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• W2029131750 abstract "The malaria sporozoite, the parasite stage transmitted by the mosquito, is delivered into the dermis and differentiates in the liver. Motile sporozoites can invade host cells by disrupting their plasma membrane and migrating through them (termed cell traversal), or by forming a parasite-cell junction and settling inside an intracellular vacuole (termed cell infection). Traversal of liver cells, observed for sporozoites in vivo, is thought to activate the sporozoite for infection of a final hepatocyte. Here, using Plasmodium berghei, we show that cell traversal is important in the host dermis for preventing sporozoite destruction by phagocytes and arrest by nonphagocytic cells. We also show that cell infection is a pathway that is masked, rather than activated, by cell traversal. We propose that the cell traversal activity of the sporozoite must be turned on for progression to the liver parenchyma, where it must be switched off for infection of a final hepatocyte. The malaria sporozoite, the parasite stage transmitted by the mosquito, is delivered into the dermis and differentiates in the liver. Motile sporozoites can invade host cells by disrupting their plasma membrane and migrating through them (termed cell traversal), or by forming a parasite-cell junction and settling inside an intracellular vacuole (termed cell infection). Traversal of liver cells, observed for sporozoites in vivo, is thought to activate the sporozoite for infection of a final hepatocyte. Here, using Plasmodium berghei, we show that cell traversal is important in the host dermis for preventing sporozoite destruction by phagocytes and arrest by nonphagocytic cells. We also show that cell infection is a pathway that is masked, rather than activated, by cell traversal. We propose that the cell traversal activity of the sporozoite must be turned on for progression to the liver parenchyma, where it must be switched off for infection of a final hepatocyte. Malaria infection is initiated when an Anopheles mosquito injects Plasmodium sporozoites into the dermis of the host (Sidjanski and Vanderberg, 1997Sidjanski S. Vanderberg J.P. Delayed migration of Plasmodium sporozoites from the mosquito bite site to the blood.Am. J. Trop. Med. Hyg. 1997; 57: 426-429Crossref PubMed Scopus (117) Google Scholar, Matsuoka et al., 2002Matsuoka H. Yoshida S. Hirai M. Ishii A. A rodent malaria, Plasmodium berghei, is experimentally transmitted to mice by merely probing of infective mosquito, Anopheles stephensi.Parasitol. Int. 2002; 51: 17-23Crossref PubMed Scopus (58) Google Scholar, Vanderberg and Frevert, 2004Vanderberg J.P. Frevert U. Intravital microscopy demonstrating antibody-mediated immobilisation of Plasmodium berghei sporozoites injected into skin by mosquitoes.Int. J. Parasitol. 2004; 34: 991-996Crossref PubMed Scopus (238) Google Scholar, Amino et al., 2006Amino R. Martin B. Thiberge S. Celli S. Shorte S. Frischknecht F. Ménard R. Quantitative imaging of Plasmodium transmission from mosquito to mammal.Nat. Med. 2006; 12: 220-224Crossref PubMed Scopus (378) Google Scholar, Yamauchi et al., 2007Yamauchi L.M. Coppi A. Snounou G. Sinnis P. Plasmodium sporozoites trickle out of the injection site.Cell. Microbiol. 2007; 9: 1215-1222Crossref PubMed Scopus (141) Google Scholar). Sporozoites travel from the site of mosquito bite to the liver, where they enter and settle inside hepatocytes. They then multiply and differentiate into the parasite form called merozoite, which infects red blood cells and causes the symptoms of the disease. The journey and fate of the sporozoite in the mammalian host is still a poorly documented part of the parasite life cycle. A recent quantitative in vivo imaging study has revealed that sporozoites inoculated by a mosquito reach not only the liver, via the bloodstream, but also the lymph node draining the site of the mosquito bite, where most are internalized inside dendritic cells and some can initiate development (Amino et al., 2006Amino R. Martin B. Thiberge S. Celli S. Shorte S. Frischknecht F. Ménard R. Quantitative imaging of Plasmodium transmission from mosquito to mammal.Nat. Med. 2006; 12: 220-224Crossref PubMed Scopus (378) Google Scholar). In liver sinusoids, sporozoites interact with resident macrophages, the Kupffer cells, which are thought to act as necessary gates to the underlying parenchyma (Pradel and Frevert, 2001Pradel G. Frevert U. Malaria sporozoites actively enter and pass through rat Kupffer cells prior to hepatocyte invasion.Hepatology. 