Proliferation of Toxoplasma gondii Suppresses Host Cell Autophagy

Autophagy is a process of cytoplasmic degradation of endogenous proteins and organelles. Although its primary role is protective, it can also contribute to cell death. Recently, autophagy was found to play a role in the activation of host defense against intracellular pathogens. The aims of our study was to investigate whether host cell autophagy influences Toxoplasma gondii proliferation and whether autophagy inhibitors modulate cell survival. HeLa cells were infected with T. gondii with and without rapamycin treatment to induce autophagy. Lactate dehydrogenase assays showed that cell death was extensive at 36-48 hr after infection in cells treated with T. gondii with or without rapamycin. The autophagic markers, LC3 II and Beclin 1, were strongly expressed at 18-24 hr after exposure as shown by Western blotting and RT-PCR. However, the subsequent T. gondii proliferation suppressed autophagy at 36 hr post-infection. Pre-treatment with the autophagy inhibitor, 3-methyladenine (3-MA), down-regulated LC3 II and Beclin 1. The latter was also down-regulated by calpeptin, a calpain inhibitor. Monodansyl cadaverine (MDC) staining detected numerous autophagic vacuoles (AVs) at 18 hr post-infection. Ultrastructural observations showed T. gondii proliferation in parasitophorous vacuoles (PVs) coinciding with a decline in the numbers of AVs by 18 hr. FACS analysis failed to confirm the presence of cell apoptosis after exposure to T. gondii and rapamycin. We concluded that T. gondii proliferation may inhibit host cell autophagy and has an impact on cell survival.     

Host cell actin polymerization is required for cellular retention of Trypanosoma cruzi and early association with endosomal/lysosomal compartments

One of the hallmarks of Trypanosoma cruzi invasion of non-professional phagocytes is facilitation of the process by host cell actin depolymerization. Host cell entry by invasive T. cruzi trypomastigotes is accomplished by exploiting a cellular wound repair process involving Ca(2+)-regulated lysosome exocytosis (i.e. lysosome-dependent) or by engaging a recently recognized lysosome-independent pathway. It was originally postulated that cortical actin microfilaments present a barrier to lysosome-plasma membrane fusion and that transient actin depolymerization enhances T. cruzi entry by increasing access to the plasma membrane for lysosome fusion. Here we demonstrate that cytochalasin D treatment of host cells inhibits early lysosome association with invading T. cruzi trypomastigotes by uncoupling the cell penetration step from lysosome recruitment and/or fusion. These findings provide the first indication that lysosome-dependent T. cruzi entry is initiated by plasma membrane invagination similar to that observed for lysosome-independent entry. Furthermore, prolonged disruption of host cell actin microfilaments results in significant loss of internalized parasites from infected host cells. Thus, the ability of internalized trypomastigotes to remain cell-associated and to fuse with host cell lysosomes is critically dependent upon host cell actin reassembly, revealing an unanticipated role for cellular actin remodelling in the T. cruzi invasion process.     

Brucella Modulates Secretory Trafficking via Multiple Type IV Secretion Effector Proteins

The intracellular pathogenic bacterium Brucella generates a replicative vacuole (rBCV) derived from the endoplasmic reticulum via subversion of the host cell secretory pathway. rBCV biogenesis requires the expression of the Type IV secretion system (T4SS) VirB, which is thought to translocate effector proteins that modulate membrane trafficking along the endocytic and secretory pathways. To date, only a few T4SS substrates have been identified, whose molecular functions remain unknown. Here, we used an in silico screen to identify putative T4SS effector candidate proteins using criteria such as limited homology in other bacterial genera, the presence of features similar to known VirB T4SS effectors, GC content and presence of eukaryotic-like motifs. Using β-lactamase and CyaA adenylate cyclase reporter assays, we identified eleven proteins translocated into host cells by Brucella, five in a VirB T4SS-dependent manner, namely BAB1_0678 (BspA), BAB1_0712 (BspB), BAB1_0847 (BspC), BAB1_1671 (BspE) and BAB1_1948 (BspF). A subset of the translocated proteins targeted secretory pathway compartments when ectopically expressed in HeLa cells, and the VirB effectors BspA, BspB and BspF inhibited protein secretion. Brucella infection also impaired host protein secretion in a process requiring BspA, BspB and BspF. Single or combined deletions of bspA, bspB and bspF affected Brucella ability to replicate in macrophages and persist in the liver of infected mice. Taken together, these findings demonstrate that Brucella modulates secretory trafficking via multiple T4SS effector proteins that likely act coordinately to promote Brucella pathogenesis.     

