Gut microbiota signatures in colorectal cancer: Evidence synthesis from umbrella review and probiotic intervention studies

Table 1 Quality evaluation using AMSTAR 2 (a measurement tool to assess systematic reviews)

Ref.	Q1	Q2	Q3	Q4	Q5	Q6	Q7	Q8	Q9	Q10	Q11	Q12	Q13	Q14	Q15	Q16
Jiang et al[21]	Y	Y	Y	Y	PY	PY	Y	Y	Y	Y	Y	PY	PY	PY	Y	Y
Jin et al[18]	Y	Y	Y	Y	PY	PY	Y	Y	Y	Y	Y	PY	Y	Y	Y	Y
Thomas et al[10]	Y	Y	Y	PY	N	N	Y	Y	Y	Y	Y	PY	Y	Y	Y	Y
Liu et al[11]	Y	Y	Y	N	N	N	Y	Y	Y	Y	Y	PY	Y	Y	Y	Y
Kharofa et al[13]	Y	Y	Y	PY	PY	PY	Y	Y	Y	Y	Y	PY	Y	Y	Y	Y
Li et al[12]	Y	Y	Y	PY	PY	PY	Y	Y	Y	Y	Y	PY	Y	Y	Y	Y
Lin et al[19]	Y	Y	Y	Y	PY	PY	Y	Y	Y	Y	Y	PY	Y	Y	Y	Y
Wirbel et al[20]	Y	Y	Y	Y	PY	PY	Y	Y	Y	Y	Y	PY	Y	Y	Y	Y
Piccinno et al[15]	Y	Y	PY	PY	PY	PY	Y	Y	Y	Y	Y	PY	PY	Y	Y	Y
Zhu et al[17]	Y	Y	PY	PY	PY	PY	Y	Y	Y	Y	Y	PY	PY	Y	Y	Y
Chen et al[16]	Y	Y	PY	PY	PY	PY	Y	Y	Y	Y	Y	PY	PY	Y	Y	Y

AMSTAR 2 (a measurement tool to assess systematic reviews) item definitions are provided below as annotations. Annotations for AMSTAR 2 items: Q1: Did the research questions and inclusion criteria for the review include the components of PICO? Q2: Did the report of the review contain an explicit statement that the review methods were established prior to the conduct of the review and did the report justify any significant deviations from the protocol? Q3: Did the review authors explain their selection of the study designs for inclusion in the review? Q4: Did the review authors use a comprehensive literature search strategy? Q5: Did the review authors perform study selection in duplicate? Q6: Did the review authors perform data extraction in duplicate? Q7: Did the review authors provide a list of excluded studies and justify the exclusions? Q8: Did the review authors describe the included studies in adequate detail? Q9: Did the review authors use a satisfactory technique for assessing the risk of bias (RoB) in individual studies that were included in the review? Q10: Did the review authors report on the sources of funding for the studies included in the review? Q11: If meta-analysis was performed did the review authors use appropriate methods for statistical combination of results? Q12: If meta-analysis was performed, did the review authors assess the potential impact of RoB in individual studies on the results of the meta-analysis or other evidence synthesis? Q13: Did the review authors account for RoB in individual studies when interpreting/discussing the results of the review? Q14: Did the review authors provide a satisfactory explanation for, and discussion of, any heterogeneity observed in the results of the review? Q15: If they performed quantitative synthesis did the review authors carry out an adequate investigation of publication bias (small study bias) and discuss its likely impact on the results of the review? Q16: Did the review authors report any potential sources of conflict of interest, including any funding they received for conducting the review? Y: Yes; PY: Partial yes; N: No.

Table 2 Characteristics of the included studies (n = 11)