2001; 33: 1154-1165Crossref PubMed Scopus (127) Google Scholar, Frevert et al., 2005Frevert U. Engelmann S. Zougbede S. Stange J. Ng B. Matuschewski K. Liebes L. Yee H. Intravital observation of Plasmodium berghei sporozoite infection of the liver.PLoS Biol. 2005; 3: e192https://doi.org/10.1371/journal.pbio.0030192Crossref PubMed Scopus (234) Google Scholar, Baer et al., 2007Baer K. Roosevelt M. Clarkson Jr., A.B. van Rooijen N. Schnieder T. Frevert U. Kupffer cells are obligatory for Plasmodium yoelii sporozoite infection of the liver.Cell. Microbiol. 2007; 9: 397-412Crossref PubMed Scopus (83) Google Scholar). The elongated sporozoite cell displays an active gliding locomotion on solid substrates in vitro and in host tissues, reaching speeds up to 4 μm/s, which is powered by a submembranous actin-myosin motor (Ménard, 2001Ménard R. Gliding motility and cell invasion by Apicomplexa: Insights from the Plasmodium sporozoite.Cell. Microbiol. 2001; 3: 63-73Crossref PubMed Scopus (99) Google Scholar, Kappe et al., 2004Kappe S.H. Buscaglia C.A. Nussenzweig V. Plasmodium sporozoite molecular cell biology.Annu. Rev. Cell Dev. Biol. 2004; 20: 29-59Crossref PubMed Scopus (116) Google Scholar). Using this motor, the sporozoite can invade host cells in two distinct ways. Like other invasive stages of Apicomplexa protozoa, it can penetrate the cell inside a so-called parasitophorous vacuole (PV) formed by invagination of the host cell plasma membrane. Typically, the apicomplexan zoite forms an intimate junction between its anterior pole and the contacting host cell surface, the so-called moving junction (MJ), on which it must exert force to pull itself inside the nascent vacuole (Hollingdale et al., 1981Hollingdale M.R. Leef J.L. McCullough M. Beaudouin R.L. In vitro cultivation of the exoerythrocytic stage of Plasmodium berghei from sporozoites.Science. 1981; 213: 1021-1022Crossref PubMed Scopus (61) Google Scholar, Sibley, 2004Sibley L.D. Intracellular parasite invasion strategies.Science. 2004; 304: 248-253Crossref PubMed Scopus (339) Google Scholar). This process, termed here cell infection, is a prerequisite for complete sporozoite differentiation into merozoites and occurs in vivo inside hepatocytes. The sporozoite can also disrupt host membranes and migrate through and out of the cell. The cell traversal behavior of the sporozoite was first described with macrophages (Vanderberg et al., 1990Vanderberg J.P. Chew S. Stewart M.J. Plasmodium sporozoite interactions with macrophages in vitro: A videomicroscopic analysis.J. Protozool. 1990; 37: 528-536Crossref PubMed Scopus (68) Google Scholar) and later shown to also occur with epithelial cells and fibroblasts (Mota et al., 2001Mota M.M. Pradel G. Vanderberg J.P. Hafalla J.C. Frevert U. Nussenzweig R.S. Nussenzweig V. Rodriguez A. Migration of Plasmodium sporozoites through cells before infection.Science. 2001; 291: 141-144Crossref PubMed Scopus (374) Google Scholar). In the P. berghei species that infect rodents, this activity was documented in vivo only in the liver (Frevert et al., 2005Frevert U. Engelmann S. Zougbede S. Stange J. Ng B. Matuschewski K. Liebes L. Yee H. Intravital observation of Plasmodium berghei sporozoite infection of the liver.PLoS Biol. 2005; 3: e192https://doi.org/10.1371/journal.pbio.0030192Crossref PubMed Scopus (234) Google Scholar, Mota et al., 2001Mota M.M. Pradel G. Vanderberg J.P. Hafalla J.C. Frevert U. Nussenzweig R.S. Nussenzweig V. Rodriguez A. Migration of Plasmodium sporozoites through cells before infection.Science. 