Galectin-1 on cervical epithelial cells is a receptor for the sexually transmitted human parasite Trichomonas vaginalis

The extracellular protozoan parasite Trichomonas vaginalis causes the most prevalent non-viral sexually transmitted human infection, yet the pathogenesis of infection is poorly understood, and host cell receptors have not been described. The surface of T. vaginalis is covered with a glycoconjugate called lipophosphoglycan (LPG), which plays a role in the adherence and cytotoxicity of parasites to human cells. T. vaginalis LPG contains high amounts of galactose, making this polysaccharide a candidate for recognition by the galactose-binding galectin family of lectins. Here we show that galectin-1 (gal-1) is expressed by human cervical epithelial cells and binds T. vaginalis LPG. Gal-1 binds to parasites in a carbohydrate-dependent manner that is inhibited in the presence of T. vaginalis LPG. Addition of purified gal-1 to cervical epithelial cells also enhances parasite binding, while a decrease in gal-1 expression by small interfering RNA (siRNA) transfection decreases parasite binding. In contrast, the related galectin-7 (gal-7) does not bind T. vaginalis in a carbohydrate-dependent manner, and is unable to mediate attachment of parasites to host cells. Our data are consistent with the presence of multiple host cell receptors for T. vaginalis of which gal-1 is the first to be identified and highlight the importance of glycoconjugates in host-pathogen interactions.     

VirE2: a unique ssDNA-compacting molecular machine

The translocation of single-stranded DNA (ssDNA) across membranes of two cells is a fundamental biological process occurring in both bacterial conjugation and Agrobacterium pathogenesis. Whereas bacterial conjugation spreads antibiotic resistance, Agrobacterium facilitates efficient interkingdom transfer of ssDNA from its cytoplasm to the host plant cell nucleus. These processes rely on the Type IV secretion system (T4SS), an active multiprotein channel spanning the bacterial inner and outer membranes. T4SSs export specific proteins, among them relaxases, which covalently bind to the 5' end of the translocated ssDNA and mediate ssDNA export. In Agrobacterium tumefaciens, another exported protein—VirE2—enhances ssDNA transfer efficiency 2000-fold. VirE2 binds cooperatively to the transferred ssDNA (T-DNA) and forms a compact helical structure, mediating T-DNA import into the host cell nucleus. We demonstrated—using single-molecule techniques—that by cooperatively binding to ssDNA, VirE2 proteins act as a powerful molecular machine. VirE2 actively pulls ssDNA and is capable of working against 50-pN loads without the need for external energy sources. Combining biochemical and cell biology data, we suggest that, in vivo, VirE2 binding to ssDNA allows an efficient import and pulling of ssDNA into the host. These findings provide a new insight into the ssDNA translocation mechanism from the recipient cell perspective. Efficient translocation only relies on the presence of ssDNA binding proteins in the recipient cell that compacts ssDNA upon binding. This facilitated transfer could hence be a more general ssDNA import mechanism also occurring in bacterial conjugation and DNA uptake processes.     

