanosomiasis caused by T. cruzi. However, trypanosomes infect animals from all vertebrate classes, including warm-blooded mammals and birds as well as cold-blooded amphibians, reptiles and fish (Simpson et al., 2006).

African trypanosomes, such as Trypanosoma brucei, are exclusively extracellular and are continuously exposed to the innate, humoral and cellular immune systems. T. brucei has become a textbook example of antigenic variation based on low frequency switches of the expressed variant surface glycoprotein (VSG) that dominates the cell surface and allows the population to escape the host adaptive immune system. In addition, individual cell survival is favoured by a process that cleans the cell surface of low levels of bound immunoglobulins through endocytosis and degradation (Engstler et al., 2007; Forlenza et al., 2009). Trypanosome motility is an integral part of this process as the hydrodynamic drag on antibody-VSG complexes, caused by the forward swimming motion of the cell, results in accumulation of the complexes at the posterior pole, close to the flagellar pocket, where endocytosis occurs (Engstler et al., 2007). In addition, motility is essential for successful cell division, immune evasion and development in the host (Broadhead et al., 2006; Griffiths et al., 2007; Ralston et al., 2006; Shimogawa et al., 2018).

In vitro studies using African trypanosomes have focused on the characterization of qualitative and quantitative parameters of trypanosome morphology and motility and emphasized the ability of trypanosome species to adapt to the various environments of their mammalian hosts (Alizadehrad et al., 2015; Bargul et al., 2016; Heddergott et al., 2012; Krüger and Engstler, 2015). Thus far, detailed analysis of swimming behaviour, morphology and trypanosome-host cell interaction has been restricted to ex vivo, using conditions that best mimic the host environment, for example in blood taken from naïve or infected animals (Bargul et al., 2016; Beattie and Gull, 1997; Engstler et al., 2007; Heddergott et al., 2012; Hemphill and Ross, 1995; Shimogawa et al., 2018; Skalický et al., 2017; Sunter and Gull, 2016; Wakid and Bates, 2004). It has not been feasible to recreate complex in vivo microenvironments, which, in the case of the vertebrate host, includes the streaming of the blood, vessels with heterogeneous size and endothelium composition, as well as changes occurring during the course of an infection, for example from the onset of anaemia. It still remains a challenge to mimic in vitro the dynamic conditions of the crowded and fast-moving bloodstream in which trypanosomes live. Given the importance of motility for trypanosome survival, analysis of trypanosome swimming behaviour in vivo, in a vertebrate host environment, is important to fully understand trypanosome biology and pathogenesis.

Quantitative and qualitative approaches in vivo are hindered by the lack of transparency of mammalian vertebrate hosts. Nevertheless, using transgenic trypanosomes expressing a luciferase reporter protein it has been possible to monitor overall trypanosome distribution and to quantify total trypanosome load in mice (Burrell-Saward et al., 2015; Capewell et al., 2016; Goyard et al., 2014; McLatchie et al., 2013). Fluorescent trypanosomes have been used in vivo to visualize their presence in mouse skin, a possible reservoir of trypanosomes in the late phase of infection (Capewell et al., 2016). Although the sensitivity of both systems allows for indirect visualization of trypanosome location or distribution, low spatio-temporal resolution and the lack of transparency of deep tissues limits the possibility for qualitative and quantitative analysis of trypanosome swimming behaviour in vivo. Finally, using a combination of multicolour light sheet fluorescence microscopy and high-speed fluorescence microscopy it has been possible to analyse the infection process and swimming behaviour of T. brucei ex vivo, in dissected tissues of the partially transparent tsetse fly vector (Gibson and Peacock, 2019; Schuster et al., 2017; Wang and Belosevic, 1994). Nevertheless, to date, no method is available to study with sufficient resolution trypanosome swimming behaviour in vivo in a vertebrate host environment.

In the current study, we have used Trypanosoma carassii infection of zebrafish, Danio rerio, to visualize trypanosome movement in the bloodstream and tissues of a vertebrate host. T. carassii infects a broad range of cyprinid fish (Overath et al., 1998), is transmitted by blood-sucking leeches (i.e., Hemiclepsis marginata) (Lom and Dyková, 1992), and lives extracellularly in the blood and tissue fluids of the fish (Haag et al., 1998; Lom and Dyková, 1992). T. carassii can establish a long-term infection characterized by polyclonal B cell activation and recurrent waves of parasitaemia, without expression of a uniform VSG-like surface coat (Agüero et al., 2002; Joerink et al., 2007; Overath et al., 2001; Overath et al., 1999). Phylogenetically, T. carassii belongs to the aquatic clade of Trypanosoma, a sister group that diverged from the Trypanosomatid lineage prior to the divergence of the Stercorarian and Salivarian trypanosomes that infect mammals (Simpson et al., 2006; Stevens, 2008). Morphologically, T. carassii isolated from fish has a single flagellum that emerges from the flagellar pocket at the cell posterior, is attached to the cell body and extends free at the anterior end, defining the anterior-posterior axis.

Zebrafish have been used as a model for developmental biology, as well as biomedical and neurobiology research (Asnani and Peterson, 2014; Blackburn and Langenau, 2014; Cronan and Tobin, 2014; Goessling and North, 2014; Miyares et al., 2014; Nguyen-Chi et al., 2014; Phillips and Westerfield, 2014; Renshaw and Trede, 2012; Schartl, 2014; Torraca et al., 2014; Veinotte et al., 2014; Zon and Cagan, 2014), and are a fresh water cyprinid fish closely related to many of the natural hosts of T. carassii. The great advantage of zebrafish is the transparency of the larvae, and of some juvenile and adult stages. Furthermore, mutant and transgenic lines including those marking blood vessels and relevant immune cell lineages are available (Benard et al., 2015; Bertrand et al., 2010; Ellett et al., 2011; Langenau et al., 2004; Lawson and Weinstein, 2002; Page et al., 2013; Petrie-Hanson et al., 2009; Renshaw et al., 2006).

