Thursday, September 22, 2016

Peroxiredoxin 3 promotes IL-12 production from macrophages and partially protects mice against infection with Toxoplasma gondii

 2016 Sep 16. pii: S1383-5769(16)30208-2. doi: 10.1016/j.parint.2016.09.008. [Epub ahead of print]

Abstract

Toxoplasmosis remains a life-threatening infection of humans and various domestic and wild animals worldwide. It is caused by the obligatory intracellular protozoan parasite Toxoplasma gondii. Peroxiredoxins (Prxs) are a family of antioxidant enzymes that protect cells from oxidative stress from hydroperoxides. In the recent years, several studies have reported the potential use of T. gondii-derived enzymes in triggering protective immunity against T. gondii infection. Therefore, this study was conducted to investigate the immunogenicity and protective efficacy of TgPrx3. In vitro stimulation of peritoneal macrophages with recombinant TgPrx3 protein fused to glutathione-S transferase (TgPrx3-GST) enhanced IL-12p40 production, indicating the immune-stimulating potentials of TgPrx3. Next, protective efficacy was investigated by subcutaneous inoculation of mice with TgPrx3-GST (25pmol), and recombinant GST or PBS were used as the controls. Mice immunized with TgPrx3-GST exhibited a significant elevation of specific antibodies in terms of IgG1 and IgG2c isotypes. Moreover, interferon-gamma production and spleen cell proliferation dramatically increased in the TgPrx3-GST-sensitized cells from mice immunized with the same antigen. The severity of the T. gondii infections tended to be attenuated in the TgPrx3-GST-immunized mice, as evidenced by their higher survival rates and lower parasite burdens in the brain. Altogether, TgPrx3 immunization induced specific humoral and cellular immune responses and partially protected the mice against lethal toxoplasmosis. Our results suggest the possible use of TgPrx3 as a vaccine candidate against T. gondii infections.
Copyright © 2016. Published by Elsevier Ireland Ltd.

KEYWORDS: 

Immunization; Peroxiredoxin 3; Toxoplasma gondii; Vaccine

Evaluation of Propranolol Effect on Experimental Acute and Chronic Toxoplasmosis using Q-PCR

 2016 Sep 19. pii: AAC.01323-16. [Epub ahead of print]

Abstract

Current therapies against toxoplasmosis are limited and drugs have significant side effects and low efficacies. We evaluated the potential anti-Toxoplasma activity of propranolol 2, 3 mg/kg/day in vivo in acute and chronic phases. Propranolol as the stabilizing cell membrane is a suitable drug for inhibiting the entrance of Toxoplasma gondii (T. gondii) tachyzoites into cells. The acute phase was performed using propranolol, pyrimethamine, propranolol plus pyrimethamine before (pre-treatment) and after (post-treatment) intraperitoneally challenge with 1×103 tachyzoites of the virulent RH strain of T. gondii in Balb/c mice. Also in the chronic phase, treatment was performed 12 hours before intraperitoneally challenge with 1×106 tachyzoites of the virulent RH strain of T. gondii in rats. One week (in acute phase) and two months (in chronic phase) after post infection, tissues were isolated and DNA was extracted. Subsequently parasite load was calculated using Q-PCR. In acute phase, in both groups, significant anti-Toxoplasma activity was observed using propranolol (P < 0.001). Propranolol in pre-treatment group showed higher anti-Toxoplasma activity than propranolol in post-treatment in brain tissues displaying therapeutic efficiency on toxoplasmosis. Also, propranolol combined with pyrimethamine reduced the parasite load as well as significantly increased survival of mice in pre-treatment group. In the chronic phase, anti-Toxoplasma activity was observed with propranolol and decreased parasite load in tissues. In conclusion, the presented results demonstrate that propranolol, as an orally available drug, is effective against acute and latent murine toxoplasmosis at low doses and increase efficiency of drug in combination therapy with propranolol-pyrimethamine.
Copyright © 2016, American Society for Microbiology. All Rights Reserved.
PMID:
 
