Psychobiotics in pediatric autism spectrum disorder: A systematic review of efficacy, mechanisms, and clinical translation

Table 1 Population, intervention, comparison, and outcome framework for the systematic review on psychobiotics in children with autism spectrum disorder

Component	Description
Population	Children and adolescents aged ≤ 18 years with a clinical diagnosis of ASD based on recognized diagnostic criteria (e.g., DSM or ICD) or validated diagnostic tools
Intervention	Psychobiotic interventions intended to modulate the gut-brain axis, including probiotics, prebiotics, synbiotics, and postbiotics, are administered as supplements, fortified foods, or dietary formulations
Comparator	Placebo, standard/usual care, dietary control, or no intervention
Outcomes	Primary outcomes: Changes in core ASD symptoms measured by validated behavioral scales (e.g., ADOS, CARS, SRS). Secondary outcomes: Gastrointestinal symptoms, quality of life, gut microbiota composition or diversity, immune or metabolic biomarkers, and safety outcomes, including adverse events
Study design	Randomized controlled trials, non-randomized controlled studies, cohort studies, and pre-post intervention studies evaluating psychobiotic interventions in pediatric ASD populations

DSM: Diagnostic and Statistical Manual of Mental Disorders; ICD: International Statistical Classification of Diseases; ADOS: Autism Diagnostic Observation Schedule; CARS: Childhood Autism Rating Scale; SRS: Social Responsiveness Scale.