The transparency of the zebrafish allows high-resolution, real-time imaging of trypanosome movement and host-parasite interaction in vivo, under conditions that exactly represent the natural environment during infection. In this study, by combining T. carassii infection of transparent zebrafish with high-speed microscopy, we provide the first description of trypanosome swimming behaviour in vivo in a vertebrate host. Our observations reveal that trypanosomes can rapidly adapt their swimming behaviour to the heterogeneous host environments. It was not possible to assign one preferred swimming behaviour to trypanosomes in either the bloodstream, tissues or other body fluids. Conditions such as the presence or absence of the blood flow or of red blood cells, the speed of the flow, the size of the blood vessel, the type of endothelium or epithelium lining the vessels or the tissues, as well as the compactness of the tissues, all influenced the swimming behaviour. Furthermore, we show that trypanosomes can change direction through backwards swimming and through a ‘whip-like’ motion. Finally, we were able to capture a novel mechanism through which trypanosomes attach to host cells or tissues. These observations greatly expand our knowledge on trypanosome swimming behaviour and show that trypanosomes can rapidly adapt to match the host environment.

In this study we describe a trypanosome infection model in zebrafish. By combining the transparency of zebrafish larvae with high spatio-temporal resolution microscopy, we were able to describe in detail the in vivo swimming behaviour of T. carassii in blood, tissues and tissue fluids of a vertebrate host. In addition to non-directional tumbling and directional forward swimming, we also describe how in vivo trypanosomes can reverse direction through a ‘whip-like’ motion or by swimming backwards. Finally, we report a novel observation that the posterior end of T. carassii, possibly coinciding with the flagellum base or flagellar pocket’s neck, can act as an anchoring site. To our knowledge, this is the first report of the swimming behaviour of trypanosomes in vivo in a vertebrate host environment.

Knowledge of trypanosome-host cell interaction, movement, and tropism in vertebrate hosts is important to understand trypanosome biology and pathology. To date, detailed analysis of trypanosome swimming behaviour and interaction with vertebrate host cells has only been possible using isolated blood or conditions that best mimic those of the host blood or tissues. Although the current in vitro approaches have brought a wealth of information on the quantitative aspects of trypanosome motility and potential immune evasion strategies, they could not fully reproduce the streaming nature of the blood, the heterogeneity of the bloodstream between and within vessels, the different sizes of blood vessels between which trypanosomes regularly alternate, the different types of endothelium lining arteries and veins and, finally, the changes that occur in the blood caused by the infection itself.

In this study, we first investigated the motion of T. carassii in infected carp blood. Based on previously proposed descriptions of the swimming behaviour of salivarian trypanosomes in mouse blood (Bargul et al., 2016; Shimogawa et al., 2018), clearly more than 90% of T. carassii could be classified as tumblers, and this was independent of whether they were in whole blood, diluted in serum or culture medium. Our observations are in agreement with a previous study reporting that directional persistent swimming was not a prominent feature of T. brucei in whole blood from an immunocompetent infected mouse (Shimogawa et al., 2018), but are in contrast to the study by Bargul and colleagues in which up to 30% persistent swimmers and 45% intermediate swimmers could be observed in blood films from immunosuppressed infected mice (Bargul et al., 2016). Differences in T. brucei strains, host immune status as well as the use of whole blood or blood films may account for the observed discrepancies. Nevertheless, both studies could not reproduce the streaming of the blood and, thus, the observations were made under static conditions.

Our initial in vitro observations led to the suggestion that the tumbling behaviour might be an intrinsic property of T. carassii in the bloodstream. However, our subsequent in vivo observations showed that this applies only to trypanosomes swimming in fluids with highly reduced or absent flow, for example the peritoneal fluid or blood in vessels in which the flow was slow or absent (Figure 7A and Video 9). In contrast, analysis of the swimming behaviour of T. carassii in the zebrafish bloodstream revealed that it is not possible to assign a single or predominant swimming behaviour. In vessels with a normal blood flow and in the presence of red blood cells, trypanosomes were dragged passively by the flow along with red blood cells. In the centre of large vessels, such as the cardinal artery or cardinal vein, the velocity of the flow does not allow either tumbling or directional swimming against or faster than the flow. Similarly, in narrow iISCs, trypanosomes are forced to move forward because of streaming of the blood or the presence of colliding red blood cells. Under these conditions, trypanosomes were never seen swimming faster than the passive movement of red blood cells as previously suggested (Heddergott et al., 2012; Langousis and Hill, 2014). Furthermore, because trypanosomes are carried along with red blood cells by the flow, it is difficult to envisage how the red blood cells could represent anything more than very occasional obstacles or surfaces for mechanical interactions that would favour forward swimming (Bargul et al., 2016; Heddergott et al., 2012). At the periphery of the cardinal vein, there is a cell-free layer where the number of red blood cells and the speed of the blood flow is reduced (Bagchi, 2007). Here, trypanosomes could clearly slow themselves down by crawling, rolling or by temporarily anchoring themselves to the dorsal endothelium (further discussed below). However, in arteries or small capillaries, adherence to the epithelium was never observed, even when the flow was reduced or completely absent. This suggests that not only the velocity of the flow but also the type of endothelium influences trypanosome swimming behaviour.

Physiological changes occur as the infection progresses that modify conditions within blood vessels. The flow is strongly reduced or interrupted and the density of red blood cells is reduced by obstruction of blood vessels or anaemia. In these conditions, directional swimmers were observed that swam faster than the flow and others that swam against the flow, or that repeatedly changed direction within blood vessels (Video 4). Taken together, our observations show that it is not possible to generalize the swimming behaviour of trypanosomes in blood as they can rapidly adopt different swimming behaviours: tumbling, directional swimming or anchoring. These behaviours are all largely influenced by the size and type of vessel, the speed of the flow, the presence, or not, of red blood cells as well as the highly dynamic microenvironment within a vessel, for example the centre compared to the periphery, and the type of endothelium.

