The role of the microbiome in aggression. Gut microbe imbalances that predict autism and ADHD. Biogaia trial for Autism.

 

By December 2020 7.3% of the Swedish cohort born in 1997-9 had been diagnosed with a Neurodevelopmental Disorder (ND). This can be predicted by samples previously collected.

Today’s post is all about the microbiome and covers three different areas covered recently in the research. Eight years after I wrote a post about our informal trial of Biogaia probiotics for autism, we now have a published paper.

Aggression and self injurious behavior (SIB) affects at least half of those diagnosed with level 3 autism at some point in their lives. SIB can become the overriding concern for care givers.

Our first paper looks at the role of the microbiome in aggression.

Gut-brain axis appears to play a critical role in aggression

A series of experiments on mice has found that they become more aggressive when their gut microbiome is depleted. Additionally, transplanting gut microbiota from human infants exposed to antibiotics led to heightened aggression in mice compared to those receiving microbiome transplants from non-exposed infants. The research was published in Brain, Behavior, and Immunity.

In the past decade, scientists have discovered a complex communication pathway linking gut microbiota—the trillions of microorganisms living in the human gut—with the brain. This pathway is called the microbiota-gut-brain axis. It regulates various physiological functions, including digestion and immunity, but also affects mood and behavior. The gut microbiota produces neurotransmitters and other metabolites that can influence brain function through neural, immune, and endocrine pathways.

Recent studies have demonstrated that symptoms of various disorders, once considered primarily psychological or neurological, can be transferred to rodents by transplanting gut microbiota from humans with these disorders. For example, researchers have shown that transplanting gut microorganisms from people with Alzheimer’s disease into mice (whose gut microbiota had been depleted to enhance transplant effectiveness) resulted in cognitive impairments in the mice. Similarly, symptoms of anxiety have been induced in mice by transplanting gut microbiota from humans with social anxiety.

For the humanized mice, the researchers obtained fecal samples from infants who had been exposed to antibiotics shortly after birth, as well as from unexposed infants. These samples were transplanted into five-week-old germ-free mice. The researchers then waited for four weeks before testing the mice for aggression.

To measure aggression, the researchers employed the resident-intruder test, a well-established behavioral assay in which a male mouse (the “resident”) is introduced to another unfamiliar male mouse (the “intruder”) in its home cage. Aggression was quantified based on the latency to the first attack (how quickly the resident mouse attacked the intruder) and the total number of attacks during a 10-minute period.

The results showed that mice raised without gut bacteria (germ-free) and those treated with antibiotics exhibited higher levels of aggression compared to the control group. These mice attacked more frequently and were quicker to initiate aggressive behavior in the resident-intruder test.

The researchers found that humanized mice receiving fecal microbiota from antibiotic-exposed infants were significantly more aggressive than those receiving transplants from non-exposed infants. Even though the infants’ microbiomes had a month to recover after antibiotic exposure, the aggressive behavior was still evident in the recipient mice.

Biochemical analyses revealed that aggressive mice (both germ-free and antibiotic-treated) had distinct metabolite profiles compared to control mice. Specifically, levels of tryptophan—a precursor to serotonin, a neurotransmitter associated with mood and behavior—were elevated in these mice. Additionally, the levels of certain metabolites associated with microbial activity, such as indole-3-lactic acid, were reduced in the aggressive mice, suggesting that the absence of a healthy microbiome might alter key biochemical pathways involved in aggression.

Here is the link to the original paper:

A gut reaction? The role of the microbiome in aggression

Recent research has unveiled conflicting evidence regarding the link between aggression and the gut microbiome. Here, we compared behavior profiles of control, germ-free (GF), and antibiotic-treated mice, as well as re-colonized GF mice to understand the impact of the gut microbiome on aggression using the resident-intruder paradigm. Our findings revealed a link between gut microbiome depletion and higher aggression, accompanied by notable changes in urine metabolite profiles and brain gene expression. This study extends beyond classical murine models to humanized mice to reveal the clinical relevance of early-life antibiotic use on aggression. Fecal microbiome transplant from infants exposed to antibiotics in early life (and sampled one month later) into mice led to increased aggression compared to mice receiving transplants from unexposed infants. This study sheds light on the role of the gut microbiome in modulating aggression and highlights its potential avenues of action, offering insights for development of therapeutic strategies for aggression-related disorders