Table 2 Effects of probiotic supplementation on core autism symptoms

Ref.	Study design	Sample size/age	Probiotic intervention	Duration	Outcomes measured	Main findings
Shaaban et al[28]	Prospective open-label study	n = 30; 5-9 years	Lactobacillus acidophilus, Lactobacillus rhamnosus, Bifidobacterium longum (100 × 106 CFU/g)	3 months	ATEC; 6-GSI; stool microbiota	Significant improvement in autism severity and GI symptoms; increased Lactobacillus and Bifidobacterium levels. Probiotic
Narula Khanna et al[25]	Single-blind randomized placebo-controlled trial	n = 180; 2-9 years	Multi-strain probiotic (12 strains; 9 billion CFU per sachet)	3 months	SRS-2; ABC-2; GSI	Significant reductions in behavioral symptoms vs placebo (SRS-2 improvement 47.77% vs 23.33%, P < 0.001); improvements in constipation and diarrhea. Probiotic
Arnold et al[19]	Randomized placebo-controlled crossover pilot trial	n = 13 (10 completed); 3-12 years	VISBIOME probiotic (8 species, mainly Lactobacillus and Bifidobacterium)	8 weeks per treatment phase	Pediatric Quality of Life Inventory (PedsQL-GI); PRAS-ASD; microbiota analysis	Significant improvement in GI complaints vs placebo (P = 0.02); moderate effect size for GI quality of life. Probiotic
Schmitt et al[29]	Randomized controlled phase Ib trial	n = 15 Participants with ASD (18-22 years)	SB-121 investigational probiotic formulation (Lactobacillus reuteri)	4 weeks	Vineland-3 Adaptive Behavior; social preference (eye-tracking)	Significant increase in Vineland-3 Adaptive Behavior Composite score (P = 0.03); treatment was well-tolerated. Probiotic
Sanctuary et al[21]	Randomized double-blind crossover pilot study	n = 8; 2-11 years with ASD and GI symptoms	Bifidobacterium infantis + bovine colostrum product	12 weeks	GI symptoms, immune markers, behavioral symptoms	Treatment was well-tolerated; reductions in some GI symptoms and certain aberrant behaviors were observed. Probiotic
Wang et al[20], 2020	Interventional study (probiotics + prebiotic)	n = 26 children with ASD	Probiotics + fructo-oligosaccharides	30, 60, 108 days	Gut microbiota composition; SCFAs; neurotransmitters; autism severity	Increased beneficial bacteria (Bifidobacterium longum) and reduction in autism and GI symptom severity after intervention. Probiotic
Mazzone et al[32]	Double-blind randomized placebo-controlled pilot trial	n = 43 Children with ASD (21 received probiotic with a mean age of 5.8 ± 1.3 years)	Lactobacillus reuteri combination (strains ATCC-PTA-6475 and DSM-17938)	6 months	Social functioning measures, autism severity, microbiome composition, and immune profile	Probiotic treatment significantly improved social behavior but did not change overall autism severity, repetitive behaviors, microbiome composition, or immune markers
Santocchi et al[33]	Randomized controlled trial protocol	n = 100 preschool children with ASD	Multi-strain probiotic mixture (Vivomixx®)	6 months	GI symptoms, autism severity, cognitive and language development, biomarkers, neurophysiology	Designed to evaluate effects of probiotics on GI symptoms, autism severity, and neurophysiological patterns; results intended to clarify gut-brain axis mechanisms
Santocchi et al[30]	Double-blind randomized placebo-controlled trial	n = 85 preschoolers with ASD (mean age 42 years)	De Simone Formulation (multi-strain probiotic)	6 months	ADOS-CSS, GI symptoms, adaptive functioning, sensory profile	No overall improvement in autism severity; subgroup without GI symptoms showed improvement in ADOS scores, while children with GI symptoms showed improvements in GI symptoms and adaptive functioning.
Kong et al[26]	Randomized double-blind placebo-controlled pilot trial	n = 35 individuals with ASD aged 3-20 years	Lactobacillus plantarum PS128 (6 × 1010 CFU) followed by PS128 + oxytocin combination therapy	28 weeks (oxytocin added at week 16)	SRS, ABC, CGI, microbiome and inflammatory markers	Combination therapy significantly improved SRS and ABC scores and CGI improvement compared with placebo; microbiome network hubs associated with social cognition improved
Li et al[27]	Prospective randomized controlled trial	n = 41 children with ASD	Oral probiotics + ABA therapy vs ABA alone	3 months	ATEC; gut microbiota composition	Both groups improved, but ATEC scores decreased significantly more in the probiotic + ABA group; probiotics increased beneficial bacteria (Bifidobacterium, Lactobacillus) and reduced Shigella and Clostridium
Meguid et al[34]	Interventional clinical study	n = 40 children with ASD aged 2-5 years	Nutritional supplement containing Bifidobacterium spp. and Lactobacillus spp.	3 months	CARS; ADI-R; GI symptom questionnaire; stool microbiota	Significant increases in probiotic bacterial counts with improvement in sleep, anxiety, and clinical symptoms; findings suggest probiotics may reduce ASD severity and correct dysbiosis
Darwesh et al[35]	Observational microbiome study	Children with ASD vs typically developing controls	No intervention (microbiome analysis of Lactobacillus plantarum, Lactobacillus reuteri, Bifidobacterium longum)	Cross-sectional	CARS, SSP, microbiome PCR analysis	Lower abundance of psychobiotic bacteria in ASD children; correlations found between psychobiotic abundance and sensory and behavioral scores
Billeci et al[31]	Randomized double-blind placebo-controlled trial	Preschool children with ASD	Multi-strain probiotic (same formulation as Santocchi trial)	6 months	EEG parameters, clinical measures, and inflammatory markers	Probiotic treatment modified brain electrical activity (reduced beta/gamma power and increased coherence), suggesting normalization of neural connectivity; EEG changes correlated with clinical measures
Liu et al[24]	Randomized double-blind placebo-controlled trial	n = 80 boys aged 7-15 years (71 completed)	Lactobacillus plantarum PS128	4 weeks	ABC-T, SRS, SNAP-IV, CBCL, CGI	PS128 showed improvements in opposition/defiance, hyperactivity, anxiety, and SNAP-IV scores, particularly in younger children; overall behavioral trends improved
Mensi et al[23]	Real-world observational study	n = 131 children and adolescents with ASD	Lactobacillus plantarum PS128 (3 × 1010-6 × 1010 CFU) vs other probiotics	6 months	CGI	77% showed clinical improvement, particularly younger children; PS128 produced greater improvements and fewer side effects compared with other probiotics

