[38] | ASD (n = 23) divided into ASD w GI symptoms (12/23) and ASD w/o GI symptoms (11/23) Range Age 3–10 y (15/23 aged 3–5 y; 6/23 aged 6–7 y; 2/23 aged 8–10 y) All males | HC w GI symptoms (n = 9) Range Age 3–10 y (7/9 aged 3–5 y; 1/9 aged 6–7 y; 1/9 aged 8–10 y). All males | Pyrosequencing of the 16S rRNA gene pan-bacterial (V2 region) on ileal and ceca mucosal biopsy samples PCR-based detection of Sutterella 16S rRNA gene sequences (V6–V8 region and C4–V8 region) on ileal and ceca mucosal biopsy samples | Sutterella ↑ in ASD w GI symptoms vs. HC w GI symptoms (ilea p = 0.022/ceca p = 0.037) | No correlations between GM and GI symptoms were studies | ASD severity was not indicated Was not indicated whether the ASD subjects had intellectual disability |
[39] | ASD (n = 51) divided into ASD w GI symptoms (28/51) and ASD w/o GI symptoms (23/51) Age 2–12 y 42 males 9 females | NT Sib (n = 53) Age 2–12 y 19 males 34 females | Pyrosequencing (bTEFAP) of the 16S rRNA gene bacterial (V1–V3 regions) on fecal samples | No difference in microbiome between total ASD vs. NT Sib. No difference regarding severity ASD in the bacterial composition. No difference regarding GI symptoms dysfunction in the bacterial composition (No differences within the ASD group compared to both with and without GI symptoms groups) | No metabolites were studied | Was not indicated whether the ASD subjects had intellectual disability |
[27] | ASD w GI symptoms (n = 20) Age 3–16 y 18 males 2 females | NT w GI symptoms (n = 20) Age 3–16 y 17 males 3 females | Pyrosequencing (bTEFAP) of the 16S rRNA gene bacterial (V2–V3 regions) on fecal samples Quantitative real-time PCR for Prevotella | Prevotella (p = 0.04), Coprococcus (p ≤ 0.06) and unclassified Veillonellaceae (p = 0.04) ↓ in ASD vs. NT. Microbial changes were more closely linked to the presence of autistic symptoms rather than to the severity of GI symptoms and specific diet/supplement regimens | No correlations between GM and GI symptoms were studies | Was not indicated whether the ASD subjects had intellectual disability |
[40] | ASD (n = 23) w or w/o GI symptoms Age 3–18 y Unrevealed sex | HC (n = 9) and NT sib (n = 22) w or w/o GI symptoms Age 3–18 y Unrevealed sex | qPCR for Sutterella, Ruminococcus torques and R. gnavus on fecal samples. | Sutterella ↑ (p = 0.044) in ASD vs. NT Sib. Sutterella ↑ (p = 0.05) in ASD vs. HC. Sutterella ↑ (p = 0.047) in NT Sib vs. HC. Ruminococcus gnavus ↑ (p = 0.046) in NT Sib vs. HC. Ruminococcus torques ↑ (p = 0.008) in ASD w GI symptoms (n = 9) vs. ASD w/o GI symptoms (n = 14). | No correlations between GM and GI symptoms were studies | ASD severity was not indicated Was not indicated whether the ASD subjects had intellectual disability |
[41] | ASD (n = 59) divided into ASD w GI symptoms (25/59) and ASD w/o GI symptoms (34/59) Age 7–14 y 52 males 7 females | NT Sib (n = 44) divided into NT Sib w GI symptoms (13/44) and NT Sib w/o GI symptoms (31/44) Age 7–14 y 21 males 23 females | qPCR for total bacteria, Sutterella subgroup, Bacteroidetes subgroup, Prevotella, C. coccoides-E. rectales subgroup, Faecalibacterium prausnitzii and Escherichia coli subgroup on fecal samples Sequencing of the 16S rRNA gene bacterial (V1–V2 and V1–V3 regions) on fecal samples | No difference between total ASD vs NT Sib in bacterial frequency. Increased prevalence of functional constipation GI symptoms in ASD children compared to NT siblings | Increased prevalence of functional constipation GI symptoms in ASD children compared to NT siblings. | ASD severity was not indicated Was not indicated whether the ASD subjects had intellectual disability |
