TOXOPLASMOSIS

Role of Monocytes and Neutrophils

Monocytes play a key role in mucosal immunity to T. gondii (Dunay, Fuchs et al. 2010). Inflammatory monocytes are recruited from the bone marrow to the site of infection in the gut. Inflammatory monocytes can restrict the growth of T. gondii through the production of nitric oxide (NO) and recent studies indicate monocytes are necessary in control of T. gondii proliferation in the lamina propria via a NO-dependent mechanism (Mordue and Sibley 2003; Dunay and Sibley 2010). Neutrophils are also one of the first cell types to arrive at the site of infection with T. gondii. They secrete IL-12 and can directly kill the parasite via oxygen-dependent and independent mechanisms and thus have also been touted as having a protective role in mucosal immunity (Denkers, Butcher et al. 2004). More recent studies however have shown neutrophils to be involved in adverse pathological events (Dunay and Sibley 2010).

Role of Dendritic Cells (DCs)

Dendritic cells (DCs) play a central role in orchestrating the innate immune response and bridging innate and adaptive immunity. DCs are the initial antigen-presenting cell at the site of infection and they are either directly infected by the parasite where they multiply and are processed for antigen presentation, or ingest apoptotic enterocytes that are then digested and processed for antigen presentation (Dunay and Sibley 2010). While macrophages and neutrophils are also infected with T. gondii and can secrete IL-12, recent evidence indicates DC production of IL-12, but not macrophages or neutrophils, is essential for IFN-γ activation of NK cells and recruitment of inflammatory monocytes (Hou, Benson et al. 2011). Both conventional and plamacytoid DC subsets are activated by T. gondii infection, with the plasmacytoid DC subset inducing high levels of IL-12, upregulating MHC I and MHC II, and accessory molecules, such as CD86, indicating their ability to prime CD4⁺ T and suggesting the plasmacytoid DC subpopulation may be particularly important in the initial stages of the immune response to T. gondii (Pepper, Dzierszinski et al. 2008).

DCs are also involved in priming CD8+ T cells and shaping the early T cell response. Cognate formation between DCs and CD8+ T cells occurs in the draining lymph nodes early in the immune response (John, Harris et al. 2009). The DC/CD8+ interaction only occurs only in the presence of cognate antigen and not infected DCs, suggesting that cross presentation by bystander DCs rather than infected DCs is an important route of antigen presentation during acute toxoplasmosis. An enhanced migratory response is induced in infected DCs, which likely facilitates parasite dissemination and DCs trafficking to the draining lymph nodes (Lambert, Dellacasa-Lindberg et al. 2011). Additionally, the infected DC also contribute to parasite dissemination in a strain-specific manner with Type II avirulent strains more efficiently exploiting DCs for parasite dissemination than the virulent, Type I strains. (Lambert, Vutova et al. 2009). Avirulent strains induce a more robust activation of DCs at the site of infection and the draining lymph node than the virulent strains, further indicating the role of DCs in shaping the immune response to T. gondii (Tait, Jordan et al. 2010).

Role of TLR's in Innate Immune Response

Several Toll like receptors (TLRs) function in recognition of T. gondii (Yarovinsky 2008; Denkers 2010). TLR11 is required for DC production of IL-12 production and thus is important in induction of innate immunity (Yarovinsky 2008). TLR11 recognizes the parasite protein profilin, which is an actin-binding protein that is used in gliding motility, involved in tissue migration, host cell invasion and egress (Yarovinsky, Zhang et al. 2005). In addition to triggering host immune response, TLR11 may also be involved in the prevention of immunopathology via regulating IFN-γ secretion by NK cells (Yarovinsky, Hieny et al. 2008). TLR2 also contributes to the host response to T. gondii, inducing macrophage activation and NO production and secretion of the chemokine, CCL2 (Del Rio, Butcher et al. 2004). The ligands for TLR2 are glycophosphatidylinositol (GPI) anchors of the parasite (Debierre-Grockiego, Campos et al. 2007). TLR4 also recognizes GPIs of the parasite, with some suggestion that TL2 and TLR4 may work cooperatively during T. gondii infection. These studies indicate TLRs may be involved in a cooperative regulation of the innate immune response against T. gondii.