ABA: Applied Behavior Analysis; ABC: Aberrant Behavior Checklist; ABC-2: Aberrant Behavior Checklist, second edition; ABC-T: Aberrant Behavior Checklist-total score; ADI-R: Autism diagnostic interview-revised; ADOS-CSS: Autism diagnostic observation schedule-calibrated severity score; ASD: Autism spectrum disorder; ATEC: Autism Treatment Evaluation Checklist; CBCL: Child Behavior Checklist; CARS: Childhood Autism Rating Scale; CFU: Colony-forming units; CGI: Clinical global impression; EEG: Electroencephalogram; GI: Gastrointestinal; GSI: Gastrointestinal severity index; PedsQL-GI: Pediatric Quality of Life Inventory–gastrointestinal module; PRAS-ASD: Pediatric rating of autism severity-autism spectrum disorder; SB-121: Investigational probiotic (synbiotic) formulation SB-121; SCFAs: Short-chain fatty acids; SNAP-IV: Swanson, Nolan, and Pelham Rating Scale–version IV; SRS: Social Responsiveness Scale.

Table 3 Effects of probiotic supplementation on gastrointestinal symptoms in children with autism spectrum disorder

Ref.	Country	Study design	Participants (n)	Probiotic intervention	Duration	Gastrointestinal outcome measures	Main gastrointestinal findings
Santocchi et al[30]	Italy	Randomized controlled trial	100	Multi-strain probiotic	6 months	GSI	Significant improvements in GI symptoms, particularly in children with baseline gastrointestinal disturbances
Shaaban et al[28]	Egypt	Randomized placebo-controlled trial	30	Multi-strain probiotic	3 months	GI symptom questionnaire	Significant reductions in constipation, abdominal pain, and diarrhea compared with placebo
Narula Khanna et al[25]	India	Randomized placebo-controlled trial	180	Multi-strain probiotic	3 months	GSI	Probiotic supplementation significantly improved constipation and diarrhea scores, with concurrent behavioral improvement
Liu et al[24] (PS128 trial)	Taiwan	Randomized double-blind trial	71	Lactobacillus plantarum PS128	4 weeks	GI symptoms (secondary outcomes)	No major GI changes reported; probiotic effects were primarily observed in behavioral domains

GI: Gastrointestinal; GSI: Gastrointestinal severity index.

Table 4 Probiotic strains, dose, and treatment duration across included autism spectrum disorder studies

Ref.	Probiotic strain(s)	Formulation type	Dose	Treatment duration
Mazzone et al[32]	Lactobacillus reuteri ATCC PTA-6475 + Lactobacillus reuteri DSM-17938	Two-strain probiotic	2 × 108 CFU chewable tablets	6 months
Narula Khanna et al[25]	Multi-strain probiotic formulation containing Lactobacillus and Bifidobacterium species	2-10 × 109 CFU/day	Oral sachet	3 months
Shaaban et al[28]	Multi-strain probiotic (Lactobacillus acidophilus, Lactobacillus rhamnosus, Bifidobacterium longum, Bifidobacterium bifidum)	5 × 109 CFU/day	Oral capsule	3 months
Santocchi et al[30]	De Simone Formulation (Vivomixx®): Lactobacillus plantarum, Lactobacillus paracasei, Lactobacillus acidophilus, Lactobacillus delbrueckii subsp. bulgaricus, Bifidobacterium breve, Bifidobacterium longum, Bifidobacterium infantis, Streptococcus thermophilus	Santocchi multi-strain probiotic	450 billion CFU/day	6 months
Kong et al[26]	Lactobacillus plantarum PS128	Single-strain psychobiotic	6 × 1010 CFU/day	28 weeks
Li et al[27]	2 g/packet, containing at least 2.0 × 107 CFU of live Bifidobacterium longum, Lactobacillus acidophilus, and Enterococcus faecalis)	Multi-strain probiotic	0.5 packets orally three times daily	3 months
Meguid et al[34]	Lactobacillus spp. + Bifidobacterium spp.	Multi-strain nutritional probiotic supplement	108 CFU/g	3 months
Liu et al[24]	Lactobacillus plantarum PS128	Single-strain probiotic	3 × 1010 CFU/ capsule	4 weeks
Liu et al[22]	Lactobacillus plantarum PS128	Single-strain probiotic	6 × 1010 CFU	2 and 4 months
Mensi et al[23]	Lactobacillus plantarum PS128	Single-strain psychobiotic	3 × 1010-6 × 1010 CFU/day	6 months
Billeci et al[31]	Same De Simone multi-strain formulation used in Santocchi trial	Multi-strain probiotic	450 billion CFU/day	6 months
Shaaban et al[28]	Lactobacillus acidophilus, Lactobacillus rhamnosus, Bifidobacterium longum (100 × 106 CFU/g)	Multi-strain probiotic	5 gm/day	3 months
Arnold et al[19]	VISBIOME probiotic (8 species, four strains of lactobacilli (Lactobacillus casei, Lactobacillus plantarum, Lactobacillus acidophilus, and Lactobacillus delbrueckii subsp. bulgaricus), three strains of bifidobacteria (Bifidobacterium longum, Bifidobacterium infantis, and Bifidobacterium breve), and one strain of Streptococcus thermophilus	Multi-strain probiotic	900 billion CFU/day	8 weeks
Schmitt et al[29]	Lactobacillus reuteri	Single-strain probiotic	2 × 1010 CFU/day	4 weeks
Sanctuary et al[21]	Bifidobacterium infantis + bovine colostrum product	Single-strain probiotic	20 billion CFU/day	5 weeks
Wang et al[20]	Bifidobacterium infantis Bi-26, Lactobacillus rhamnosus HN001, Bifidobacterium lactis BL-04, and Lactobacillus paracasei	Multi-strain probiotic	1010 CFU/day	30, 60, 108 days