Leukocyte infiltration and tumor cell plasticity are parameters of aggressiveness in primary cutaneous melanoma

Various clinical and experimental observations detected an immunological host defense in cutaneous melanoma. In order to investigate the prognostic value of leukocyte effector mechanisms, we examined the presence of different subsets of leukocytes in tumor samples of 58 patients diagnosed with primary cutaneous melanoma. The presence of T lymphocytes, cytotoxic T lymphocytes, B lymphocytes, CD16+ cells and macrophages was correlated to Breslow depth. A significantly higher amount of several subsets of leukocytes was found in samples with a more progressed tumor stage and survival analysis demonstrated that a higher amount of T lymphocytes and CD16+ cells was associated with a short survival. The amount of FOXP3+ regulatory T lymphocytes did not correlate with survival, nevertheless, it correlated with the amount of total infiltrate. In contrast, analysis of the expression of CD69, a marker for activated lymphocytes, demonstrated that patients with a higher amount of CD69+ lymphocytes had a better survival. In addition, a new parameter for aggressiveness of melanoma, tumor cell plasticity [i.e., the presence of periodic acid Schiff’s (PAS) reagent positive loops], also predicted short survival and a trend of a higher amount of tumor infiltrating leukocytes in tumors with PAS positive loops was observed. These findings demonstrate that leukocyte infiltration and the presence of PAS loops is a sign of tumor aggressiveness and may have prognostic value.     

Theileria annulata promotes Src kinase-dependent host cell polarization by manipulating actin dynamics in podosomes and lamellipodia

Theileria annulata is an intracellular protozoan parasite that infects B cells and macrophages of ruminants. Macrophages infected with T. annulata are de-differentiated and display tumour cell properties and a metastatic behaviour. How parasitized cells adapt their morphology, motility and invasive behaviour has not yet been addressed in detail. In this study, I investigated the regulation of host cell actin dynamics in T. annulata-transformed macrophages and how this affects host cell morphology and motility. T. annulata was found to promote the formation of filamentous-actin-rich podosome-type adhesions (PTAs) and lamellipodia, and to establish a polarized morphology of the infected cell. Characteristic for parasite-dependent host cell polarization is that infected cells display a single, persistent lamellipodium. Src kinases--in particular Hck--are required for the polar extension of this lamellipodium. Hck does so by promoting the clustered assembly of PTAs and accumulation of proteins of the Ezrin, Radixin, Moesin (ERM) family in lamellipodia. Polar accumulation of PTAs and ERM proteins correlates with focal matrix degradation underneath lamellipodia. These findings suggest that T. annulata equips its host cell with properties to adhere and invade. These properties are likely to promote the motile behaviour required for dissemination of infected cells in vivo.     

Interleukin-2 Inhibits HIV-1 Replication in Some Human T Cell Lymphotrophic Virus-1-infected Cell Lines via the Induction and Incorporation of APOBEC3G into the Virion

IL-2 has been used in culture of primary T cells to maintain cell proliferation. We have previously reported that IL-27 inhibits HIV-1 replication in primary T cells in the presence of IL-2. To gain a better understanding of the mechanisms involved in this inhibitory effect, we attempted to investigate in detail the effects of IL-27 and IL-2 using several cell lines. Unexpectedly, IL-27 did not inhibit HIV-1 in T cell lines, whereas IL-2 inhibited HIV-1 replication in the human T cell lymphotrophic virus (HTLV)-1-transformed T cell lines, MT-2, MT-4, SLB-1, and ATL-2. No effects were seen in HTLV-1-negative cell lines. Utilizing MT-2 cells, we demonstrated that IL-2 treatment inhibited HIV-1 syncytia-inducing ability and dose-dependently decreased supernatant p24 antigen levels by >90%. Using real time PCR and Western blot analysis, we observed that IL-2 treatment induced the host restriction factor, APOBEC3G with accumulation into the lower molecular mass active form as characterized by FPLC. Further analysis revealed that the virus recovered from IL-2-treated MT-2 cells had impaired replication competency. This was found to be due to incorporation of APOBEC3G into the virion despite the presence of Vif. These findings demonstrate a novel role for IL-2 in regulating production of infectious HIV-1 virions in HTLV-1-infected cells through the induction of APOBEC3G.     