Role of CD8+ T cells

CD8+ T cells are a key mediator of the adaptive immune response to T. gondii. The role of CD8+ T cells in the protective response to T. gondii was established in early studies in which transfer of CD8+ T cells from immunized mice provided protection to naïve mice challenged with T. gondii and correspondingly protection was ablated by the depletion of CD8+ T cells (Suzuki and Remington 1988; Parker, Roberts et al. 1991; Shirahata, Yamashita et al. 1994). CD8+ T cells are the principle T lymphocyte population involved in the control of the chronic infection and prevention of reactivated infection in the brain (Suzuki and Remington 1988; Gazzinelli, Xu et al. 1992; Wang, Claflin et al. 2005). CD4⁺ T cells, are not required for induction or maintenance of CD8+ T cells but are necessary to regulate the functional activity of intracerebral CD8⁺ T cells (Lutjen, Soltek et al. 2006).

CD8+ T cells are activated via presentation of major histocompatibility complex class I (MHC-I)-restricted parasite antigens. Activation of T cells occurs via direct interaction with infected host cells as opposed to cross-presentation as has been found in the initial priming of CD8+ T cells by DC in the lymph nodes (Dzierszinski, Pepper et al. 2007). Professional APCs, such as dendritic cells and macrophages, are involved as in direct antigen presentation, but non-professional APCs, such as endothelial cells, epithelial cells and astrocytes, are equally as efficient in activating CD8+ T cells in this MHC-I restricted response.

The anti-parasitic effector mechanisms mediated by activated CD8+ T cells include production of cytokines, the predominant of which is IFN-γ, which stimulates anti-parasitic IFN-γ effector mechanisms in a variety of cell types including macrophages, microglia, and astrocytes. The importance of IFN-γ-mediated mechanism is illustrated by the loss of protection against both the acute and chronic stages of infection when IFN-γ is deficient or absent. CD8+ T cells are also known to produce the anti-inflammatory cytokines, IL-17 and IL-27, both of which are associated with down regulation of the inflammatory response to T. gondii during infection (Johnson 1992; Stumhofer, Laurence et al. 2006).

Activated CD8+ T cells can also mediate cytotoxic activity (CTL) against Toxoplasma- infected cells in the context of MHC presenting parasite antigen on their cell surface. The ability of parasite-specific CD8+ T cells to lyse Toxoplasma-infected cells has been demonstrated in in vitro studies (Hakim, Gazzinelli et al. 1991; Khan, Ely et al. 1991; Subauste, Koniaris et al. 1991; Denkers, Sher et al. 1993). In vivo studies using mice deficient in the cytotoxic molecule, perforin, demonstrated the CTL response did not affect resistance to the acute stage but was necessary for resistance to the chronic infection (Denkers, Sher et al. 1993; Denkers, Yap et al. 1997). CD8+ T cells may also be able to eliminate Toxoplasma cysts in the brain (Suzuki, Wang et al. 2010). Surprisingly the CD8+ T cells did not require production of IFN-γ but did require perforin-mediated cytotoxic activity.

IFN-γ Effector mechanisms and IFN-γ Effector Cells

IFN-γ activated macrophages acquire anti-microbicidal activity that is largely dependent upon the production of reactive nitrogen intermediates. The Interferon Response Genes (IRGs), are a family of IFN-γ induced proteins, that have recently been recognized as an important resistance system to target vacuolar pathogens including T. gondii (Taylor 2007; Zhao, Rohde et al. 2009). In macrophages the IRG mechanism induces disruption of the Toxoplasma vacuole leading to degradation of intracellular parasites via autophagomal delivery to the lysosomes (Butcher, Greene et al. 2005; Ling, Shaw et al. 2006). The IRG mechanism involves a coordinated loading of IRG GTPases on the Toxoplasma vacuole (Khaminets, Hunn et al. 2010). Virulent strains of T. gondii evade vacuolar disruption by IRG proteins via phosphorylation of several IRG proteins, via the parasite kinase, ROP18, thereby inhibiting the accumulation of IRG proteins on the vacuole (Zhao, Ferguson et al. 2009; Steinfeldt, Konen-Waisman et al. 2010). IFN-γ induced damage to the T. gondii vacuole can also occur via an autophagosome-independent process involving the autophagy protein, Atg5 (Zhao, Fux et al. 2008).