2001; 291: 141-144Crossref PubMed Scopus (374) Google Scholar). Based on work performed with P. berghei-hepatocyte in vitro systems, the current view is that, in vivo, sporozoites traverse several hepatocytes before infecting a final hepatocyte, and that the former step has a dual activating role on the latter. First, it appeared to render the sporozoite competent for infecting a final cell inside a PV, by inducing progressive exocytosis of the parasite proteins specifically involved in this process (Mota et al., 2002Mota M.M. Hafalla J.C.R. Rodriguez A. Migration through host cells activates Plasmodium sporozoites for infection.Nat. Med. 2002; 8: 1318-1322Crossref PubMed Scopus (149) Google Scholar). Second, it was found to cause the release of hepatocyte growth factor (HGF) from traversed cells, and HGF to promote parasite development in infected cells via cMET-dependent signaling pathways (Carrolo et al., 2003Carrolo M. Giordano S. Cabrita-Santos L. Corso S. Vigario A.M. Silva S. Leiriao P. Carapau D. Armas-Portela R. Comoglio P.M. et al.Hepatocyte growth factor and its receptor are required for malaria infection.Nat. Med. 2003; 9: 1363-1369Crossref PubMed Scopus (113) Google Scholar). More recent work, however, has challenged these conclusions. Inactivation in P. berghei of the genes named spect (Ishino et al., 2004Ishino T. Yano K. Chinzei Y. Yuda M. Cell-passage activity is required for the malarial parasite to cross the liver sinusoidal cell layer.PLoS Biol. 2004; 2: 77-84Crossref Scopus (186) Google Scholar) or spect2 (Ishino et al., 2005aIshino T. Chinzei Y. Yuda M. A Plasmodium sporozoite protein with a membrane attack complex domain is required for breaching the liver sinusoidal cell layer prior to hepatocyte infection.Cell. Microbiol. 2005; 7: 199-208Crossref PubMed Scopus (157) Google Scholar) impaired the sporozoite capacity to traverse hepatoma cells lines without affecting its ability to develop inside these cells. Host cell traversal was found to be important for sporozoite crossing the liver sinusoid barrier, possibly by migrating through Kupffer cells (Ishino et al., 2004Ishino T. Yano K. Chinzei Y. Yuda M. Cell-passage activity is required for the malarial parasite to cross the liver sinusoidal cell layer.PLoS Biol. 2004; 2: 77-84Crossref Scopus (186) Google Scholar, Ishino et al., 2005aIshino T. Chinzei Y. Yuda M. A Plasmodium sporozoite protein with a membrane attack complex domain is required for breaching the liver sinusoidal cell layer prior to hepatocyte infection.Cell. Microbiol. 2005; 7: 199-208Crossref PubMed Scopus (157) Google Scholar). SPECT and SPECT2 are structurally unrelated secretory proteins, and SPECT2 possesses a typical membrane-attack/perforin (MACPF)-like domain, found in pore-forming proteins such as components of the mammalian complement system and perforin. Here, to further study the role of host cell traversal in vivo, we rendered spect(−) and spect2(−) sporozoites fluorescent and characterized their behavior by real-time imaging both in vivo and in vitro. We first generated a fluorescent P. berghei ANKA clone, named ConF, by integrating at the DHFR-TS locus of the wild-type the GFP gene fused to HSP70 regulatory sequences (see Figure S1 available online). Erythrocytic stages of the P. berghei spect(−) and spect2(−) clones (Ishino et al., 2004Ishino T. Yano K. Chinzei Y. Yuda M. Cell-passage activity is required for the malarial parasite to cross the liver sinusoidal cell layer.PLoS Biol. 2004; 2: 77-84Crossref Scopus (186) Google Scholar, Ishino et al., 2005aIshino T. Chinzei Y. Yuda M. A Plasmodium sporozoite protein with a membrane attack complex domain is required for breaching the liver sinusoidal cell layer prior to hepatocyte infection.Cell. Microbiol. 