Repopulation of host lymphohematopoietic systems by donor cells during graft-versus-host reaction in unirradiated adult F1 mice injected with parental lymphocytes.

The graft-vs-host reaction (GVHR) generated by the injection of parental lymphocytes into unirradiated immune-competent F1 hosts is characterized by an acute loss of immune functions, an attack on host tissues, and a gradual recovery of function. Flow cytometric analysis of the donor- and host-derived splenic populations during the course of acute dysfunction and gradual recovery revealed a complex pattern of changes in lymphoid and myeloid populations that resulted in the repopulation of the host with donor-derived cells. Initially, donor-derived T cell populations expanded, particularly CD8+ T cells. Next, host T cell and B cell populations disappeared. Finally, donor-derived cells repopulated the lymphohematopoietic system in the sequence myeloid populations, B cells, and, after a protracted period, T cells. The recovery of immune functions following GVHR-induced immune deficiency was associated with this repopulation of the spleen by donor-derived cells. Donor repopulation of the host lymphohematopoietic system required the presence of both CD4 and CD8 cells in the original donor inoculum. Depletion of donor CD4 populations precluded development of GVHR or any donor engraftment; depletion of CD8 cells resulted in engraftment solely of donor CD4 populations.     

Proliferative responses of T cells from SJL----F1 and F1----SJL bone marrow chimeras to SJL lymphoma cells

RCS tumor cells induce marked proliferation of syngeneic SJL T cells in vivo and in vitro. Certain F1 hybrids of SJL mice give high proliferative responses to gamma-RCS, while other F1 hybrids give low responses. SJL----"non-responder" F1 and "non-responder" F1----SJL semiallogeneic bone marrow chimeras were prepared to study how the host environment affects the ability of T cells to give a proliferative response to gamma-RCS. The results indicate that T cells educated in an SJL host become responsive to RCS cells, while T cells educated in an (SJL X BALB/c)F1 host become unresponsive. This finding applies to both thymus and lymph node T cells. The unresponsiveness in F1 mice is not due to suppressor cells, since added F1 cells do not affect the proliferative response of SJL cells to gamma-RCS. Instead, it appears that RCS-specific T cells are either deleted in (SJL X BALB/c)F1 mice, or expanded in SJL mice as they develop. These findings are discussed in relation to the specificity of the responding T cells, for LPS activated syngeneic B cell blasts as well as RCS cells, and to the presence of a "leaky" thymus barrier in SJL mice for B cells.     

The effect of host mycorrhizal status on host plant–parasitic plant interactions

Two pot experiments were conducted to examine three-level interactions between host plants, mycorrhizal fungi and parasitic plants. In a greenhouse experiment, Poa annua plants were grown in the presence or absence of an AM fungus (either Glomus lamellosum V43a or G. mosseae BEG29) and in the presence or absence of a root hemiparasitic plant (Odontites vulgaris). In a laboratory experiment, mycorrhizal infection (Glomus claroideum BEG31) of Trifolium pratense host plants (mycorrhizal versus non-mycorrhizal) was combined with hemiparasite infection (Rhinanthus serotinus) of the host (parasitized versus non-parasitized). Infection with the two species of Glomus had no significant effect on the growth of P. annua, while hemiparasite infection caused a significant reduction in host biomass. Mycorrhizal status of P. annua hosts (i.e. presence/absence of AM fungus) affected neither the biomass nor the number of flowers produced by the attached O. vulgaris plants. Infection with G. claroideum BEG31 greatly increased the biomass of T. pratense, but hemiparasite infection had no effect. The hemiparasitic R. serotinus plants attached to mycorrhizal hosts had higher biomass and produced more flowers than plants growing with non-mycorrhizal hosts. Roots of T. pratense were colonized by the AM fungus to an extent independent of the presence or absence of the hemiparasite. Our results confirm earlier findings that the mycorrhizal status of a host plant can affect the performance of an attached root hemiparasite. However, improvement of the performance of the parasitic plant following attachment to a mycorrhizal host depends on the extent to which the AM fungi is able to enhance the growth of the host. Accepted: 23 February 2001     