IFN-γ also induces anti-Toxoplasma activity in non-phagocytic cells such as fibroblasts, epithelial cells, endothelial cells and astrocytes and in these nonphagocytic cells, the mechanism of IFNγ inhibition is via nitric oxide independent mechanisms. The known mechanisms of IFNγ inhibition in non-phagocytic cells includes induction of indoleamine 2,3-dioxygenase (IDO), which degrades intracellular tryptophan, increases in reactive oxygen species, and iron deprivation (Pfefferkorn 1984; Pfefferkorn, Rebhun et al. 1986; Daubener, Remscheid et al. 1996; Nagineni, Pardhasaradhi et al. 1996; Brunton, Wallace et al. 2000). The IRG mediated parasite degradation has also been found in fibroblasts and astrocytes (Martens, Parvanova et al. 2005; Zhao, Rohde et al. 2009).

Role of Microglial Cells

Microglia are the resident macrophage population of the central nervous system and are rapidly activated in murine Toxoplasmic Encephalitis (TE). Activated microglia express elevated levels of the cytokines, IL-12, IL-1β1, TNFα, and the cell surface ligands, MHC I and II and ICAM-1, indicating their role in recruitment and activation of T cells in the brain and ability to act as an APC in the brain (Schluter, Meyer et al. 2001; Wang, Michie et al. 2007). CD8+ T cells regulate cytokine production of microglia, while cell surface molecules are less dependent upon T cells (Schluter, Meyer et al. 2001). Macrophages and microglia can also be source of IFN-γ in the brain and produce IFN-γ independently of T cells during acute toxoplasmosis in the brain (Suzuki, Claflin et al. 2005; Wang and Suzuki 2007). Microglial production of IFN-γ independent of T cells, may be important in the early stages of tachyzoite proliferation in the brain, serving to limiting parasite growth, prior to the recruitment of CD8 T cells into the brain. Microglia can also serve as IFN-γ activated immune effector cells in the brain (Deckert-Schluter, Bluethmann et al. 1999; Suzuki 2002; Suzuki, Claflin et al. 2005). The mechanism of IFNγ inhibition of T. gondii infection in murine microglia is primarily mediated via inducible nitric oxide synthase (iNOS) and the production of nitric oxide (Suzuki 2002b). Nitric oxide (NO) is believed to be toxoplasmacidal, and in sufficient amounts may produce stasis and/or intracellular killing of parasites (Chao, Gekker et al. 1994; Gross, Bohne et al. 1996). Activated microglial cells can inhibit growth of the parasite via CD40-autophagy pathway in microglia (Portillo 2010). Microglia can also function in the down-regulation of the intracerebral immune response via the secretion of TGF-β which prevents neuron degeneration, indicating microglial play an important neuroprotective function in the brain in chronic TE (Rozenfeld, Martinez et al. 2005).

Role of Astrocytes

Astrocytes are also important immune effector cells in the brain, controlling parasite growth and immunopathological responses to the chronic infection (Wilson and Hunter 2004). IFN-γ activated astrocytes significantly inhibit the growth of T. gondii (Halonen, Taylor et al. 2001; Scheidegger, Vonlaufen et al. 2005). IFN-γ inhibition in astrocytes in mice is via the IRG-mediated mechanism while inhibition in human astrocytes is via nitric oxide (Peterson, Gekker et al. 1995; Halonen, Chiu et al. 1998; Halonen and Weiss 2000; Halonen, Taylor et al. 2001; Martens, Parvanova et al. 2005). Astrocytes, are efficient in MHC I presentation, indicating they may be an important antigen presenting cell controlling T. gondii in the brain, capable of stimulating CD8⁺ T cells (Dzierszinski, Pepper et al. 2007). Soluble factors produced by secreted by infected astrocytes down modulate NO-production by IFN-γ activated microglia and prevent neuronal degeneration (Rozenfeld, Martinez et al. 2003). Finally, astrocytes have been shown to be essential for control of Toxoplasma encephalitis, serving to contain sites of parasite replication and the inflammatory lesions in the brain (Drogemuller, Helmuth et al. 2008).