27645234
 
DOI:
 
10.1128/AAC.01323-16

Tuesday, September 20, 2016

Macrophages facilitate the excystation and differentiation of Toxoplasma gondii sporozoites into tachyzoites following oocyst internalisation

 2016 Sep 19;6:33654. doi: 10.1038/srep33654.

Abstract

Toxoplasma gondii is a common parasite of humans and animals, which is transmitted via oocysts in cat faeces or tissue cysts in contaminated meat. The robust oocyst and sporocyst walls protect the infective sporozoites from deleterious external attacks including disinfectants. Upon oocyst acquisition, these walls lose their integrity to let the sporozoites excyst and invade host cells following a process that remains poorly understood. Given the resistance of the oocyst wall to digestive enzymes and the ability of oocysts to cause parenteral infections, the present study investigated the possible contribution of macrophages in supporting sporozoite excystation following oocyst internalisation. By using single cell micromanipulations, real-time and time-point imaging techniques, we demonstrated that RAW macrophages could interact rapidly with oocysts and engulfed them by remodelling of their actin cytoskeleton. Internalised oocysts were associated to macrophage acidic compartments and showed evidences of wall disruption. Sporozoites were observed in macrophages containing oocyst remnants or in new macrophages, giving rise to dividing tachyzoites. All together, these results highlight an unexpected role of phagocytic cells in processing T. gondii oocysts, in line with non-classical routes of infection, and open new perspectives to identify chemical factors that lead to oocyst wall disruption under physiological conditions. 
PMID:
 
27641141
 
DOI:
 
10.1038/srep33654

Thursday, September 15, 2016

Binding of Toxoplasma gondii AMA1 to RON2 during Invasion Protects AMA1 from Rhomboid-Mediated Cleavage and Leads to Dephosphorylation of Its Cytosolic Tail

 2016 Sep 13;7(5). pii: e00754-16. doi: 10.1128/mBio.00754-16.

Abstract

Apical membrane antigen 1 (AMA1) is a receptor protein on the surface of Toxoplasma gondii that plays a critical role in host cell invasion. The ligand to which T gondii AMA1 (TgAMA1) binds, TgRON2, is secreted into the host cell membrane by the parasite during the early stages of invasion. The TgAMA1-TgRON2 complex forms the core of the "moving junction," a ring-shaped zone of tight contact between the parasite and host cell membranes, through which the parasite pushes itself during invasion. Paradoxically, the parasite also expresses rhomboid proteases that constitutively cleave the TgAMA1 transmembrane domain. How can TgAMA1 function effectively in host cell binding if its extracellular domain is constantly shed from the parasite surface? We show here that when TgAMA1 binds the domain 3 (D3) peptide of TgRON2, its susceptibility to cleavage by rhomboid protease(s) is greatly reduced. This likely serves to maintain parasite-host cell binding at the moving junction, a hypothesis supported by data showing that parasites expressing a hypercleavable version of TgAMA1 invade less efficiently than wild-type parasites do. Treatment of parasites with the D3 peptide was also found to reduce phosphorylation of S527 on the cytoplasmic tail of TgAMA1, and parasites expressing a phosphomimetic S527D allele of TgAMA1 showed an invasion defect. Taken together, these data suggest that TgAMA1-TgRON2 interaction at the moving junction protects TgAMA1 molecules that are actively engaged in host cell penetration from rhomboid-mediated cleavage and generates an outside-in signal that leads to dephosphorylation of the TgAMA1 cytosolic tail. Both of these effects are required for maximally efficient host cell invasion.

