A Toxoplasma Patatin-Like Protein Changes Localization and Alters the Cytokine Response during Toxoplasmic Encephalitis

ABSTRACTToxoplasma gondiiis an obligate intracellular parasite that forms a lifelong infection within the central nervous system of its host. TheT. gondiigenome encodes six members of the patatin-like phospholipase family; related proteins are associated with host-microbe interactions in bacteria.T. gondiipatatin-like protein 1 (TgPL1) was previously determined to be necessary for parasites to suppress nitric oxide and prevent degradation in activated macrophages. Here, we show that in the rapidly replicating tachyzoite stage, TgPL1 is localized within vesicles inside the parasite that are distinct from the dense granules; however, in the encysted bradyzoite stage, TgPL1 localizes to the parasitophorous vacuole (PV) and cyst wall. While we had not previously seen a defect of the TgPL1 deletion mutant (ΔTgPL1) during acute and early chronic infection, the localization change of TgPL1 in bradyzoites caused us to reevaluate the ΔTgPL1 mutant during late chronic infection and in a toxoplasmic encephalitis (TE) mouse model. Mice infected with ΔTgPL1 are more resistant to TE and have fewer inflammatory lesions than mice infected with the wild type and ΔTgPL1 genetically complemented withTgPL1. This increased resistance to TE could result from several contributing factors. First, we found that ΔTgPL1 bradyzoites did not convert back to tachyzoites readily in tissue culture. Second, a subset of cytokine levels were higher in ΔTgPL1-infected mice, including gamma interferon (IFN-γ), tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6), and monocyte chemotactic protein 1 (MCP-1). These studies suggest that TgPL1 plays a role in the maintenance of chronicT. gondiiinfection.

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A Family of Toxoplasma gondii Genes Related to GRA12 Regulate Cyst Burdens and Cyst Reactivation

mSphere ◽

10.1128/msphere.00182-21 ◽

2021 ◽

Vol 6 (2) ◽

Author(s):

Rebekah B. Guevara ◽

Barbara A. Fox ◽

David J. Bzik

Keyword(s):

Toxoplasma Gondii ◽

Cyst Wall ◽

Chronic Infection ◽

Acute Infection ◽

Dense Granule ◽

Toxoplasmic Encephalitis ◽

Content Type ◽

The Central Nervous System ◽

Related Proteins ◽

Cyst Development

ABSTRACT Toxoplasma gondii causes a chronic infection that renders the immunocompromised human host susceptible to toxoplasmic encephalitis triggered by cyst reactivation in the central nervous system. The dense granule protein GRA12 is a major parasite virulence factor required for parasite survival during acute infection. Here, we characterized the role of four GRA12-related genes in acute and chronic stages of infection. While GRA12A, GRA12B, and GRA12D were highly expressed in asexual stage tachyzoites and bradyzoites, expression of GRA12C appeared to be restricted to the sexual stages. In contrast to deletion of GRA12 (Δgra12), no major defects in acute virulence were observed in Δgra12A, Δgra12B, or Δgra12D parasites, though Δgra12B parasites exhibited an increased tachyzoite replication rate. Bradyzoites secreted GRA12A, GRA12B, and GRA12D and incorporated these molecules into the developing cyst wall, as well as the cyst matrix in distinct patterns. Similar to GRA12, GRA12A, GRA12B, and GRA12D colocalized with the dense granules in extracellular tachyzoites, with GRA2 and the intravacuolar network in the tachyzoite stage parasitophorous vacuole and with GRA2 in the cyst matrix and cyst wall. Chronic stage cyst burdens were decreased in mice infected with Δgra12A parasites and were increased in mice infected with Δgra12B parasites. However, Δgra12B cysts were not efficiently maintained in vivo. Δgra12A, Δgra12B, and Δgra12D in vitro cysts displayed a reduced reactivation efficiency, and reactivation of Δgra12A cysts was delayed. Collectively, our results suggest that a family of genes related to GRA12 play significant roles in the formation, maintenance, and reactivation of chronic stage cysts. IMPORTANCE If host immunity weakens, Toxoplasma gondii cysts recrudesce in the central nervous system and cause a severe toxoplasmic encephalitis. Current therapies target acute stage infection but do not eliminate chronic cysts. Parasite molecules that mediate the development and persistence of chronic infection are poorly characterized. Dense granule (GRA) proteins such as GRA12 are key virulence factors during acute infection. Here, we investigated four GRA12-related genes. GRA12-related genes were not major virulence factors during acute infection. Instead, GRA12-related proteins localized at the cyst wall and cyst matrix and played significant roles in cyst development, persistence, and reactivation during chronic infection. Similar to GRA12, the GRA12-related proteins selectively associated with the intravacuolar network of membranes inside the vacuole. Collectively, our results support the hypothesis that GRA12 proteins associated with the intravacuolar membrane system support parasite virulence during acute infection and cyst development, persistence, and reactivation during chronic infection.

