Prime-Boost Vaccination with Toxoplasma Lysate Antigen, but Not with a Mixture of Recombinant Protein Antigens, Leads to Reduction of Brain Cyst Formation

PLoS One. 2015 May 26;10(5):e0126334. doi: 10.1371/journal.pone.0126334.

 

Wagner A¹, Schabussova I¹, Ruttkowski B², Peschke R², Kur J³, Kundi M⁴, Joachim A², Wiedermann U⁵.

INTRODUCTION:

Infection with the ubiquitous parasite Toxoplasma gondii is a threat for immunocompromised patients and pregnant women and effective immune-prophylaxis is still lacking.

METHODS:

Here we tested a mixture of recombinant T. gondii antigens expressed in different developmental stages, i.e., SAG1, MAG1 and GRA7 (SMG), and a lysate derived from T. gondii tachyzoites (TLA) for prophylactic vaccination against cyst formation. Both vaccine formulations were applied systemically followed by an oral TLA-booster in BALB/c mice.

RESULTS:

Systemic priming with SMG and oral TLA-booster did not show significant induction of protective immune responses. In contrast, systemic priming and oral booster with TLA induced higher levels of Toxoplasma-specific IgG, IgG1 and IgG2a in sera as well as high levels of Toxoplasma-specific IgG1 in small intestines. Furthermore, high levels of Toxoplasma-specific Th1-, Th17- and Th2-associated cytokines were only detected in restimulated splenocytes of TLA-vaccinated mice. Importantly, in mice orally infected with T. gondii oocysts, only TLA-vaccination and booster reduced brain cysts. Furthermore, sera from these mice reduced tachyzoites invasion of Vero cells in vitro, indicating that antibodies may play a critical role for protection against Toxoplasma infection. Additionally, supernatants from splenocyte cultures of TLA-vaccinated mice containing high levels of IFN-γ lead to substantial production of nitric oxide (NO) after incubation with macrophages in vitro. Since NO is involved in the control of parasite growth, the high levels of IFN-γ induced by vaccination with TLA may contribute to the protection against T. gondii.

CONCLUSION:

In conclusion, our data indicate that prime-boost approach with TLA, but not with the mixture of recombinant antigens SMG, induces effective humoral and cellular Toxoplasma-specific responses and leads to significant reduction of cerebral cysts, thereby presenting a viable strategy for further vaccine development against T. gondii infection.

PMID:

26010355

[PubMed - as supplied by publisher]

New views of the Toxoplasma gondii parasitophorous vacuole as revealed by Helium Ion Microscopy (HIM)

J Struct Biol. 2015 May 21. pii: S1047-8477(15)00105-7. doi: 10.1016/j.jsb.2015.05.003. [Epub ahead of print]

 

de Souza W¹, Attias M².

 

The Helium Ion Microscope (HIM) is a new technology that uses a highly focused helium ion beam to scan and interact with the sample, which is not coated. The images have resolution and depth of field superior to field emission scanning electron microscopes. In this paper, we used HIM to study LLC-MK2 cells infected with Toxoplasma gondii. These samples were chemically fixed and, after critical point drying, were scraped with adhesive tape to expose the inner structure of the cell and parasitophorous vacuoles. We confirmed some of the previous findings made by field emission-scanning electron microscopy and showed that the surface of the parasite is rich in structures suggestive of secretion, that the nanotubules of the intravacuolar network (IVN) are not always straight, and that bifurcations are less frequent than previously thought. Fusion of the tubules with the parasite membrane or the parasitophorous vacuole membrane (PVM) was also infrequent. Tiny adhesive links were observed for the first time connecting the IVN tubules. The PVM showed openings of various sizes that even allowed the observation of endoplasmic reticulum membranes in the cytoplasm of the host cell. These findings are discussed in relation to current knowledge on the cell biology of T. gondii.
Copyright © 2015. Published by Elsevier Inc.

KEYWORDS:

Intravacuolar network; Parasitophorous vacuole; Scanning electron microscopy; Toxoplasma; Ultrastructure

PMID:

26004092

[PubMed - as supplied by publisher]

The distribution pattern of α2,3- and α2,6-linked sialic acids affects host cell preference in Toxoplasma gondii

Exp Parasitol. 2015 May 20. pii: S0014-4894(15)00129-0. doi: 10.1016/j.exppara.2015.05.005. [Epub ahead of print]

Baba M1, Sato M2, Kitoh K1, Takashima Y3.

