Urinary p-Cresol in Autism Spectrum Disorder

Background:  Autism is a neurodevelopmental disorder, yet the elevated incidence of immune abnormalities, gastrointestinal (GI) symptoms, macrosomy, as well as peripheral markers of disease such as elevated 5-HT blood levels underscore that autism is a systemic disease. We found significantly elevated urinary amounts of p-cresol (4-methylphenol) in autistic children up until 8 years of age. P-cresol can be absorbed through the skin, GI or respiratory tracts, but its main source is represented by some gut bacteria including several clostridia, which express synthetic enzymes able to push the fermentation of tyrosine up to the formation of p-cresol.

Objectives:  (1) To measure urinary amounts of total p-cresol and of its components p-cresylsulphate, p-cresylglucuronate and free p-cresol, in an independent sample of French autistic children and controls, while assessing also correlations with autism severity; (2) to study the behavioral effects of acute p-cresol administration in BTBR mice; (3) to investigate the origin of elevated urinary p-cresol in small autistic children.

Methods: (1) Recruitment of 34 French children (N=17 ASD and 17 matched controls, final N=25 each) and psychodiagnostic battery;  measurement of total p-cresol and components using HPLC–ultraviolet diode array detection; (2) Acute i.v. injection of twelve male BTBR mice at P60 with p-cresol 1 mg/Kg (N=4), 10 mg/Kg (N=4), or vehicle (N=4) and behavioural assessment in the open field, elevated plus maze and object recognition test; (3) parallel assessment of urinary p-cresol, intestinal permeability using the LA/MA test, presence of Clostridial species in the feces as well as toxinA and calprotectin, stool habits and recent antibiotic use in 41 Italian ASD children and 16 controls.

 Results: (1) urinary p-cresol is significantly higher among French ASD cases compared to controls (p<0.05), only prior to age 8 (P<0.01) and with a significant positive correlation with clinical severity; (2) acute p-cresol largely increases time spent in the open arms of the plus maze and decreases exploration of novel objects at the object recognition test (P<0.05 or 0.01), with no effect in the open field; (3) elevated urinary p-cresol does not appear associated with enhanced gut permeability, constipation or presence/titre of fecal Clostridial species, whereas there appears to be an association with recurrent ear infections and recent antibiotic use.  

Conclusions: These preliminary studies replicate and largely extend our initial findings (Altieri et al., Biomarkers 16:252-260, 2011), showing that p-cresol is elevated in a substantial group of small autistic children, it is correlated with more severe autistic behaviors, it is behaviorally active in BTBR mice, and may possibly stem from frequent antibiotic use in part due to recurrent ear infections. If confirmed, they will support the testing of probiotics designed using cresol-resistant species in small autistic children with elevated urinary p-cresol. Furthermore, they will strengthen initial converging evidence pointing toward possible roles for postnatal environmental factors in modulating the severity of ASD, while not causing the disease.