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Introduction
Why do some individuals get sick after infection while others do not? We believe that this is determined by two major factors: 1) Some parasite strains are more virulent than others and can therefore cause more disease, 2) Some individuals are more susceptible than others and therefore get more sick. We use the obligate intracellular parasite Toxoplasma gondii to study the molecular basis for strain and species differences in susceptibility to infection and parasite differences in virulence.
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Education
January 1998 - September 2002
September 1992 - September 1997
Publications
Publications (207)
The obligate intracellular parasite Toxoplasma gondii causes life-threatening toxoplasmosis to immunocompromised individuals. The pathogenesis of Toxoplasma relies on its swift dissemination to the central nervous system through a ‘Trojan Horse’ mechanism using infected leukocytes as carriers. Previous work found Tg WIP, a protein secreted from Tox...
Toxoplasma gondii is a parasite that replicates within a specialized compartment called the parasitophorous vacuole (PV), which is surrounded by the PV membrane (PVM). To obtain essential nutrients, Toxoplasma must transport molecules across the PVM, a process mediated by the secreted parasite proteins GRA17 and GRA23. These proteins form pores in...
Apicomplexan parasites have specialized secretory organelles called rhoptries, micronemes, and dense granules that are essential for host infection. Here, we show that TgREMIND, a Toxoplasma gondii protein containing a membrane phospholipid interacting domain, is required for the biogenesis of rhoptries and dense granules. TgREMIND contains a Fes/C...
Toxoplasma virulence depends on its ability to evade or survive the toxoplasmacidal mechanisms induced by interferon gamma (IFNγ). While many Toxoplasma genes involved in the evasion of the murine IFNγ response have been identified, genes required to survive the human IFNγ response are largely unknown. In this study, we used a genome-wide loss-of-f...
Toxoplasma gondii oocysts, which are shed in large quantities in the feces from infected felines, are very stable in the environment, resistant to most inactivation procedures, and highly infectious. The oocyst wall provides an important physical barrier for sporozoites contained inside oocysts, protecting them from many chemical and physical stres...
An adult Indian ringneck parakeet (Psittacula krameri manillensis) from an outdoor aviary in Sacramento, California was found dead on the nest box. Postmortem examination showed firm, enlarged, yellow‑tinged liver and splenomegaly. Multifocal to coalescing, acute necrosis with macrophages, lymphocytes, plasma cells, and periportal ductular reaction...
Upon pathogen detection, macrophages normally stay sessile in tissues while dendritic cells (DCs) migrate to secondary lymphoid tissues. The obligate intracellular protozoan Toxoplasma gondii exploits the trafficking of mononuclear phagocytes for dissemination via unclear mechanisms. We report that, upon T. gondii infection, macrophages initiate th...
The intraspecific variability among Neospora caninum isolates in their in vitro behaviour and in vivo virulence has been widely studied. In particular, transcriptomic and proteomic analyses have shown a higher expression/abundance of specific genes/proteins in high-virulence isolates. Consequently, the dense granule protein NcGRA7 and the rhoptry p...
The obligate intracellular parasite Toxoplasma gondii makes use of infected leukocytes for systemic dissemination. Yet, how infection impacts the processes of leukocyte diapedesis has remained unresolved. Here, we addressed the effects of T. gondii infection on the trans-endothelial migration (TEM) of dendritic cells (DCs) across polarised brain en...
Toxoplasmosis is a worldwide disease affecting all warm-blooded animals, including humans. Vaccination strategies aimed at inducing an efficient immune response while preventing transmission have been attempted in the past. While many different approaches can partially protect immunized animals against subsequent infections, full and lasting protec...
Outbreaks of neurological disease associated with Sarcocystis calchasi have been observed in captive and free-ranging rock pigeons (Columba livia) in Europe and the United States as well as in wild Brandt's cormorants (Phalacrocorax penicillatus) and captive psittacines in California, USA. Experimental and field studies have identified northern gos...
Toxoplasmic encephalitis can develop in individuals infected with the protozoan parasite Toxoplasma gondii and is typified by parasite replication and inflammation within the brain. Patients often present with seizures, but the parasite genes and host pathways involved in seizure development and/or propagation are unknown. We previously reported th...
The virulence of eukaryotic parasites like Toxoplasma gondii depends on their capacity to escape from the host immune response and disseminate throughout the host organism. However, Toxoplasma gene products essential for its in vivo pathogenesis remain uncharacterized. Here, we present the complete workflow of a CRISPR-Cas9 in vivo loss-of-function...