Outside of the blood, it has been suggested that the directionality of swimming is influenced by the density or compactness of the tissue (Bargul et al., 2016; Sun et al., 2018; Wheeler et al., 2013). Our observation of swimming behaviour in various tissues revealed that it is the size of the interstitial space through which the trypanosome has to swim that largely determines whether it will swim directionally, tumble or both. For example, within the compact tissue of the fins, where the space below epithelial cells is very narrow (mesenchyme) and the basement membrane is not deformable (Mateus et al., 2012), the vast majority of trypanosomes swim directionally. This observation is in agreement with the swimming behaviour described for procyclic forms of T. brucei swimming through microfluidic devices smaller than their maximum cell diameter, mimicking potential size-limiting environments within host tissues (Sun et al., 2018). In zebrafish fins, the space can be so restrictive that to invert direction the trypanosome is obliged to swim backwards (Video 7). Backward swimming was previously observed in vitro or ex vivo for T. brucei motility mutants or for wild type trypanosomes swimming in high-density medium or in whole blood (Bargul et al., 2016; Baron et al., 2007; Branche et al., 2006; Engstler et al., 2007; Heddergott et al., 2012; Shimogawa et al., 2018), and ex vivo for procyclic and mesocyclic stages of T. brucei parasites swimming in confined spaces within the midgut of the tsetse fly (Schuster et al., 2017). The observation of T. carassii swimming in zebrafish tissues is the first report of backward swimming in a vertebrate host environment.

In the less compact areas of the fins or in the tissue lining the blood vessels, in addition to directional swimmers (forward or backwards), tumblers and trypanosomes that repeatedly inverted direction through a whip-like motion were observed (Video 8). When directional swimmers reached dead ends, the persistent forward swimming translated into either a drilling movement similar to the movement described for T. brucei in dead-end spaces within the midgut of the tsetse fly (Schuster et al., 2017), or backwards swimming as observed in the fins. Altogether, these observations indicate that trypanosomes can adopt several swimming behaviours and that these are largely influenced by the compactness and confinement offered by the tissue.

Perhaps one of the most interesting observations is the discovery that trypanosomes can anchor themselves to the vein endothelium by their posterior end, leaving the flagellum and the entire cell body free to move. Anchoring was observed as soon as 30 min to 1 h after T. carassii injection into zebrafish larvae, and by 1 dpi extravasation was observed. Under the current conditions, it was not possible to ascertain whether this adhesion mechanism favours or is even required for extravasation. So far, we have been unable to capture the exact moment of extravasation.

The anchoring site on the posterior of the trypanosome leaves the cell body and flagellum free to move (Video 2 and Video 5), so it seems likely that the adhesion on the trypanosome occurs via the flagellum base or the neck of the flagellar pocket itself. The presence of specific adhesion molecules would at least partly explain how T. carassii can very rapidly anchor themselves, upon sudden collision with the endothelium, and remain in position for more than 15 s despite the very rapid blood flow and collisions with blood cells. Anchoring was only observed in the cardinal vein but trypanosomes were seen crawling on vein endothelium (Video 5) through dynamic interactions that involved the cell membrane, not just the flagellum membrane, suggesting that the molecules required for whole-cell adherence might be different from those required for anchoring through the posterior end. This is reminiscent of leukocyte rolling, which also occurs only in veins and not in arteries. Altogether, the possibility to observe T. carassii behaviour both in vitro and in vivo, in the presence or absence of a strong hydrodynamic flow, and at various locations within the vertebrate host, demonstrated how the environmental conditions, especially the presence or absence of a flow, strongly influence the ability of the trypanosome to attach and the duration of the attachment.

T. carassii anchors to zebrafish cells in a manner clearly distinct from that described for other trypanosomes. The major difference being the lack of extensive interaction between the trypanosome’s flagellum membrane and the host cell or artificial surface. Stable interaction involving large portions of the flagellum membrane has been described among others for haptomonads stages (surface-attached) of Paratrypanosoma confusum, or Leishmania promastigotes (Figure 11A) (Skalický et al., 2017; Wakid and Bates, 2004). In these liberform parasites (flagellum not laterally attached to the cell), adhesion occurs through an attachment pad forming from the bulge at the base of the flagellum. At least in vitro, the formation of the pad takes approximately 1 h, causing extensive remodelling of the flagellum itself, and effectively anchors the parasites to the surface, and in the case of P. confusum, also favours their division (Skalický et al., 2017). Similarly, T. brucei epimastigotes divide while attached to the brush border of the salivary gland epithelium through extensive outgrowths of the non-cell-attached anterior part of the flagellum membrane (Figure 11B) (Beattie and Gull, 1997; Langousis and Hill, 2014; Schuster et al., 2017). Such an adhesion mechanism was only recently captured ex vivo through high-speed videography of dissected tsetse fly salivary glands; however, the exact moment of attachment and the time required to establish the stable interaction in vivo were not reported (Schuster et al., 2017). T. congolense was reported to adhere in vitro to bovine aorta endothelial (BAE) cell monolayers via extensive membrane protrusions (filopodia) of the membrane-attached flagellum (Figure 11C) (Beattie and Gull, 1997; Hemphill and Ross, 1995), an interaction that was shown to involve sialic acid residues on BAE cells (Hemphill et al., 1994). Although adhesion was observed already at 1 h, the filopodia increased in size over a period of 24–48 h. Whether such a type of interaction occurs with similar kinetics also in vivo, in vessels with an intact blood flow, is yet to be confirmed.

