Welcome to the ICOPA 2022 Online Program

Abstract Body

Ocular disease is the major clinical manifestation of toxoplasmosis. However, this subject is still surrounded with controversy. The aim of this presentation is to provide a panorama on ocular toxoplasmosis, dissecting its importance as the main etiology of infectious posterior uveitis worldwide and an important cause of visual disability in endemic areas.

Clinical aspects of ocular toxoplasmosis will be dissected, with emphasis on diagnostic advances brought by multimodal imaging and by molecular biology. Epidemiology in the context of congenital/postnatally acquired disease will also be presented. Immunopathogenetic aspects will be briefly reviewed, discussing influence of host genetics/immune response, and of parasite genotype/virulence. Finally, recent trends in treatment of ocular toxoplasmosis will be discussed, including the role of local therapy and of secondary prophylaxis.

Introduction

Toxoplasma gondii's prevalence is estimated as about 30%. A strategy of propagation has therefore been suspected, consisting in the manipulation of host behavior. Such idea was confirmed in rodents: latent T. gondii infection attenuates or even suppresses the innate fear of Felidae, their natural predators. In humans, several studies have suggested that infected individuals may present positive links with suicidal behavior, traffic accidents or schizophrenia. Since sleep represents a highly sensitive index of brain functions, as sleep disorders often precedes the development of symptoms in several neurodegenerative and psychiatric disorders, we studied the effects of chronic infection by T. gondii on the sleep/wake cycle in the mouse.

Methods

CBA/Jrj mice were infected using T. gondii cysts of the type II Prugniaud strain. We examined quantitative and qualitative changes of sleep/wake cycle and spectral characteristics of electroencephalographic waveforms specific for each state (wake, slow wave sleep and paradoxical sleep).

Results

We found that infected mice exhibited a chronic sleep-wake alteration over months, characterized by a more than 20% increase of wakefulness (785.7±8.8 vs 634.8±7,6 minutes per 24h; p<0.0001), enhanced EEG θ power and concomitantly decreased slow wave sleep, effects alleviated by an anti-inflammatory treatment using corticosteroids.

Conclusions

This is the first study that reports the impact of chronic toxoplasmosis on the sleep wake/cycle. Such change in consciousness may represent another component of behavioral manipulation of the host, by which T. gondii would promote its dissemination by increasing the frequencies of encounter and interaction with host’s environment.

Introduction

Toxoplasma gondii is a zoonotic pathogen with up to 60% of acquired infections associated with foodborne transmission. Consumption of raw fresh produce (FP) contaminated with T. gondii oocysts is one infection route. The relative importance of FP as source for human T. gondii infection is underestimated as standardized detection method(s) are lacking. We developed a standard operating procedure (SOP) for molecular detection of T. gondii oocysts in leafy green salads, validated it by a ring trial (RT) and are currently applying it in a multicentre survey (MS) on ready-to-eat (RTE) salad at European level.

Methods

The SOP was implemented in 7 laboratories using video tutorials. A RT was organized to evaluate laboratory performance and efficiency of different steps of the procedure. An evidence-based sampling strategy was designed to conduct a MS.

Results

Implementation in the laboratories was successful and allowed identification and resolution of procedure limitations. The expected LoD (10 ocysts/30g salad) was reached. RT analysis confirmed robustness of the procedure and comparability of results among participants. The MS on two types of RTE mixed salads (baby leaves and cut-leaf mixes) in 10 European countries has started and preliminary analysis will be presented.

Conclusions

The application of a well-validated SOP is proving to be a useful tool to investigate the occurrence of T. gondii contamination in RTE-salad, which is needed to assess the associated potential risk for humans.

Acknowledgments: RT and survey participants. This work was done as part of TOXOSOURCES project, EU Horizon 2020 Research and Innovation programme under grant agreement No 773830: One Health European Joint Programme. NMLU is granted by a UCM-Santander/2018 predoctoral fellowship.

Introduction

This case contributes to the limited literature on central nervous system (CNS) involvement of blastic plasmacytoid dendritic cell neoplasm (BPDCN) and raises suspicion of a possible association between toxoplasmosis and hematological malignancies.

Methods

A 63-year-old male presented to the department of neurology with a three-day history of rapidly progressing headache, fatigue, and confusion. Medical history was unremarkable except for a spinal disc herniation. The patient was unable to speak in coherent sentences, remaining neurological examination was normal. General physical examination revealed multiple bruise-like lesions on the body with no prior history of trauma.