Our studies on novel cyst wall proteins serendipitously led us to the discovery that cyst wall and vacuolar matrix protein MAG1, first identified a quarter of a century ago, functions as a secreted immunomodulatory effector. MAG1 is a dense granular protein that is found in the parasitophorous vacuolar matrix in tachyzoite vacuoles and the cyst wal...
The severity of toxoplasmosis depends on a combination of host and parasite factors. Among them, the Toxoplasma strain causing the infection is an important determinant of the disease outcome. Type 2 strains dominate in Europe, whereas in North America type 2, followed by type 3 and 12 strains are commonly isolated from wildlife and patients. To id...
As an immune-privileged organ, the placenta can tolerate the introduction of
antigens without inducing a strong inflammatory response that would lead
to abortion. However, for the control of intracellular pathogens, a strong Th1
response characterized by the production of interferon-γ is needed. Thus, invasion
of the placenta by intracellular paras...
Macrophages play an essential role in the early immune response against Toxoplasma and are the cell type preferentially infected by the parasite in vivo. Interferon gamma (IFNγ) elicits a variety of anti-Toxoplasma activities in macrophages. Using a genome-wide CRISPR screen we identify 353 Toxoplasma genes that determine parasite fitness in naїve...
Toxoplasma gondii is an exceptionally successful parasite that infects a very broad host range, including humans, across the globe. The outcome of infection differs remarkably between hosts, ranging from acute death to sterile infection. These differential disease patterns are strongly influenced by both host- and parasite-specific genetic factors....
Host resistance to Toxoplasma gondii relies on CD8 T cell IFNγ responses, which if modulated by the host or parasite could influence chronic infection and parasite transmission between hosts. Since host-parasite interactions that govern this response are not fully elucidated, we investigated requirements for eliciting naïve CD8 T cell IFNγ response...
Many intracellular pathogens, including the protozoan parasite Toxoplasma gondii, live inside a vacuole that resides in the host cytosol. Vacuolar residence provides these pathogens with a defined niche for replication and protection from detection by host cytosolic pattern recognition receptors. However, the limiting membrane of the vacuole, which...
Between March and May 2019, wildlife rehabilitation centers along coastal southern California admitted increased numbers of Brandt's cormorants (Phalacrocorax penicillatus) with neurological disease including head tilt, nystagmus, torticollis, tremors, paresis, paralysis, and ataxia. Seven cormorants from Los Angeles County and one cormorant from O...
The murine innate immune response against Toxoplasma gondii is predominated by the interaction of TLR11/12 with Toxoplasma profilin. However, mice lacking Tlr11 or humans, who do not have functional TLR11 or TLR12, still elicit a strong innate immune response upon Toxoplasma infection. The parasite factors that determine this immune response are la...
The protozoan parasite Toxoplasma gondii lives inside a vacuole in the host cytosol where it is protected from host cytoplasmic innate immune responses. However, IFNγ-dependent cell-autonomous immunity can destroy the vacuole and the parasite inside. Toxoplasma strain differences in susceptibility to human IFNγ exist, but the Toxoplasma effector(s)...
The biological variability among Neospora caninum isolates has been widely shown, however, the molecular basis that determines this diversity has not been thoroughly elucidated to date. The latest studies have focused on a limited number of isolates. Therefore, the goal of the present study was to compare the proteome of a larger number of N. canin...
The protozoan parasite Toxoplasma gondii lives inside a vacuole in the host cytoplasm where it is protected from host cytoplasmic innate immune responses. However, IFNγ-dependent cell-autonomous immunity can destroy the vacuole and the parasite inside. Toxoplasma strain differences in susceptibility to human IFNγ exist but the Toxoplasma effector(s...
The murine innate immune response against Toxoplasma gondii is predominated by the interaction of TLR11/12 with Toxoplasma profilin. However, mice lacking Tlr11 or humans, who do not have functional TLR11 or TLR12, still elicit a strong innate immune response upon Toxoplasma infection. The parasite factors that determine this immune response are la...
Host resistance to Toxoplasma gondii relies on CD8 T cell IFNγ responses, which if modulated by the host or parasite could influence chronic infection and parasite transmission between hosts. Since host-parasite interactions that govern this response are not fully elucidated, we investigated requirements for eliciting naïve CD8 T cell IFNγ response...
Toxoplasma gondii chronically infects a quarter of the world's population, and its recrudescence can cause life-threatening disease in immunocompromised individuals and recurrent ocular lesions in the immunocompetent. Acute-stage tachyzoites differentiate into chronic-stage bradyzoites, which form intracellular cysts resistant to immune clearance a...