Figure 11

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All the above attachment mechanisms are clearly geared towards creating a very stable interaction with a surface to either establish a permanent infection in the salivary glands of the insect host, or to possibly adhere to the vertebrate host endothelium. They all involve extensive modifications of the flagellum membrane that occur over time to increase the contact area between the (para)trypanosomes and the surface. Most of these interactions, however, were observed in vitro for (para)trypanosomes cultivated on glass or plastic surfaces or endothelial cell monolayers and studied by means of scanning or transmission electron microscopy, as well as in vitro binding assays (Beattie and Gull, 1997; Hemphill and Ross, 1995; Hemphill et al., 1994; Skalický et al., 2017; Vickerman, 1969; Wakid and Bates, 2004). Therefore, to what extent the kinetics of interaction and the size of the contact area described also apply to the more dynamic in vivo environment is yet to be ascertained. The interactions between T. carassii and zebrafish cells were limited to the tip of the posterior end of the trypanosome body (Figure 11D), leaving the entire flagellum and trypanosome cell body free to rapidly move (Video 2). Despite the small surface involved, anchoring occurred very rapidly, suggesting a very strong, yet dynamic, type of interaction, the duration of which was influenced by the presence or absence of a strong hydrodynamic flow as well as colliding red blood cells. Given the in vivo dynamic conditions within a blood vessel, it seems unlikely that T. carassii would establish stable interactions that involve large portions of the trypanosome cell surface, as for example described in vitro for T. congolense. We were unable to determine whether the anchoring area of T. carassii corresponds to the flagellum base, flagellar pocket neck or to the cell body membrane. Nevertheless, because T. carassii adherence to red blood cells and to glass surfaces was also observed in vitro, it will be possible to investigate the adhesion mechanism at high resolution in the future.

In conclusion, we describe here for the first time the swimming behaviour of a trypanosome in vivo in the natural environment of a vertebrate host. We report the complex and heterogeneous environment in which the trypanosomes reside and how this highly influences the swimming behaviour. We describe how it is not possible to assign specific behaviours to trypanosomes swimming in any of the host compartments, as the trypanosomes were extremely effective in rapidly adapting their motion to the highly dynamic host environment. We describe backward swimming and whip-like movements that allowed trypanosomes to invert direction as well as identifiedying the posterior end as a novel anchoring site that allows the cell to adhere to host cells in a manner different from those described to date for other trypanosomes. Altogether, establishment of the T. carassii zebrafish infection model in combination with the genetic tractability of the zebrafish and of trypanosomes, represent a unique possibility to address questions related to: 1) trypanosome swimming behaviour in vivo in the natural environment of a vertebrate host, 2) host-pathogen interaction, 3) trypanosome biology, 4) the effect of specific immune factors on the progression of the infection, 5) the effect of drugs on both the trypanosome and the host, and 6) immune evasion strategies of trypanosomes that do not present antigenic variation. For all these reasons, the T. carassii-zebrafish model holds the promise to become a valuable complementary model to those currently available, to study the complex biology of trypanosomes and their interaction with the vertebrate host.

All data generated or analysed during this study are included in the manuscript. Source data files have been provided for Figures 1, 2 and 8.

1. 1. Agüero F
   2. Campo V
   3. Cremona L
   4. Jäger A
   5. Di Noia JM
   6. Overath P
   7. Sánchez DO
   8. Frasch AC

   (2002) Gene discovery in the freshwater fish parasite Trypanosoma carassii: identification of trans-sialidase-like and mucin-like genes

   Infection and Immunity 70:7140–7144.

   https://doi.org/10.1128/IAI.70.12.7140-7144.2002

   • PubMed
   • Google Scholar
2. 1. Alizadehrad D
   2. Krüger T
   3. Engstler M
   4. Stark H

   (2015) Simulating the complex cell design of Trypanosoma brucei and its motility

   PLOS Computational Biology 11:e1003967.

   https://doi.org/10.1371/journal.pcbi.1003967

   • PubMed
   • Google Scholar
3. 1. Asnani A
   2. Peterson RT

   (2014) The zebrafish as a tool to identify novel therapies for human cardiovascular disease

   Disease Models & Mechanisms 7:763–767.

   https://doi.org/10.1242/dmm.016170

   • PubMed
   • Google Scholar
4. 1. Bagchi P

   (2007) Mesoscale simulation of blood flow in small vessels

   Biophysical Journal 92:1858–1877.

   https://doi.org/10.1529/biophysj.106.095042

   • PubMed
   • Google Scholar
5. 1. Bargul JL
   2. Jung J
   3. McOdimba FA
   4. Omogo CO
   5. Adung'a VO
   6. Krüger T
   7. Masiga DK
   8. Engstler M

   (2016) Species-Specific adaptations of trypanosome morphology and motility to the mammalian host

   PLOS Pathogens 12:e1005448.

   https://doi.org/10.1371/journal.ppat.1005448

   • PubMed
   • Google Scholar
6. 1. Baron DM
   2. Kabututu ZP
   3. Hill KL

   (2007) Stuck in reverse: loss of LC1 in Trypanosoma brucei disrupts outer dynein arms and leads to reverse flagellar beat and backward movement

   Journal of Cell Science 120:1513–1520.

   https://doi.org/10.1242/jcs.004846

   • PubMed
   • Google Scholar
7. 1. Beattie P
   2. Gull K

   (1997) Cytoskeletal architecture and components involved in the attachment of Trypanosoma congolense epimastigotes

   Parasitology 115 ( Pt 1:47–55.

   https://doi.org/10.1017/s0031182097001042

   • PubMed
   • Google Scholar
8. 1. Benard EL
   2. Racz PI
   3. Rougeot J
   4. Nezhinsky AE
   5. Verbeek FJ
   6. Spaink HP
   7. Meijer AH

   (2015) Macrophage-expressed perforins mpeg1 and mpeg1.2 have an anti-bacterial function in zebrafish

   Journal of Innate Immunity 7:136–152.