Ref.	First author	Journal and year	k meta	CRC cases (n)	Controls (n)	Country	Objective of study	Outcome
Liu et al[11]	Ning-Ning Liu	Nature Microbiology, 2022	5	491	494	Austria, France, Germany, China, Japan, Italy, and United States	To analyze multi-kingdom gut microbiome alterations in CRC and identify reproducible diagnostic biomarkers	CRC showed multi-kingdom dysbiosis, with 16 validated biomarkers and altered D-amino acid and butanoate metabolism
Li et al[12]	Tian-Hui Li	Gastroenterology, 2025	11	738	896	China, Austria, United States, Italy, Japan, Germany, and France	To characterize CRC-associated gut archaea and assess their interactions with bacterial communities	CRC showed reduced archaeal diversity, altered methanogenesis-related taxa, and improved diagnostic performance in combined archaea-bacteria models
Jiang et al[21]	Pu-Zi Jiang	mSystems, 2021	7	354	357	Japan, China, United States, Germany, France, Australia, and Italy	To identify shared and disease-specific microbial signatures across CD (Crohn’s disease), UC (ulcerative colitis), and CRC	Species- and pathway-level biomarkers were identified, with UC and CRC showing greater microbial similarity and strong diagnostic potential
Jin et al[18]	Dong-Min Jin	mSystems, 2024	3 (CRC only)	141	141	Germany, Austria, and France	To develop an interpretable meta-analysis workflow for comparing microbiome features across diseases	Disease pairs showed varying microbiome similarities, reflecting both pathogen enrichment and depletion of beneficial microbes
Lin et al[19]	Yu-Feng Lin	Gastroenterology, 2022	8	454	525	Canada, United States, Germany, France, Italy, China, Japan, and Australia	To investigate CRC-associated gut mycobiota and evaluate fungal biomarkers for diagnosis	CRC showed fungal dysbiosis, and fungal-bacterial biomarkers improved diagnostic performance over bacterial markers alone
Wirbel et al[20]	Jakob Wirbel	Nature Medicine, 2019	8	386	392	France, Austria, China, United States, Germany, Italy, and Japan	To identify robust taxonomic and functional microbial signatures of CRC across heterogeneous cohorts	CRC was associated with 29 core enriched species, increased protein and mucin degradation, and elevated secondary bile acid production
Kharofa et al[13]	Jordan Kharofa	Cancer Medicine, 2023	11	692	609	Austria, Canada, China, Germany, France, India, Italy, Japan, and United States	To compare microbial features between early-onset and late-onset CRC, including virulence factors	Early-onset CRC showed distinct microbial and virulence profiles, including associations with colibactin, Fusobacterium adhesin A, and Fusobacterium nucleatum
Thomas et al[10]	Andrew Maltez Thomas	Nature Medicine, 2019	7	313	308	Austria, France, Germany, Italy, Denmark, United States, China, and Japan	To identify reproducible CRC-associated biomarkers and explore related functional mechanisms	CRC showed increased microbial richness, reproducible markers such as Fusobacterium nucleatum, and altered amino acid, gluconeogenesis, and choline metabolism
Piccinno et al[15]	Gianmarco Piccinno	Nature Medicine, 2025	18	1471	1568	Austria, China, Czech Republic, France, Germany, India, Italy, Japan, Spain, Turkey, and United States	To examine gut microbiome changes across adenoma-carcinoma progression, CRC stage, and tumor location	CRC microbiomes were enriched with oral taxa, and cross-cohort classifiers showed strong predictive performance with reproducible species- and strain-level markers
Zhu et al[17]	Xin-Yue Zhu	Communications Biology, 2024	8	485	496	Austria, China, France, Germany, Italy, and United States	To compare multi-kingdom microbial signatures between excess body weight CRC (EBW-CRC) and lean CRC	EBW-CRC and lean-CRC showed distinct microbial and functional patterns, with the EBW-CRC classifier achieving strong validation performance
Chen et al[16]	Fang Chen	Journal of Advanced Research, 2023	9	554	546	France, China, Austria, United States, Canada, Italy, Germany, and Japan	To identify reproducible gut virome signatures in CRC and adenoma and assess their diagnostic potential	CRC and adenoma showed altered viral profiles, and viral biomarkers demonstrated diagnostic potential across cohorts

CRC: Colorectal cancer; CD: Crohn’s disease; UC: Ulcerative colitis; EBW: Excess body weight.

Table 3 Characteristics of the included studies (n = 6)