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Neighbors Working Together: a Toxoplasma Rhoptry Protein That Facilitates Dense Granule Protein Translocation into the Host Cell

mSphere ◽

10.1128/msphere.00523-19 ◽

2019 ◽

Vol 4 (4) ◽

Author(s):

Gustavo Arrizabalaga

Keyword(s):

Host Cell ◽

Protein Translocation ◽

Parasitophorous Vacuole ◽

Opportunistic Pathogen ◽

Dense Granule ◽

Effector Proteins ◽

Content Type ◽

Dense Granules ◽

Cell Functions ◽

Host Signaling

ABSTRACT The opportunistic pathogen Toxoplasma gondii is highly adept at manipulating host cell functions. While inside a host cell, Toxoplasma divides within a parasitophorous vacuole from which it secretes numerous effector proteins from its dense granules. Many of these so-called GRA proteins are translocated from the parsitophorous vacuole into the host cell where they directly disrupt host signaling pathways. The machinery that drives the translocation of GRA proteins across the parasitophorous vacuole membrane is being elucidated through both genetic and biochemical approaches. A new mSphere research article (M. W. Panas, A. Ferrel, A. Naor, E. Tenborg, et al., mSphere 4:e00276-19, 2019, https://doi.org/10.1128/mSphere.00276-19) describes how the kinase ROP17, which is secreted from the parasite’s rhoptries into the host cell during invasion, regulates the translocation of GRA effectors.

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Multifaceted Role of Heme during Severe Plasmodium falciparum Infections in India

Infection and Immunity ◽

10.1128/iai.00531-15 ◽

2015 ◽

Vol 83 (10) ◽

pp. 3793-3799 ◽

Cited By ~ 12

Author(s):

Esther Dalko ◽

Bidyut Das ◽

Fabien Herbert ◽

Constantin Fesel ◽

Sulabha Pathak ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Severe Malaria ◽

Rank Correlation ◽

Interleukin 10 ◽

Necrosis Factor Alpha ◽

Content Type ◽

Canonical Correlations ◽

Cytokines And Chemokines ◽

Chemotactic Protein ◽

Mild Malaria

Several immunomodulatory factors are involved in malaria pathogenesis. Among them, heme has been shown to play a role in the pathophysiology of severe malaria in rodents, but its role in human severe malaria remains unclear. Circulating levels of total heme and its main scavenger, hemopexin, along with cytokine/chemokine levels and biological parameters, including hemoglobin and creatinine levels, as well as transaminase activities, were measured in the plasma of 237Plasmodium falciparum-infected patients living in the state of Odisha, India, where malaria is endemic. All patients were categorized into well-defined groups of mild malaria, cerebral malaria (CM), or severe noncerebral malaria, which included acute renal failure (ARF) and hepatopathy. Our results show a significant increase in total plasma heme levels with malaria severity, especially for CM and malarial ARF. Spearman rank correlation and canonical correlation analyses have shown a correlation between total heme, hemopexin, interleukin-10, tumor necrosis factor alpha, gamma interferon-induced protein 10 (IP-10), and monocyte chemotactic protein 1 (MCP-1) levels. In addition, canonical correlations revealed that heme, along with IP-10, was associated with the CM pathophysiology, whereas both IP-10 and MCP-1 together with heme discriminated ARF. Altogether, our data indicate that heme, in association with cytokines and chemokines, is involved in the pathophysiology of both CM and ARF but through different mechanisms.