   https://doi.org/10.1159/000366103

   • PubMed
   • Google Scholar
9. 1. Bertrand JY
   2. Chi NC
   3. Santoso B
   4. Teng S
   5. Stainier DY
   6. Traver D

   (2010) Haematopoietic stem cells derive directly from aortic endothelium during development

   Nature 464:108–111.

   https://doi.org/10.1038/nature08738

   • PubMed
   • Google Scholar
10. 1. Blackburn JS
    2. Langenau DM

    (2014) Zebrafish as a model to assess Cancer heterogeneity, progression and relapse

    Disease Models & Mechanisms 7:755–762.

    https://doi.org/10.1242/dmm.015842

    • PubMed
    • Google Scholar
11. 1. Branche C
    2. Kohl L
    3. Toutirais G
    4. Buisson J
    5. Cosson J
    6. Bastin P

    (2006) Conserved and specific functions of axoneme components in trypanosome motility

    Journal of Cell Science 119:3443–3455.

    https://doi.org/10.1242/jcs.03078

    • PubMed
    • Google Scholar
12. 1. Broadhead R
    2. Dawe HR
    3. Farr H
    4. Griffiths S
    5. Hart SR
    6. Portman N
    7. Shaw MK
    8. Ginger ML
    9. Gaskell SJ
    10. McKean PG
    11. Gull K

    (2006) Flagellar motility is required for the viability of the bloodstream trypanosome

    Nature 440:224–227.

    https://doi.org/10.1038/nature04541

    • PubMed
    • Google Scholar
13. 1. Burrell-Saward H
    2. Rodgers J
    3. Bradley B
    4. Croft SL
    5. Ward TH

    (2015) A sensitive and reproducible in vivo imaging mouse model for evaluation of drugs against late-stage human african trypanosomiasis

    Journal of Antimicrobial Chemotherapy 70:510–517.

    https://doi.org/10.1093/jac/dku393

    • PubMed
    • Google Scholar
14. 1. Capewell P
    2. Cren-Travaillé C
    3. Marchesi F
    4. Johnston P
    5. Clucas C
    6. Benson RA
    7. Gorman TA
    8. Calvo-Alvarez E
    9. Crouzols A
    10. Jouvion G
    11. Jamonneau V
    12. Weir W
    13. Stevenson ML
    14. O'Neill K
    15. Cooper A
    16. Swar NK
    17. Bucheton B
    18. Ngoyi DM
    19. Garside P
    20. Rotureau B
    21. MacLeod A

    (2016) The skin is a significant but overlooked anatomical reservoir for vector-borne african trypanosomes

    eLife 5:e17716.

    https://doi.org/10.7554/eLife.17716

    • PubMed
    • Google Scholar
15. 1. Cronan MR
    2. Tobin DM

    (2014) Fit for consumption: zebrafish as a model for tuberculosis

    Disease Models & Mechanisms 7:777–784.

    https://doi.org/10.1242/dmm.016089

    • PubMed
    • Google Scholar
16. 1. Ellett F
    2. Pase L
    3. Hayman JW
    4. Andrianopoulos A
    5. Lieschke GJ

    (2011) mpeg1 promoter transgenes direct macrophage-lineage expression in zebrafish

    Blood 117:e49–e56.

    https://doi.org/10.1182/blood-2010-10-314120

    • PubMed
    • Google Scholar
17. 1. Engstler M
    2. Pfohl T
    3. Herminghaus S
    4. Boshart M
    5. Wiegertjes G
    6. Heddergott N
    7. Overath P

    (2007) Hydrodynamic flow-mediated protein sorting on the cell surface of trypanosomes

    Cell 131:505–515.

    https://doi.org/10.1016/j.cell.2007.08.046

    • PubMed
    • Google Scholar
18. 1. Forlenza M
    2. de Carvalho Dias JD
    3. Veselý T
    4. Pokorová D
    5. Savelkoul HF
    6. Wiegertjes GF

    (2008) Transcription of signal-3 cytokines, IL-12 and IFN alpha beta, coincides with the timing of CD8 alpha beta up-regulation during viral infection of common carp (Cyprinus carpio L)

    Molecular Immunology 45:1531–1547.

    https://doi.org/10.1016/j.molimm.2007.10.010

    • PubMed
    • Google Scholar
19. 1. Forlenza M
    2. Nakao M
    3. Wibowo I
    4. Joerink M
    5. Arts JA
    6. Savelkoul HF
    7. Wiegertjes GF

    (2009) Nitric oxide hinders antibody clearance from the surface of Trypanoplasma borreli and increases susceptibility to complement-mediated lysis

    Molecular Immunology 46:3188–3197.

    https://doi.org/10.1016/j.molimm.2009.08.011

    • PubMed
    • Google Scholar
20. 1. Forlenza M
    2. Kaiser T
    3. Savelkoul HFJ
    4. Wiegertjes GF

    (2012) The use of real-time quantitative PCR for the analysis of cytokine mRNA levels

    Cytokine Protocols 23:7–23.

    https://doi.org/10.1007/978-1-61779-439-1_2

    • Google Scholar
21. 1. Gibson W
    2. Peacock L

    (2019) Fluorescent proteins reveal what trypanosomes get up to inside the tsetse fly

    Parasites & Vectors 12:6.

    https://doi.org/10.1186/s13071-018-3204-y

    • PubMed
    • Google Scholar
22. 1. Goessling W
    2. North TE

    (2014) Repairing quite swimmingly: advances in regenerative medicine using zebrafish

    Disease Models & Mechanisms 7:769–776.

    https://doi.org/10.1242/dmm.016352

    • PubMed
    • Google Scholar
23. 1. Goyard S
    2. Dutra PL
    3. Deolindo P
    4. Autheman D
    5. D'Archivio S
    6. Minoprio P

    (2014) In vivo imaging of trypanosomes for a better assessment of host-parasite relationships and drug efficacy