The obligate intracellular protozoan parasite Toxoplasma gondii can infect any nucleated cell from a warm-blooded host. However, its interaction with host macrophages plays a critical role in shaping the immune response during infection. Therefore, assessing Toxoplasma–macrophage interactions at a cellular level is important. In this chapter, we de...
Apicomplexan parasites have complex life cycles involving sexual replication in a single definitive host and asexual replication in a variety of alternative hosts. Toxoplasma gondii undergoes its sexual cycle only in cats, yet it also infects a wide range of other vertebrates where it propagates asexually. In Europe and North America the bulk of T....
Macrophages play an essential role in the early immune response against Toxoplasma and are the cell type preferentially infected by the parasite in vivo . Interferon gamma (IFNγ) elicits a variety of anti- Toxoplasma activities in macrophages. Using a genome-wide CRISPR screen we identified ∼130 Toxoplasma genes that determine parasite fitness in n...
The intracellular parasite Toxoplasma gondii can cause chronic infections in most warm-blooded animals, including humans. In the USA, strains belonging to four different Toxoplasma clonal lineages (types 1, 2, 3, and 12) are commonly isolated, whereas strains not belonging to these lineages are predominant in other continents such as South America....
Toxoplasma can reach distant organs, especially the brain, leading to a lifelong chronic phase. However, genes involved in related in vivo processes are currently unknown. Here, we use focused CRISPR libraries to identify Toxoplasma genes that affect in vivo fitness. We focus on TgWIP, whose deletion affects Toxoplasma dissemination to distant orga...
The Toxoplasma gondii parasitophorous vacuole membrane (PVM) offers protection from the host immune system but is also a barrier for uptake of nutrients from the host. Previously, we showed that GRA17 mediates the tachyzoite PVM permeability to small molecules. During the conversion from tachyzoites to encysted bradyzoites, the PVM become the cyst...
The protozoan parasite Toxoplasma gondii secretes proteins from specialized organelles, the rhoptries, and dense granules, which are involved in the modulation of host cell processes. Dense granule protein GRA15 activates the nuclear factor kappa B (NF-κB) pathway, which plays an important role in cell death, innate immunity, and inflammation. Exac...
Toxoplasma gondii chronically infects a quarter of the world's population, and its recrudescence can cause life-threatening disease in immunocompromised individuals and recurrent ocular lesions in the immunocompetent. Chronic stages are established by differentiation of rapidly replicating tachyzoites into slow-growing bradyzoites, which form intra...
Inflammasomes are major components of the innate immune system and are responsible for detecting various microbial and environmental danger signals. Upon invasion of Lewis rat macrophages, the parasite rapidly activates the NLRP1 inflammasome, resulting in pyroptosis and elimination of the parasite’s replication niche. The work reported here reveal...
Transcript abundance, represented as fragments per kilobase exon per million reads (FPKM), in naïve and interferon gamma (IFNG)-stimulated bone-derived macrophages (BMDMs) from A/J, C57BL/6J, PWK/Phj, CAST/Eij, and WSB/Eij mouse strains. TNF, tumour necrosis factor alpha.
Additional South American (atypical) Toxoplasma strains (GUY-MAT, VAND, FOU, and GPHT) exhibit variable virulence in consomic mice. (A) Survival times of B6/PWD 11.1 consomic mice infected with different atypical Toxoplasma strains. (B) Molecular phylogenetic analysis of ROP17 in different Toxoplasma strains that are virulent (VAND,RUB and, FOU) or...
The severe virulence of Toxoplasma gondii in classical laboratory inbred mouse strains contradicts the hypothesis that house mice (Mus musculus) are the most important intermediate hosts for its transmission and evolution because death of the mouse before parasite transmission equals death of the parasite. However, the classical laboratory inbred m...
After invasion, Toxoplasma resides in a parasitophorous vacuole (PV) that is surrounded by the PV membrane (PVM). Once inside the PV, tachyzoites secrete dense granule proteins (GRAs) of which some, such as GRA16 and GRA24, are transported beyond the PVM likely via a putative translocon. However, once tachyzoites convert into bradyzoites within cys...
Cats are the definitive host for Toxoplasma gondii. When this parasite invades the cat intestinal epithelium, it differentiates into merozoites that will convert into male/female gametes. Upon fusion, extremely resistant and infectious oocysts will be formed and shed within cat feces. Despite the crucial epidemiological role of cat intestinal stage...
The Lewis rat is the only known warm-blooded animal that has sterile immunity to Toxoplasma . Upon invasion of Lewis rat macrophages Toxoplasma rapidly activates the nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain containing 1 (NLRP1) inflammasome resulting in interleukin (IL)-1β secretion and a form of cell death kn...