Author information

Abstract

Tachyzoites of Toxoplasma gondii, an obligate intracellular parasite, actively invade almost all types of nucleated cells. However, T. gondii tachyzoites preferentially infect particular types of animal tissue cells. The mechanism underlying the host cell preference of T. gondii is not yet known. In this study, we found that enzymatic removal of α2,3- but not α2,6-linked sialic acids on the surface of Vero cells decreased T. gondii tachyzoite adhesion or invasion to the treated cells. Although Chinese hamster ovary (CHO) cells express only α2,3-linked sialic acid, a genetically modified CHO cell line constructed by transfection with the α2,6-sialiltransferase gene contains subpopulations with a variety of expression patterns of α2,3- and α2,6-linked sialic acids. When T. gondii tachyzoites were added to the modified CHO cells, the tachyzoites preferentially attached to cells belonging to a subpopulation of cells that highly expressed α2,3-linked sialic acids. Additionally, multiple regression analysis performed to analyse the relationship between the amount of α2,3-linked/α2,6-linked sialic acids and parasite-expressed fluorescence intensity suggested that more tachyzoites adhered to individual α2,3-linked sialic acid rich-cells than to α2,3-linked sialic acid-poor/null cells. The results of confocal laser microscopy confirmed this finding. These results indicate that the host cell preference of T. gondii was, at least partially, affected by the distribution pattern of α2,3-, but almost never α2,6-linked sialic acids on host cells.

Copyright © 2015 Elsevier Inc. All rights reserved.

KEYWORDS: 

Toxoplasma gondii; cell adhesion; sialic acid; α2,3-linkage; α2,6-linkage

PMID:

 

26003519

 

[PubMed - as supplied by publisher]

Metabolic Needs and Capabilities of Toxoplasma gondii through Combined Computational and Experimental Analysis

PLoS Comput Biol. 2015 May 22;11(5):e1004261. doi: 10.1371/journal.pcbi.1004261. eCollection 2015.

Tymoshenko S1, Oppenheim RD2, Agren R3, Nielsen J3, Soldati-Favre D2, Hatzimanikatis V4.

Author information

Abstract

Toxoplasma gondii is a human pathogen prevalent worldwide that poses a challenging and unmet need for novel treatment of toxoplasmosis. Using a semi-automated reconstruction algorithm, we reconstructed a genome-scale metabolic model, ToxoNet1. The reconstruction process and flux-balance analysis of the model offer a systematic overview of the metabolic capabilities of this parasite. Using ToxoNet1 we have identified significant gaps in the current knowledge of Toxoplasma metabolic pathways and have clarified its minimal nutritional requirements for replication. By probing the model via metabolic tasks, we have further defined sets of alternative precursors necessary for parasite growth. Within a human host cell environment, ToxoNet1 predicts a minimal set of 53 enzyme-coding genes and 76 reactions to be essential for parasite replication. Double-gene-essentiality analysis identified 20 pairs of genes for which simultaneous deletion is deleterious. To validate several predictions of ToxoNet1 we have performed experimental analyses of cytosolic acetyl-CoA biosynthesis. ATP-citrate lyase and acetyl-CoA synthase were localised and their corresponding genes disrupted, establishing that each of these enzymes is dispensable for the growth of T. gondii, however together they make a synthetic lethal pair.

PMID:

 

26001086

 

[PubMed - as supplied by publisher]

Shared elements of host-targeting pathways among apicomplexan parasites of differing life styles

Cell Microbiol. 2015 May 21. doi: 10.1111/cmi.12460. [Epub ahead of print]

Pellé KG1, Jiang RH1,2, Mantel PY1, Xiao YP3, Hjelmqvist D1, Gallego-Lopez GM4, Lau AO4, Kang BH5, Allred DR3,6, Marti M1.

Author information

Abstract

Apicomplexans are a diverse group of obligate parasites occupying different intracellular niches that require modification to meet the needs of the parasite. To efficiently manipulate their environment, apicomplexans translocate numerous parasite proteins into the host cell. Whereas some parasites remain contained within a parasitophorous vacuole membrane (PVM) throughout their developmental cycle, others do not, a difference that affects the machinery needed for protein export. A signal-mediated pathway for protein export into the host cell has been characterized in malaria parasites, which maintain the PVM. Here, we functionally demonstrate an analogous host-targeting pathway involving organellar staging prior to secretion in the related bovine parasite, Babesia bovis, a parasite that destroys the PVM shortly after invasion. Taking into account recent identification of a similar signal-mediated pathway in the coccidian parasite Toxoplasma gondii, we suggest a model in which this conserved pathway has evolved in multiple steps from signal-mediated trafficking to specific secretory organelles for controlled secretion to a complex protein translocation process across the PVM.