Note the ABX means antibiotics

We include a study of humanized mice using unique fecal samples of 1-month-old infants, collected nearly a month after early-life ABX administration. In previous work (Uzan-Yulzari et al. 2021, Nat Comm), we have demonstrated that ABX in this critical period of life can have lasting effects of childhood growth. Here, we extend these findings using samples from the same cohort. Using fecal samples collected weeks after ABX administration also reduces the direct chemical effects of ABX on the host, highlighting the causative role of the dysbiotic host microbiome and associated metabolome in driving aggressive behavior. We demonstrate that infant microbiota, perturbed within the first 48 h of life, has a lasting signature through 1 month of age that, when transplanted into GF mice, results in increased aggression (3–5 weeks after transplant) when compared to effects of stools of infants not exposed to any early-life antibiotics. The findings are revolutionary as they show how ABX-altered microbiota during a critical development window can lead to persisting behavioral deficits.

 

Gut microbe imbalances could predict a child’s risk for autism, ADHD and speech disorders years before symptoms appear.

Study Identifies Gut Microbe Imbalances That Predict Autism And ADHD

We are researchers who study the role the microbiome plays in a variety of conditions, such as mental illness, autoimmunity, obesity, preterm birth and others. In our recently published research on Swedish children, we found that microbes and the metabolites they produce in the guts of infants – both found in poop and cord blood – could help screen for a child’s risk of neurodevelopmental conditions such as autism. And these differences can be detected as early as birth or within the first year of life. These markers were evident, on average, over a decade before the children were diagnosed. 

The imbalance in microbial composition – what microbiologists call dysbiosis – we observed suggests that incomplete recovery from repeated antibiotic use may greatly affect children during this vulnerable period. Similarly, we saw that repeated ear infections were linked to a twofold increased likelihood of developing autism.

Children who both repeatedly used antibiotics and had microbial imbalances were significantly more likely to develop autism. More specifically, children with an absence of Coprococcus comes, a bacterium linked to mental health and quality of life, and increased prevalence of Citrobacter, a bacterium known for antimicrobial resistance, along with repeated antibiotic use were two to four times more likely to develop a neurodevelopmental disorder.

Antibiotics are necessary for treating certain bacterial infections in children, and we emphasize that our findings do not suggest avoiding their use altogether. Parents should use antibiotics if they are prescribed and deemed necessary by their pediatrician. Rather, our study suggests that repeated antibiotic use during early childhood may signal underlying immune dysfunction or disrupted brain development, which can be influenced by the gut microbiome. In any case, it is important to consider whether children could benefit from treatments to restore their gut microbes after taking antibiotics, an area we are actively studying.

Another microbial imbalance in children who later were diagnosed with neurodevelopmental disorders was a decrease in Akkermansia muciniphila, a bacterium that reinforces the lining of the gut and is linked to neurotransmitters important to neurological health.

Even after we accounted for factors that could influence gut microbe composition, such as how the baby was delivered and breastfeeding, the relationship between imbalanced bacteria and future diagnosis persisted. And these imbalances preceded diagnosis of autism, ADHD or intellectual disability by 13 to 14 years on average, refuting the assumption that gut microbe imbalances arise from diet.

We found that lipids and bile acids were depleted in the cord blood of newborns with future autism. These compounds provide nutrients for beneficial bacteria, help maintain immune balance and influence neurotransmitter systems and signaling pathways in the brain.