After invasion, Toxoplasma resides in a parasitophorous vacuole (PV) that is surrounded by the PV membrane (PVM). Once inside the PV, tachyzoites secrete dense granule proteins (GRAs) of which some, such as GRA16 and GRA24, are transported beyond the PVM likely via a putative translocon. However, once tachyzoites convert into bradyzoites within cys...
Herein we describe, to our knowledge for the first time the use of the clustered regularly interspaced short palindromic repeats/ CRISPR-associated gene 9 (CRISPR/Cas9) system for genome editing of Neospora caninum, an apicomplexan parasite considered one of the main causes of abortion in cattle worldwide. By using plasmids containing the CRISPR/Ca...
The intracellular parasite Toxoplasma gondii resides inside a vacuole, which shields it from the host's intracellular defense mechanisms. The cytokine interferon gamma (IFNγ) upregulates host cell effector pathways that are able to destroy the vacuole, restrict parasite growth and induce host cell death. Interferon-inducible GTPases such as the Gua...
The intracellular parasite Toxoplasma gondii can cause chronic infection in most warm-blooded animals, including humans. In the USA, four different Toxoplasma strains (types I, II, III and XII) are commonly isolated from animals, whereas atypical strains not belonging to these lineages are predominant in other continents such as South America. Stra...
The zoonotic intracellular parasite, Toxoplasma gondii can cause chronic infection in many species including humans. Approximately 90% of reported Toxoplasma isolates in the U.S. are one of four types (I, II, III, or XII) the other types are categorized as non-canonical or exotic. Currently there is not a non-invasive test that can distinguish each...
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...
Toxoplasma infection in humans manifests in a wide range of ways. Host’s immune status is critical, but even immunocompetent individuals can succumb to severe disease. For an infection to be effectively established, the parasite needs to modulate the host immune response. Specialized effector proteins from subcellular organelles known as rhoptries...
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Differences among individuals in susceptibility to infectious diseases can be modulated by host genetics. Much of the research in this field has aimed to identify loci within the host genome that are associated with these differences. In mice, A/J (AJ) and C57BL/6J (B6) mice show differential susceptibilities to various pathogens, incl...
Toxoplasma gondii is among the most prevalent parasites worldwide, infecting many wild and domestic animals and causing zoonotic infections in humans. T. gondii differs substantially in its broad distribution from closely related parasites that typically have narrow, specialized host ranges. To elucidate the genetic basis for these differences, we...
Analysis of shared and specific genes for N. caninum, T. gondii and H. hammondi.
Supplementary Figures, Supplementary Tables, Supplementary Methods and Supplementary References
Supplementary Figures 1-17, Supplementary Tables 1-2, Supplementary Methods and Supplementary References
Conserved region SNPs
OrthoMCL clusters for N. caninum, H. hammondi and 16 reference genomes of T. gondii.
List of CNV and tandem arrays for 16 reference T. gondii genomes.
List of parasite specific gene families
List of SNPs found in all 62 strains
List of T. gondii strains used.
OrthoMCL clusters for E. tennella, S. neurona, N. caninum, T. gondii and H. hammondi.
List of T. gondii genes with top dN/dS ratios.
Non-conserved region SNPs
Summary of SPDs in conserved regions
Macrophages display flexible activation states that range between pro-inflammatory (classical activation) and anti-inflammatory (alternative activation). These macrophage polarization states contribute to a variety of organismal phenotypes such as tissue remodeling and susceptibility to infectious and inflammatory diseases. Several macrophage-or im...
Toxoplasmosis is the most common cause of infectious retinochoroiditis. It is caused by the parasite Toxoplasma gondii, which affects both immune compromised and immune competent patients. The cytokine interferon gamma (IFNg) plays an important role in the inhibition of Toxoplasma growth. In some cell types (such as HeLa cells) IFNg induces the enz...
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 pa...
Toxoplasma gondii is an intracellular parasite, capable of infecting all warm-blooded animals. Upon infection of a host, Toxoplasma converts into cysts in brain and muscle tissue and establishes a life-long chronic infection. Rats, like humans, are resistant, surviving infection with high doses of mouse-virulent Toxoplasma strains. The Lewis rat is...
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The intracellular parasite Toxoplasma gondii infects a wide variety of vertebrate species globally. Infection in most hosts causes a lifelong chronic infection and generates immunological memory responses that protect the host against new infections. In regions where the organism is endemic, multiple exposures to T. gondii likely occur...