Diagnosis of Acquired Toxoplasmosis

Diagnosis of a chronic infection is typically done via the serological detection of antibodies to parasite specific antigens (Montoya 2002; Remington, Thulliez et al. 2004). A combination of IgM and IgG ELISA assays are most commonly used. IgM is indicative of an acute infection, but it is now appreciated that IgM may persist for as long as a year following infection (Montoya and Remington 2008). The IgG avidity test was developed to help discriminate between past and recently acquired infections as IgG affinity results from an antigen-driven B-cell selection process, and thus an increase in affinity will occur over time (Remington, Thulliez et al. 2004). Therefore, low avidity IgG antibodies are consistent with acute infection. To confirm diagnosis, samples can be tested in a reference laboratory that provides specialized testing for this pathogen. For example, The Toxoplasma Serology Reference Laboratory of the Palo Alto Medical Foundation Research Institute (TSL-PAMFRI), conducts a serological panel that consists of ELISA for IgM, IgA and IgE, avidity testing, the Dye test which measures IgG antibodies and the differential agglutination test (Montoya 2002). The combination of these tests can be useful in determining when an infection occurred, which is of critical importance in counseling during pregnancy; as infection before pregnancy does not result in transmission in utero. Other serological tests that can be used include the indirect hemagglutination test, latex agglutination test, indirect fluorescent antibody tests.

Commercially available tests commonly use T. gondii native antigens, derived from whole cell lysate of T. gondii, and a display wide variation in test accuracy. The use of recombinant antigens has been proposed as an alternative to reduce the problems of antigen standardization and assay reproducibility currently faced by diagnostic laboratories in some parts of the world and in resource-poor settings (Kotresha and Noordin 2010). The parasite recombinant antigens shown to be more sensitive in detecting toxoplasmosis include combinations of rSAG2A, rGRA2, rGRA4, rROP2, rGRA8 and rGRA7 for use in detecting IgG antibodies in human serum and rROP1, rSAG1, rGRA7, rGRA8 and rGRA6 for detecting IgM antibodies in human serum.

Treatment of Acquired Toxoplasmosis

Many of these infections are self limited and resolve without specific treatment or with symptomatic management (i.e. ibuprofen for pain and fever). Treatment with pyrimethamine and sulfadiazine for 4 weeks is effective and is used for moderate to severe cases.

Diagnosis of Congenital Toxoplasmosis

Diagnosis of toxoplasmosis in pregnant women depends is serological testing (Montoya 2002). In acute infection, IgG and IgM levels rise during the first 2 weeks of infection. Elevation of Toxoplasma-specific IgG antibodies indicates that infection has occurred, but does not discriminate between recent and past infection. A negative IgM in combination with a positive IgG result usually indicates infection at least six months duration. However IgM antibodies can persist for up to 18 months after infection and can be non-specific generating false-positive reactions. As noted above, avidity testing, IgA, IgE and differential agglutination tests can be useful in determining when an infection occurred and are critical adjuncts in understanding if a positive IgM test in pregnancy will be associated with a risk of congenital transmission of infection. Once a diagnosis of acute Toxoplasma infection is made in a pregnant mother the diagnosis of congenital transmission of infection in the fetus in utero can be done via polymerase chain reactions (PCR)-assays from amniotic fluid. Treatment of the mother with Spiramycin is often instituted with the diagnosis of acute infection while further diagnostic testing is being done to determine if congenital infection has occurred. PCR assays from amniotic fluid, typically based on the B1 gene, have reported sensitivities of 65% - 91% (Romand, Wallon et al. 2001; Thalib et al. 2005, Gras et al. 2005; Bessieres, Berrebi et al. 2009; Eida, Eida et al. 2009). A recently reported real-time PCR with a sensitivity of 98% further improves detection of T. gondii in amniotic fluid (Wallon, Franck et al. 2010; Abdul-Ghani 2011). If congenital transmission is determined then treatment should be instituted to decrease the complications of congenital toxoplasmosis.