Ref.	Country	Design	Population	N, P/C	Intervention	Dose	Duration/sampling	Age, P/C	Sex, M/F, P/C	BMI, P/C
Rafter et al[22]	Ireland	RCT, double-blind, placebo-controlled; single-center	Postoperative colon cancer patients within 5 years of resection	37, 19/18	Bifidobacterium lactis Bb12 + Lactobacillus rhamnosus GG + inulin	Synbiotic 1 12 g/day; 1010 CFU/day	12 weeks; baseline, weeks 6, weeks 12	61.1 ± 5.55/57.0 ± 9.75	Reported values need checking	NR/NR
Park et al[26]	South Korea	RCT, double-blind, placebo-controlled; multicenter	Colon cancer patients undergoing anterior resection; perioperative intervention	60, 29/31	Bifidobacterium animalis subsp. lactis HY8002 + Lactiplantibacillus casei HY2782 + Lactiplantibacillus plantarum HY7712	2 g bid; 2 × 108 CFU/day total	4 weeks + 1-week follow-up; baseline, pre-op, 3 weeks post-op, 4 weeks post-op	60.10 ± 10.37/61.03 ± 7.02	13/18, 13/18, check P group	24.42 ± 2.90/24.04 ± 3.53
Wang et al[27]	China	Retrospective cohort; single-center	Advanced CRC with malnutrition; receiving chemotherapy or radiotherapy ± targeted therapy	80, 40/40	Streptococcus thermophilus acidophilus NCFM + Bifidobacterium lactis Bi-07 + Streptococcus thermophilus St-21 + mixed fiber	2 × 109 CFU/day + fiber 20 g/day	12 weeks; baseline, weeks 12	64.75 ± 6.22/63.94 ± 5.63	19/21, 23/17	17.56 ± 1.04/17.47 ± 1.16
Hibberd et al[25]	Sweden	Prospective randomized pilot study; single-center	Stage I-III colon cancer patients; intervention after colonoscopy and before surgery	15, 8/7	Bifidobacterium lactis Bl-04 + Lactiplantibacillus acidophilus NCFM + inulin	2 tablets/day; 2.1 × 1010 CFU/day total + inulin 0.63 g	Mean 31 ± 28 days; post-colonoscopy, pre-op	77 ± 5.2/63 ± 13.3	Reported values need checking	24.1 ± 1.7/NR
Gianotti et al[23]	Italy	RCT, double-blind, placebo-controlled, 3-arm; multicenter	Colon cancer patients undergoing elective colorectal resection; perioperative intervention	31, 11 low-dose/10 high-dose/10 control	Lactiplantibacillus johnsonii La1 + Bifidobacterium longum BB536	Low: 2 × 107 CFU/day; high: 2 × 109 CFU/day; bid	6 days; day - 4, day 0, day + 5	64.7 ± 4.8 low/62.7 ± 7.8 high/63.3 ± 10.2 control	Reported values need checking	26.5 ± 4.1 low/24.4 ± 3.7 high/25.6 ± 2.6 control
Liu et al[24]	China	RCT, double-blind, placebo-controlled; single-center	CRC patients undergoing elective radical colorectomy; perioperative intervention	100, 50/50	Lactiplantibacillus plantarum CGMCC No.1258 + Lactiplantibacillus acidophilus LA-11 + Bifidobacterium longum BL-88	2 g/day; 2.6 × 1014 CFU/day total	16 days; pre-trial, pre-op, post-op day 3, post-op day 10	65.7 ± 9.9/65.3 ± 11.0	28/22, 31/19	22.8 ± 1.8/22.6 ± 2.0

P/C: Probiotic/control; M/F: Male/female; BMI: Body mass index; CRC: Colorectal cancer; NR: Not reported; RCT: Randomized controlled trial; CFU: Colony-forming units; pre-op: Preoperative; post-op: Postoperative; bid: Twice daily.

Table 4 17 microbial genera were significantly associated with colorectal cancer risk according to second-order meta-analysis (P ≤ 0.05, k ≥ 5)

Taxon	Outcome	k meta	Effects (n)	Pooled effect	CI low	CI high	P value	I2 value	τ2 value	QE test P value
Parvimonas	CRC/control	9	9	1.227	0.09	2.365	0.034	99.999	2.97	0
Fusobacterium	CRC/control	8	8	0.917	0.139	1.696	0.021	99.999	1.129	0
Hungatella	CRC/control	5	5	0.885	0.425	1.346	1.62 × 10-4	91.939	0.248	2.76 × 10-10
Anaerococcus	CRC/control	5	5	0.756	0.22	1.292	0.006	98.816	0.349	8.30 × 10-6
Gemella	CRC/control	8	8	0.616	0.287	0.945	2.39 × 10-4	99.998	0.22	0
Clostridium	CRC/control	9	9	0.508	0.129	0.888	0.009	100	0.327	0
Prevotella	CRC/control	10	10	0.429	0.161	0.697	0.002	99.999	0.173	0
Anaerotruncus	CRC/control	7	7	0.423	0.046	0.8	0.028	99.997	0.25	0
Parabacteroides	CRC/control	7	7	0.42	0.041	0.799	0.03	99.995	0.257	0
Dialister	CRC/control	5	5	0.401	0.051	0.751	0.025	99.996	0.153	0
Peptostreptococcus	CRC/control	6	6	0.398	0.27	0.526	9.96 × 10-10	81.662	0.014	7.06 × 10-4
Methanobrevibacter	CRC/control	5	5	0.398	0.042	0.753	0.028	96.741	0.13	0
Slackia	CRC/control	5	5	0.393	0.076	0.709	0.015	98.302	0.125	0
Bilophila	CRC/control	6	6	0.384	0.068	0.7	0.017	97.399	0.147	0
Bacteroides	CRC/control	10	10	0.355	0.092	0.619	0.008	99.999	0.176	0
Alistipes	CRC/control	9	9	0.353	0.049	0.657	0.023	99.994	0.206	0
Streptococcus	CRC/control	9	9	0.296	0.047	0.545	0.02	99.997	0.134	0

CI: Confidence interval; CRC: Colorectal cancer.