The full paper is here: 

Infant microbes and metabolites point to childhood neurodevelopmental disorders 

Highlights

• Infant microbes and metabolites differentiate controls and future NDs

• Early-life otitis lowers Coprococcus and increases Citrobacter in future NDs

• Preterm birth, infection, stress, parental smoking, and HLA DR4-DQ8 increase ND risk

• Linolenic acid is lower and PFDA toxins higher in the cord serum of future ASD

Summary

This study has followed a birth cohort for over 20 years to find factors associated with neurodevelopmental disorder (ND) diagnosis. Detailed, early-life longitudinal questionnaires captured infection and antibiotic events, stress, prenatal factors, family history, and more. Biomarkers including cord serum metabolome and lipidome, human leukocyte antigen (HLA) genotype, infant microbiota, and stool metabolome were assessed. Among the 16,440 Swedish children followed across time, 1,197 developed an ND. Significant associations emerged for future ND diagnosis in general and for specific ND subtypes, spanning intellectual disability, speech disorder, attention-deficit/hyperactivity disorder, and autism. This investigation revealed microbiome connections to future diagnosis as well as early emerging mood and gastrointestinal problems. The findings suggest links to immune-dysregulation and metabolism, compounded by stress, early-life infection, and antibiotics. The convergence of infant biomarkers and risk factors in this prospective, longitudinal study on a large-scale population establishes a foundation for early-life prediction and intervention in neurodevelopment.

ABIS = All Babies in Southeast Sweden cohort

NDs = Neurodevelopmental disorders

Young children later diagnosed with ASD or exhibiting significant autistic traits tend to experience more ear and upper respiratory symptoms. In ABIS, infants who had otitis in their first year were found to be more prone to acquiring NDs if they lacked detectable levels of Coprococcus or harbored Citrobacter. The absence of Coprococcus, despite comparable levels in controls irrespective of otitis, raises questions about microbial community recovery. This potential failure of the microbiome to recover following such events may serve as a mechanism connecting otitis media to ND risk. Moreover, antibiotic-resistant Citrobacter was more prevalent in these infants. The presence of strains related  to Salmonella and Citrobacter, labeled in this investigation as SREB, was significantly higher in infants who later developed comorbid ASD/ADHD (21%), compared to controls (3%). This disruption may have consequences on neurodevelopment during a critical period. Salmonella and Citrobacter have shown the ability to upregulate the Wingless (Wnt) signaling. The Wnt pathway is vital for immune dysregulation and brain development, and its disruption has been implicated in ASD pathogenesis. 

Two fatty acid differences were notable in the stool of future ASD versus controls: omega-7 monounsaturated palmitoleic acid, (9Z)-hexadec-9-enoic acid (below the level of detection in 87.0% of future ASD but present in 43.5% of controls), and palmitic acid (elevated in future ASD). Palmitoleic acid has been associated with a decreased risk of islet and primary insulin autoimmunity. Conversely, palmitic acid, a saturated fatty acid, has been linked to neuronal homeostasis interference. Its effects are partially protected by oleic acid, which although approaching significance, was lower in the cord serum of future ASD.

Few metabolites were higher in stool of infants with future ASD, but there are a few notable examples: α-d-glucose, pyruvate, and 3-isopropylmalate. Coprococcus inversely correlated with 3-isopropylmalate, suggesting gut-brain connections and a possible imbalance in branched-chain amino acid (BCAA) pathways given the role of 3-isopropylmalate dehydrogenase in leucine and isoleucine biosynthesis. An increase in dehydroascorbate suggests potential disruptions in vitamin C metabolism, crucial for neurotransmitter synthesis and antioxidant defense, while elevated pyruvate suggests disturbance of neurotransmitter synthesis or energy production early in life. Pimelic acid elevation, found in disorders of fatty acid oxidation, suggests disruption of mitochondrial pathways for fatty acid oxidation.