[31] | ASD (n = 10) divided into ASD w GI symptoms (9/10) and ASD w/o GI symptoms (1/10) Age 2–9, 9 males and 1 female Sib (n = 9) divided into Sib w GI symptoms (7/9) and Sib w/o GI symptoms (2/9) Age 5–17, 7 males and 3 females | HC (n = 10) divided into HC w GI symptoms (6/10) and HC w/o GI symptoms (4/10) Age 2–11, 10 males | qPCR for Bacteroidetes, Firmicutes, Bifidobacterium, Lactobacillus, Clostridium cluster 1, S. thermophiles, Desulfovibrio on fecal samples | Bacteroidetes/Firmicutes ratio↓ in ASD vs. HC (p < 0.05); ↓ Sib vs. ASD (p < 0.05); ↓ Sib vs. HC (p < 0.05) Firmicutes ↑ in Sib vs. HC Lactobacillus ↑ in ASD vs. HC (p < 0.05) Bifidobacterium ↑ in ASD vs. Sib (p < 0.05) | Desulfovibrio with ASD severity in the ADI restricted/repetitive behavior subscale score. There is a correlation of the autism severity with the severity of GI dysfunction. (p = 0.01) No correlation of GI symptoms and Desulfovibrio abundance. | Was not indicated whether the ASD subjects had intellectual disability |
[42] | ASD (n = 40) divided into severe ASD (36/40) and moderately severe ASD (4/40) 5 constipated 29 non-constipated Average age 11.1 ± 6.8 31 males 9 females | NT (n = 40) 11 constipated 29 non-constipated Average age 9.2 ± 7.9 28 males 12 females | Pyrosequencing of the 16S rRNA gene bacterial (V3–V5 regions) and the internal transcribed spacer (ITS) for fungal (ITS1 rDNA region) on fecal samples | Firmicutes/Bacteroidetes ratio↑, Bacteroidetes ↓, Veillonella ↓, Alistipes ↓, Bilophila↓, Dialister↓, Parabacteroides↓ in ASD vs. NT (p < 0.005) Lactobacillus ↑, Dorea ↑, Corynebacterium ↑, Collinsella ↑, Candida ↑ in ASD vs. NT (p < 0.001) | Escherichia/Shigella and cluster XVIII with GI symptoms or constipation (p < 0.05) | Was not indicated whether the ASD subjects had intellectual disability |
[30] | Severe ASD (n = 47) w/o or w GI symptoms Average age 6 ± 2.8 y 40 males 7 females | HC (n = 33) w/o or w GI symptoms Average age 7.3 ± 3.1 y 24 males 9 females | Examination and culture of fecal samples: (a) morphological examination, (b) microscopic examination staining, (c) search for toxins a/b of Clostridium difficilis, (d) bacterial/yeast culture and (e) identification of bacteria and yeast colonies by VITEK 2 microbial identification system | Candida aggressive form (pseudo-hyphae presenting) Candida ↑ in ASD vs. HC (p = 8.67 × 10−6) Lactobacillus ↓ in ASD vs. HC (p = 7.28x10−4) Clostridium ↓ in ASD vs. HC (p = 0.01) | GI symptoms in 70.2% ASDs and no controls, with a mild correlation by multivariate analyses of constipation and alternating bowel versus increased permeability to lactulose. | Was not indicated whether the ASD subjects had intellectual disability |
[43] | ASD w GI symptoms (n = 14) Age 4–13 y 14 males | NT w GI symptoms (n = 15) Age 3–18 y 12 males 3 females NT w/o GI symptoms (n = 6) Age 3–18 y 6 males | Sequencing of the 16S rRNA gene (V1–V3 and V4 regions) on rectum mucosal biopsy samples | Blautia ↓ (p = 0.02), Dorea ↓ (p = 0.006), Sutterella ↓ (p = 0.025) in ASD w GI symptoms vs. NT w GI symptoms Clostridium lituseburense ↑ (p = 0.002), Lachnoclostridium bolteae ↑ (p = 0.017), Lachnoclostridium hathewayi ↑ (p = 0.03), Clostridium aldenense ↑ (p = 0.03), and Flavonifractor plautii ↑ (p = 0.03), in ASD w GI symptoms vs. NT w GI symptoms and NT w/o GI symptoms Faecalibacterium prausnitzii ↑, Roseburia intestinalis ↑, Oscillospira valericigenes ↑, and Bilophila wadsworthia ↑ (p < 0.05) in NT w GI symptoms vs. NT w/o GI symptoms | IBS with ↑ Clostridium aldenense (p = 0.04); Functional constipation with ↓ Flavonifractor plautii (p = 0.03), Bacteroides eggerthii (p = 0.02), Bacteroides uniformis (p = 0.04), Faecalibacterium prausnitzii (p = 0.013), Clostridium clariflavum (p = 0.03); Aerophagia with ↑ Clostridium aldenense (p = 0.03), ↓ in Blautia luti (p = 0.003), Bifidobacterium adolescentis (p = 0.01), Eubacterium ventriosum (p = 0.05), Anoxystipes fissicatena (p = 0.02), Coprococcus comes (p = 0.04), Eubacterium ramulus (p = 0.006), and Phascolarctobacterium faecium (p = 0.04); Abdominal migraine with ↓ in Akkermansia muciniphila (p = 0.03), Coprococcus catus (p = 0.007), Odoribacter splanchnicus (p = 0.05), Clostridium lactatifermentans (p = 0.03) and Ruminococcus lactaris (p = 0.03) Serotonin with Lachnoclostridium bolteae (p = 0.002), Lachnoclostridium hathewayi (p = 0.003) and Flavonifractor plautii (p = 0.001) | ASD severity was not indicated Was not indicated whether the ASD subjects had intellectual disability |
[29] | ASD w GI symptoms (n = 33) Age 2–9 y Unidentified sex | HC w/o GI symptoms (n = 13) Age 2–9 y Unidentified sex | Selective culture methods for Clostridium and for Clostridium perfringens strains on fecal samples: Brucella and CDC agar. PCR for Clostridium perfringens toxin genes: alpha (cpa), beta (cpb), beta 2 (cpb2), epsilon (ctx), iota (iA), and enterotoxin (cpe) | Clostridium perfringens ↑ in ASD w GI symptoms vs. HC (p = 0.03) | No correlations between GM and GI symptoms were studies | ASD severity was not indicated Was not indicated whether the ASD subjects had intellectual disability |
[44] | ASD (n = 21) w GI symptoms Age 14.4 ± 1.1 y 19 males 2 females | HC (n = 19) w GI symptoms Age 16 ± 1.2 y 10 males 9 females | Pyrosequencing of the 16S rRNA gene bacterial (V1–V3 regions) on duodenal biopsies samples from the second part of the duodenum | No differences in microbiome diversity (alpha and beta) Burkholderia ↑ (p = 0.03) and Neisseria ↓ (p = 0.01) in ASD vs. HC Bacteroides vulgatus ↓ (p = 0.005), unidentified Bacteroides ↓ (p = 0.04), and Escherichia coli ↓ (p = 0.05) in ASD vs. HC Oscillospira, Actinomyces, Peptostreptococcus, and Ralstonia ↑ (p < 0.05) in ASD vs. HC Devosia, Prevotella, Bacteroides, and Streptococcus ↓ (p < 0.05) in ASD vs. HC | ↑ frequency of constipation in ASD vs. HC (p < 0.005) | ASD severity was not indicated Was not indicated whether the ASD subjects had intellectual disability |
[45] | ASD (n = 29) w GI symptoms (49 isolated strains of Clostridium perfringens) Age 3.5–18 y 23 males 9 females | HC (n = 17) (30 isolated strains of Clostridium perfringens) Obese children (n = 24) (32 isolated strains of Clostridium perfringens) Unrevealed age and sex | Selective culture method for fecal samples: Columbia blood and reinforced clostridial agar under anaerobic conditions. Hemolysis test, lecithinase, and lipase production on egg yolk agar, and identified with use of ANC cards in VITEK 2 compact. Subcultured in BHI broth and Gene MATRIX DNA Purifi cation Kit by DNA Gdansk for isolation of DN Multiplex PCR for toxin alpha (cpa), toxin beta (cbp), enterotoxin (ecpe), iota-toxin (cpiA), epsilon toxin (etx) genes | The cpa gene encoding alpha toxin was present in all 111 (100%) strains The cpb2 gene encoding beta2 toxin was found in: 45/49 (91.8%) strains isolated from ASD children, 17/30 (56.7%) strains isolated from healthy subjects (p < 0.001), and 12/32 (37.5%) strains isolated from obese children (p < 0.001) Clostridium perfringens (cpb2 gene) was detected in: 27/29 (93.1%) ASD, 10/17 (58.8%) HC (p < 0.008), and 11/24 (45.8%) obese children (p < 0.001) No differences between HC and obese children (cpb2 and Clostridium perfringens with cpb2) | No correlations between GM and GI symptoms were studies | ASD severity was not indicated Was not indicated whether the ASD subjects had intellectual disability A small number of studied patients and strains |