2005; 7: 199-208Crossref PubMed Scopus (157) Google Scholar) were separately mixed with erythrocytic stages of the ConF clone and transmitted to Anopheles stephensi mosquitoes, where meiosis and random chromosome segregation occur. The double transgenic parasites spect−/gfp+ and spect2−/gfp+ emerging from cross-fertilized zygotes, named SpectF and Spect2F, respectively, were cloned as erythrocytic stages after parasite cycling (Figure S1). The ConF, SpectF, and Spect2F clones were then transmitted to Anopheles stephensi mosquitoes, and 18 days after parasite transmission, similar numbers of sporozoites in the three clones were present in the mosquito salivary glands. In matrigel, ∼80% of the sporozoites in the three clones glided with an average velocity of ∼1.4 μm/s for up to 30 min at 37°C (Figure 1A and Figure S2), typically following a corkscrew path while occasionally moving randomly. To examine sporozoite capacity to wound host cell plasma membranes, sporozoites were recorded for 30 min inside matrigels containing host cells in the presence of SYTOX Orange, a nucleic acid stain that penetrates cells with compromised plasma membranes and fluoresces in the nucleus. Cells fluorescent after 30 min were individually examined and scored as wounding events when cell fluorescence started less than 2 min after sporozoite contact. ConF sporozoites wounded all cell types tested, i.e., mast cells and dermal fibroblasts (data not shown), primary hepatocytes (Figure 1B), and HepG2 hepatoma cells (Figure 1C and Movie S1). In primary hepatocytes, ConF sporozoites (multiplicity of infection = 1) provoked an average of 1.8 wounding events/sporozoite/hr. In contrast, SpectF and Spect2F sporozoites did not induce host cell fluorescence in any of the cell types tested. The spect and spect2 genes are thus both dispensable for sporozoite gliding in three-dimensional (3D) matrices but individually critical for the membrane-damaging capacity of the sporozoite. To test whether the traversal activity was important during the skin phase of the sporozoite's journey, we compared infectivity of intravenously or subcutaneously injected wild-type or spect(−) sporozoites in animals treated with liposome-encapsulated clodronate. Clodronate destroys macrophages in the liver and, to a lesser extent, in the spleen, but not in other tissues (van Rooijen et al., 1997van Rooijen N. Bakker J. Sanders A. Transient suppression of macrophage functions by liposome-encapsulated drugs.Trends Biotechnol. 1997; 15: 178-185Abstract Full Text PDF PubMed Scopus (99) Google Scholar). In these animals, intravenously injected wild-type and mutant sporozoites had similar infectivity, whereas subcutaneously injected mutant sporozoites were ∼5- to 10-fold less infective than the wild-type (Figure 2A). This suggested that the traversal activity was important during the sporozoite transit in the skin. We then examined by intravital imaging the fate of sporozoites in the dermis of mice after natural transmission. A single mosquito was allowed to probe the ear of an anesthetized Hairless mouse for 1 min, and the probed site was observed by spinning disk confocal microscopy (Amino et al., 2007Amino R. Thiberge S. Blazquez S. Baldacci P. Renaud O. Shorte S. Ménard R. Imaging malaria sporozoites in the dermis of the mammalian host.Nat. Protocols. 2007; 2: 1705-1712Crossref PubMed Scopus (58) Google Scholar) at various times postinfection (p.i.). Like P. berghei NK65 sporozoites (Amino et al., 2006Amino R. Martin B. Thiberge S. Celli S. Shorte S. Frischknecht F. Ménard R. Quantitative imaging of Plasmodium transmission from mosquito to mammal.Nat. Med. 2006; 12: 220-224Crossref PubMed Scopus (378) Google Scholar), most (∼80%) ANKA ConF sporozoites glided at an average speed of ∼1–2 μm/s, following a tortuous path (Figures 2B and 2C; Figure S3). In contrast, most mutant sporozoites were immotile, with only ∼10% of the sporozoites in the two mutant clones gliding between 15 and 30 min p.i. (Figures 2B and 2C; Figure S3). In agreement with their normal gliding in 3D matrices, the path and speed of the few mutant sporozoites that were motile were similar