2 (PDD-BI) [7], the Autism Treatment Evaluation Checklist (ATEC) [8], and the Severity of Autism Scale (SAS). The individual burden of toxic metals was assessed based on urinary excretion, both before and after taking oral dimer- captosuccinic acid (DMSA). DMSA is a licensed medication for treating lead poisoning and indicated in cases meeting toxic criteria. DMSA is, however, widely used oﬀ-label for other metal exposures, for example, mercury. It acts by forming sulfhydryl linkages to divalent metal cations, forming a chelated metal complex which is then excreted in the urine [9]. Urine measurements before and after taking DMSA provide an indication of both ongoing environmental exposures (before DMSA provocation) and the accumulated or relative body burden (postprovocation with DMSA). Red blood cell (RBC) glutathione was measured because it is one of the body’s primary means for excretion of toxic metals. This paper investigates the possible relationship of the severity of autism to the body burden of toxic metals and RBC glutathione levels. This paper is part of a larger study which investigates the safety and eﬃcacy of DMSA therapy, including both the biological consequences [10] and the DMSA associated behavioural eﬀects [11]. The larger study involves a 3-day round of DMSA, to screen for admission into a 3-month DMSA treatment study; only children with high levels of urinary toxic metals were admitted into the long-term 3-month treatment study. 2. Methods The methodology is discussed in detail in the companion paper [10]. Brieﬂy, this study was conducted with the ap proval of the Human Subjects Institutional Review Board of Southwest College of Naturopathic Medicine. All parents and (where possible) children signed informed consent/assent forms. The study participants were recruited in Arizona, with the help of the Autism Society of America—Greater Phoenix Chapter and the Arizona Division of Developmental Disabilities. The entry criteria were the following. (1) Children with autism spectrum disorder, diagnosed by a psychiatrist, psychologist, or developmental pediatrician. (2) Age 3–8 years. (3) No mercury amalgam dental ﬁllings (due to a concern of their interaction with DMSA). (4) No previous use of DMSA or other prescription chelators. (5) No anemia or currently being treated for anemia due to low iron. (6) No known allergies to DMSA. (7) No liver or kidney disease. (8) Children are well hydrated (receiving adequate daily intake of water). Journal of Toxicology Four metrics were employed to assess the severity of autism: the PDD-BI, ATEC, SAS, and ADOS. Multiple assessment instruments were selected because they each provide insights into various aspects of autism. The ATEC was completed approximately 2-3 weeks prior to taking the DMSA, and the other three instruments were completed approximately 2–4 weeks after the initial 3-day round of DMSA, for children whose excretion of toxic metals was deemed high enough to warrant continuation in the long-term treatment study. The ATEC, PDD-BI, and SAS were assessed by the participant’s parents, and the ADOS evaluation was performed by a certiﬁed ADOS evaluator. It should be noted that the ADOS was developed primarily for diagnosing autism, whereas the other tools were developed for assessing changes in autistic symptoms during treatment studies. DMSA was administered orally in 9 doses of 10 mg/kg, 3 times daily, over 3 days. Urine was collected for approxi- mately 8 hours prior to taking the DMSA, and for approxi- mately 8 hours immediately after the 9th dose, in a process similar to a previous retrospective study of relative body burden of heavy metals [2]. RBC glutathione was measured approximately 1-2 weeks prior to taking the DMSA. The details of measuring the urinary metals and RBC glutathione are given in [10]. The PDD-BI is composed of many subscales. One of the subscales, the Semantic/Pragmatic Problems (SPPs), was diﬃcult to interpret, since children with no spoken language inappropriately scored as less severely aﬀected than those with limited language. Therefore, we exclude the SPP sub- scale in the Autism Composite score, resulting in a modiﬁed Autism Composite score consisting of Sensory/Perceptual Approach, Ritualisms/Resistance to Change, Social Prag- matic Problems, Social Approach Behaviors, Phonological and Semantic Pragmatic subscales. This modiﬁed Autism Composite score was discussed with I. Cohen, the developer of the PDD-BI. We believe that this modiﬁed subscale is more useful because several children initially without speech began talking after DMSA treatment in the study. The development of speech led to a worsening of their score on the SPP, because a nonverbal child is given a score of zero (indicating no semantic/pragmatic problems, which is the same score a typically developed child would receive) compared to a child with limited speech but major semantic/pragmatic problems who would receive a high score on the SPP. Thus, we think the modiﬁed Autism subscale (without the SPP) is more useful for children with very limited or no language. In order to assess global changes in autism severity, a new metric was developed for this study. The Severity of Autism Scale (SAS) is introduced for the ﬁrst time in this series of papers. It is essentially a Clinical Global Impression scale using a 0–10 severity scale, with the diﬀerence being that the scale was made speciﬁc to autism by deﬁning the numeric values (see below). The purpose of the tool is to provide a simple, overall assessment of the severity of the symptoms of autism. In this study we will analyze the correlation of this scale with the other more established assessment tools.