A randomized clinical trial reveals that six months of probiotic supplementation significantly reduced autism severity scores in children without gastrointestinal symptoms, with changes tracked through fecal metabolomics.
For years, parents of children with Autism Spectrum Disorder (ASD) have reported that dietary changes and probiotic supplements seem to help their children. But science has been slow to catch up. A new study published in the journal Metabolites provides some of the strongest evidence yet that the gut-brain connection is real—and that probiotic supplementation may meaningfully reduce autism severity in a specific subgroup of children.
The study, titled “Effects of Probiotic Supplementation on Gut Microbiota and Fecal Metabolome in Autism Spectrum Disorders: A Secondary Analysis of a Randomized Clinical Trial in Preschoolers,” followed 57 children with ASD over six months. Half received a high-potency probiotic mixture (the De Simone Formulation, containing 450 billion live bacteria from eight strains), while the other half received a placebo.
The results were striking—but only for children who did not have gastrointestinal (GI) symptoms at the start of the study. In these children, the probiotic group showed a significant decrease in ADOS-2 scores (the gold-standard measure of autism severity), while the placebo group showed a slight increase. In other words: probiotics helped, and the effect was measurable.
The Study Design: A Closer Look at the Science
The original randomized controlled trial (RCT) included 85 preschool children with ASD. For this secondary analysis, researchers focused on the 57 children for whom complete fecal samples and metabolomics data were available at both the beginning (T0) and end (T2) of the six-month intervention.
The children were divided into two groups based on whether they had gastrointestinal symptoms (GI) or not (NGI) at baseline. GI symptoms included constipation, diarrhea, and abdominal pain—conditions that are significantly more common in children with ASD than in typically developing peers.
The probiotic used was the De Simone Formulation (marketed as Vivomixx® in the EU and Visbiome® in the USA). Each packet contains 450 billion live bacteria from eight strains:
- Streptococcus thermophilus (1 strain)
- Bifidobacterium (3 strains: B. breve, B. longum, B. lactis)
- Lactobacillus (4 strains: L. acidophilus, L. plantarum, L. paracasei, L. helveticus)
The researchers analyzed two things: the children’s fecal microbiome (which bacteria were present) and their fecal metabolome (which small molecules were produced by those bacteria). They then correlated these biological measures with changes in ADOS-2 severity scores.
The Key Finding: Probiotics Work for NGI Children
When all children were analyzed as a single group, there was no significant change in ADOS-2 scores. But when the researchers separated children by GI status, a clear pattern emerged.
For children without gastrointestinal symptoms (NGI):
- The placebo group showed an increase in ADOS-2 scores of +0.19 on average.
- The probiotic group showed a decrease in ADOS-2 scores of -0.68 on average.
- This difference was statistically significant (p = 0.044).
For children with gastrointestinal symptoms (GI):
- No significant difference was found between placebo and probiotic groups.
Changes in Autism Severity (ADOS-2) After 6 Months
| Group | Treatment | Average Change in ADOS-2 Score | Statistical Significance |
|---|---|---|---|
| NGI (No GI Symptoms) | Placebo | +0.19 (worsening) | p = 0.044 (significant) |
| NGI (No GI Symptoms) | Probiotic | -0.68 (improvement) | |
| GI (With GI Symptoms) | Placebo | -0.5 | p = 0.51 (not significant) |
| GI (With GI Symptoms) | Probiotic | -0.22 |
What does this mean? In NGI children, probiotics were associated with a meaningful reduction in autism severity. In GI children, the probiotic did not outperform placebo. This suggests that children with ASD and GI symptoms may have a different underlying gut profile—and may require different interventions.
The Microbiome Shift: What Changed in the Gut?
The researchers measured the relative abundance of several key bacterial groups. Two showed significant changes in response to probiotic administration:
1. Lactobacilli increased significantly with probiotics.
- In NGI children, probiotics increased lactobacilli abundance by 2.10 log10 (median), while placebo showed no change.
- This difference was highly significant (p = 0.002 for Wilcoxon test; p = 0.018 for Chi test).
Lactobacilli are “good bacteria” known to:
- Compete against pathogenic microorganisms
- Improve gastrointestinal barrier function
- Support immune homeostasis
- Protect against inflammatory diseases
The researchers note that “the abundance of lactobacilli is negatively correlated with molecules that characterize high ADOS-2 scores.” In other words, more lactobacilli = less severe autism symptoms.
2. Sutterella decreased significantly with probiotics in NGI children.
- Sutterella abundance decreased in NGI children treated with probiotics, while it increased in all other groups.
- This difference was significant (p = 0.035 for Wilcoxon test; p = 0.028 for Chi test).
Sutterella is a bacterium that has been associated with:
- Infections under certain conditions
- Proteolytic functions that may contribute to gut inflammation
- Potentially harmful effects on the gut barrier
The researchers suggest that “the potential competition of species induced by probiotics might have reduced the proliferation of Sutterella.”
Changes in Gut Bacteria After 6 Months of Probiotics vs. Placebo (NGI Children)
| Bacterial Group | Change with Placebo | Change with Probiotic | Statistical Significance |
|---|---|---|---|
| Lactobacilli | No significant change | Significant increase (p = 0.002) | Probiotics increased beneficial bacteria |
| Sutterella | Increase | Significant decrease (p = 0.035) | Probiotics reduced potentially harmful bacteria |
| Bifidobacteria | No change | No change | Not significant |
| Akkermansia | No change | No change | Not significant |
| Bacteroides | No change | No change | Not significant |
| Prevotella | No change | No change | Not significant |
The Metabolome: Which Molecules Changed?