to those of ConF sporozoites. Therefore, most of the mutants, despite normally gliding in 3D matrices, were rapidly immobilized in the dermis, presumably by host cells they cannot traverse. We next tested whether host leukocytes were involved in the arrest of cell traversal-deficient mutants in the dermis. Spect2F sporozoites were transmitted to Hairless mice by mosquito bite, and the site of bite was extracted, stained with antibodies to CD11b, a leukocyte-specific antigen, and examined by confocal microscopy. At 5 and 30 min p.i., when ∼75% of the control sporozoites are motile, about 10% and 50% of the mutant sporozoites, respectively, were associated with CD11b+ cells (Figure 3A), and ∼25% of the mutants could be detected inside these cells after 30 min (Figure 3B). Similar results were obtained using SpectF sporozoites (data not shown). Interactions between SpectF sporozoites and host phagocytes were then imaged in real-time in lys-gfp mice (Faust et al., 2000Faust N. Varas F. Kelly L.M. Heck S. Graf T. Insertion of enhanced green fluorescent protein into the lysozyme gene creates mice with green fluorescent granulocytes and macrophages.Blood. 2000; 96: 719-726Crossref PubMed Google Scholar), in which myelomonocytic cells (macrophages, neutrophil granulocytes, and dendritic leuykocytes) express GFP (Figures 3C and 3D). At the mosquito bite sites, weakly fluorescent, resident phagocytes were present, while brightly fluorescent cells were recruited starting at ∼25 min p.i. (Figure 3C). Mutant sporozoites were rapidly immobilized, as early as 3 min p.i., and were frequently seen in contact with dermal cells, labeled by red fluorescent BSA, or green fluorescent phagocytic cells. The fluorescence of many mutant sporozoites gradually faded during 1 hr observation periods (Figure 3D, blue inset). Importantly, however, although cell traversal-deficient sporozoites could be destroyed by phagocytic cells, the fluorescence of some sporozoites remained unchanged with time (Figure 3D, red inset), suggesting that immobilized mutants could also escape degradation. The fact that a proportion of mutant sporozoites immobilized in the dermis were not associated with CD11b+ cells and were not destroyed by phagocytes suggested that mutants might also invade nonprofessional phagocytes. To test this, we examined sporozoite interactions with fibroblasts, a major nonphagocytic cell type in the dermis. ConF, SpectF, or Spect2F sporozoites were mixed with either human foreskin fibroblasts (HFF) or mouse dermal fibroblasts (ATCC CRL-2017) in matrigel, the trajectories of gliding sporozoites were visualized as maximal intensity projections, and the proportion of immotile sporozoites was counted at various times (Figures 4A–4D). For up to 30 min, ∼80% of the ConF sporozoites moved in a pattern indistinguishable from that in cell-free matrigel, showing that the presence of host cells did not affect motility of normal sporozoites. In contrast, less than 20% of the sporozoites in both mutant clones were still motile after 30 min (Figures 4C and 4D), most mutant sporozoites being immobilized in association with a cell (Figure 4B). Time-lapse imaging showed that mutant sporozoites were suddenly arrested upon the first contact with a fibroblast, frequently remaining bound to the host cell surface for extended periods of time (Figure 4E and Movie S2). To test whether mutant sporozoites could penetrate fibroblasts, SpectF sporozoites were incubated with HFF cells, and after 30 min thin sections were examined by transmission electron microscopy (TEM). SpectF sporozoites were detected inside cells and surrounded by a membrane (Figure 4F), showing that mutant sporozoites could also be arrested by and invade dermal fibroblasts. Next, we tested whether host cell traversal played a role during sporozoite crossing endothelial barriers in the dermis. We have shown that sporozoites can actively cross the walls of both blood and lymphatic vessels in the dermis, and that ∼1% of the P. berghei