This article is protected by copyright. All rights reserved.

KEYWORDS: 

Bioinformatics; Diversity; Microbial-cell interaction; Protozoa

PMID:

 

25996544

 

[PubMed - as supplied by publisher]

Infection of male rats with Toxoplasma gondii results in enhanced delay aversion and neural changes in the nucleus accumbens core

Proc Biol Sci. 2015 Jun 7;282(1808). pii: 20150042. doi: 10.1098/rspb.2015.0042.

Tan D1, Soh LJ1, Lim LW1, Daniel TC2, Zhang X3, Vyas A4.

Author information

Abstract

Rats infected with the protozoan parasite Toxoplasma gondii exhibit reduced avoidance of predator odours. This behavioural change is likely to increase transmission of the parasite from rats to cats. Here, we show that infection with T. gondii increases the propensity of the infected rats to make more impulsive choices, manifested as delay aversion in an intertemporal choice task. Concomitantly, T. gondii infection causes reduction in dopamine content and neuronal spine density of the nucleus accumbens core, but not of the nucleus accumbens shell. These results are consistent with a role of the nucleus accumbens dopaminergic system in mediation of choice impulsivity and goal-directed behaviours. Our observations suggest that T. gondii infection in rats causes a syndromic shift in related behavioural constructs of innate aversion and making foraging decisions. 

© 2015 The Author(s) Published by the Royal Society. All rights reserved.

KEYWORDS: 

behavioural manipulation; brain; delay discounting; dopamine; monoamines; parasites

PMID:

 

25994671

 

[PubMed - in process]

The Toxoplasma Dense Granule Proteins GRA17 and GRA23 Mediate the Movement of Small Molecules between the Host and the Parasitophorous Vacuole

Cell Host Microbe. 2015 May 13;17(5):642-652. doi: 10.1016/j.chom.2015.04.003.

Gold DA1, Kaplan AD2, Lis A2, Bett GC3, Rosowski EE1, Cirelli KM1, Bougdour A4, Sidik SM5, Beck JR6, Lourido S7, Egea PF8, Bradley PJ6, Hakimi MA4, Rasmusson RL2, Saeij JP9.

Author information

Abstract

Toxoplasma gondii is a protozoan pathogen in the phylum Apicomplexa that resides within an intracellular parasitophorous vacuole (PV) that is selectively permeable to small molecules through unidentified mechanisms. We have identified GRA17 as a Toxoplasma-secreted protein that localizes to the parasitophorous vacuole membrane (PVM) and mediates passive transport of small molecules across the PVM. GRA17 is related to the putative Plasmodium translocon protein EXP2 and conserved across PV-residing Apicomplexa. The PVs of GRA17-deficient parasites have aberrant morphology, reduced permeability to small molecules, and structural instability. GRA17-deficient parasites proliferate slowly and are avirulent in mice. These GRA17-deficient phenotypes are rescued by complementation with Plasmodium EXP2. GRA17 functions synergistically with a related protein, GRA23. Exogenous expression of GRA17 or GRA23 alters the membrane conductance properties of Xenopus oocytes in a manner consistent with a large non-selective pore. Thus, GRA17 and GRA23 provide a molecular basis for PVM permeability and nutrient access.

Copyright © 2015 Elsevier Inc. All rights reserved.

PMID:

 

25974303

 

[PubMed - as supplied by publisher]

The zebrafish, Danio rerio, as a model for Toxoplasma gondii: an initial description of infection in fish

J Fish Dis. 2015 May 7. doi: 10.1111/jfd.12393. [Epub ahead of print]

Sanders JL1, Zhou Y, Moulton HM, Moulton ZX, McLeod R, Dubey JP, Weiss LM, Kent ML.

Author information

KEYWORDS: 

Danio rerio ; infection experiment

PMID:

 

25951508

 

[PubMed - as supplied by publisher]

Ciprofloxacin Derivatives Affect Parasite Cell Division and Increase the Survival of Mice Infected with Toxoplasma gondii

PLoS One. 2015 May 7;10(5):e0125705. doi: 10.1371/journal.pone.0125705.

Martins-Duarte ES1, Dubar F2, Lawton P3, França da Silva C4, C Soeiro MN4, de Souza W5, Biot C6, Vommaro RC1.