Prenatal, Neonatal and Postnatal Treatment of Congenital Toxoplasmosis

In the US, routine pre-natal testing is not routinely done. In contrast in countries with a higher incidence, universal prenatal screening procedures have been adopted. In France, for example, screening of pregnant women, in utero diagnosis and treatment are routine with the effect that severe disease has been reduced as indicated by several studies (Bessieres, Berrebi et al. 2001; Bessieres, Berrebi et al. 2009; Garcia-Meric, Franck et al. 2010). Routine prenatal treatment has been advocated given the suggested improved clinical outcome in reducing transplacental transmission and disease (McLeod, Boyer et al. 2006; McLeod, Kieffer et al. 2009). Spiramycin is often used for treatment while waiting for diagnostic testing for prenatal transmission and in some studies is continued for women in whom acute toxoplasmosis is documented even if tests for prenatal transmission are negative. Spiramycin is stopped and treatment with pyrimethamine and sulfadizine is recommended for documented prenatal transmission (McLeod, Boyer et al. 2006; McLeod, Kieffer et al. 2009).

Treatment of the neonatal infant during the first year of life and longer has been demonstrated to significantly improve clinical outcome (Montoya and Remington 2008). Treatment with pyrimethamine and sulfadiazine during at least the first year of life reduces parasite burden and may help prevent adverse sequelae such as retinal damage and loss of vision (Kieffer, Wallon et al. 2008; Phan, Kasza et al. 2008; Phan, Kasza et al. 2008). In severe cases steroids may be useful to decrease inflammation. Results from long-term studies found individuals with treated congenital toxoplasmosis had no significant reduction in visual function, as compared to uninfected individuals (Peyron, Garweg et al. 2011).

Diagnosis and Treatment

In most cases, diagnosis is based upon characteristic finding on the ocular examination and the presence circulating specific antibodies against T. gondii. PCR, which can determine the parasite directly, can be utilized to demonstrate this organism in vitreous fluid samples. Analysis of intraocular fluids, such as aquaeous, vitreous and chorioretinal biopsies is usually reserved for difficult cases in which the differential diagnosis is broad, e.g. immune compromised hosts such as those with AIDS, and includes other infections such as cytomegalovirus and Herpes virus where a delay in diagnosis and treatment may have significant adverse outcomes. This is especially true in children, when vision is often lost because of retinochoroiditis or retinal complications. Disease should be monitored and treated through the first 2 years of life (Kieffer, Wallon et al. 2008).

Anti-toxoplasmic therapy is effective with active lesions becoming quiescent in ten to 14 days (Mets, Holfels et al. 1997). Pyrimethamine and sulfadiazine as well as Trimethoprim/sulfamethoxazole have been effective therapeutic agents. Systemic steroids can be added to avoid further damage of the retina by the inflammation. Treatment needs to be started before necrosis and damage to the retina, caused by recurrent bouts of the retinochroiditis occurs (Montoya and Rosso 2005) (Mets, Holfels et al. 1997; Phan, Kasza et al. 2008; Phan, Kasza et al. 2008). (Pereira-Chioccola, Vidal et al. 2009). Retinochroiditis occurs in clusters over time, with recurrence influenced by age and duration of infection (Holland, Crespi et al. 2008). Treatment does not avoid recurrences and long-tem follow-up is recommended into the second decade of life (Phan, Kasza et al. 2008b).