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VCAM-1/α4β1 Integrin Interaction Is Crucial for Prompt Recruitment of Immune T Cells into the Brain during the Early Stage of Reactivation of Chronic Infection with Toxoplasma gondii To Prevent Toxoplasmic Encephalitis

Infection and Immunity ◽

10.1128/iai.01494-13 ◽

2014 ◽

Vol 82 (7) ◽

pp. 2826-2839 ◽

Cited By ~ 21

Author(s):

Qila Sa ◽

Eri Ochiai ◽

Tomoko Sengoku ◽

Melinda E. Wilson ◽

Morgan Brogli ◽

...

Keyword(s):

T Cells ◽

Toxoplasma Gondii ◽

Chronic Infection ◽

Early Stage ◽

Toxoplasmic Encephalitis ◽

Mrna Levels ◽

Wild Type ◽

Content Type ◽

Ifn Γ ◽

The Brain

ABSTRACTReactivation of chronic infection withToxoplasma gondiican cause life-threatening toxoplasmic encephalitis in immunocompromised individuals. We examined the role of VCAM-1/α4β1 integrin interaction in T cell recruitment to prevent reactivation of the infection in the brain. SCID mice were infected and treated with sulfadiazine to establish a chronic infection. VCAM-1 and ICAM-1 were the endothelial adhesion molecules detected on cerebral vessels of the infected SCID and wild-type animals. Immune T cells from infected wild-type mice were treated with anti-α4 integrin or control antibodies and transferred into infected SCID or nude mice, and the animals received the same antibody every other day. Three days later, sulfadiazine was discontinued to initiate reactivation of infection. Expression of mRNAs for CD3δ, CD4, CD8β, gamma interferon (IFN-γ), and inducible nitric oxide synthase (NOS2) (an effector molecule to inhibitT. gondiigrowth) and the numbers of CD4+and CD8+T cells in the brain were significantly less in mice treated with anti-α4 integrin antibody than in those treated with control antibody at 3 days after sulfadiazine discontinuation. At 6 days after sulfadiazine discontinuation, cerebral tachyzoite-specific SAG1 mRNA levels and numbers of inflammatory foci associated with tachyzoites were markedly greater in anti-α4 integrin antibody-treated than in control antibody-treated animals, even though IFN-γ and NOS2 mRNA levels were higher in the former than in the latter. These results indicate that VCAM-1/α4β1 integrin interaction is crucial for prompt recruitment of immune T cells and induction of IFN-γ-mediated protective immune responses during the early stage of reactivation of chronicT. gondiiinfection to control tachyzoite growth.

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Toxoplasma gondiiParasitophorous Vacuole Membrane-Associated Dense Granule Proteins Orchestrate Chronic Infection and GRA12 Underpins Resistance to Host Gamma Interferon

mBio ◽

10.1128/mbio.00589-19 ◽

2019 ◽

Vol 10 (4) ◽

Cited By ~ 15

Author(s):

Barbara A. Fox ◽

Rebekah B. Guevara ◽

Leah M. Rommereim ◽

Alejandra Falla ◽

Valeria Bellini ◽

...

Keyword(s):

