The causal relationship between Eubacterium and constipation: a Mendelian randomization study

The causal relationship between Eubacterium and constipation: a Mendelian randomization study | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article The causal relationship between Eubacterium and constipation: a Mendelian randomization study Dingwen Fan, Liangliang Zhang, Hongchang Liu, Dan Zhu This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4482347/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background: Constipation may be related to an imbalance in flora imbalance. In this study, univariate and multivariate Mendelian randomization (MR) studies were used to further understand the causal relationship between different members of genus Eubacteriumand constipation. Methods: The constipation and genus Eubacterium datasets were obtained from the Integrative Epidemiology Unit (IEU) Open Genome-Wide Association Study (GWAS) database. The causal effects between constipation and genus Eubacterium were analyzed using MR-Egger, Weighted median, Simple mode, Weighted mode and Inverse variance weighted (IVW). In univariate MR analysis (UVMR), genus Eubacillus was the exposure factor, constipation was the outcome. In addition, the reliability of UVMR analysis was assessed by sensitivity analyses (heterogeneity tests, horizontal pleiotropy tests and leave-one-out (LOO) analysis). Finally, the MVMR analysis was performed. Results: In the UVMR analysis, genus Eubacterium rectale group id.14374 (p=0.043, OR=0.805) and genus Eubacterium brachy group id.11296 (p=0.036, OR=0.904) were all protective factors on constipation. The results of sensitivity analysis showed that UVMR analysis was reliable. In the MVMR analysis, genus Eubacterium rectalegroup was still a significant protective factor (p=0.007, OR=0.80), while genus Eubacterium brachy group was not a significant protective factor for constipation (p=0.151, OR=0.94) . Conclusion: The results of study supported that genus Eubacterium rectale group had a significant causal relationship with constipation, and the increase of genus Eubacterium rectale group could reduce the risk of constipation. Figures Figure 1 Figure 2 Figure 3 Introduction Constipation is frequently encountered in the realm of digestive issues and is characterized by challenges in stool evacuation, discomfort during bowel movements, or the need for marked exertion to eliminate feces(Bharucha 2007 ; Rao 2009 ; Rao and Singh 2010 ). The global prevalence of constipation is approximately 14%, with the incidence varying across different demographics(Black and Ford 2018 ). Specifically, the incidence rates in children and the elderly (aged 60 and over) are about 9.5% and 33.5%, respectively(Forootan et al. 2018 ; Koppen et al. 2018 ). While constipation is not inherently life-threatening, chronic constipation can substantially degrade the quality of life(Bharucha and Wald 2019 ). Furthermore, constipation markedly elevates the risk of coronary heart disease and ischemic stroke and increases all-cause mortality among the elderly, thus imposing a significant health burden on this age group(Sumida et al. 2019 ). Substantial current evidence suggests that the gut microbiome, bile secretion, lifestyle, and behavioral psychology are all factors influencing constipation.(Cha et al. 2023 ; Erhardt et al. 2023 ; Neves et al. 2023 ; Wang et al. 2023 ). Central to intestinal well-being and digestion is the gut microbiome—a rich ecosystem of microorganisms such as bacteria, fungi, and viruses that inhabit the human gastrointestinal tract(Wu et al. 2024 ). In the past few years, there has been a growing focus on investigating the correlation between gut microorganisms and illnesses(Cui et al. 2024 ; Gilbert et al. 2018 ; Harnett et al. 2017 ; Kong et al. 2023 ); however, substantial knowledge regarding the direct impact of gut microbes on constipation is still lacking(Oliphant and Allen-Vercoe 2019 ; Quigley 2019 ). The Eubacterium genus, comprising prevalent anaerobic bacteria within the gastrointestinal tracts of both humans and animals(Mukherjee et al. 2020 ), is essential for the metabolic processing of carbohydrates(Pessoa et al. 2023 ), amino acids(Jin et al. 2023 ), and fatty acids(Yan et al. 2023 ), as well as for normal digestive processes(Gomes et al. 2018 ). Crucial to the integrity of intestinal health is the presence of an appropriate population of Eubacterium in the gut's microbial ecosystem. Studies have revealed a clear association between a diminished presence or complete lack of Eubacterium strains and the development of inflammatory bowel conditions and constipation(El-Salhy 2023 ; Hong and Rhee 2014 ; Rendeli et al. 2023 ). In theory, Eubacterium could influence constipation through various mechanisms. Initially, Eubacterium strains can be involved in carbohydrate