Diagnosis

Diagnosis depends on a combination of clinical, serological and radiological information (Mamidi, DeSimone et al. 2002). A presumptive diagnosis is often confirmed with a positive response to anti-Toxoplasma therapy. A favorable clinical response is expected within 14 days of specific treatment. Molecular diagnosis using PCR can be a useful tool for diagnosis of T. gondii (Pereira-Chioccola, Vidal et al. 2009). Imaging using computed tomography (CT) of MRI is essential for the presumptive diagnosis of cerebral toxoplasmosis. Single or multiple focal lesions are usually observed with CT or MRI, with a typical pattern of a hypodense lesion with ring-enhancing and perilesional edema evident (Pereira-Chioccola, Vidal et al. 2009). The differential diagnosis includes primary lymphoma of the CNS, which constitutes the most common differential diagnosis of cerebral toxoplasmosis in developed countries, while in developing countries focal forms of cerebral TB is also a common alternative diagnosis (Manzardo, Del Mar Ortega et al. 2005; Trujillo, Jaramillo-Rangel et al. 2005). In addition to these neurologic complications, the differential diagnosis of brain lesions in HIV-infected patients also includes infections such as cryptococcosis, aspergillosis, microsporidiosis and Trypanosoma cruzi or metastases of disseminated lymphomas and glioblastoma tumors.

Immunological diagnosis using antibodies in serum is complicated by the fact that healthy individuals with chronic Toxoplasma infection maintain T. gondii IgG antibodies. Thus, serological testing cannot discriminate between reactivated versus latent infection. The presence of a negative serology would make cerebral toxoplasmosis less likely and is an indication for brain biopsy in cases consistent with cerebral toxoplasmosis as the probability of an alternative diagnosis is higher. Nonetheless, it should be appreciated that a negative serological result or low titer, does not exclude a positive diagnosis for cerebral toxoplasmosis especially in the context of compatible clinical and radiological findings (Antinori, Larussa et al. 2004).

Over the last two decades, the development of PCR based molecular diagnostic methodologies has allowed detection of T. gondii in a variety of clinical specimens. PCR-based methods using cerebrospinal fluid (CSF) or peripheral blood samples have been demonstrated to have utility in the diagnosis of cerebral toxoplasmosis (Bastien 2002). Molecular methods are particularly appropriate for AIDS patients since these methods are not affected by immunological status and are rapid, sensitive and specific (Colombo, Vidal et al. 2005; Pereira-Chioccola, Vidal et al. 2009). PCR using CSF has been reported to have a high specificity of 96 to 100% (Pereira-Chioccola, Vidal et al. 2009; Correia, Melo et al. 2010). However collection of CSF is invasion and may be inappropriate for patients with extensive cerebral lesions with extensive edema. Reports on the utility of PCR using peripheral blood samples for diagnosing cerebral toxoplasmosis have been variable (Correia, Melo et al. 2010).

The wide range of variation in the sensitivities of PCR methods is a reflection of the variations in PCR protocols and the target gene. Conventional PCR (cnPCR), nested PCR (nPCR), and quantitative real time PCR (gPCR) methods have all been used to detect T. gondii. The most common targets used include the B1 gene and the 529 bp repeat element. A European study found the B1 gene was more sensitive than the 529-bp sequence while conversely a study conducted in Brazil found that the B1 gene was more sensitive than the 529 bp sequence for diagnosis of both cerebral and congenital toxoplasmosis in both cnPCR and qPCR (Pereira-Chioccola, Vidal et al. 2009; Mesquita, Vidal et al. 2010). Thus uncertainty exists surrounding the best target(s) and PCR methods for accurate molecular diagnosis of cerebral toxoplasmosis. Some of this variation may also depend upon the dominant strain of the parasite present in geographic location. Molecular characterization techniques are also used for genetic characterization and identification of parasite strains in epidemiological and genetic studies. The commonly used methods for these purposes are multilocus PCR-RFLP, microsatellite, or multilocus sequence typing (MLST). An integrated approach for the molecular detection and genetic characterization of T. gondii has recently been propsed (Su, Shwab et al. 2010). This approach consists of detection of T. gondii infection in biological samples by nPCR or qPCR of repetitive DNA sequences using the B1 gene or the 529-bp repeat element, followed by multiplexed multilocus nested-PCR-RFLP technique for the subsequent genetic identification of T. gondii.