ATCC PTA: American type culture collection, patent and trademark office designation; Bi-26: Strain designation Bi-26; BL-04: Strain designation BL-04; CFU: Colony-forming units; DSM: Deutsche Sammlung von Mikroorganismen und Zellkulturen; HN001: Strain designation HN001; PS128: Strain designation PS128; spp: Species (plural).

Table 5 Mechanistic effects of psychobiotics in autism spectrum disorder

Ref.	Psychobiotic intervention	Microbiome changes	Metabolic/neurochemical changes	Immune/inflammatory effects	Proposed clinical impact
Santocchi et al[30]	Multi-strain probiotic (Lactobacillus, Bifidobacterium, Streptococcus)	Increased abundance of beneficial gut bacteria; improved microbial diversity	Modulation of SCFA production	Reduction in gut inflammation and improved intestinal barrier function	Improvement in gastrointestinal symptoms and potential behavioral benefits in subgroups
Liu et al[24] (PS128)	Lactobacillus plantarum PS128	Increased Lactobacillus colonization	Modulation of neurotransmitter-related metabolites, including serotonin and dopamine pathways	Possible modulation of neuroinflammatory signaling	Improvements in hyperactivity, anxiety, and behavioral regulation
Shaaban et al[28]	Multi-strain probiotic (Lactobacillus, Bifidobacterium)	Increased beneficial bacterial taxa and a reduction in potentially pathogenic microbes	Normalization of microbial metabolic products	Decreased systemic inflammatory markers	Reduced ASD symptom severity and improved gastrointestinal function
Narula Khanna et al[25]	Multi-strain probiotic formulation	Increased Bifidobacterium and Lactobacillus species	Increased production of SCFAs is associated with gut barrier integrity	Reduced pro-inflammatory cytokine activity	Improvement in social responsiveness, stereotypic behavior, and gastrointestinal symptoms
Other pilot studies	Various probiotic combinations	Improved microbial diversity and reduction of dysbiosis	Altered microbial metabolite profiles affecting gut-brain signaling	Modulation of immune responses and gut permeability	Potential improvements in both behavioral and gastrointestinal outcomes

ASD: Autism spectrum disorder; SCFAs: Short chain fatty acids.

Table 6 Behavioral scales used across included studies evaluating psychobiotics in children with autism spectrum disorder