Glucocorticosteroid modification of lymphocyte blastogenesis in tumor-bearing hosts

The present study was done to evaluate glucocorticosteroid modulation of the individual cells contributing to tumor-induced dualistic (macrophage and suppressor T-cell) suppression in mitogen-induced lymphocyte proliferation. Steroid-sensitive T cells from normal mice showed a characteristic decrease in phytohemagglutinin responsiveness in the in vitro presence of 0.1 μg hydrocortisone sodium-21 succinate (HC), while tumor-bearing host (TBH) T cells demonstrated little or no decrease. This correlates well with kinetic study results demonstrating decreased HC-mediated suppression of mitogen-induced TBH T-cell DNA synthesis at 8 to 10 days posttumor cell inoculation. Hydrocortisone-treated cells from mice with advanced tumors had significantly higher degrees of blastogenesis than untreated TBH cells. In the optimal presence of both macrophages (Mφ) and HC, TBH cells had significantly greater blastogenesis than their normal counterparts. Alone, Mφ or HC depressed normal host PHA responsiveness. Experiments were carried out to determine if the increased blastogenesis was due to the additional Mφ presence in TBH. In vivo administration of methylprednisolone acetate demonstrated that steroids can inhibit tumor growth. A significant delay in tumor appearance occurred when the steroid was administered 4 to 7 days after tumor transplant perhaps due to the measurable lack of suppressor T cells. These studies indicated that steroid suppression was directed against a sensitive suppressor T cell and possibly acts by preventing its differentiation. The role of the Mφ in the afferent limb of the cell-mediated immune response seems less clear, but the results suggest nonspecific inhibition of TBH T-cell response to blastogenic stimulus.     

iTRAQ-based phosphoproteomic analysis reveals host cell's specific responses to Toxoplasma gondii at the phases of invasion and prior to egress

Protein phosphorylation plays a key role in host cell-T. gondii interaction. However, the phosphoproteome data of host cell at various phases of T. gondii infection has not been thoroughly described. In this study, we assessed the host phosphoproteome data with isobaric tags for relative and absolute quantification (iTRAQ) method during the phases of T. gondii invasion (30 min post infection, PI) and prior to egress (28 h PI). Our iTRAQ analysis revealed a total of 665 phosphoproteins, among which the significantly regulated phosphoproteins in different between-group comparisons were further analyzed. Functional analysis of these significantly regulated phosphoproteins suggested that T. gondii modulated host cell processes through phosphorylation including cell cycle regulation, inducing apoptosis, blocking the synthesis of some inflammatory factors, mediating metabolism to support its proliferation at the infection phase prior to egress, and utilizing membrane and energy from host cell, reorganizing cytoskeleton to favor its invasion and PV formation at the phase of invasion. The phosphorylation level of Smad2, CTNNA1, and HSPB1 identified with western blot revealed a consistent trend of change with iTRAQ result. These newly identified and significantly regulated phosphoproteins from our phosphoproteome data may provide new clues to unravel the host cell's complex reaction against T. gondii infection and the interaction between the host cell and T. gondii.     

Molecular chaperone function of stress inducible Hsp70 is critical for intracellular multiplication of Toxoplasma gondii