NK65 sporozoites inoculated in the mouse footpad by subcutaneous injection terminate their journey and accumulate in the first draining (popliteal) lymph node (Amino et al., 2006Amino R. Martin B. Thiberge S. Celli S. Shorte S. Frischknecht F. Ménard R. Quantitative imaging of Plasmodium transmission from mosquito to mammal.Nat. Med. 2006; 12: 220-224Crossref PubMed Scopus (378) Google Scholar). Similarly, after injection of 104 ConF sporozoites in the footpad of mice, an average of ∼1%–2% was counted in the popliteal node after 2 hr. To compare the capacity of mutant and control sporozoites to reach the popliteal node, we injected the same number of SpectF or Spect2F sporozoites in the footpad of a mouse and of ConF sporozoites contralaterally and counted the number of sporozoites in the popliteal nodes 2 hr p.i. (Figure 2D). The numbers of SpectF and Spect2F sporozoites were only 25% and 32% of that of ConF sporozoites, respectively. The similar decrease in the proportion of mutant sporozoites in the lymph node at 2 hr p.i. (Figure 2D) and in the proportion that initially glided in the dermis (Figure 2B) thus suggests that mutant sporozoites have no specific defect in crossing the wall of lymph vessels. The residual infectivity of mutant sporozoites delivered to normal rats by mosquito bite (data not shown) or injected subcutaneously in normal (data not shown) or clodronate-treated animals (Figure 2A) also indicates that cell traversal is not essential for crossing the wall of dermal blood vessels. Therefore, immobilization of mutant sporozoites inside leukocytes or other cell types in the dermis seems to constitute a primary defect, rather than a consequence of an inability to cross endothelial barriers in the dermis. We next investigated sporozoite traversal of hepatocytes. Previous studies have suggested that sporozoites traverse several hepatocytes in vivo before final infection (Frevert et al., 2005Frevert U. Engelmann S. Zougbede S. Stange J. Ng B. Matuschewski K. Liebes L. Yee H. Intravital observation of Plasmodium berghei sporozoite infection of the liver.PLoS Biol. 2005; 3: e192https://doi.org/10.1371/journal.pbio.0030192Crossref PubMed Scopus (234) Google Scholar, Mota et al., 2001Mota M.M. Pradel G. Vanderberg J.P. Hafalla J.C. Frevert U. Nussenzweig R.S. Nussenzweig V. Rodriguez A. Migration of Plasmodium sporozoites through cells before infection.Science. 2001; 291: 141-144Crossref PubMed Scopus (374) Google Scholar). In agreement with this, ConF sporozoites were found to glide extensively in the liver parenchyma before finally arresting, as exemplified in Figure S4 (Movie S4). To study cell traversal in the liver parenchyma, we first compared the differentiation of ConF, SpectF, and Spect2F sporozoites in rodent primary hepatocytes. P. berghei sporozoites develop into exoerythrocytic forms (EEF) that yield after ∼60 hr thousands of mature merozoites, the erythrocyte-infecting stage (Sturm et al., 2006Sturm A. Amino R. van de Sand C. Regen T. Retzlaff S. Rennenberg A. Krueger A. Pollok J.M. Ménard R. Heussler V.T. Manipulation of host hepatocytes by the malaria parasite for delivery into liver sinusoids.Science. 2006; 313: 1287-1290Crossref PubMed Scopus (349) Google Scholar). No difference was noticed between the three clones in the number, size, and fluorescence intensity of EEF at 4, 12, 24, or 48 hr in rat (Figure 5A) or mouse (data not shown) primary hepatocytes. The three clones generated merozoites with similar infectivity to rats, as measured by prepatent periods of infection (data not shown). Therefore parasite development inside hepatocytes does not appear to depend on prior traversal of these cells. To examine sporozoite entry into primary hepatocytes, infection events (parasites internalized inside a vacuole) were measured after 1 hr incubation, when sporozoites are no longer motile and invasive. For this, samples fixed at 1 hr were labeled with antibodies to the sporozoite CS surface protein to discriminate extracellular (red) from intracellular (green) parasites, and infection events counted as green parasites (see Experimental Procedures). After 1 hr incubation with primary hepatocytes, ∼20% of the initial sporozoites in the three clones were scored as infection events (Figure 5B). Similar results were obtained using CRL-2017 dermal fibroblasts (Figure 5B). The kinetics of cell invasion by mutant and control sporozoites were then compared by TEM analysis of primary hepatocytes fixed after 10 or 30 min incubation. Using Spect2F sporozoites, at both 10 and 30 min a high proportion of cells (37%) contained an intracellular parasite, and as expected, 100% of the intracellular Spect2F sporozoites were surrounded by a membrane (Figure 5C and Figure S5). Instead, using ConF sporozoites, a lower proportion of cells (∼15%) contained an intracellular parasite, and the proportion of intracellular parasites surrounded by a membrane increased from 35% to 60% after 10 and 30 min, respectively (Figure 5C and Figure S5). The rapid kinetics of hepatocyte invasion by mutant sporozoites was confirmed by CS staining assays (Figures 6A and 6B). Using Spect2F, the first intracellular sporozoites were detected after only 2 min incubation, >2% of the sporozoites appeared bicolor (half red/CS-half green/GFP) during the first 4 min, presumably fixed during cell penetration, and the final levels of 20% of internalized sporozoites were reached after only 10 min. Similar results were obtained during entry of Spect2F sporozoites into dermal fibroblasts (Figure S6) and with SpectF sporozoites in both cell types (data not shown). Time-lapse imaging of Spect2F sporozoites incubated with primary hepatocytes readily showed sporozoites displaying and moving through a constriction, suggestive of a MJ, at the site of host cell contact (Figure 6C and Movie S3). Finally, TEM analysis of samples fixed at 3 min detected Spect2F sporozoites entering primary hepatocytes while forming a MJ with the host cell surface (Figure 6D), which is seen here for the first time between a Plasmodium sporozoite and a mammalian cell. Finally, we addressed the possibility that the “rapid invader” phenotype of the mutants might be due to secondary changes on their surfaces, conferring gain-of-function infective capacities that would normally be activated by traversal of host cells. We first compared adhesion of control and mutant sporozoites to confluent CRL-2017 monolayers in the presence of 1 μg/ml cytochalasin (Figures 7A and 7B), which prevents parasite internalization into but not attachment to host cells, or using sporozoites metabolically inhibited by 0.03% azide (data not shown). No significant difference between adhesion of control and mutant sporozoites to cells was noticed in any of these conditions. Also, using real-time qPCR, we compared in salivary gland sporozoites the levels of transcripts encoding the 13 parasite products currently known or suspected to be involved in sporozoite adhesion and/or invasion of host cells, listed in Figure 7C. No significant difference was observed in expression of any of these genes in ConF, SpectF, and Spect2F sporozoites (Figure 7C). These data confirm the view that the “rapid invader” phenotype of the mutants is a direct consequence of their lack of cell traversal. The primary function of the host cell traversal capacity of the Plasmodium sporozoite, first described by Vanderberg and collaborators in 1990, remains controversial. Because the traversal activity in vivo was documented first in hepatocytes in rodents, it was presumed that traversing hepatocytes was an important step that would directly favor the final infection step in a PV (Mota et al., 2001Mota M.M. Pradel G. Vanderberg J.P. Hafalla J.C. Frevert U. Nussenzweig R.S. Nussenzweig V. Rodriguez A. Migration of Plasmodium sporozoites through cells before infection.Science. 