IMPORTANCE: 

Nearly one-third of the world's population is infected with the protozoan parasite Toxoplasma gondii, which causes life-threatening disease in neonates and immunocompromised individuals. T. gondii is a member of the phylum Apicomplexa, which includes many other parasites of veterinary and medical importance, such as those that cause coccidiosis, babesiosis, and malaria. Apicomplexan parasites grow within their hosts through repeated cycles of host cell invasion, parasite replication, and host cell lysis. Parasites that cannot invade host cells cannot survive or cause disease. AMA1 is a highly conserved protein on the surface of apicomplexan parasites that is known to be important for invasion, and the work presented here reveals new and unexpected insights into AMA1 function. A more complete understanding of the role of AMA1 in invasion may ultimately contribute to the development of new chemotherapeutics designed to disrupt AMA1 function and invasion-related signaling in this important group of human pathogens.
Copyright © 2016 Krishnamurthy et al.
PMID:
 
27624124
 
DOI:
 
10.1128/mBio.00754-16
[PubMed - in process]

Tuesday, September 13, 2016

Huntingtons Disease Mice Infected with Toxoplasma gondii Demonstrate Early Kynurenine Pathway Activation, Altered CD8+ T-Cell Responses, and Premature Mortality

2016 Sep 9;11(9):e0162404. doi: 10.1371/journal.pone.0162404. eCollection 2016.


Huntington's disease (HD) is a progressive neurodegenerative disorder caused by a polyglutamine-repeat expansion in the huntingtin protein. Activation of the kynurenine pathway of tryptophan degradation is implicated in the pathogenesis of HD. Indoleamine-2,3-dioxygenase (IDO) catalyzes the oxidation of tryptophan to kynurenine, the first step in this pathway. The prevalent, neuroinvasive protozoal pathogen Toxoplasma gondii (T. gondii) results in clinically silent life-long infection in immune-competent individuals. T. gondii infection results in activation of IDO which provides some protection against the parasite by depleting tryptophan which the parasite cannot synthesize. The kynurenine pathway may therefore represent a point of synergism between HD and T. gondii infection. We show here that IDO activity is elevated at least four-fold in frontal cortex and striata of non-infected N171-82Q HD mice at 14-weeks corresponding to early-advanced HD. T. gondii infection at 5 weeks resulted in elevation of cortical IDO activity in HD mice. HD-infected mice died significantly earlier than wild-type infected and HD control mice. Prior to death, infected HD mice demonstrated decreased CD8+ T-lymphocyte proliferation in brain and spleen compared to wild-type infected mice. We demonstrate for the first time that HD mice have an altered response to an infectious agent that is characterized by premature mortality, altered immune responses and early activation of IDO. Findings are relevant to understanding how T. gondii infection may interact with pathways mediating neurodegeneration in HD.

Thursday, September 08, 2016

A Review of Experimental Compounds Demonstrating Anti-Toxoplasma Activity

 2016 Sep 6. pii: AAC.01176-16. [Epub ahead of print]

Abstract

Toxoplasma gondii is a ubiquitous apicomplexan parasite capable of infecting humans and other animals. Current treatment options for T. gondii infection are limited and most have drawbacks, including high toxicity and low tolerability. Additionally, no FDA-approved treatments are available for pregnant women, a high-risk population due to transplacental infection. Therefore, the development of novel treatment options is needed. To aid this effort, this review highlights experimental compounds that, at a minimum, demonstrate inhibition of in vitro growth of T. gondii When available, host cell toxicity and in vivo data is also discussed. The purpose of this review is to facilitate additional development of anti-Toxoplasmacompounds, and potentially to extend our knowledge of the parasite.
Copyright © 2016, American Society for Microbiology. All Rights Reserved.
PMID:
 
27600037
 
DOI:
 
10.1128/AAC.01176-16
[PubMed - as supplied by publisher]

Wednesday, September 07, 2016

A Genome-wide CRISPR Screen in Toxoplasma Identifies Essential Apicomplexan Genes