Chronic Infection ◽

Parasitophorous Vacuole ◽

Dense Granule ◽

Gamma Interferon ◽

Type I ◽

Type Ii ◽

Content Type ◽

Vacuole Membrane ◽

Ifn Γ

ABSTRACTToxoplasma gondiievades host immunity to establish a chronic infection. Here, we assessed the role of parasitophorous vacuole (PV) membrane (PVM)- and intravacuolar network (IVN) membrane-localized dense granule (GRA) proteins in the development of acute and chronicToxoplasmainfection. Deletion of PVM-associated GRA3, GRA7, GRA8, and GRA14 or IVN membrane-associated GRA2, GRA9, and GRA12 in the low-virulence type II Prugniaud (Pru) strain induced severe defects in the development of chronic-stage cystsin vivowithout affecting the parasite growth rate or the ability to differentiate into cystsin vitro. Acute virulence of the PruΔgra2, PruΔgra3, and PruΔgra4mutants was reduced but not abolished. In contrast, the PruΔgra12mutant was avirulent in mice and PruΔgra12parasites failed to establish a chronic infection. High-virulence type I strain RHΔgra12parasites also exhibited a major defect in acute virulence. In gamma interferon (IFN-γ)-activated macrophages, type I RHΔgra12and type II PruΔgra12parasites resisted the coating of the PVM with host immunity-related GTPases as effectively as the parental type I RHΔku80and type II PruΔku80strains, respectively. Despite this resistance, Δgra12PVs ultimately succumbed to IFN-γ-activated host cell innate immunity. Our findings uncover a key role for GRA12 in mediating resistance to host IFN-γ and reveal that many other IVN membrane-associated GRA proteins, as well as PVM-localized GRA proteins, play important roles in establishing chronic infection.IMPORTANCEToxoplasma gondiicysts reactivate during immune deficiency and cause fatal encephalitis. Parasite molecules that coordinate the development of acute and chronic infection are poorly characterized. Here, we show that many intravacuolar network membrane and parasitophorous vacuole membrane-associated dense granule (GRA) proteins orchestrate the development of chronic cystsin vivo. A subset of these GRA proteins also modulate acute virulence, and one protein that associates with the intravacuolar network membranes, namely GRA12, was identified as a major virulence factor required for parasite resistance to host gamma interferon (IFN-γ). Our results revealed that many parasitophorous vacuole membrane and intravacuolar network membrane-associated GRA proteins are essential for successful chronic infection.

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Discerning the Role of Bacteroides fragilis in Celiac Disease Pathogenesis

Applied and Environmental Microbiology ◽

10.1128/aem.00563-12 ◽

2012 ◽

Vol 78 (18) ◽

pp. 6507-6515 ◽

Cited By ~ 46

Author(s):

E. Sánchez ◽

J. M. Laparra ◽

Y. Sanz

Keyword(s):

Celiac Disease ◽

Virulence Genes ◽

Cell Monolayer ◽

Bacteroides Fragilis ◽

Necrosis Factor Alpha ◽

Gelatinase Activity ◽

Content Type ◽

Immunogenic Peptides ◽

Host Microbe Interactions

ABSTRACTCeliac disease (CD) is associated with intestinal dysbiosis, which can theoretically lead to dysfunctions in host-microbe interactions and contribute to the disease. In the present study, possible differences inBacteroidesspp. and their pathogenic features between CD patients and controls were investigated.Bacteroidesclones (n= 274) were isolated, identified, and screened for the presence of the virulence genes (bftandmpII) coding for metalloproteases. The proteolytic activity of selectedBacteroides fragilisstrains was evaluated by zymography and, after gastrointestinal digestion of gliadin, by high-pressure liquid chromatography/electrospray ionization/tandem mass spectrometry. The effects ofB. fragilisstrains on Caco-2 cell culture permeability and inflammatory response to digested gliadin were determined.B. fragiliswas more frequently identified in CD patients than in healthy controls, in contrast toBacteroides ovatus.B. fragilisclones carrying virulence genes coding for metalloproteases were more abundant in CD patients than in controls.B. fragilisstrains, representing the isolated clones and carrying metalloprotease genes, showed gelatinase activity and exerted the strongest adverse effects on the integrity of the Caco-2 cell monolayer. AllB. fragilisstrains also showed gliadin-hydrolyzing activity, and some of them generated immunogenic peptides that preserved or increased inflammatory cytokine production (tumor necrosis factor alpha) and showed increased ability to permeate through Caco-2 cell cultures. These findings suggest that increased abundance ofB. fragilisstrains with metalloprotease activities could play a role in CD pathogenesis, although furtherin vivostudies are required to support this hypothesis.