    Parasitology International 63:260–268.

    https://doi.org/10.1016/j.parint.2013.07.011

    • PubMed
    • Google Scholar
24. 1. Griffiths S
    2. Portman N
    3. Taylor PR
    4. Gordon S
    5. Ginger ML
    6. Gull K

    (2007) RNA interference mutant induction in vivo demonstrates the essential nature of trypanosome flagellar function during mammalian infection

    Eukaryotic Cell 6:1248–1250.

    https://doi.org/10.1128/EC.00110-07

    • PubMed
    • Google Scholar
25. 1. Haag J
    2. O'hUigin C
    3. Overath P

    (1998) The molecular phylogeny of trypanosomes: evidence for an early divergence of the salivaria

    Molecular and Biochemical Parasitology 91:37–49.

    https://doi.org/10.1016/S0166-6851(97)00185-0

    • PubMed
    • Google Scholar
26. 1. Heddergott N
    2. Krüger T
    3. Babu SB
    4. Wei A
    5. Stellamanns E
    6. Uppaluri S
    7. Pfohl T
    8. Stark H
    9. Engstler M

    (2012) Trypanosome motion represents an adaptation to the crowded environment of the vertebrate bloodstream

    PLOS Pathogens 8:e1003023.

    https://doi.org/10.1371/journal.ppat.1003023

    • Google Scholar
27. 1. Hemphill A
    2. Frame I
    3. Ross CA

    (1994) The interaction of Trypanosoma congolense with endothelial cells

    Parasitology 109 ( Pt 5:631–641.

    https://doi.org/10.1017/s0031182000076514

    • PubMed
    • Google Scholar
28. 1. Hemphill A
    2. Ross CA

    (1995) Flagellum-mediated adhesion of Trypanosoma congolense to bovine aorta endothelial cells

    Parasitology Research 81:412–420.

    https://doi.org/10.1007/BF00931503

    • PubMed
    • Google Scholar
29. 1. Irnazarow I

    (1995) Genetic variability of polish and hungarian carp lines

    Aquaculture 129:215.

    https://doi.org/10.1016/0044-8486(95)91961-T

    • Google Scholar
30. 1. Joerink M
    2. Groeneveld A
    3. Ducro B
    4. Savelkoul HF
    5. Wiegertjes GF

    (2007) Mixed infection with Trypanoplasma borreli and Trypanosoma carassii induces protection: involvement of cross-reactive antibodies

    Developmental & Comparative Immunology 31:903–915.

    https://doi.org/10.1016/j.dci.2006.12.003

    • PubMed
    • Google Scholar
31. 1. Krüger T
    2. Engstler M

    (2015) Flagellar motility in eukaryotic human parasites

    Seminars in Cell & Developmental Biology 46:113–127.

    https://doi.org/10.1016/j.semcdb.2015.10.034

    • PubMed
    • Google Scholar
32. 1. Langenau DM
    2. Ferrando AA
    3. Traver D
    4. Kutok JL
    5. Hezel J-PD
    6. Kanki JP
    7. Zon LI
    8. Look AT
    9. Trede NS
    10. Alt FW
    11. Travert D
    12. Zont L

    (2004) In vivo tracking of T cell development, ablation, and engraftment in transgenic in vivo tracking of T cell development, ablation, and engraftment in transgenic zebrafish

    PNAS 101:7369–7374.

    https://doi.org/10.1073/pnas.0402248101

    • Google Scholar
33. 1. Langousis G
    2. Hill KL

    (2014) Motility and more: the flagellum of Trypanosoma brucei

    Nature Reviews Microbiology 12:505–518.

    https://doi.org/10.1038/nrmicro3274

    • PubMed
    • Google Scholar
34. 1. Lawson ND
    2. Weinstein BM

    (2002) In vivo imaging of embryonic vascular development using transgenic zebrafish

    Developmental Biology 248:307–318.

    https://doi.org/10.1006/dbio.2002.0711

    • PubMed
    • Google Scholar
35. 1. Lom J
    2. Dyková I

    (1992)

    Developments in Aquaculture and Fisheries Science

    Protozoan Parasites of Fishes, Developments in Aquaculture and Fisheries Science, Amsterdam, Elsevier.

    • Google Scholar
36. 1. Mateus R
    2. Pereira T
    3. Sousa S
    4. de Lima JE
    5. Pascoal S
    6. Saúde L
    7. Jacinto A

    (2012) In vivo cell and tissue dynamics underlying zebrafish fin fold regeneration

    PLOS ONE 7:e51766.

    https://doi.org/10.1371/journal.pone.0051766

    • PubMed
    • Google Scholar
37. 1. McLatchie AP
    2. Burrell-Saward H
    3. Myburgh E
    4. Lewis MD
    5. Ward TH
    6. Mottram JC
    7. Croft SL
    8. Kelly JM
    9. Taylor MC

    (2013) Highly sensitive in vivo imaging of Trypanosoma brucei expressing "red-shifted" luciferase

    PLOS Neglected Tropical Diseases 7:e2571.

    https://doi.org/10.1371/journal.pntd.0002571

    • PubMed
    • Google Scholar
38. 1. Miyares RL
    2. de Rezende VB
    3. Farber SA

    (2014) Zebrafish yolk lipid processing: a tractable tool for the study of vertebrate lipid transport and metabolism

    Disease Models & Mechanisms 7:915–927.

    https://doi.org/10.1242/dmm.015800

    • PubMed
    • Google Scholar
39. 1. Nguyen-Chi M
    2. Phan QT
    3. Gonzalez C
    4. Dubremetz JF
    5. Levraud JP
    6. Lutfalla G

    (2014) Transient infection of the zebrafish notochord with E. coli induces chronic inflammation

    Disease Models & Mechanisms 7:871–882.