Ref.	Behavioral scale	Full name	Main domains assessed	Clinical relevance in ASD trials
Santocchi et al[30]	ADOS-CSS	Autism Diagnostic Observation Schedule-Calibrated Severity Score	Social communication, restricted and repetitive behaviors	Considered a gold-standard observational measure of ASD symptom severity and is commonly used in clinical trials
Narula Khanna et al[25]; Liu et al[24]; Kong et al[26]	SRS/SRS-2	Social Responsiveness Scale	Social awareness, social cognition, social communication, social motivation, restricted interests	Widely used caregiver-reported scale for measuring social impairment and treatment response in ASD
Narula Khanna et al[25]; Liu et al[24]	ABC	Aberrant Behavior Checklist	Irritability, hyperactivity, stereotypy, lethargy/social withdrawal, inappropriate speech	Frequently used in pharmacological and behavioral intervention trials to evaluate behavioral changes
Shaaban et al[28]	CARS	Childhood Autism Rating Scale	Overall autism severity, including social interaction, communication, emotional response, and sensory behaviors	Clinician-rated scale used for diagnostic assessment and monitoring changes in autism severity
Shaaban et al[28]	ATEC	Autism Treatment Evaluation Checklist	Speech/language communication, sociability, sensory/cognitive awareness, health/behavior	Commonly used in intervention studies to assess treatment-related changes in multiple ASD domains
Liu et al[24]	CBCL	Child Behavior Checklist	Emotional and behavioral problems including anxiety, depression, and social functioning	Measures broader behavioral and psychological symptoms that may co-occur with ASD
Liu et al[24]	SNAP-IV	Swanson, Nolan, and Pelham Rating Scale	Attention deficit, hyperactivity, and impulsivity symptoms	Useful for assessing ADHD-related behaviors often present in children with ASD

ABC: Aberrant Behavior Checklist; ADI-R: Autism diagnostic interview-revised; ADOS-CSS: Autism diagnostic observation schedule-calibrated severity score; ADHD: Attention-deficit/hyperactivity disorder; ASD: Autism spectrum disorder; ATEC: Autism treatment evaluation checklist; CBCL: Child Behavior Checklist; CARS: Childhood Autism Rating Scale; CGI: Clinical global impression; SNAP-IV: Swanson, Nolan, and Pelham Rating Scale-version IV; SRS: Social Responsiveness Scale.

Table 7 Characteristics of included prebiotic studies in children with autism spectrum disorder

Ref.	Study design	Sample characteristics	Prebiotic type	Dose	Duration	Microbiome outcomes	Clinical outcomes
Grimaldi et al[36], 2018	Dietary intervention study	30 children with ASD	B-GOS	1.8 g (80% GOS content)	6 weeks	Increased Lachnospiraceae; altered fecal and urinary metabolites	Improvement in anti-social behavior
Palmer et al[37], 2025	Double-blind randomized placebo-controlled trial	33 children with ASD (4-10 years)	GOS	2.4 g/day	6 weeks	Threefold increase in Bifidobacterium (1.4%-5.9%, P < 0.001)	No significant behavioural differences vs placebo; moderate improvement in GI symptoms (effect size d = 0.47)
Raghavan et al[38], 2022	Randomized parallel-group pilot study	18 children with ASD	β-glucan (Nichi Glucan)	0.5 g twice daily	90 days	Not evaluated	Significant reduction in CARS scores (P = 0.034); increased plasma α-synuclein levels
Inoue et al[39], 2019	Clinical dietary supplementation study	Children with ASD and constipation	PHGG	6 g/day	2-15 (median = 2 months)	Altered gut microbiota composition	Increased defecation frequency, reduced IL-1β and TNF-α, decreased behavioral irritability
Saxami et al[40], 2023	In vitro microbiome fermentation study	Fecal samples from autistic and neurotypical children	Mushroom-derived prebiotics (Pleurotus eryngii, Pleurotus ostreatus)	Not applicable	24-hour fermentation model	Increased Bifidobacterium, Bacteroides, and Faecalibacterium prausnitzii; increased butyrate production	Mechanistic study (no direct clinical outcomes)
Grimaldi et al[41], 2017	In vitro gut model study	ASD and neurotypical fecal microbiota	B-GOS prebiotic	Not applicable	Gut model simulation	Increased Bifidobacterium and Lactobacillus; altered SCFA production	Mechanistic microbiome outcomes only

ASD: Autism spectrum disorder; B-GOS: Bimuno^® galacto-oligosaccharide; GOS: Galacto-oligosaccharides; IL-1β: Interleukin-1 beta; PHGG: Partially hydrolyzed guar gum; SCFAs: Short-chain fatty acids; TNF-α: Tumor necrosis factor alpha.