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Succinylated Wheat Germ Agglutinin Colocalizes with the Toxoplasma gondii Cyst Wall Glycoprotein CST1

mSphere ◽

10.1128/msphere.00031-20 ◽

2020 ◽

Vol 5 (2) ◽

Author(s):

Rebekah B. Guevara ◽

Barbara A. Fox ◽

David J. Bzik

Keyword(s):

Toxoplasma Gondii ◽

Cyst Wall ◽

Wheat Germ Agglutinin ◽

Wheat Germ ◽

Parasitophorous Vacuole ◽

Biological Information ◽

Toxoplasmic Encephalitis ◽

Content Type ◽

Cyst Development

ABSTRACT The glycosylated mucin domain of the Toxoplasma gondii cyst wall glycoprotein CST1 is heavily stained by Dolichos biflorus agglutinin, a lectin that binds to N-acetylgalactosamine. The cyst wall is also heavily stained by the chitin binding lectin succinylated wheat germ agglutinin (s-WGA), which selectively binds to N-acetylglucosamine-decorated structures. Here, we tracked the localization of N-acetylglucosamine-decorated structures that bind to s-WGA in immature and mature in vitro cysts. s-WGA localization was observed at the cyst periphery 6 h after the differentiation of the tachyzoite-stage parasitophorous vacuole. By day 1 and at all later times after differentiation, s-WGA was localized in a continuous staining pattern at the cyst wall. Coinciding with the maturation of the cyst matrix by day 3 of cyst development, s-WGA also localized in a continuous matrix pattern inside the cyst. s-WGA localized in both the outer and inner layer regions of the cyst wall and in a continuous matrix pattern inside mature 7- and 10-day-old cysts. In addition, s-WGA colocalized in the cyst wall with CST1, suggesting that N-acetylglucosamine- and N-acetylgalactosamine-decorated molecules colocalized in the cyst wall. In contrast to CST1, GRA4, and GRA6, the relative accumulation of the molecules that bind s-WGA in the cyst wall was not dependent on the expression of GRA2. Our results suggest that GRA2-dependent and GRA2-independent mechanisms regulate the trafficking and accumulation of glycosylated molecules that colocalize in the cyst wall. IMPORTANCE Chronic Toxoplasma gondii infection is maintained in the central nervous system by thick-walled cysts. If host immunity wanes, cysts recrudesce and cause severe and often lethal toxoplasmic encephalitis. Currently, there are no therapies to eliminate cysts, and little biological information is available regarding cyst structure(s). Here, we investigated cyst wall molecules recognized by succinylated wheat germ agglutinin (s-WGA), a lectin that specifically binds to N-acetylglucosamine-decorated structures. N-Acetylglucosamine regulates cell signaling and plays structural roles at the cell surface in many organisms. The cyst wall and cyst matrix were heavily stained by s-WGA in mature cysts and were differentially stained during cyst development. The relative accumulation of molecules that bind to s-WGA in the cyst wall was not dependent on the expression of GRA2. Our findings suggest that glycosylated cyst wall molecules gain access to the cyst wall via GRA2-dependent and GRA2-independent mechanisms and colocalize in the cyst wall.

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Additive Manufacturing of Astragaloside-Containing Polyurethane Nerve Conduits Influenced Schwann Cell Inflammation and Regeneration

Processes ◽

10.3390/pr9020353 ◽

2021 ◽

Vol 9 (2) ◽

pp. 353

Author(s):

Yueh-Sheng Chen ◽

Shih-Sheng Chang ◽

Hooi Yee Ng ◽

Yu-Xuan Huang ◽

Chien-Chang Chen ◽

...

Keyword(s):

Nervous System ◽

Cellular Proliferation ◽

Neural Regeneration ◽

Nerve Grafting ◽

Necrosis Factor Alpha ◽

Digital Light Processing ◽

Nerve Conduits ◽

Tnf Α ◽

The Central Nervous System ◽

Factor Alpha

The peripheral nervous system is the bridge of communication between the central nervous system and other body systems. Autologous nerve grafting is the mainstream method for repair of nerve lesions greater than 20 mm. However, there are several disadvantages and limitations of autologous nerve grafting, thus prompting the need for fabrication of nerve conduits for clinical use. In this study, we successfully fabricated astragaloside (Ast)-containing polyurethane (PU) nerve guidance conduits via digital light processing, and it was noted that the addition of Ast improved the hydrophilicity of traditional PU conduits by at least 23%. The improved hydrophilicity not only led to enhanced cellular proliferation of rat Schwann cells, we also noted that levels of inflammatory markers tumor necrosis factor-alpha (TNF-α) and cyclooxygenase-2 (COX-2) significantly decreased with increasing concentrations of Ast. Furthermore, the levels of neural regeneration markers were significantly enhanced with the addition of Ast. This study demonstrated that Ast-containing PU nerve conduits can be potentially used as an alternative solution to regenerate peripheral nerve injuries.