2016 Sep 1. pii: S0092-8674(16)31070-4. doi: 10.1016/j.cell.2016.08.019. [Epub ahead of print]


Apicomplexan parasites are leading causes of human and livestock diseases such as malaria and toxoplasmosis, yet most of their genes remain uncharacterized. Here, we present the first genome-wide genetic screen of an apicomplexan. We adapted CRISPR/Cas9 to assess the contribution of each gene from the parasite Toxoplasma gondii during infection of human fibroblasts. Our analysis defines ∼200 previously uncharacterized, fitness-conferring genes unique to the phylum, from which 16 were investigated, revealing essential functions during infection of human cells. Secondary screens identify as an invasion factor the claudin-like apicomplexan microneme protein (CLAMP), which resembles mammalian tight-junction proteins and localizes to secretory organelles, making it critical to the initiation of infection. CLAMP is present throughout sequenced apicomplexan genomes and is essential during the asexual stages of the malaria parasite Plasmodium falciparum. These results provide broad-based functional information on T. gondii genes and will facilitate future approaches to expand the horizon of antiparasitic interventions.
Copyright © 2016 Elsevier Inc. All rights reserved.

KEYWORDS:

Apicomplexan parasites; eukaryotic pathogen; genome-wide CRISPR screen; host-cell invasion; host-pathogen interactions; malaria; toxoplasmosis
[PubMed - as supplied by publisher]

Toxoplasma gondii and Epilepsy

2016 Jun;40(2):90-6. doi: 10.5152/tpd.2016.4708.


Toxoplasma gondii is a zoonotic parasite can be seen in all the vital organ; in the acute phase, it can be found in the blood, cerebrospinal fluid, semen, tears, saliva, urine, and in almost all body fluids. Transplasental infection can lead to fetal damage and miscarriage. Its last hosts are felines and intermediate hosts are all mammals, including humans. People infected by the ingestion of meat containing cysts in undercooked or raw, are thrown oocysts with cat felines By taking in water and food, from mother to fetus transplacental way, the infected organ transplantation, blood transfusion, laboratory accidents and kaprofaj transmitted by mechanical vectors of the invertebrates. Suppression of the immune system is being transformed to the shape and texture of the cysts with bradyzoite. The parasite settles in the cells of the tissue cysts and causes change in the cellular mechanisms, such as cytokinin task. Depending on changes and type of neurotransmitter (GABA, glutamate, serotonin, dopamine) levels in CSF in ions (Ca, K, Cl, Mg), it is believed that there is a change in their concentration. In this review, literature about the relationship between T. gondii and epilepsy and epileptiform activity the importance of parasites, which settle in the brain, will be highlighted.
[PubMed - in process]

Wednesday, August 31, 2016

Reassessing the mechanics of parasite motility and host-cell invasion

2016 Aug 29;214(5):507-15. doi: 10.1083/jcb.201605100.


The capacity to migrate is fundamental to multicellular and single-celled life. Apicomplexan parasites, an ancient protozoan clade that includes malaria parasites (Plasmodium) and Toxoplasma, achieve remarkable speeds of directional cell movement. This rapidity is achieved via a divergent actomyosin motor system, housed within a narrow compartment that lies underneath the length of the parasite plasma membrane. How this motor functions at a mechanistic level during motility and host cell invasion is a matter of debate. Here, we integrate old and new insights toward refining the current model for the function of this motor with the aim of revitalizing interest in the mechanics of how these deadly pathogens move.
© 2016 Tardieux and Baum.

Repurposing of conserved autophagy-related protein ATG8 in a divergent eukaryote

2016 Jul 1;9(4):e1197447. doi: 10.1080/19420889.2016.1197447.

Abstract

Toxoplasma gondii and other apicomplexan parasites contain a peculiar non-photosynthetic plastid called the apicoplast, which is essential for their survival. The localization of autophagy-related protein ATG8 to the apicoplast in several apicomplexan species and life stages has recently been described, and we have shown this protein is essential for proper inheritance of this complex plastid into daughter cells during cell division. Although the mechanism behind ATG8 association to the apicoplast in T. gondii is related to the canonical conjugation system leading to autophagosome formation, its singular role seems independent from the initial catabolic purpose of autophagy. Here we also discuss further the functional evolution and innovative adaptations of the autophagy machinery to maintain this organelle during parasite division.

KEYWORDS:

ATG8; Toxoplasma; apicomplexa; apicoplast; non canonical autophagy; plastid