Author information

Abstract

Toxoplasmosis, caused by the protozoan Toxoplasma gondii, is a worldwide disease whose clinical manifestations include encephalitis and congenital malformations in newborns. Previously, we described the synthesis of new ethyl-ester derivatives of the antibiotic ciprofloxacin with ~40-fold increased activity against T. gondii in vitro, compared with the original compound. Cipro derivatives are expected to target the parasite's DNA gyrase complex in the apicoplast. The activity of these compounds in vivo, as well as their mode of action, remained thus far uncharacterized. Here, we examined the activity of the Cipro derivatives in vivo, in a model of acute murine toxoplasmosis. In addition, we investigated the cellular effects T. gondii tachyzoites in vitro, by immunofluorescence and transmission electron microscopy (TEM). When compared with Cipro treatment, 7-day treatments with Cipro derivatives increased mouse survival significantly, with 13-25% of mice surviving for up to 60 days post-infection (vs. complete lethality 10 days post-infection, with Cipro treatment). Light microscopy examination early (6 and 24h) post-infection revealed that 6-h treatments with Cipro derivatives inhibited the initial event of parasite cell division inside host cells, in an irreversible manner. By TEM and immunofluorescence, the main cellular effects observed after treatment with Cipro derivatives and Cipro were cell scission inhibition - with the appearance of 'tethered' parasites - malformation of the inner membrane complex, and apicoplast enlargement and missegregation. Interestingly, tethered daughter cells resulting from Cipro derivatives, and also Cipro, treatment did not show MORN1 cap or centrocone localization. The biological activity of Cipro derivatives against C. parvum, an apicomplexan species that lacks the apicoplast, is, approximately, 50 fold lower than that in T. gondii tachyzoites, supporting that these compounds targets the apicoplast. Our results show that Cipro derivatives improved the survival of mice acutely infected with T. gondii and inhibited parasite replication early in the first cycle of infection in vitro, highlighting their therapeutic potential for the treatment of toxoplasmosis.

PMID:

 

25950173

 

[PubMed - as supplied by publisher] 

Towards a molecular understanding of the apicomplexan actin motor: on a road to novel targets for malaria remedies?

Acta Crystallogr F Struct Biol Commun. 2015 May 1;71(Pt 5):500-513. Epub 2015 Apr 16

 

Kumpula EP¹, Kursula I¹.

 

Apicomplexan parasites are the causative agents of notorious human and animal diseases that give rise to considerable human suffering and economic losses worldwide. The most prominent parasites of this phylum are the malaria-causing Plasmodium species, which are widespread in tropical and subtropical regions, and Toxoplasma gondii, which infects one third of the world's population. These parasites share a common form of gliding motility which relies on an actin-myosin motor. The components of this motor and the actin-regulatory proteins in Apicomplexa have unique features compared with all other eukaryotes. This, together with the crucial roles of these proteins, makes them attractive targets for structure-based drug design. In recent years, several structures of glideosome components, in particular of actins and actin regulators from apicomplexan parasites, have been determined, which will hopefully soon allow the creation of a complete molecular picture of the parasite actin-myosin motor and its regulatory machinery. Here, current knowledge of the function of this motor is reviewed from a structural perspective.

KEYWORDS:

actin polymerization; cytoskeleton; drug design; gliding motility; malaria; parasitology; regulation

PMID:

25945702

[PubMed - as supplied by publisher]

CD8+ T cell immunity in an encephalitis model of Toxoplasma gondii infection

Semin Immunopathol. 2015 May 6. [Epub ahead of print]

 

Hwang S¹, Khan IA.

 

Toxoplasma gondii infection induces a robust CD8 T cell immunity in the infected host, which is critical for keeping chronic infection under control. IFNγ production and cytolytic activity exhibited by CD8 T cells are critical functions needed to prevent the reactivation of latent infection. Paradoxically, the susceptible mice infected with the parasite develop encephalitis irrespective of the presence of vigorous CD8 T cell immunity. Recent studies from our laboratory have demonstrated that these animals have defect in the memory CD8 T cell population, which become dysfunctional due to exhibition of inhibitory receptors like PD-1. Although the blockade of PD-1-PDL-1 pathway rescues the CD8 response, PD-1^hi expressing cells are refractory to the treatment. In this review, we discuss the development of CD8 memory response during chronic infection, mechanism responsible for their dysfunctionality, and possible therapeutic measures that can be taken to reverse the process.

PMID:

25944514

[PubMed - as supplied by publisher]