    https://doi.org/10.1242/dmm.014498

    • PubMed
    • Google Scholar
40. 1. Overath P
    2. Ruoff J
    3. Stierhof YD
    4. Haag J
    5. Tichy H
    6. Dyková I
    7. Lom J

    (1998) Cultivation of bloodstream forms of Trypanosoma carassii, a common parasite of freshwater fish

    Parasitology Research 84:343–347.

    https://doi.org/10.1007/s004360050408

    • PubMed
    • Google Scholar
41. 1. Overath P
    2. Haag J
    3. Mameza MG
    4. Lischke A

    (1999) Freshwater fish trypanosomes: definition of two types, host control by antibodies and lack of antigenic variation

    Parasitology 119 ( Pt 6:591–601.

    https://doi.org/10.1017/S0031182099005089

    • PubMed
    • Google Scholar
42. 1. Overath P
    2. Haag J
    3. Lischke A
    4. O'hUigin C

    (2001) The surface structure of trypanosomes in relation to their molecular phylogeny

    International Journal for Parasitology 31:468–471.

    https://doi.org/10.1016/S0020-7519(01)00152-7

    • PubMed
    • Google Scholar
43. 1. Page DM
    2. Wittamer V
    3. Bertrand JY
    4. Lewis KL
    5. Pratt DN
    6. Delgado N
    7. Schale SE
    8. McGue C
    9. Jacobsen BH
    10. Doty A
    11. Pao Y
    12. Yang H
    13. Chi NC
    14. Magor BG
    15. Traver D

    (2013) An evolutionarily conserved program of B-cell development and activation in zebrafish

    Blood 122:e1–e11.

    https://doi.org/10.1182/blood-2012-12-471029

    • Google Scholar
44. 1. Petrie-Hanson L
    2. Hohn C
    3. Hanson L

    (2009) Characterization of rag1 mutant zebrafish leukocytes

    BMC Immunology 10:8.

    https://doi.org/10.1186/1471-2172-10-8

    • Google Scholar
45. 1. Phillips JB
    2. Westerfield M

    (2014) Zebrafish models in translational research: tipping the scales toward advancements in human health

    Disease Models & Mechanisms 7:739–743.

    https://doi.org/10.1242/dmm.015545

    • Google Scholar
46. 1. Ralston KS
    2. Lerner AG
    3. Diener DR
    4. Hill KL

    (2006) Flagellar motility contributes to cytokinesis in Trypanosoma brucei and is modulated by an evolutionarily conserved dynein regulatory system

    Eukaryotic Cell 5:696–711.

    https://doi.org/10.1128/EC.5.4.696-711.2006

    • PubMed
    • Google Scholar
47. 1. Renshaw SA
    2. Loynes CA
    3. Trushell DMI
    4. Elworthy S
    5. Ingham PW
    6. Whyte MKB

    (2006) A transgenic zebrafish model of neutrophilic inflammation

    Blood 108:3976–3978.

    https://doi.org/10.1182/blood-2006-05-024075

    • Google Scholar
48. 1. Renshaw SA
    2. Trede NS

    (2012) A model 450 million years in the making: zebrafish and vertebrate immunity

    Disease Models & Mechanisms 5:38–47.

    https://doi.org/10.1242/dmm.007138

    • Google Scholar
49. 1. Schartl M

    (2014) Beyond the zebrafish: diverse fish species for modeling human disease

    Disease Models & Mechanisms 7:181–192.

    https://doi.org/10.1242/dmm.012245

    • Google Scholar
50. 1. Schuster S
    2. Krüger T
    3. Subota I
    4. Thusek S
    5. Rotureau B
    6. Beilhack A
    7. Engstler M

    (2017) Developmental adaptations of trypanosome motility to the tsetse fly host environments unravel a multifaceted in vivo microswimmer system

    eLife 6:e27656.

    https://doi.org/10.7554/eLife.27656

    • PubMed
    • Google Scholar
51. 1. Shimogawa MM
    2. Ray SS
    3. Kisalu N
    4. Zhang Y
    5. Geng Q
    6. Ozcan A
    7. Hill KL

    (2018) Parasite motility is critical for virulence of African trypanosomes

    Scientific Reports 8:9122.

    https://doi.org/10.1038/s41598-018-27228-0

    • Google Scholar
52. 1. Simpson AGB
    2. Stevens JR
    3. Lukeš J

    (2006) The evolution and diversity of kinetoplastid flagellates

    Trends in Parasitology 22:168–174.

    https://doi.org/10.1016/j.pt.2006.02.006

    • Google Scholar
53. 1. Skalický T
    2. Dobáková E
    3. Wheeler RJ
    4. Tesařová M
    5. Flegontov P
    6. Jirsová D
    7. Votýpka J
    8. Yurchenko V
    9. Ayala FJ
    10. Lukeš J

    (2017) Extensive flagellar remodeling during the complex life cycle of Paratrypanosoma , an early-branching trypanosomatid

    PNAS 114:11757–11762.

    https://doi.org/10.1073/pnas.1712311114

    • Google Scholar
54. 1. Stevens JR

    (2008) Kinetoplastid phylogenetics, with special reference to the evolution of parasitic trypanosomes

    Parasite 15:226–232.

    https://doi.org/10.1051/parasite/2008153226

    • Google Scholar
55. 1. Sun SY
    2. Kaelber JT
    3. Chen M
    4. Dong X
    5. Nematbakhsh Y
    6. Shi J
    7. Dougherty M
    8. Lim CT
    9. Schmid MF
    10. Chiu W
    11. He CY
    12. Cy H

    (2018) Flagellum couples cell shape to motility in Trypanosoma brucei

    PNAS 115:E5916–E5925.

    https://doi.org/10.1073/pnas.1722618115

    • Google Scholar
56. 1. Sunter JD
    2. Gull K

    (2016) The Flagellum Attachment Zone: ‘The Cellular Ruler’ of Trypanosome Morphology

