The correlation between Toxoplasma gondii infection and Parkinson’s disease: a case-control study

Abstract

Toxoplasma gondii is an obligate intracellular parasite that infects all nucleate cells of vertebrates. Human infected by vertical transmission and also using raw or undercooked meat or food and water that contaminated with mature oocysts. Parkinson‘s disease as neurodegenerative disease affects people above 60 years. Due to high prevalence of toxoplasmosis in Iran and evidence about effects of T. gondii on neurodegenerative diseases, this study has been conducted to investigate possible correlation between Toxoplasma and Parkinson’s disease in Iran. Seventy five Parkinson’s patients and equal healthy volunteers were enrolled. After obtaining informed consent and sociodemographic features, 5 ml blood sample were collected and then anti-Toxoplasma IgG and IgM levels were examined by ELISA method. Data was analyzed with Chi-squre and Fisher’s test by usig stata 11 software. Binary logistic regression was used for multivariate analysis in assessing the correlation between toxoplasmosis and Parkinson. Eighty five percent of Parkinson’s group and 90.3 % of control group were positive for anti-Toxoplasma IgG antibody. In this investigation no statically differences were observed between groups and age, gender, residency and using raw or undercooked meat. There is no significant association between IgG positive titer and Parkinson’s disease. However, statistically significant association was found between Parkinson and keeping cat (P = 0.03) as well as the using of undercooked egg (P = 0.004). Although there is high level of anti-Toxoplasma IgG antibody in Parkinson’s patients which reflects chronic Toxoplasma infection; we couldn’t detect any statistical association between T. gondii infection and Parkinson’s disease.

Introduction

Toxoplasma gondii is an obligate intracellular parasite in coccidian subclass that infects all nucleate cells of vertebrates. This Protozoa has 3 forms; tachyzoite or active form which is observed during acute infection phase, tissue cyst that is seen in latent phase and plays a role in disease transmission and oocyst form that is found in cat feces. Humans are infected by vertical transmission and also consuming tissue cysts using raw or undercooked meat (especially beef and pork) or food and water that contaminated with mature oocysts (Ferreira and Borges 2002). Because of active immune responses, T. gondii remains in muscles and nervous tissues, multiplies in host cells and causes chronic phase of infection (Yazar et al. 2004). In this condition, tissue cyst containing bradyzoites may be ruptured spontaneously and then increased antibody titer (Barragan and Sibley 2002).

Parkinson‘s disease (PD) is the main neurodegenerative disease after Alzheimer. The disease is clinically recognized by symptoms such as tremor at rest, rigidity and bradykinesia or pathologically by degeneration of the dopamine producing neurons in the substantia nigra and presence of lewy bodies as protein deposits in cytoplasm of neurons (Guttmacher et al. 2003; Pandey 2012; Gazewood et al. 2013). Prevalence of the disease is 0.5–1 % in 65–69 years old people and reaches to 1–3 % in those over 80 years (Guttmacher et al. 2003; De Lau and Breteler 2006; Gazewood et al. 2013). Although the clinical signs is the base of diagnostic method for Parkinson‘s disease but some of its symptoms are similar to other diseases that causing confusion in diagnosis (Newman et al. 2009), for this reason, autopsy has been selected as definitive diagnosis method (Guttmacher et al.2003). Central nervous system (CNS) is the most infected site in latent toxoplasmosis. Several epidemiological studies on central nervous system diseases represent hypothesis of toxoplasmosis as a pathogenic factor in development of psychotic symptoms in some patients. Clinical evidence has shown that Toxoplasma infection in CNS causes brain damage (Zhu et al. 2007). Researchers are shown the effect of Toxoplasma on behavior of infected rats (Fleger 2007; Zhu et al. 2007). Dopamine as a neurotransmitter plays the important role in etiology of neurologic diseases such as Alzheimer and Parkinson and its amount is reduced in these diseases (Pearce et al. 2012). By activation of immune system which is caused by T. gondii infection, inflammatory cytokines such as IFNγ and toxic metabolites e.g. nitric oxide, are released and lead to degeneration of dopamine-producing neurons (Çelik et al. 2010). In Alzheimer and Parkinson‘s disease, dopamine level is decreased and when dopamine-producing neurons are destroyed, nervous system is not able to control movements (Jung et al. 2012).