Treatment of Cerebral Toxoplasmosis

Therapy with pyrimethamine and sulfadiazine is usually effective. In patients with sulfadiazine intolerance either clindamycin or atovaquone can be used in combination with pyrimethamine. Therapy is continued for at least 6 weeks and if immune suppression is present secondary prophylaxis with trimethropim/sulfamethoxazole or pyrimethamine/sulfadiazine is continued indefinitely. Recent data suggests that. trimethoprim/sulfamethoxazole can be used for primary therapy in place of pyrimethamine/sulfadiazine with good outcomes (particularly in resource poor settings)(Aberg and Powderly 2010). Steroids may be useful in cases with significant cerebral edema. Neurological imaging should be done 2 weeks after initiating therapy should be done to assess efficacy of treatment. If patients fail to respond to therapy, a biopsy of the lesion is indicated to look for an alternative diagnosis.

Primary prophylaxis with trimethoprim/sulfamethoxazole is effective in preventing cerebral toxoplasmosis in T. gondii seropositive patients with AIDS who have a CD4+ T-cell count below100 cells/mm³. Primary prophylaxis can be discontinued in patients with a good response to cART, defined as a CD4+ T-cell count above 200 cells/mm³ after 3 months. Secondary prophylaxis can be discontinued when HIV-infected patients receiving cART have a sustained increase of CD4+ T-cell count above 200 cell/mm³ after about 6 months of cerebral toxoplasmosis drug therapy (Lejeune, Miro et al. 2011).

Direct Effect

The parasite exhibits a neurotrophism forming cysts predominantly in the brain during a chronic infection and thus it is well placed anatomically to mediate direct effects on neurological functions in the host. The parasite encysts in neurons, glia cells and astrocytes, potentially affecting all these cell type (Halonen, Lyman et al. 1996; Carruthers and Suzuki 2007). Furthermore, in chronic infection in mice, cysts are found predominantly in neurons, occurring in all parts of the neuron (dendrites, axons and the cell body), indicating the parasite could directly impact neuronal function (Ferguson and Hutchison 1987). While cysts persist in the brain for the lifetime of the host, studies in chronically infected mice found microglial nodules containing parasites are present throughout the chronic infection, indicating cysts might periodically degenerate releasing parasites during the latent phase of infection (Frenkel 1988; Ferguson, Huskinson-Mark et al. 1994). Damage to the brain in congenital and reactivated infections in immunocompromised individuals is characterized by necrosis and microglia nodules in the cortex, basal ganglia and other areas of the brain, indicating the potential direct impact the parasite may have on brain tissue (Fekadu, Shibre et al. 2010).

Neuroimmunological Mechanisms

Cytokines may influence mood and behavior by modulating dopaminergic and glutamatergic neurotransmission (Deutsch, Mastropaolo et al. 1989; Lang, Puls et al. 2007). It has been suggested that proinflammatory cytokines produced by infected astrocytes and microglia, may influence mood and behavior by modulating these monoamine and glutamate pathways in the brain (Carruthers and Suzuki 2007; Fekadu, Shibre et al. 2010). Activated astrocytes are a hallmark of Toxoplasma encephalitis and have also been suggested to affect neurotransmitters via the production of kynurenic acid (KYNA) which may contribute to excessive inhibition of glutamine and nicotine neurotransmitter receptors, a effect that is believed to cause the cognitive impairment observed in schizophrenic patients (Schwarcz and Hunter 2007) Finally, the parasite has two genes encoding tyrosine hydroxylase that produce L-DOPA, one which is induced during the formation of bradyzoites in the brain, and thus it is plausible the parasite can directly affect dopamine and/or serotonin biosynthesis, which are neurotransmitter know to be important in psychotic and mood disorders (Gaskell, Smith et al. 2009; Henriquez, Brett et al. 2009). Dopamine has been found to play a role in behavioral changes in infected rodents and is suggested to play a role in behavioral changes of infected individuals (Novotna, Hanusova et al. 2005; Webster and McConkey 2010).