Intracellular pathogens like Toxoplasma gondii often target proteins and pathways critical for host cell survival and stress response. Molecular chaperones encoded by the evolutionary conserved Heat shock proteins (Hsps) maintain proteostasis and are vital to cell survival following exposure to any form of stress. A key protein of this family is Hsp70, an ATP-driven molecular chaperone, which is stress inducible and often indiscernible in normal cells. Role of this protein with respect to intracellular survival and multiplication of protozoan parasite like T. gondii remains to be examined. We find that T. gondii infection upregulates expression of host Hsp70. Hsp70 selective inhibitor 2-phenylethynesulfonamide (PES) attenuates intracellular T. gondii multiplication. Biotinylated PES confirms selective interaction of this small molecule inhibitor with Hsp70. We show that PES acts by disrupting Hsp70 chaperone function which leads to dysregulation of host autophagy. Silencing of host Hsp70 underscores its importance for intracellular multiplication of T. gondii, however, attenuation achieved using PES is not completely attributable to host Hsp70 indicating the presence of other intracellular targets of PES in infected host cells. We find that PES is also able to target T. gondii Hsp70 homologue which was shown using PES binding assay. Detailed molecular docking analysis substantiates PES targeting of TgHsp70 in addition to host Hsp70. While establishing the importance of protein quality control in infection, this study brings to the fore a unique opportunity of dual targeting of host and parasite Hsp70 demonstrating how structural conservation of these proteins may be exploited for therapeutic design.     

Infection by Trypanosoma cruzi Enhances Anion Conductance in Rat Neonatal Ventricular Cardiomyocytes

Recent studies on malaria-infected erythrocytes have shown increased anion channel activity in the host cell membrane, increasing the exchange of solutes between the cytoplasm and exterior. In the present work, we addressed the question of whether another intracellular protozoan parasite, Trypanosoma cruzi, alters membrane transport systems in the host cardiac cell. Neonatal rat cardiomyocytes were cultured and infected with T. cruzi in vitro. Ion currents were measured by patch-clamp technique in the whole-cell configuration. Two small-magnitude instantaneous anion currents, outward- and inward-rectifying, were recorded in all noninfected cardiomyocytes. In addition, ~10% of cardiomyocytes expressed a large anion-preferable, time-dependent current activated at positive membrane potentials. Hypotonic (230?mOsm) treatment resulted in the disappearance of the time-dependent current but provoked a dramatic increase of the instantaneous outward-rectifying one. Both instantaneous currents were suppressed by intracellular Mg(2+). T. cruzi infection did not provoke new anion currents in the host cells but caused an increase of the density of intrinsic swelling-activated outward current, up to twice in heavily infected cells. The occurrence of a time-dependent current dramatically increased in infected cells in the presence of Mg(2+) in the intracellular solution, from ~10 to ~80%, without a significant change of the current density. Our findings represent one further, besides the known Plasmodium falciparum, example of an intracellular parasite which upregulates the anionic currents expressed in the host cell.     

Bringing down the host: enteropathogenic and enterohaemorrhagic Escherichia coli effector‐mediated subversion of host innate immune pathways

Enteric bacterial pathogens commonly use a type III secretion system (T3SS) to successfully infect intestinal epithelial cells and survive and proliferate in the host. Enteropathogenic and enterohaemorrhagic Escherichia coli (EPEC; EHEC) colonize the human intestinal mucosa, form characteristic histological lesions on the infected epithelium and require the T3SS for full virulence. T3SS effectors injected into host cells subvert cellular pathways to execute a variety of functions within infected host cells. The EPEC and EHEC effectors that subvert innate immune pathways – specifically those involved in phagocytosis, host cell survival, apoptotic cell death and inflammatory signalling – are all required to cause disease. These processes are reviewed within, with a focus on recent work that has provided insights into the functions and host cell targets of these effectors.     

TIM-3 drives temporal differences in restimulation-induced cell death sensitivity in effector CD8+ T cells in conjunction with CEACAM1