Epidemiological studies on prevalence of toxoplasmosis indicate global spread of this parasite with dissimilar prevalence in different countries (Dubey 2004). According to the studies, prevalence of this parasite in different provinces of Iran is 18–70 % (Mostafavi and Jalali Monfared 2012). Due to high prevalence of toxoplasmosis in Iran and evidence about effects of T. gondii on neurodegenerative diseases and lack of study about influence of this parasite on Parkinson’s disease in Iran, this study has been conducted to investigate possible correlation between Toxoplasma and Parkinson’s disease in Iran.

Materials and methods

Patients

In this case–control study, 75 Parkinson’s patients diagnosed based on the United Kingdom Parkinson’s disease Society Brain Bank Clinical Diagnostic Criteria (Hughes et al. 1992) and 75 healthy volunteers were selected and matched in terms of age and gender. Patients were selected from people who were referred to neurologist’s clinics in Tabriz, Northwest of Iran. Patients with other neurological diseases such as MS, epilepsy, migraine, primary encephalitis and people with history of brain surgery were excluded from the study. Healthy volunteers were selected from people who had been admitted in different wards of hospital and were devoid any neurological diseases. Sociodemographic features were obtained from patients and control group as confounding factors (age, gender, residency, education, cat keeping, using raw or undercooked meat and egg).

ELISA method

Five milliliters blood samples were collected in sterile condition. After obtaining serum by centrifugation blood at 3,000 rpm, they were stored in −20 °C. Sera were brought room temperature in time of assay. Anti-Toxoplasma IgG and IgM levels were examined on sera by ELISA method (ACON ELISA Kit, China) according to the manufacturer instructions. For obtaining anti-Toxoplasma IgM result, we calculated Index value by dividing the specimen absorbance by cut-off value based on manufacturer recommendations. For anti-Toxoplasma IgG, calibration curve was drawn to obtain quantitative results from their absorbance. Index value <0.9 IU/ml was interpreted as negative, 0.9–1.1 as Equivocal and >1.1 as positive.

Statistical analysis

Data was analyzed with Chi-squre and Fisher’s test by usig stata 11 software. Then binary logistic regression was used for multivariate analysis in assessing the relationship between toxoplasmosis and Parkinson’s disease. The default level of statistical significance was based on P < 0.05. Adjusted odds ratio (OR) and 95 % confidence interval were calculated.

Ethical aspects

This study has been approved by the ethics committee of Tabriz University of Medical Sciences, Iran. Informed consent was obtained from all participants after explain the study purpose.

Results

In this study, mean age of patient and control groups were 63.7 ± 11.3 (35–82 years) and 63.4 ± 11.6 (34–89 years) respectively. The mean duration of disease was 56 months. Most of patients were in 51–60 age group and 58 patients (77.3 %) were men. Only 4 % of patients were resident of rural region. Most of patients were undereducated. 52 % of patients and 44 % of control group were habit for using of raw and undercooked meat. 15 % of patients who used undercooked egg were positive for anti-Toxoplasma IgG antibody.

In this investigation, 85.3 % of Parkinson group and 90.3 % of control group were positive for anti-Toxoplasma IgG antibody. All subjects were found as negative for IgM. There was no significant difference between Parkinson’s and control group in term of anti-Toxoplasma IgG antibody (P = 0.2). No statically differences were observed between groups and age, gender, residency and consumption of raw or undercooked meat using Chi square test. However, statistically significant association was found between Parkinson’s disease and keeping cat (P = 0.03) as well as the using of undercooked egg (P = 0.004). There was no equivocal sample in this study. Statistically significant association between IgG positive titer and Parkinson’s disease was not observed. The effect of potential confounding variables was adjusted using multivariate logistic regression analysis (Table 1).

Table 1.

Multivariate analysis of the characteristics in Parkinson’s disease and their association with Toxoplasma gondii infection

Variable	Adjusted OR	95 % CI	P value
IgG positive	0.59	0.2–1.6	0.3
Keeping cat	2.8	0.97–8.4	0.05
Using undercooked egg	5.2	1.4–19.3	0.01

Discussion

According to the previous studies, genetic and environmental factors plays an important role in etiology and development of PD (Nuti et al. 2004). The effect of environmental factors such as exposure to chemicals, metals, nonmetallic toxins, infections, head injuries, rural living, well-water drinking, diet, alcohol, coffee and smoking have been investigated (Priyadarshi et al. 2001; Chen et al. 2004; Logroscino 2005). Role of infections in the occurrence of Parkinson’s disease is already approved. Many studies were planned in order to explore possible association between PD and variety of bacteria and viruses. In the study conducted in Belgrade during 2001–2005, Parkinson’s disease was significantly related to mumps, scarlet fever, influenza, herpes simplex and whooping cough infections but not related to measles, chicken pox and tuberculosis (Vlajinac et al. 2013).

