. 2021 Jun 29;10(13):2916. doi: 10.3390/jcm10132916

Table 2.

Characteristics and summary, of studies included for research question 3.

(a) Obesity in Women and Gut Microbiome
Study	Sample Size	Patient Characteristics	Gut Microbiota Analysis	Main Findings
Menni et al. (2016)	n = 544 women with weight loss: BMI from 25.4 to 24.4 (group 1) n = 544 women with little weight gain: BMI from 24 to 25.2 (group 2) n = 544 women with heavy weight gain BMI from 25.4 to 28.8 (group 3)	Group 1 Age (yrs) 49.91 ± 9.49 Group 2 Age (yrs) 50.11 ± 5.54 Group 3 Age (yrs) 49.25 ± 8.48 All groups 15% smokers, further no exclusions.	V4 region of the 16S ribosomal RNA gene was amplified and sequenced on Illumina. De novo OTU clustering was carried across all reads using Sumaclust within QIIME 1.9.0. Alpha diversities → Shannon index, OTU counts.	Alpha diversity (Shannon index): Group 1 (weight loss) 5.21 Group 2 (weight gain) 5.19 Group 3 (heavy weight gain) 5.07 (p < 0.05) Alpha diversity (OTU): Group 1 346.3 Group 2 348 Group 3 331.8 (p < 0.05) Family Bacteriodes - Positive correlation weight gain (OR = 1.18 (0.04) p = 0.002). Negative correlation microbiome diversity Ruminococcaceae (firmicutes phyla) - Nominally protective of weight gain (OR = 0.89 (0.05), p = 0.038)
Chavez-Carbajal et al. (2019)	n = 25 control women n = 17 obese women n = 25 obese women with metabolic syndrome	Controls Age (yrs) 23.3 ± 3.1 BMI (kg/m²) 21.4 ± 1.9 Obesity Age (yrs) 28.8 ± 8.4 BMI (kg/m²) 34.8 ± 6.1 Obesity + metabolic syndrome (ms) Age (yrs) 40.5 ±10.3 BMI (kg/m²) 35.8 ± 5.1 Only women to avoid gender bias Controls significant different in age and bmi from other 2 groups	V3 region of the 16S rDNA Amplicon PCR amplification using PCR GeneAmp System 2700 Thermal Cycler. Determine with an open reference the OTUs and using a 97% similarity using QIIME pipeline (v1.9.0) and Geengenes database v13.8. Alpha diversity → Observed Species, Chao1, Shannon, Simpson.	Alpha diversity (Shannon index) Controls 4.9 Obesity 5.23 Obesity + MS 5.15 Dominant phyla in all groups: Firmicutes, Bacteroidetes, Proteobacteria, and Actinobacteria Phyla Frimicutes - Controls 57.0% - Obesity 73.0% - Obesity + MS 73.3% (p = 0.003) Bacteroidetes - Controls 36.2% - Obesity 22.5% - Obesity + MS 23.4% (p = 0.7) Firmicutes to Bacteroidetes ratio - Controls 1.57 - Obesity 3.24 - Obesity + MS 3.13 Genus Obesity and obesity + MS ↓ Bacteroides, compared to controls (p < 0.0001) Faecalibacterium (phyla firmicutes) Controls 0.55% ↑ Obesity 1.2% ↑ Obesity + MS 1.2% (p = 0.0003) Roseburia (phyla firmicutes) Controls 0.89% ↑ Obesity 2.72% ↑ Obesity + MS 2.14% (p = 0.0002) Lachnospira (phyla firmicutes) Controls 0.99% ↑ Obesity 3.24% ↑ Obesity + MS 3.79% (p < 0.0001) Coprococcus, (phyla firmicutes) Controls 2.18% ↑ Obesity 4.55% ↑ Obesity + MS 4.51% (p = 0.0002) family Erysipelotrichaceae (firmicutes) Controls 1.74% ↓ Obesity 0.38% ↓ Obesity + MS 0.36% (p < 0.0001)
Miranda er al. (2017) Observational study	n = 31 controls n = 32 normal BMI but high body fat percentage. n = 33 obesity	Controls Age (yrs) 16.3 ± 0.8 Gynoid fat (%) 34.5 (30.6–36.7) High body fat Age (yrs) 16.5 ± 0.9 Gynoid fat (%) 39.7 (37.9–46.9) Obesity Age (yrs) 16.2 ±1.3 Gynoid fat (%) 48.0 (45.5–54.1)	RT-qPCR to analyze microbiota CFX96 Touch™ detection system (Bio-Rad, Hercules, CA, USA) Alfa diversity → Shannon index
