Evaluation of antibodies to cytomegalovirus and Epstein-Barr virus in patients with autism spectrum disorder

1. Introduction

Autism Spectrum Disorders (ASD) is a range of complex heterogeneous disorders characterized by social impairment and communication defects [1]. ASD is one of the most fast-growing neurodevelopmental disorders with prevalence around 1%–2% worldwide [2]. Although ASD is strongly considered as a genetic-based disorder, there are several environmental related causes that are reported to be associated with ASD etiology [3]. Besides, accumulating evidence show that immune dysregulations such as inflammation, cytokine dysfunctions, and anti-brain autoantibodies can lead to ASD. Different autoimmune disorders were reported in ASD patients such as T regulatory cell defects [4, 5, 6], IgA deficiency, suppressed cellular immune system through reduction in induced lymphocytes and decreased activity of natural killer cells [6, 7]. In this regards, one of the suggested environmental causes of ASD is viral infection during pregnancy or early infancy [8, 9]. Since the central nervous system (CNS) is not completely developed during early childhood, it is more susceptible to viral damages. However, the exact mechanism by which viral infection causes ASD is steel unclear, there might be three different possibilities. Viruses may infect CNS directly or infect other parts of the body that subsequently trigger the disease in the CNS. The other possibility is to change immune response in mother or fetus itself, moreover, a combination of these three possible mechanisms may work simultaneously to initiate the disease. Several studies have been conducted either to prove the association between viral infection and underlying etiology of ASD or to refuse it. So far the best association have been shown between maternal Rubella infection and ASD [10]. Also, other viruses such as Herpesviruses [11] and cytomegaloviruses [12] are suggested to cause ASD [13]. According to previous studies, exposure to CMV and Rubella viruses during first trimester of pregnancy increased the risk of developing ASD. Probably the observed immune defects in Autistic children is related to viral exposure during pregnancy. Some researchers believe that viral infection may be responsible for the decreased T helper cells in autistic patients. Other findings revealed that primary responses to the viral infection, either in embryo or in fetal may lead to immune system defects and autoantibody production against brain tissue in the embryo or fetus and therefore, cause ASD related brain symptoms [14]. Based on these evidence in the current study we analyzed the amount of IgG and IgM antibodies against CMV and EBV in the serum sample of ASD patients to investigate the difference between ASD patients in comparison to healthy controls.

2. Material and methods

3. Result

In this study, 45 ASD patients were included based on DSM5 criteria which were diagnosed by a neurologist. The average age at onset of patients was 3.5 years and with no family history of ASD in siblings and relatives. The male to female ratio in both case and control groups was 38.7 that is according to the sex ratio for disease reported in literature around 5/1. In this study, the mean age (years) ± SD was 5.89 for patients and 6.07 years for control individuals. Based on the t-test which was performed for matching the age in case and control group, there was not a signi?cant difference between control and case groups.

Statistical analysis showed that anti-cytomegalov-irus IgG antibody average value in sera of ASD patients compared to antibody average value in the blood of control individuals was not significantly different either between male or female subgroups or in total samples (0.4, 0.8, and 0.3 respectively) Table 1.

The results of Table 2 showed that anti-cytomegal-ovirus IgM antibody average value in the sera of patients (in male and female) comparing with antibody average value in sera of control individuals only increased but it was not statistically significant ( $p>$ 0.05: 0.6, 0.65 and 0.4 for male, female and total individuals respectively).

The results of Table 3 showed a significant increase in the amount of anti-Epstein-Barr virus IgM antibody average value in sera samples of ASD patients (in male and female) comparing with antibody average value in the blood of control individuals ( 0.05). The obtained P-values respectively for male, female and overall were 0.002, 0.008 and 0.009.

In addition, the evaluated amount of anti-Epstein-Barr virus IgG antibody show an increased amount in patients in comparison to control individuals; however, the results were not statistically significant. The obtained P-value for male, female and total individual were respectively 0.09, 0.1 and 0.05 (Table 4).

Autism Spectrum Disorder is a childhood neurodevelopmental disease that various genetic and environmental factors are involved in its etiology [15]. One of the strongest causative factors is most of the neurological disease such as schizophrenia and autism is viral infection that could affect a child during pregnancy or early infancy [16]. The precise mechanism of viral infection to cause ASD is not completely understood, however, the increased amount of anti-brain autoantibodies are reported in ASD patients.

The development of brain is strongly influenced by immune system and it is showed that dysregulation of neuroendocrine pathways due to immune defects may lead to behavioral disorders [17]. In particular, recently increasing amount of evidence indicate that aberrant immune functions in ASD patients such as reduced number of lymphocytes and natural killer cells [7], abnormal cytokine profiles, imbalance level of serum immunoglobulins [18] and also, the association between autism and immune-related genes include complement C4 alleles and human leukocyte antigen (HLA) [19] may cause neurological manifestations of ASD especially if they occurred during critical neurodevelopmental periods of pregnancy and early infancy [20].

The results of the present study indicate an increased amount of serum level of anti-CMV IgG and IgM antibodies in ASD patients in comparison to healthy controls, however, these incensement was not statistically significant. In contrast, the anti-EBV IgM level in the serum of ASD patients was reported significantly more than the amount in matched controls. Besides, the amount of anti-EBV IgG level of patients was just increased in comparison to healthy group but it show no significant difference.

The previous studies have investigated the association of ASD with family members of herpesviruses such as EBV and CMV. However some evidence reported autistic cases with congenital cytomegalovirus infection [8, 9, 12, 21], evaluation of IgG and IgM antibodies against CMV have shown no significant difference between ASD cases and healthy children. In this regards, the result of one study investigated the antibodies against CMV in the blood samples of 40 ASD patients reported no significant difference between IgM and IgG amount in the serum of ASD patients in comparison to healthy controls [14]. Accordingly, our results showed an insignificant incensement in the serum level of IgG and IgM antibodies against CMV in ASD patients compare to healthy controls. In another study Gentile and their colleagues by considering CMV and EBV as effective factor in ASD disease, had evaluating the antibody titer of these viruses in the 54 children with autism in comparison to 46 healthy matched children. The obtained data from this study indicated that the serum level of anti-CMV and anti-EBV antibodies were equal between case and control groups [22]. In contrast, our results indicate a statistically significant incensement in the serum level of IgM antibody against EBV in cases, however, the IgG level was not different between cases and controls. These results may considered as a guide to investigating the possible role of EBV infection in the ASD etiology.

5. Conclusion

According to the present study, there is a significant increment in the level of anti-EBV IgM in autism patients in comparison to healthy matched controls, while neither anti-EBV IgG nor anti-CMV IgG and IgM antibodies were significantly different between two groups. Altogether, this results could be used as a guide for other studies to investigate the correlation between EBV infection and the ASD disease in larger sample size.