Table 8 Characteristics of included synbiotic studies in autism spectrum disorder

Ref.	Study design	Participants (n)	Age range	Synbiotic intervention	Duration	Main outcomes assessed	Key findings
Sanctuary et al[21]	Randomized double-blind crossover pilot study	8 children with ASD and GI symptoms	2-11 years	Bifidobacterium infantis + bovine colostrum product (prebiotic oligosaccharides)	12 weeks (5 weeks synbiotic + washout + 5 weeks prebiotic alone)	Gastrointestinal symptoms, behavioral symptoms, and inflammatory markers	Combination treatment was well tolerated and associated with reduced GI symptoms and some improvements in aberrant behaviors; decreases in IL-13 and TNF-α were observed in some participants. Symbiotics
Wang et al[20]	Controlled intervention study	26 children with ASD (16 synbiotic, 10 placebo)	2-8 years	Probiotics + FOS	30, 60, and 108 days	Autism severity, gut microbiota composition, SCFAs, neurotransmitters	Synbiotic treatment increased beneficial bacteria (Bifidobacterium longum), elevated SCFAs, normalized serotonin and dopamine metabolites, and significantly reduced autism and GI symptom severity. Symbiotics
Schmitt et al[29]	Randomized double-blind placebo-controlled crossover trial	15 participants with ASD	15-45 years	SB-121 synbiotic (Limosilactobacillus reuteri + dextran microparticles + maltose)	28-day treatment periods	Adaptive behavior (Vineland-3), social preference, safety	Significant improvement in Vineland-3 Adaptive Behavior Composite score (P = 0.03) and trends toward improved social preference; treatment was safe and well tolerated. Symbiotics
Phan et al[42]	Open-label clinical study	170 ASD participants completed the study	10.4 ± 7.1 years	Precision synbiotic formulation targeting microbiome diversity	3 months	ASD symptoms, GI symptoms, and metagenomic microbiome profiling	Synbiotic supplementation increased microbiome diversity and improved GI discomfort and several ASD-related symptoms including language, cognition, and communication. Symbiotics
Mitchell et al[44]	Randomized pilot trial	40 children with ASD	5-10 years	Synbiotics alone vs synbiotics + gut-directed hypnotherapy	12 weeks (24-week follow-up)	GI symptom scores, behavior, anxiety, microbiome composition	Both groups showed significant reductions in GI symptoms (P < 0.001); the combined therapy group also showed additional reductions in anxiety and irritability. Symbiotics
Wong et al[43]	Open-label pilot study	30 children with ASD	Mean age 82 years	SCM06 synbiotic formulation	12 weeks	Anxiety, sensory hyperresponsiveness, abdominal pain, microbiome metabolomics	Significant improvements in anxiety, sensory hyperresponsiveness, and abdominal pain; increases in Bifidobacterium pseudocatenulatum and SCFAs were observed. Symbiotics

ASD: Autism spectrum disorder; FOS: Fructo-oligosaccharides; GI: Gastrointestinal; IL-13: Interleukin-13; SB-121: Investigational synbiotic formulation SB-121; SCFAs: Short-chain fatty acids; TNF-α: Tumor necrosis factor alpha.

Table 9 Synbiotic strains, prebiotic components, dose, treatment duration, and reported outcomes in autism spectrum disorder studies