[28] | ASD (n = 23) divided into ASD w GI symptoms (21/23) and ASD w/o GI symptoms (2/23) Age 4–17 y 15 males 6 females | NT (n = 21) divided into NT w GI symptoms (10/21) and NT w/o GI symptoms (11/21) Age 4–17 y 22 males 1 female | Pyrosequencing of the 16S rRNA gene bacterial (V2–V3 regions) on fecal samples | Gut microbial diversity (alpha) ↓ (p < 0.001) and relative abundances of phylotypes most closely related to Prevotella copri ↓ (p < 0.04) in ASD Faecalibacterium ↓ (p < 0.01) and Haemophilus ↓ (p < 0.05) in ASD vs. NT Feacalibacterium prausnitzii ↓ (p < 0.01) and Haemophilus parainfluenzae ↓ (p < 0.05) in ASD vs. NT | GI symptoms were significantly more severe for children with ASD compared to controls (ATEC subscale = p < 0.01) | ASD severity was not indicated Was not indicated whether the ASD subjects had intellectual disability |
[46] | ASD (n = 30) divided into severe ASD (28/30) and moderate ASD (2/30) w GI symptoms Age 3–16 y 28 males 2 females BMI 6.9–20.5 | HC mostly Sib or blood relatives to the ASD children (n = 24) w/o GI symptoms Age 3.5–16 y 15 males 9 females BMI 13.4–31 | Sequencing of the 16S rRNA gene bacterial (V3 region) on fecal samples | Firmicutes ↑ in ASD vs. HC Sib (p < 0.05) Prevotellaceae ↓ and Veillonelleaceae ↑ in ASD vs. HC Sib Lactobacillaceae ↑ (p = 0.018), Bifidobacteriaceae ↑ (p = 0.0054), and Veillonellaceae ↑ (p = 0.008) in ASD vs. HC Sib Erysipelotrichaceae ↑ (p = 0.0005), Enterococcaceae ↑ (p = 0.0127), and Desulfovibrionaceae ↑ (p = 0.03) in ASD vs. HC Sib Bifidobacterium ↑ (p = 0.005), Lactobacillus ↑ (p = 0.018), Megasphaera ↑ (p = 0.0008), and Mirsuokella ↑ (p = 0.007) in ASD vs. HC Sib 99% of Lactobacillus was Lactobacillus ruminis in ASD group | No correlations between GM and GI symptoms were studies | Was not indicated whether the ASD subjects had intellectual disability |
[47] | ASD w GI symptoms (blood: n = 20, 15 males, 5 females) (stool: n = 21, 17 males, 4 females) ASD w/o GI symptoms (blood: n = 26, 19 males, 7 females) (stool: n = 29, 25 males, 4 females) Age 3–12 y | HC w GI symptoms (blood: n = 6, 5 males, 1 female) (stool: n = 7, 6 males, 1 female) HC w/o GI symptoms (blood: n = 35, 24 males, 11 females) (stool: n = 34, 32 males, 2 females) Age 3–12 y | Sequencing of the 16S rRNA gene bacterial (V3–V4 regions) on fecal samples | Bacteroidaceae ↑, Lachnospiraceae ↑, Ruminococcaceae ↑, and Prevotellaceae ↑ in ASD w GI symptoms vs. HC w GI symptoms Il-5, IL-15, and IL-17 ↑ in ASD w GI symptoms vs. ASD w/o GI symptoms (after exposure to the TLR-4 agonist LPS) TGFbeta1↓ in ASD w GI symptoms vs. ASD w/o GI symptoms and HC w/o GI symptoms (p < 0.05) (under the majority of conditions examined) | Differences in the microbiome composition of children with ASD vs HC groups, irrespective of GI symptoms | ASD severity was not indicated. Was not indicated whether the ASD subjects had intellectual disability Limited sample size and younger age of the HC with GI symptoms group |
[48] | ASD w GI symptoms (n = 35, 29 males, 6 females) Age 3–8 y | HC (n = 6, 5 males, 1 females) Age 3–8 y | Sequencing of the 16S rRNA gene bacterial (V3–V4 regions) on fecal samples. | Firmicutes/Bacteroidetes ratio↑ in ASD w GI symptoms vs. HC (p < 0.05) Bacteroidetes ↑ in ASD w GI symptoms vs. HC (p < 0.05) Firmicutes ↓ in ASD w GI symptoms vs. HC (p < 0.05) Veillonella, Streptococcus, Escherichia, Actinomyces, Parvimonas, Bulleida, and Peptoniphilus ↓ in ASD w GI symptoms vs. HC (p < 0.05) | Positive microbe-based link between periodontitis and ASD Negative microbe-based link between type 1 diabetes, constipation (p < 0.05), IBS, psoriasis, and ASD | ASD severity was not indicated. Was not indicated whether the ASD subjects had intellectual disability A small number of studied patients and strains |