The researchers also analyzed the fecal metabolome—the collection of small molecules produced by gut bacteria. Several key metabolites changed significantly in NGI children treated with probiotics:
Short-Chain Fatty Acids (SCFAs): Acetate, propionate, butyrate, and isobutyrate all decreased in the probiotic group compared to placebo. This was unexpected because SCFAs are generally considered beneficial. However, the researchers explain that “the intake of lactobacilli through the probiotic administration does not promote a positive effect through the production of SCFAs, but the potential beneficial effects of probiotics should be attributed to other mechanisms of action.”
Amino Acids: Aspartate, leucine, and valine increased in the probiotic group, while tryptophan decreased. Tryptophan is particularly interesting because it is the precursor to serotonin—the “happiness neurotransmitter”—and its metabolism is linked to the gut-brain axis.
The researchers note that “low amounts of both tryptophan and nicotinate might indicate an under-regulation of [the kynurenine] pathway in NGI subjects treated with probiotics.” This pathway is connected to cellular energy metabolism and has been implicated in neuropsychiatric disorders.
Predicting Improvement: A Metabolic Fingerprint
One of the most exciting findings is that the researchers could identify which children were likely to improve—regardless of treatment—based on their baseline (T0) fecal metabolome.
Children who later showed reduced ADOS-2 scores (improvement) had, at the start of the study:
- Lower levels of 4-hydroxyphenylacetate and tyramine
- Higher levels of proline and galactose
These four molecules formed a predictive model. When the researchers projected the samples onto a principal component analysis (PCA) model, children destined to improve appeared at negative PC1 scores, while those destined to worsen appeared at positive PC1 scores. Children whose scores stayed the same appeared in between.
This suggests that the fecal metabolome contains a “fingerprint” that predicts how a child with ASD will respond over time—not just to probiotics, but to the natural course of development and other interventions.
Why This Matters: The Gut-Brain Axis in Autism
The gut-brain axis is the bidirectional communication system between the gastrointestinal tract and the central nervous system. It involves neural pathways (the vagus nerve), immune signaling, and metabolic pathways (bacterial metabolites entering the bloodstream).
In autism, multiple studies have shown that the gut microbiome is altered. Children with ASD often have:
- Reduced microbial diversity
- Lower levels of Prevotella and other fermenters
- Higher levels of certain pathogenic bacteria
This study adds to the evidence by showing that modulating the gut microbiome with probiotics can produce measurable changes in autism severity—at least in children without pre-existing GI symptoms.
The researchers are careful to note that this is an exploratory secondary analysis, not a primary outcome. The sample size is small (57 children), and some subgroups (like GI children) were particularly small. Nevertheless, the consistency of the findings across microbiome, metabolome, and clinical measures is compelling.
Practical Implications for Families
What does this mean for parents of children with ASD?
1. If your child has GI symptoms: This study did not find a benefit of probiotics for autism severity in this subgroup. However, probiotics may still help with the GI symptoms themselves. The original trial did find improvements in GI symptoms (reported elsewhere).
2. If your child does NOT have GI symptoms: This study suggests that a high-potency, multi-strain probiotic (like the De Simone Formulation) may reduce autism severity over six months. The effect size (-0.68 on ADOS-2) is modest but meaningful, especially given that the placebo group worsened slightly (+0.19).
3. The microbiome matters: The significant increase in lactobacilli and decrease in Sutterella in the probiotic group provides a biological mechanism. If you are considering probiotics, look for products containing Lactobacillus strains (especially L. acidophilus, L. plantarum, L. paracasei, and L. helveticus).
4. The metabolome can guide treatment: The finding that baseline levels of 4-hydroxyphenylacetate, tyramine, proline, and galactose predict improvement suggests that, in the future, we may be able to test a child’s fecal metabolome to determine whether probiotics are likely to help.
Limitations and Future Directions
This study has several limitations:
- Small sample size, especially for subgroup analyses.
- Exploratory secondary analysis, not a primary outcome. The findings need replication in larger, prospective trials.
- No dietary control. Diet strongly influences the gut microbiome. The study did not control for what children ate.
- No long-term follow-up. We do not know if the benefits persisted after stopping the probiotic.
- Mechanisms remain unclear. While the study identified correlations, it did not prove causation.
Future research should:
- Conduct larger RCTs powered specifically for NGI children.
- Include dietary tracking and standardization.
- Follow children for longer periods (12-24 months).
- Measure additional outcomes like language, social communication, and adaptive behavior.
A Final Word of Hope
This study is not a cure for autism. It does not suggest that probiotics alone can “treat” ASD. But it does provide rigorous, peer-reviewed evidence that the gut-brain axis is a valid therapeutic target—and that a relatively simple, low-risk intervention (probiotics) can produce measurable improvements in a subgroup of children.
For parents who have been told that dietary and microbiome interventions are “unscientific” or “alternative,” this study offers validation. The science is catching up to what many families have observed: gut health matters for brain health.
And for children with ASD who do not have GI symptoms, a six-month trial of a high-quality, multi-strain probiotic may be a reasonable, evidence-based addition to their care plan.
The researchers conclude: “This work suggests that fecal metabolomics may be useful for monitoring and potentially for predicting changes in ASD severity following probiotic administration, while also providing insights into the underlying biological mechanisms driving these changes.”
It is a small step—but in the right direction.
Reference: here
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