Ref.	Probiotic strain(s)	Prebiotic component	Dose	Treatment duration	Microbiome outcomes	Clinical outcomes
Sanctuary et al[21]	Bifidobacterium longum subsp. infantis	Bovine colostrum product containing oligosaccharides	0.15 g/Lb body weight/day + FOS	5 weeks synbiotic phase (within 12-week crossover design)	Changes in gut microbial composition; modulation of immune markers, including decreased IL-13 and TNF-α	Reduction in gastrointestinal symptoms and modest improvements in aberrant behavior scores in some participants. Symbiotics
Wang et al[20]	Multi-strain probiotic mixture (including Bifidobacterium longum)	FOS	1010 CFU/pack/day	30, 60, 108 days	Increased abundance of beneficial bacteria (Bifidobacterium longum); increased SCFAs	Significant reduction in autism severity scores and gastrointestinal symptoms; normalization of serotonin and dopamine metabolites. Symbiotics
Schmitt et al[29]	Limosilactobacillus reuteri (SB-121 formulation)	Dextran microparticles with maltose carrier	2 × 1010 CFU of Limosilactobacillus reuteri, + 200 mg Sephadex, + 74 mmol/L of maltose in a final volume of 10.8 mL	28-day treatment periods (crossover design)	Designed to enhance bacterial adherence and colonization	Significant improvement in Vineland-3 Adaptive Behavior Composite score; improved social preference trends; treatment well tolerated
Phan et al[42]	Multi-strain probiotic formulation	Precision synbiotic formulation (specific prebiotic composition not specified)	Personalized	3 months	Increased microbial diversity; restoration of beneficial taxa such as Faecalibacterium and Prevotella; reduction of pathogenic taxa including Shigella and Klebsiella	Improvements in gastrointestinal discomfort and ASD-related symptoms including language, cognition, and communication
Mitchell et al[44]	Multi-strain probiotic blend	Prebiotic component included within the synbiotic formulation	5 g of PHGG and a probiotic mixture	12 weeks	Increased abundance of beneficial taxa including Bifidobacterium animalis and Dialister	Significant reductions in gastrointestinal symptom scores; improvements in anxiety and irritability when combined with hypnotherapy
Wong et al[43]	Synbiotic formulation SCM06, including probiotic strains	Prebiotic component included in SCM06	Maltodextrin + galactooligosaccharide + 5 × 109 CFU of 4 probiotic species: Bifidobacterium bifidum, Bifidobacterium longum, Lactobacillus plantarum, and Streptococcus thermophilus	12 weeks	Increased abundance of Bifidobacterium pseudocatenulatum and increased SCFAs (butyrate and valeric acid)	Improvements in anxiety, sensory hyperresponsiveness, and abdominal pain in children with ASD

ASD: Autism spectrum disorder; CFU: Colony-forming units; FOS: Fructo-oligosaccharides; IL-13: Interleukin-13; PHGG: Partially hydrolyzed guar gum; SB-121: Investigational synbiotic formulation SB-121; SCFAs: Short-chain fatty acids; TNF-α: Tumor necrosis factor alpha.

Table 10 Characteristics of included studies investigating fecal microbiota transplantation in autism spectrum disorder

Ref.	Study design	Participants	FMT type/delivery	Outcomes assessed	Main findings
Wang et al[45]	Randomized double-blind placebo-controlled trial	41 children with ASD (39 boys and 3 girls, aged 4-12 years)	FMT vs placebo	GSRS, CARS, ABC, SRS; urinary metabolites	Significant improvements after FMT: GSRS decreased (30.17 → 19), CARS (36.22 → 33.33), SRS (151.17 → 137.5), ABC (76.39 → 53.17). Urinary 5-HIAA decreased, suggesting altered serotonin metabolism
Liu et al[53]	Prospective interventional study	24 ASD children initially; 18 second course; 13 third; 8 fourth	Fresh WMT	ASD symptoms, sleep disorder, constipation, microbiome metabolites	WMT improved behavioral symptoms, sleep disturbances, and constipation. Microbiome shifts included ↓Bacteroides, Flavonifractor, Parasutterella and ↑Prevotella
Chen et al[54]	Microbiome mechanistic study within FMT clinical dataset	ASD patients with GI comorbidities	Encapsulated FMT	Metagenomic microbial dynamics	Donor-recipient microbial interactions influenced subspecies transfer and clinical response, suggesting microbial compatibility affects FMT success
Li et al[47]	Prospective single-arm study	98 children with ASD	FMT via capsules, transendoscopic enteral tube, or nasojejunal tube	ABC, CARS, SRS, GSRS, SDSC; adverse events	Improvements in ASD symptoms, GI symptoms, and sleep disturbances. Capsules and nasojejunal tube showed greater symptom reduction than transendoscopic enteral tube. No serious adverse events
Li et al[48]	Prospective clinical study	38 ASD children; 30 healthy controls	Oral lyophilized FMT (every 4 weeks for 12 weeks)	ABC, CARS, SRS, SDSC; microbiome composition	After treatment: ABC ↓23%, CARS ↓10%, SRS ↓6%, SDSC ↓10%. Gut bacterial and fungal composition shifted toward healthier profiles
Li et al[49]	Open-label clinical trial	40 children with ASD (age 3-17 years)	FMT	GI symptoms, ASD symptoms, gut microbiota, neurotransmitters	FMT improved GI and behavioral symptoms, altered serum neurotransmitters, and promoted colonization of donor microbes
Pan et al[50]	Retrospective study	55 Children with ASD (median age: 6 years, 80.95% were male)	Repeated WMT	ABC, CARS, SDSC, GI symptoms, and inflammatory markers	WMT improved ASD symptoms, GI symptoms, and sleep disorders. Multiple treatment courses produced greater improvements
Zhang et al[51]	Retrospective observational study	49 children with ASD	WMT	Sleep disturbance (SDSC), stool characteristics (BSFS)	WMT improved sleep disorders and constipation; behavioral symptoms also improved with no serious adverse events
Kang et al[46]	Open-label clinical trial	18 children with ASD	Microbiota transfer therapy (antibiotics + bowel cleanse + FMT)	GI symptoms, ASD behavioral measures, microbiome	80% reduction in GI symptoms and significant improvements in ASD behaviors; increased bacterial diversity and beneficial taxa
Kang et al[52]	Long-term follow-up study	Same 18 participants from the previous trial	Microbiota transfer therapy	GI symptoms, ASD symptoms, gut microbiota	Improvements in GI symptoms and ASD behaviors were maintained for 2 years, with sustained microbiome changes