Pekkala et al. (2015)	n = 4 women with high TLR gene expression (BMI 31) n = 4 women with low TLR gene expression (BMI 28)	High TLR gene expression Age (yrs) 35.5 ± 6.0 BMI (kg/m²) 31 ± 2.0 Low TLR gene expression Age (yrs) 56.9 ± 6.4 BMI (kg/m²) 28 ± 2.5 BMI significantly higher in High TLR group.	Real-time PCR analysis was performed using in-house designed primers, iQ SYBR Supermix and CFX96 TM Real-time PCR Detection System (Bio-Rad Laboratories) Real-time PCR analysis was performed using in-house designed primers, iQ SYBR Supermix and CFX96 TM Real-time PCR Detection System (Bio-Rad Laboratories) Real-time PCR analysis was performed using in-house designed primers, iQ SYBR Supermix and CFX96 TM Real-time PCR Detection System (Bio-Rad Laboratories) RNA extraction and rt-PCR analysis using in-house designed primers.	Alpha diversity High TLR group: significant dysbiosis. Phyla Firmicutes to Bacteroidetes ratio - Low TLR 5% (p < 0.05) - High TLR 15% Cluster - High TLR: ↓ flagellated clostridium (p = 0.029) Genus - High TLR: ↓ 20% Bifidobacterium
Ott et al. (2018)	n = 20 women (own controls) n = 20 after diet n = 20 14 days after diet	Women Age (yrs) 46.8 ± 11.5 Before diet BMI (kg/m²) 34.9 ± 3.8 After diet BMI (kg/m²) 32.5 ± 3.5 14 dys after diet BMI (kg/m²) 32.6 ± 3.8	16 S rRNA gene amplicons were sequenced in paired-end modus (PE275) using a MiSeq system (Illumina)	Alpha diversity No differences Phyla Protobacteria ↓ after decrease BMI (p < 0.05) Firmicutes (after decrease BMI) ↓ Tree OTUs from family Lachnospiraceae ↓ Ruminococcaceae Actinobacteria (after decrease BMI) ↑ Bifdobacteriaceas
Choi et al. (2017) Animal study	n = 3 SHAM mice n = 3 SHAM-HF n = 5 ovariectomized mice (OVX) n = 5 OVX-HF	SHAM Weight (g) 29.96 ± 2.13 LDL (mg/dL) 30.9 ± 5.1 SHAM-HF Weight (g) 53.13 ± 3.88 LDL (mg/dL) 78 ± 4.4 OVX Weight (g) 41.44 ± 1.52 LDL (mg/dL) 45.1 ± 9.1 OVX-HF Weight (g) 57.54 ± 3.84 LDL (mg/dL) 95.7 ± 12.3 Weight significantly different	V3-V4 16S rRNA amplification following the 16S Metagenomic Sequencing Library Preparation guide by Illumina. Gene-enrichment and functional annotation analysis performed using gene ontology and KEGG pathway analysis.	Alpha diversity (Shannon index) - SHAM 3.3 - SHAM-HF 2.4 (significant reduction) - OVX 2.4 (significant reduction) - OVX-HF 2.7 Phyla Firmicutes - SHAM mice: 20% - OVX mice: 90% Bacteroidetes - SHAM mice: 78% - OVX mice: 2% Verrucomicrobia Proteobacteria - Increase in OVX-HF mice Genus and species SHAM ↑ Prevotella (p 0.036) ↑ Bacteroides (p 0.036) ↑ Bacteroidales (p 0.036) SHAM-HF ↑ Lactobacillus species ↑ Clostridiales Link gut microbiome and estradiol - estrogen signaling pathways not different in the OVX-HF vs. SHAM-HF Microbiome host interaction in OVX - OVX Lactobacillus species interaction metabolic pathways, antibiotic biosynthesis pathways, FoxO signaling pathway, glycerophospholipid metabolism pathway, and steroid hormone biosynthesis pathway. Akkermansia muciniphila related to - Pik3ca and Lgf1 → estrogen signaling pathway and ovarian steroidogenesis pathway - Cyp26b1, Nnmt, Pnpla3, and Ptgds, → metabolic pathways. - OVX-HF Ruminococcus, Dorea species, and A. muciniphila correlation with metabolic pathway, MAPK signaling pathway, AMPK signaling pathway, and FoxO signaling pathway.
