Relationship between Toxoplasma Gondii and Psychotic Disorders with Implications toward a Brain-based Diagnostic System and Novel Treatment Approaches: A Study Protocol

What role infectious agents play in the causation of psychotic disorders? To investigate this area, we have aimed to investigate the relationship between Toxoplasma gondii and psychotic disorders. A hospital-based cross-sectional study is designed. IgM and IgG antibodies to T. gondii in patients with psychotic disorders will be measured and presented in result. Seropositivity rates will be compared with first-degree relatives and healthy volunteers. Also, types of psychotic disorders and seropositivity rate will be compared. Here, we are presenting the study protocol with implications toward a brain-based diagnostic system and novel treatment approaches.


Background and Rationale
The role that infectious agents play in the aetiology of psychotic disorders is an area of interest. Among such candidates, Toxoplasma gondii becomes a prominent one. Evidences are emerging for this line of thinking [1]. Antibodies to T. gondii are more in patients with psychotic disorders. Some adults with toxoplasmosis exhibit psychotic symptoms. Epidemiological similarities are observed in toxoplasmosis and psychotic disorders. Antipsychotic agents inhibit T. gondii [2]. Toxoplasma raises dopamine levels in animals. Childhood exposure to cats is high in patients with psychotic disorders.
Abdollahian et al. [3] found more prevalence of T. gondii infection among schizophrenia patients compared to control in Iran. Chen et al. [4] found increased seropositivity of anti-Toxoplasma immunoglobulin G (IgG) and immunoglobulin M (IgM) not only with schizophrenia but also with bipolar disorder in China. By demonstrating elevated Toxoplasma exposure in recent onset psychosis, Yolken et al. [5] gave new insight to temporal relationship between exposure and disease onset.
The relationship of T. gondii and psychotic disorders has the potential to establish a brainbased diagnostic system in psychiatry and pave the way for novel therapeutic options with disease modifying effect.

1.
Measurement of IgM and IgG antibodies to T. gondii in patients with psychotic disorders.

2.
Comparison of the seropositivity rate for anti-Toxoplasma IgG and IgM antibodies among patients with psychotic disorders with that of first-degree relatives (FDR) as well as healthy volunteers (HV).

3.
Comparison of the types of psychotic disorders and seropositivity rate.

Hypotheses
A group of patients with psychotic disorders have serological evidence of Toxoplasma infection. There are certain characteristic clinical and demographic variables among patients with psychotic disorders who have serological evidence of Toxoplasma infection.
There might be a relationship between toxoplasmosis and the aetiology of schizophrenia, and an understanding of the pathogenesis of Toxoplasma infections in individuals with schizophrenia might lead to new approaches to the management of this disorder.

Design
The study will be a hospital-based cross-sectional study.

Inclusion criteria for participants-Patients
with psychosis as a defining feature (schizophrenia, acute and transient psychotic disorder, and delusional disorder) and psychosis as an associated symptom (mood disorder and substance use disorder).

Exclusion criteria for participants-Delirium
, dementia, mental retardation, and neurological disorders that would affect cognitive performance including epilepsy, a history of encephalitis or head trauma, or any other reported disorder of the central nervous system.

Eligibility criteria for study centres-Patients with psychotic disorders attend
for diagnosis and treatment, e.g. in Department of Psychiatry, GMCH.
IgG and IgM antibodies to T. gondii are measured, e.g. in Department of Microbiology, GMCH.

Eligibility criteria for who will perform-Competency in diagnosis and
treatment of psychotic disorders, e.g. second, fourth, fifth, and seventh authors are psychiatrists.

3.2.5
Outcomes-Demography of participants. Clinical information of patients. IgM and IgG antibodies to T. gondii.

Sample Size
The sample size is calculated taking the prevalence to be estimated at 60% that gives the maximum sample size, with 95% levels of confidence and 20% bound on error of estimation. Taking this prevalence and stated permissible level of error, the sample size for the study is calculated using the formula where n = required sample size, p = 0.6, q = 0.4, d = 20%.

Recruitment
Laboratory technician attending the individuals for collection of blood sample, without the individuals having to go to the laboratory is a strategy to improve adherence and procedure for monitoring adherence.

Data collection methods-After written consent by the participant and her
family, demographic and clinical information as well as 5 ml of blood samples will be taken from patients with psychotic disorders, first-degree relatives (FDR), and healthy volunteers (HV). The samples will be centrifuged for 5 minutes at 3,500 rpm; separated, quantitated, and stored at −20°C until later analysis. The serum obtained will be screened for IgM and IgG anti-Toxoplasma antibodies using the Enzyme Linked Fluorescent Assay (ELFA) in the mini VIDAS system (BioMérieux).The samples will be processed according to manufacturer's instruction. The diagnoses of the patients with psychotic disorders will be made according to the criteria of the tenth revision of the World Health Organizatioin's International Statistical Classification of Diseases and Related Health Problems (ICD-10) [6].
Relevant care and interventions for which the patients with psychotic disorders have attended Department of Psychiatry, GMCH are permitted to continue concomitantly. Participant/guardian request and worsening disease leading to clinically uncooperative state are criteria for discontinuing. Fig. 1. schematically represents the study procedure. Participants would constitute of patients, FDR, and HV. Their demography and clinical data would be collected. Sex and locality would be the demography while clinical data would be the antibodies to T. gondii.

DISCUSSION
We have presented here the study protocol of the investigation of the relationship between Toxoplasma gondii and psychotic disorders with implications toward a brain-based diagnostic system and novel treatment approaches.
This line of investigation is not restricted to psychosis among adults. Researchers have explored this subject in different neuropsychiatric disorders as well as in the young population. Abd El-Aal et al. [7] found "significant associations between T. gondii seropositivity of epilepsy and depression groups where youth and adults had the highest sero-T. gondii infection especially male in rural areas with low social class". Among children and adolescents, Yalın Sapmaz et al. [8] found higher T.gondii IgG seropositivity in depression, especially with suicidality. But, exploring suicidality among adolescents and T. gondii infection, Sari and Kara [9] found no significant differences between the patient and the control groups.

CONCLUSION
This study protocol outlines how the investigation of the relationship between Toxoplasma gondii and psychotic disorders is planned that has implications toward a brain-based diagnostic system and novel treatment approaches.  Table 1.  Table 3.    Table 5.