Results

Initial laboratory workup raised suspicion of acute leukemia, and a brain computer tomography identified several hyperdense processes, initially interpreted as metastases or vascular malformations. Acute chemotherapy and supportive treatment were initiated. A bone marrow biopsy gave the final diagnosis BPDCN. The patient’s cerebral status worsened despite clearance of leukemic cells in the cerebrospinal fluid (CSF) and regression of the cerebral lesions on scans. Considering the immunosuppressive treatment alternative causes were examined and toxoplasma gondii DNA was detected in the CSF in a sample obtained 16 days after chemotherapy was initiated. Despite intensive treatment with systemic and intrathecal chemotherapy the patient died 25 days later due to multi-organ failure.

Conclusions

While CNS involvement of BPDCN was proven by CSF flow cytometry analysis, it remains unclear whether the preliminary neurological condition and the findings on the brain scans can be attributed to cerebral dissemination of BPDCN, cerebral toxoplasmosis or both.

Introduction

Toxoplasma gondii, the causative agent of toxoplasmosis, relies on successful cell division for pathogenesis. The centrosome of T. gondii, its main microtubule(MT)-organizing center, plays central roles orchestrating the temporal and physical coordination of major events during division. It is constituted by two domains; an outer and inner core. Homeostasis of the outer core has been shown critical for proper assembly of daughter cells. However, the role of the inner core remains unexplored. We focus on understanding the function of the inner core by studying the role of its only known molecular marker; TgCEP250L1.

Methods

We generated TgCEP250L1-mAID-3HA, a strain in which TgCEP250L1 is rapidly degraded upon IAA addition to the growth medium. We addressed the impact of TgCEP250L1’s degradation by functional and ultrastructural analyses.

Results

Mutants exhibit nuclear segregation defects whilst normally forming daughter cells. In addition, the outer core of the centrosome disconnects from the nucleus. High resolution microscopy reveals structural defects underlying these phenotypes. We show that TgCEP250L1’s location varies along the forming mitotic spindle, and that in the absence of TgCEP250L1, the MT binding protein TgEB1, fails to translocate to the spindle.

Conclusions

Our data supports a model in which the inner core of T. gondii critically participates in cell division by impacting the formation or stability of the mitotic spindle.

Introduction

Toxoplasmosis can be particularly severe when opportunistic or congenital. Toxoplasma gondii is successively encountered in its hosts as a proliferative tachyzoite stage in acute infection and as a latent encysted bradyzoite stage in chronic infection. While serology is a key element in the diagnosis of this parasitosis and although the contribution of the bradyzoite to the pathophysiology of the infection is undeniable, current tests are based only on tachyzoite antigens or antigens expressed during both tachyzoite and bradyzoite development. Our aim was to identify proteins that are immunogenic and bradyzoite-specific.

Methods

We used the MORC-Auxin-Inducible Degron strategy to obtain a parasite culture enriched in bradyzoite-specific proteins in-vitro, and compared the immunogenicity of this parasite extract versus tachyzoite extract on serum from chronically infected mice by western blot to identify candidate proteins. We have thus identified the protein BSM (bradyzoite serological marker). We used this recombinantly produced protein to develop an ELISA assay to identify cyst carriage associated with chronic infection in 82 mice.

Results

BSM serology discriminates cyst-carrying mice with a sensitivity of 97.7%, a specificity and a positive predictive value of 100% and a negative predictive value of 95%.

Conclusions

The use of this serological test in humans could improve the risk assessment of post-transplant or congenital toxoplasmosis and enhance the understanding of the pathophysiology of cysts.

Introduction

Toxoplasmosis is an important and potentially severe opportunistic infection in heart transplant recipients, due to tropism of Toxoplasma for the cardiac muscle. We conducted an eight-year-long prospective study on the diagnosis and monitoring of Toxoplasma infection (TI) in orthotopic heart transplant (OHT) recipients on continuous trimethoprim-sulfamethoxazole (TMP-SMX) prophylaxis. The objective was to determine the incidence of TI before it evolves into clinical, potentially fatal Toxoplasma disease (TD).

Methods

Pre-transplantation serological screening of the recipients (and some donors) was followed by post-transplantation peripheral blood (PB)-based qPCR monitoring targeting the Toxoplasma 529 bp gene. In OHT recipients qPCR was performed monthly for two months post-OHT and then yearly. TI was diagnosed based on a positive PCR result in at least one PB sample.

Results

Regardless of the TMP-SMX prophylaxis, TI was diagnosed in 3/37 (8.1%) OHT recipients. Of the three patients that developed TI, infection was graft-transmitted in 2 seronegative OHT recipients and reactivated in 1 seropositive recipient of a seronegative donor’s heart transplant.

Conclusions

The presented results raise awareness of reactivated toxoplasmosis even in the setting of continuous TMP-SMX prophylaxis in seropositive OHT recipients, suggesting a possible drug compliance issue. Hence, we advocate systematic PB-based qPCR monitoring of TI in all OHT recipients, regardless of their serological status, the frequency of which should be adjusted according to the clinical data.