(b) Obesity and Gut Microbiome: Sex Differences
Study	Sample Size	Patient Characteristics	Gut Microbiota Analysis	Main Findings
Haro et al. (2016)	n = 39 men n = 13 men < BMI 30 n = 13 BMI 30–33 n = 13 men BMI > 33 n = 36 women n = 13 BMI < 30 n = 10 BMI 30–33 n = 23 BMI > 33	Men BMI < 30 Age (yrs) 63.2 ± 2.0 BMI (kg/m²) 27.6 ± 0.6 LDL (mg/dL) 76.6 ± 4.2 BMI 30–33 Age (yrs) 58.9 ± 2.4 BMI (kg/m²) 31.4 ± 0.3 LDL (mg/dL) 95.3 ± 6.0 BMI > 33 Age (yrs) 61.3 ± 2.2 BMI (kg/m²) 35.3 ± 0.7 LDL (mg/dL) 87.8 ± 2.1 Women BMI < 30 Age (yrs) 60.1 ± 2.6 BMI (kg/m²) 27.0 ± 0.8 LDL (mg/dL) 94.2 ± 9.4 BMI 30–33 Age (yrs) 62.4 ±2.3 BMI (kg/m²) 31.4 ± 0.3 LDL (mg/dL) 87.1 ± 7.6 BMI > 33 Age (yrs) 58.9 ± 2.3 BMI (kg/m²) 36.7 ± 1.4 LDL (mg/dL) 80.4 ± 4.4	Sequencing V4 16S microbial rRNA on the Illumina MiSeq. Taxonomy assigned to OTUs against the Greengenes v13-8 preclustered at 97% identity. Alpha diversities → observed OTU counts, Shannon, Simpson.	Alpha diversity similar men and women and comparing BMI Phyla Firmicutes to Bacteroidetes ratio - BMI < 33: Men higher ratio - BMI > 33: Women higher ratio (p = 0.018) Genus Women BMI > 33 ↑ Bilophila (p = 0.002) ↑ Veillonella (p = 0.001) Men BMI > 33 ↑ Methanobrevibacter (p = 0.002) Bacterial species Women BMI > 33 ↑ Bacteroides caccae (p = 0.009) Men BMI > 33 ↑ Bacteroides plebeius (p = 0.001) ↑ Coprococcus catus
Min et al. (2019)	n = 116 women n = 96 men	Women Age (yrs) 50.7 ± 14.1 BMI (kg/m²) 23.0 ± 3.0 Gynoid fat 15.9 ± 3.0 Android fat 12.5 ± 1.2 LDL (mmol/L) 2.7 ± 0.7 Men Age (yrs) 50.7 ± 14.5 BMI (kg/m²) 23.6 ± 3.0 Gynoid fat 17.7 ± 3.0 (p < 0.005) Android fat 9.9 ± 1.4 (p < 0.005) LDL (mmol/L) 2.8 ± 0.7	16S rRNA V4 region sequencing The denoised sequences are mapped to the GreenGenes reference database43. Taxonomy is assigned at 97% identity. Alfa diversity → Shannon index	Alpha diversity potential negative association between gynoid fat ratio and microbiome abundance in both sexes. In women compared to men different taxa responsible for relation between fat distribution and diversity. Gynoid fat ratio positive correlation Women: - Provotellaceae family (effect size 9.6) - Ruminococcaceae family Men: - Lachnospiraceae family - Clostridium_XlVa (effect size 10) Gynoid fat ratio negative correlation - Bacteroidaceae family, Bacteroides genus (effect size of −24.2) - Ruminococcaceae family No taxa associated with android fat ratio.