    Trends in Parasitology 32:309–324.

    https://doi.org/10.1016/j.pt.2015.12.010

    • Google Scholar
57. 1. Torraca V
    2. Masud S
    3. Spaink HP
    4. Meijer AH

    (2014) Macrophage-pathogen interactions in infectious diseases: new therapeutic insights from the zebrafish host model

    Disease Models & Mechanisms 7:785–797.

    https://doi.org/10.1242/dmm.015594

    • Google Scholar
58. 1. Veinotte CJ
    2. Dellaire G
    3. Berman JN

    (2014) Hooking the big one: the potential of zebrafish xenotransplantation to reform cancer drug screening in the genomic era

    Disease Models & Mechanisms 7:745–754.

    https://doi.org/10.1242/dmm.015784

    • Google Scholar
59. 1. Vickerman K

    (1969)

    On the surface coat and flagellar adhesion in trypanosomes

    Journal of Cell Science 5:163.

    • PubMed
    • Google Scholar
60. Book

    1. Vickerman K
    2. Tetley L

    (1990) Flagellar Surfaces of Parasitic Protozoa and Their Role in Attachment

    In: Bloodgood R. A, editors. Ciliary and Flagellar Membranes. Boston: Springer US. pp. 267–304.

    https://doi.org/10.1007/978-1-4613-0515-6_11

    • Google Scholar
61. 1. Wakid MH
    2. Bates PA

    (2004) Flagellar attachment of Leishmania promastigotes to plastic film in vitro

    Experimental Parasitology 106:173–178.

    https://doi.org/10.1016/j.exppara.2004.03.001

    • Google Scholar
62. 1. Wang R
    2. Belosevic M

    (1994) Cultivation of Trypanosoma danilewskyi (Laveran & Mesnil, 1904) in serum-free medium and assessment of the course of infection in goldfish, Carassius auratus (L.)

    Journal of Fish Diseases 17:47–56.

    https://doi.org/10.1111/j.1365-2761.1994.tb00344.x

    • Google Scholar
63. 1. Wheeler RJ
    2. Gluenz E
    3. Gull K

    (2013) The Limits on Trypanosomatid Morphological Diversity

    PLOS ONE 8:e79581.

    https://doi.org/10.1371/journal.pone.0079581

    • Google Scholar
64. 1. White RM
    2. Sessa A
    3. Burke C
    4. Bowman T
    5. LeBlanc J
    6. Ceol C
    7. Bourque C
    8. Dovey M
    9. Goessling W
    10. Burns CE
    11. Zon LI

    (2008) Transparent Adult Zebrafish as a Tool for In Vivo Transplantation Analysis

    Cell Stem Cell 2:183–189.

    https://doi.org/10.1016/j.stem.2007.11.002

    • Google Scholar
65. 1. Zon L
    2. Cagan R

    (2014) From fish tank to bedside in ancer therapy: an interview with eonard Zon

    Disease Models & Mechanisms 7:735–738.

    https://doi.org/10.1242/dmm.016642

    • PubMed
    • Google Scholar

Author details

1. Éva Dóró

   Department of Animal Sciences, Cell Biology and Immunology Group, Wageningen University & Research, Wageningen, Netherlands

   Present address

   Institute of Transdisciplinary Discoveries, Medical School, Faculty of Medicine, University of Pécs, Pécs, Hungary

   Contribution

   Conceptualization, Data curation, Formal analysis, Supervision, Funding acquisition, Methodology, Writing—original draft, Project administration, Writing—review and editing

   Competing interests

   No competing interests declared

2. Sem H Jacobs

   Department of Animal Sciences, Cell Biology and Immunology Group, Wageningen University & Research, Wageningen, Netherlands

   Contribution

   Conceptualization, Data curation, Formal analysis, Methodology, Writing—review and editing

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0001-7482-3438

3. Ffion R Hammond

   Department of Animal Sciences, Cell Biology and Immunology Group, Wageningen University & Research, Wageningen, Netherlands

   Present address

   Department of Infection, Immunity and Cardiovascular Disease, University of Sheffield, Sheffield, United Kingdom

   Contribution

   Conceptualization, Data curation, Formal analysis, Methodology, Writing—original draft, Writing—review and editing

   Competing interests

   No competing interests declared

4. Henk Schipper

   Department of Animal Sciences, Experimental Zoology Group, Wageningen University & Research, Wageningen, Netherlands

   Contribution

   Data curation, Software, Formal analysis, Methodology, Writing—review and editing

   Competing interests

   No competing interests declared

5. Remco PM Pieters

   Department of Animal Sciences, Experimental Zoology Group, Wageningen University & Research, Wageningen, Netherlands

   Contribution

   Conceptualization, Data curation, Software, Formal analysis, Methodology, Writing—review and editing

   Competing interests

   No competing interests declared

6. Mark Carrington

   Department of Biochemistry, University of Cambridge, Cambridge, United Kingdom

   Contribution

   Conceptualization, Software, Methodology, Writing—review and editing

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0002-6435-7266

7. Geert F Wiegertjes

   1. Department of Animal Sciences, Cell Biology and Immunology Group, Wageningen University & Research, Wageningen, Netherlands
   2. Department of Animal Sciences, Aquaculture and Fisheries Group, Wageningen University & Research, Wageningen, Netherlands

   Contribution

   Conceptualization, Software, Supervision, Funding acquisition, Methodology, Writing—original draft, Project administration, Writing—review and editing

   Competing interests

   No competing interests declared

8. Maria Forlenza

   Department of Animal Sciences, Cell Biology and Immunology Group, Wageningen University & Research, Wageningen, Netherlands

   Contribution

   Conceptualization, Data curation, Formal analysis, Supervision, Funding acquisition, Methodology, Writing—original draft, Project administration, Writing—review and editing

   For correspondence

   maria.forlenza@wur.nl

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0001-9026-7320

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