ABC: Aberrant Behavior Checklist; ASD: Autism spectrum disorder; BSFS: Bristol Stool Form Scale; CARS: Childhood Autism Rating Scale; FMT: Fecal microbiota transplantation; GI: Gastrointestinal; GSRS: Gastrointestinal Symptom Rating Scale; MTT: Microbiota transfer therapy; SDSC: Sleep Disturbance Scale for Children; SRS: Social Responsiveness Scale; WMT: Washed microbiota transplantation.

Table 11 Overall quantitative synthesis of fecal microbiota transplantation effects on autism spectrum disorder core symptoms and gastrointestinal manifestations

Outcome	Number of studies	Total participants	Pooled SMD	Interpretation
GSRS	2	37	-2.71	Very large GI improvement
CARS	3	135	-1.60	Large improvement in ASD severity
ABC	2	116	-3.00	Very large behavioral improvement
SRS	3	135	-4.06	Very large improvement in social responsiveness

GI: Gastrointestinal; ASD: Autism spectrum disorder; ABC: Aberrant Behavior Checklist; CARS: Childhood Autism Rating Scale; GSRS: Gastrointestinal Symptom Rating Scale; SMD: Standardized mean difference; SRS: Social Responsiveness Scale.

Table 12 Evidence-based selection of psychobiotic strains and formulations for targeted symptom management in children with autism spectrum disorder

Clinical target	Evidence-based psychobiotic	Key mechanism
Social functioning	Limosilactobacillus reuteri (combined strains)	Oxytocin signaling modulation via the Vagus nerve
Anxiety/mood	Lactiplantibacillus plantarum PS128	Modulation of dopamine and serotonin metabolism
Behavioral irritability	Multi-strain Lacto/Bifido blends	reduction in systemic inflammation/pro-inflammatory cytokines
GI severity	De Simone formulation/Bifidobacterium infantis	Enhancement of intestinal barrier and SCFA production
Global improvement	Fecal microbiota transplantation	Ecosystem-wide restoration of microbial diversity

This table provides a clinical translation of the systematic review findings, mapping specific microbial interventions to their corresponding therapeutic targets within the ASD phenotype. Recommendations are categorized by the primary clinical presentation: (1) Strain specificity: Highlights the transition toward “precision psychobiotics”, where specific strains like Limosilactobacillus reuteri and Lactiplantibacillus. plantarum PS128 are utilized for neurological and emotional modulation rather than general gut health; (2) Clinical metrics: Effects are based on improvements in validated instruments, including the Autism Diagnostic Observation Schedule, Social Responsiveness Scale, Aberrant Behavior Checklist, and the Gastrointestinal Severity Index; and (3) Formulations: Includes single-strain psychobiotics, standardized multi-strain blends (e.g., De Simone Formulation), and ecosystem-wide interventions like Fecal Microbiota Transplantation. These interventions are intended as adjunctive therapies alongside standard behavioral and educational supports. Specific dosing should be adjusted based on individual patient tolerance and clinical response. GI: Gastrointestinal; SCFA: Short-chain fatty acids.