Akkermansia and Coprococcus, absent or reduced in infants with future NDs, positively correlated with signals in stool representing neurotransmitter precursors and essential vitamins in stool. Specifically, Akkermansia correlated with tyrosine and tryptophan (i.e., catecholamine and serotonin precursors, respectively) and Coprococcus with riboflavin. Disruption of BCAA metabolism in ASD has been documented, involving coding variants in large amino acid transporters (LATs) and reduced utilization of trypotphan and large aromatic amino acids along with increased glutamate and decreases in tyrosine, isoleucine, phenylalanine, and tryptophan in children with ASD. Oxidative stress, a diminished capacity for efficient energy transport, and deficiencies in vitamins (like vitamin B2) essential for neurotransmitter synthesis and nerve cell maintenance have been implicated. Riboflavin as an antioxidant reduces oxidative stress and inflammation, demonstrating neuroprotective benefits in neurological disorders, possibly through maintenance of vitamin B6, which is necessary for glutamate conversion to glutamine and 5-hydroxytryptophan to serotonin.

Together, these findings support a hypothesis of early-life origins of NDs, mediated by gut microbiota. This provides a foundation for research and for developing early interventions for NDs.

 

Today’s final paper was highlighted recently in a comment on a post I wrote eight years ago, when we were trialing Biogaia probiotics. This original interest was prompted by a reader sharing her successful experiences of treating her son with severe autism. Perhaps she left the recent comment?

The two bacteria involved are both types of L. reuteri.

L. reuteri 6475 is sold as Biogaia Osfortis

L. reuteri 17938 is sold Biogaia Protectis

The combination of L. reuteri 17938 and L. reuteri 6475 is sold as Biogaia Gastrus.

My old post from 2016:-

Epiphany: Biogaia Trial for Inflammatory Autism Subtypes

The recently published trial:

Precision microbial intervention improves social behavior but not autism severity: A pilot double-blind randomized placebo-controlled trial -

Highlights

• L. reuteri (6475 + 17938) improves social functioning in children with autism

• L. reuteri does not improve overall autism severity or repetitive behaviors

• L. reuteri does not significantly alter microbiome composition or immune profile

•  Only the 6475 strain reverses the social deficits in a mouse model for autism

we performed a double-blind, randomized, placebo-controlled, parallel-design pilot trial in children with ASD. Importantly, we found that L. reuteri, compared with placebo, significantly improved social functioning, both in terms of reducing social deficits, as measured by the social responsiveness scale (SRS^31,32), and increasing adaptive social functioning, as measured by the social adaptive composite score of the Adaptive Behavior Assessment System, Second Edition (ABAS-2³³). L. reuteri did not improve overall autism severity, restricted and repetitive behaviors, and co-occurring psychiatric and behavioral problems, nor did it significantly modulate the microbiome or immune response. Thus, this safe microbial manipulation has the potential for improving social deficits associated with ASD in children.

I had to amend my old post with a warning long ago.

UPDATE: A significant minority of parents report negative reaction to Bio Gaia, this seems to relate to histamine; but more than 50% report very positive effects without any side effects; so best to try a very small dose initially to see if it is not well tolerated. 

Histamine Reaction to BioGaia gastrus

Conclusion

The gut microbiota does indeed play a key role in how your brain functions, but the gut-brain axis works in both directions. What goes on in your brain can affect your gut and not just the other way around. It is called bidirectional signaling.

Antibiotics taken during pregnancy, or during early childhood, will have unintended consequences. Often there is no choice, like for those readers whose baby experienced sepsis at birth (bacterial blood stream infection); you have to give antibiotics to avoid death.

In today’s second paper we see that the researchers are thinking about therapeutical implications. Perhaps the newborn’s gut flora should be repopulated during the weeks after the antibiotic treatment?

I receive many questions about how to treat self injurious behavior that does not respond to anything the doctor has prescribed. Rifaximin, an antibiotic used to treat irritable bowel syndrome with diarrhea, is one therapy that does help some types of SIB (and SIBO, small intestinal bacterial overgrowth, of course). This probably would not surprise the authors of today’s first paper.

Biogaia Gastrus (L. reuteri 6475 + 17938) from today’s third paper worked wonders for the SIB of one reader’s child.

Not surprisingly fecal microbiota transplantation (FMT) can improve SIB in some people.

The Swedish data shows interesting insights such as that lipids and bile acids were depleted in the cord blood of newborns with future autism. The researchers think they can predict the diagnosis of autism or ADHD. The question is and then what? Even when there is a diagnosis of autism, not much changes for most children.