Immune homeostasis depends upon effective clearance of pathogens while simultaneously preventing autoimmunity and immunopathology in the host. Restimulation-induced cell death (RICD) is one such mechanism where by activated T cells receive subsequent antigenic stimulation, reach a critical signal threshold through the T cell receptor (TCR), and commit to apoptosis. Many details of this process remain unclear, including the role of co-stimulatory and co-inhibitory proteins that influence the TCR signaling cascade. Here we characterize the role of T cell immunoglobulin and mucin domain containing 3 (TIM-3) in RICD regulation. TIM-3 protected newly activated CD8⁺ effector T cells from premature RICD during clonal expansion. Surprisingly, however, we found that TIM-3 potentiated RICD in late-stage effector T cells. The presence of TIM-3 increased proximal TCR signaling and proapoptotic protein expression in late-stage effector T cells, with no consistent signaling effects noted in newly activated cells with or without TIM-3. To better explain these differences in TIM-3 function as T cells aged, we characterized the temporal pattern of TIM-3 expression in effector T cells. We found that TIM-3 was expressed on the surface of newly activated effector T cells, but remained largely intracellular in late-stage effector cells. Consistent with this, TIM-3 required a ligand to prevent early RICD, whereas ligand manipulation had no effects at later stages. Of the known TIM-3 ligands, carcinoembryonic antigen‐related cell adhesion molecule (CEACAM1) showed the greatest difference in surface expression over time and also protected newly activated cells from premature RICD, with no measurable effects in late-stage effectors. Indeed, CEACAM1 enabled TIM-3 surface expression on T cells, implying a co-dependency for these proteins in protecting expanding T cells from premature RICD. Our findings suggest that co-signaling proteins like TIM-3 and CEACAM1 can alter RICD sensitivity at different stages of the effector T cell response, with important implications for checkpoint blockade therapy.Subject terms: Cell death and immune response, Immune cell death, Cytotoxic T cells     

Toxoplasma gondii Proliferation Require Down-Regulation of Host Nox4 Expression via Activation of PI3 Kinase/Akt Signaling Pathway

Toxoplasma gondii results in ocular toxoplasmosis characterized by chorioretinitis with inflammation and necrosis of the neuroretina, pigment epithelium, and choroid. After invasion, T. gondii replicates in host cells before cell lysis, which releases the parasites to invade neighboring cells to repeat the life cycle and establish a chronic retinal infection. The mechanism by which T. gondii avoids innate immune defense, however, is unknown. Therefore, we determined whether PI3K/Akt signaling pathway activation by T. gondii is essential for subversion of host immunity and parasite proliferation. T. gondii infection or excretory/secretory protein (ESP) treatment of the human retinal pigment epithelium cell line ARPE-19 induced Akt phosphorylation, and PI3K inhibitors effectively reduced T. gondii proliferation in host cells. Furthermore, T. gondii reduced intracellular reactive oxygen species (ROS) while activating the PI3K/Akt signaling pathway. While searching for the main source of these ROS, we found that NADPH oxidase 4 (Nox4) was prominently expressed in ARPE-19 cells, and this expression was significantly reduced by T. gondii infection or ESP treatment along with decreased ROS levels. In addition, artificial reduction of host Nox4 levels with specific siRNA increased replication of intracellular T. gondii compared to controls. Interestingly, these T. gondii-induced effects were reversed by PI3K inhibitors, suggesting that activation of the PI3K/Akt signaling pathway is important for suppression of both Nox4 expression and ROS levels by T. gondii infection. These findings demonstrate that manipulation of the host PI3K/Akt signaling pathway and Nox4 gene expression is a novel mechanism involved in T. gondii survival and proliferation.     

New data on Oncophora melanocephala (Nematoda: Camallanidae), a little-known parasite of scombrid fishes

The camallanid nematode Oncophora melanocephala (Rudolphi, 1819) is redescribed from specimens collected in tunas, Thunnus thynnus (L.), T. albacares (Bonnaterre), T. atlanticus (Lesson) and Auxis thazard (Lacépède) from the Atlantic littoral of Rio de Janeiro, Brazil. In addition to a specific structure of the buccal capsule, the species is characterized by the presence of seven pairs of preanal and five pairs of postanal subventral papillae in the male and by an unusual shape of body in the adult female; deirids are described for the first time in O. melanocephala. The structure of the buccal capsule of O. melanocephala, particularly the presence of a large posterior portion of capsule and a colourless connecting ring between buccal capsule and oesophagus, shows close affinities of Oncophora Diesing, 1851 with Paracamallanus Yorke et Maplestone, 1928. The findings of O. melanocephala in T. atlanticus and A thazard represent new host records.