Many studies have been performed around the world for investigation about T. gondii effect as an infectious agent on pathogenesis of Parkinson’s disease. According to the studies, there was possible relationship between latent toxoplasmosis and mental or nervous diseases. Based on the studies that conducted for possible relationship between Toxoplasma infection and schizophrenia, most results have been shown high anti-Toxoplasma IgG antibody in patients compared with control group (Cetinkaya et al. 2007; Torrey et al. 2007; Alipour et al. 2011; Alvardo-Esquivel et al. 2011). However according to the same study in Iran, didn’t found any association between Toxoplasma and schizophrenia (Daryani et al. 2010). Also there was possible association between Toxoplasma infection and bipolar disorder type I (Pearce et al. 2012), suicide attempt (Arling et al. 2009; Yagmur et al. 2010; Ling et al. 2011), migraine (Koseoglu et al. 2009), obsessive—compulsive disorder (Miman et al. 2010b), psychiatric inpatients (Alvarado-Esquivel et al. 2006) and Alzheimer’s diseases (Kusbce et al. 2011).

The CNS is the most affected organ in the latent toxoplasmosis (Zhu et al. 2007). T. gondii invades in brain cells including glia and neurons (Cetinkaya et al. 2007; Daryani et al. 2010). T. gondii can also affect neurotransmitters such as dopamine and norepinephrine (Skallova et al. 2006; Cetinkaya et al. 2007; Yolken et al. 2009; Alipour et al. 2011). It has 2 genes in its genome, encoding two enzymes that producing dopamine in brain (Çelik et al. 2010; Okusaga et al. 2011). These enzymes are expressed in chronic phase and bradzoites form of parasite (Gaskell et al. 2009). In Parkinson’s disease, patients’s neurons have been degenerated and they have lost dopamine producing neurons (Çelik et al. 2010). Degeneration of this neurons by T. gondii and contribution of proinflammatory cytokines may be causes Parkinson disease (Obeso et al. 2008; Olanow et al. 2009).

Present study is the first report on investigation of probable correlation between T. gondii infection and Parkinson’s disease in Iran. In this case–control study, there was no significant difference between Parkinson’s disease and control group in term of anti-Toxoplasma IgG antibody. We couldn’t observe any association between toxoplasmosis and Parkinson’s disease. There are very little researches about evaluation of this correlation. Mean age of patients in our study was 63.7 ± 11.3 years that is consistent with two other studies (Çelik et al. 2010; Miman et al. 2010a). Parkinson’s disease founds in one percent of people that older than 60 years. 85.3 % of Parkinson group was positive for anti-Toxoplasma IgG antibody which shows chronic infection and presence of cyst in the tissues. 15 % of patients who had habit using undercooked egg were IgG positive. Significant association was found between the Parkinson’s disease and keeping cat as well as the eating undercooked egg. These factors are important in Toxoplasma infection. People who kept cat at home could be infected with oocyst form of parasite that cat excrete by feces. Infection by using raw or undercooked meat is probably less important than infection with oocyst in toxoplasmosis. In a serological study in Turkey, there was statistically significant difference between seroposivity of anti-Toxoplasma IgG antibody in Parkinson and control groups while all subjects were negative for IgM antibody. Effect of confounding factors were not discussed in the article (Miman et al. 2010a). However, according to another study in Turkey, didn’t found any relationship between toxoplasmosis and Parkinson’s disease by Sabin––Feldman dye test. Also there was no significant differences between control and Parkinson’s groups in term of occupation, residency, keeping cat and using of undercooked meat (Çelik et al. 2010). Our study is in agreement with the results of this study. Previous studies had some limitations including number of participants and laboratory method. Currently, Sabin-Feldman not be used as safe and standard test in laboratories because this method has high risk of infection for experimenter and also is the less sensitive and specific in comparison with other serologic tests such as ELISA (Robert-Gangneux and Dardé 2012). Although there is high level of anti-Toxoplasma IgG antibody in Parkinson’s patients which reflects chronic Toxoplasma infection; we couldn’t detect any statistical association between latent toxoplasmosis and Parkinson’s disease. Further studies such as cohort study would be helpful to assess the exact impact of T. gondii as the infectious agent in the etiology of Parkinson’s disease.