Sample
The study sample was recruited from our child and adolescent psychiatry department between March 2014 and April 2015.
The inclusion criteria for the patient group were as follows: 1) ASD diagnosis according to DSM-V. 2) An age range of 2–18 years. 3) Not having an infection, inflammatory conditions, or chronic physical illness. 4) Not receiving iron supplementation and going on a diet. ASD diagnosis was made using the information gathered by history taking, clinical symptoms and observations, CARS, and checklists. One of the authors, who is a faculty member of the same hospital and has an extensive experience in ASD, reviewed all the data and confirmed the ASD diagnoses. Initially, 116 children with DSM-V ASD diagnosis were recruited. From these children, 16 were excluded for the following reasons: Six parents of children with ASD did not want to participate in the study, four children had a continuing infection, two children had a chronic medical illness, and four children were excluded by the faculty member for not meeting ASD diagnostic criteria. So, the final sample for the patient group included 100 children with ASD.
The inclusion criteria for the control group were as followings: 1) No psychiatric disorder diagnosis according to DSM-V 2) An age range of 2–18 years. 3) Not having an infection, inflammatory conditions, chronic physical or mental illness. 4) Not receiving iron supplementation and going on a diet. 5) For preschoolers, normal developmental history and normal Denver II Test. 6) For school-aged children, normal intelligence based on either a WISC-R full scale IQ score above 80 or the average/above academic functioning documented with the last year’s final school grades. For the control group, 100 children (an equal number with patients), who referred to the department for counseling about child development, school adjustment and performance, teenage problems, family and friend relations, were recruited.
Intellectual evaluation
Intellectual capacities of children in both patient and control groups were examined. A child development specialist performed Denver II Test to children under 6 years. An experienced psychologist performed WISC-R full scale to children over 6 years. Children who could not adapt to Denver II Test and WISC-R full scale were evaluated with developmental history, clinical symptoms and observations, and the average academic functioning documented with the last year’s final school grades. In the patient group, children with normal intellectual capacity and border intellectual disability were accepted as normal and children with mild, moderate, and severe intellectual disability were accepted as intellectual disability. In the control group, 13 children were preschoolers, 72 children were school-aged children, and 15 children were over 16 years. Sixty-six of school-aged children were administered the WISC-R full scale and 6 children were not. For 21 children, normal intellectual capacity was established based on the average academic functioning documented with the last year’s final school grades.
Instruments
Childhood autism rating scale
The Childhood Autism Rating Scale (CARS) consists of 15 items and all the items contribute equally to one total score. Each item is rated by half scoring between 1 and 4 [12]. 15–29 points represent the child does not have autism, 30–36.5 points represent mild-moderate autism, and 37–60 points represent severe autism. For the Turkish version of the scale, item analysis shows that all items (except item 14) differentiate children with mild to severe autism [13]. According to CARS total score, children with ASD were divided into two groups: children with mild-moderate ASD and children with severe ASD.
Autism behavior checklist
The Autism Behavior Checklist (AuBC) is especially used in determining the frequency and severity of autistic symptoms [14]. The checklist includes 57 questions divided into five categories. The validity and the reliability of AuBC were satisfying for the Turkish sample [15].
Aberrant behavior checklist
The Aberrant Behavior Checklist (AbBC) is useful for evaluating behavioral problems in children [4]. It consists of 58 items and each item is rated on a four-point scale ranging from 0 (not a problem) to 3 (the problem is severe). The Turkish translation and adaptation of the checklist were conducted by Sucuoglu [16] and Karabekiroglu and Aman [17].
Laboratory measurements
Serum ferritin level was used as an indicator for ID since it is a precursor for ID and represents iron levels in body tissues including brain [4]. We used ferritin cutoff < 10 ng/mL for preschool children and < 12 ng/mL for school-aged children to evaluate ID [3, 5, 6]. Anemia was defined as hemoglobin < 11.0 g/dL for preschool children and < 12.0 g/dL for school-aged children [3]. The following cutoffs were used based on our hospital laboratory values: iron < 50 μg/dL, hematocrit < 32%, MCV < 75 fL, and RDW > 14.5%. Hemoglobin, hematocrit, iron, ferritin, MCV, and RDW values were measured in fasting blood in the morning at our hospital biochemistry laboratory using standard measurement assays.
Statistical analysis
The collected data were analyzed by using the SPSS version 16.0 (SPSS, Inc., Chicago, IL, USA). Clinical data were shown as means and SD. Categorical variables were compared by using chi-square test. Normally distributed parametric variables were compared between groups by using Independent Samples t test. The Mann–Whitney U test was used to compare the age difference between ASD children and healthy controls. Pearson’s correlation analysis was used in order to examine the correlation between hemoglobin, hematocrit, iron, ferritin, MCV, RDW values and CARS, AuBC, and AbBC total scores. The p value < 0.05 was accepted to be statistically significant.