2001; 291: 141-144Crossref PubMed Scopus (374) Google Scholar). In vitro data have suggested that traversal of hepatocytes was essential in two distinct ways: by rendering the sporozoite competent for entering a cell inside a PV (Mota et al., 2002Mota M.M. Hafalla J.C.R. Rodriguez A. Migration through host cells activates Plasmodium sporozoites for infection.Nat. Med. 2002; 8: 1318-1322Crossref PubMed Scopus (149) Google Scholar) and by modifying the infected hepatocyte for optimal development of he parasite in the PV (Carrolo et al., 2003Carrolo M. Giordano S. Cabrita-Santos L. Corso S. Vigario A.M. Silva S. Leiriao P. Carapau D. Armas-Portela R. Comoglio P.M. et al.Hepatocyte growth factor and its receptor are required for malaria infection.Nat. Med. 2003; 9: 1363-1369Crossref PubMed Scopus (113) Google Scholar). The results presented here, along with previous studies (Ishino et al., 2004Ishino T. Yano K. Chinzei Y. Yuda M. Cell-passage activity is required for the malarial parasite to cross the liver sinusoidal cell layer.PLoS Biol. 2004; 2: 77-84Crossref Scopus (186) Google Scholar, Ishino et al., 2005aIshino T. Chinzei Y. Yuda M. A Plasmodium sporozoite protein with a membrane attack complex domain is required for breaching the liver sinusoidal cell layer prior to hepatocyte infection.Cell. Microbiol. 2005; 7: 199-208Crossref PubMed Scopus (157) Google Scholar), suggest a different contribution of the cell traversal activity. We show here that this activity is first important in the dermis of the host, where it primarily prevents sporozoite destruction by phagocytic cells. A secondary effect of cell traversal might be to avoid infection of cells that sporozoites can penetrate inside a vacuole but are not their final destination, such as dermal fibroblasts. These two roles might serve the sporozoite at other steps of its journey to the liver parenchyma, including, as previously suggested, for resistance to clearance by Kupffer cells in liver sinusoids (Ishino et al., 2004Ishino T. Yano K. C" @default.
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• W2029131750 date "2008-02-01" @default.
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• W2029131750 title "Host Cell Traversal Is Important for Progression of the Malaria Parasite through the Dermis to the Liver" @default.
• W2029131750 cites W1522914336 @default.
• W2029131750 cites W1583188198 @default.
• W2029131750 cites W1586954533 @default.
• W2029131750 cites W175241905 @default.
• W2029131750 cites W1905994259 @default.
• W2029131750 cites W1973529656 @default.
• W2029131750 cites W1979084140 @default.
• W2029131750 cites W1983531823 @default.
• W2029131750 cites W1986974838 @default.
• W2029131750 cites W1990402424 @default.
• W2029131750 cites W1992040269 @default.
• W2029131750 cites W1992797356 @default.
• W2029131750 cites W1996871916 @default.
• W2029131750 cites W1999475846 @default.
• W2029131750 cites W2007759522 @default.
• W2029131750 cites W2008667436 @default.
• W2029131750 cites W2013222017 @default.
• W2029131750 cites W2014416747 @default.
• W2029131750 cites W2015186773 @default.
• W2029131750 cites W2021528981 @default.
• W2029131750 cites W2029748270 @default.
• W2029131750 cites W2033113133 @default.
• W2029131750 cites W2039481952 @default.
• W2029131750 cites W2052272127 @default.
• W2029131750 cites W2060564531 @default.
• W2029131750 cites W2060650331 @default.
• W2029131750 cites W2064348801 @default.
• W2029131750 cites W2069999816 @default.
• W2029131750 cites W2077675327 @default.
• W2029131750 cites W2093071092 @default.
• W2029131750 cites W2100775932 @default.
• W2029131750 cites W2111146118 @default.
• W2029131750 cites W2113424859 @default.
• W2029131750 cites W2130373733 @default.
• W2029131750 cites W2138623091 @default.
• W2029131750 cites W2139267818 @default.
• W2029131750 cites W2141722810 @default.
• W2029131750 cites W2142106325 @default.
• W2029131750 cites W2155731278 @default.
• W2029131750 cites W2170963096 @default.
• W2029131750 cites W2409659455 @default.
• W2029131750 doi "https://doi.org/10.1016/j.chom.2007.12.007" @default.
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