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Spinal cord arteriovenous malformation with an associated lymphatic anomaly

Journal of Neurosurgery ◽

10.3171/jns.1990.73.5.0768 ◽

1990 ◽

Vol 73 (5) ◽

pp. 768-773 ◽

Cited By ~ 4

Author(s):

Gary E. Kraus ◽

Richard D. Bucholz ◽

Thomas R. Weber

Keyword(s):

Spinal Cord ◽

Lymphatic System ◽

Vascular Lesions ◽

Dietary Manipulation ◽

Paravertebral Muscle ◽

Content Type ◽

Muscle Fascia ◽

Spinal Osteomyelitis ◽

The Central Nervous System ◽

Spinal Cord Arteriovenous Malformation

✓ Spinal cord arteriovenous malformations (AVM's), like other vascular anomalies of the central nervous system, can be associated with similar vascular lesions of the skin and viscera. A 7-year-old girl, who presented with rapidly progressing paraplegia, was found to have a spinal cord AVM, cutaneous angioma, and a chylous malformation of the lymphatic system. She had previously undergone treatment for a posterior thoracic cutaneous angioma. At surgery, upon incision of the paravertebral muscle fascia, viscous pale fluid was encountered emanating from a foramen in the thoracic lamina. The spinal AVM was resected in spite of concern that the abnormality represented spinal osteomyelitis. Postoperatively, there was full return of function in the lower extremities, along with recurrent episodes of chylothorax, which slowly came under control with dietary manipulation. A review of the anatomy of the thoracic duct and nontraumatic causes of chylothorax is presented, and the association of cutaneous and central angiomas is discussed. Finally, the treatment of chylothorax is delineated.

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Changes in Antigen-Specific Cytokine and Chemokine Responses to Plasmodium falciparum Antigens in a Highland Area of Kenya after a Prolonged Absence of Malaria Exposure

Infection and Immunity ◽

10.1128/iai.01924-14 ◽

2014 ◽

Vol 82 (9) ◽

pp. 3775-3782 ◽

Cited By ~ 6

Author(s):

Lyticia A. Ochola ◽

Cyrus Ayieko ◽

Lily Kisia ◽

Ng'wena G. Magak ◽

Estela Shabani ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Severe Disease ◽

Clinical Malaria ◽

Necrosis Factor Alpha ◽

Content Type ◽

Highland Area ◽

Cytokine Responses ◽

Increased Risk ◽

Tnf Α ◽

Ifn Γ

ABSTRACTIndividuals naturally exposed toPlasmodium falciparumlose clinical immunity after a prolonged lack of exposure.P. falciparumantigen-specific cytokine responses have been associated with protection from clinical malaria, but the longevity ofP. falciparumantigen-specific cytokine responses in the absence of exposure is not well characterized. A highland area of Kenya with low and unstable malaria transmission provided an opportunity to study this question. The levels of antigen-specific cytokines and chemokines associated in previous studies with protection from clinical malaria (gamma interferon [IFN-γ], interleukin-10 [IL-10], and tumor necrosis factor alpha [TNF-α]), with increased risk of clinical malaria (IL-6), or with pathogenesis of severe disease in malaria (IL-5 and RANTES) were assessed by cytometric bead assay in April 2008, October 2008, and April 2009 in 100 children and adults. During the 1-year study period, none had an episode of clinicalP. falciparummalaria. Two patterns of cytokine responses emerged, with some variation by antigen: a decrease at 6 months (IFN-γ and IL-5) or at both 6 and 12 months (IL-10 and TNF-α) or no change over time (IL-6 and RANTES). These findings document thatP. falciparumantigen-specific cytokine responses associated in prior studies with protection from malaria (IFN-γ, TNF-α, and IL-10) decrease significantly in the absence ofP. falciparumexposure, whereas those associated with increased risk of malaria (IL-6) do not. The study findings provide a strong rationale for future studies of antigen-specific IFN-γ, TNF-α, and IL-10 responses as biomarkers of increased population-level susceptibility to malaria after prolonged lack ofP. falciparumexposure.

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