metabolism and generate short-chain fatty acids, which are crucial for preserving the health of the intestinal lining and enhancing the movement of the intestines(Shoaie et al. 2015 ). Additionally, these strains of Eubacterium may collaborate with other intestinal microbes to maintain a healthy microbial balance in the gut and affect the activities of the intestinal immune system(Hiippala et al. 2018 ; Magrone and Jirillo 2013 ). Nevertheless, even though there is research backing the link between Eubacterium and constipation(Galkin et al. 2018 ; Odamaki et al. 2016 ), the gut microbial environment is intricate and affected by various elements, such as diet, lifestyle, and genetics. Consequently, the exact nature of the cause-and-effect relationship between Eubacterium and constipation remains uncertain. Mendelian randomization (MR) represents a technique for identifying causal links between exposure factors and outcomes(Burgess and Thompson 2015 ). MR analyses are contingent upon three pivotal assumptions. The first is the relevance assumption, signifying that the chosen instrumental variables (IVs) should have a robust correlation with the exposure of interest, such as a specific biomarker or behavior. Next, the independence assumption dictates that the IVs need to be free from any confounding factors. Lastly, the exclusion-restriction assumption demands that the influence of the instrumental variable on the disease outcome should only be mediated through its effect on the exposure, and not via alternative pathways(Burgess et al. 2015 ). This method capitalizes on the naturally random distribution of genetic variations, which are unconnected to ethnicity, gender, or status at birth, to offer a robust analysis of clinical intervention efficacy while reducing the risks of reverse causality and confounding variables(Burgess et al. 2012 ; Richmond and Davey Smith 2022 ). Our study applied MR with the intention of clarifying the causative link between Eubacterium and constipation, in order to provide a theoretical basis for the improvement of clinical practices. Methods Source of data and study analysis The Integrative Epidemiology Unit (IEU) Open Genome-Wide Association Study (GWAS) database ( https://gwas.mrcieu.ac.uk/ ) was used to search for genus Eubacterium and constipation, and the traitID and GWAS data were obtained. The 11 species of genus Eubacillus was the exposure factor, constipation was the outcome. The datasets of 11 species of genus Eubacillus shown in Supplementary Table 1 , the ebi-a-GCST90017003 of constipation included 5,708,381 single nucleotide polymorphisms (SNPs) from 14,306 samples. Univariable Mendelian randomization (UVMR) analysis The extract_instruments in TwoSampleMR package (Hemani et al. 2018 ) was used to read exposure factors and filter instrument variables (IVs) with p < 5*10 − 6. The linkage disequilibrium analysis (LDA) were performed to ensure independence (clump = TRUE, r2 = 0.001 and kb = 100). The mr function of TwoSampleMR package combined with five algorithms (MR Egger (Bowden et al. 2015 ), Weighted median(Bowden et al. 2016 ), Inverse variance weighted (IVW)(Burgess et al. 2015 ), Simple mode, Weighted mode(Hartwig et al. 2017 )) to carry out UVMR analysis. Results were mainly referred to IVW. The foundational assumptions of this MR analysis were based on three key principles: a strong correlation between the IVs and the exposure, the independence of IVs from potential confounders, and the exclusive influence of IVs on the outcome through the exposure without involvement in alternative pathways. Multivariable-MR (MVMR) analysis The input data in MVMR analysis were outcome and exposure factors that were significantly associated with constipation in UVMR analysis. The SNPs significantly associated with exposure factors were searched by the TwoSampleMR package “mv extract exposures” (screening criteria: p = 5e-08, clump = TRUE, r2 = 0.001 and kb = 10000). The TwoSampleMR package “extract outcome data” was used to read exposure factors and screen SNPs (screening criteria: proxies = TRUE; rsq = 0.8). Finally, MVMR was performed through the TwoSampleMR package function mv_harmonise_data Sensitivity analysis To assess the reliability of MR analysis, sensitivity analysis were performed. In the heterogeneity test, Q pvalue greater than 0.05 indicated that there was no heterogeneity. In pleiotropy test, p value greater than 0.05 indicated that there was no horizontal pleiotropy. Leave-one-out (LOO) sensitivity test was mainly to judge the impact of the remaining SNPs on the outcome after removing the SNP one by one. If the result changed significantly, the outcome was sensitive to the SNP(Xu et al. 2022 ) . Result Eubacterium rectale group id.14374/Eubacterium rectale group id.14374 had a significant causal relationship with constipation inUVMR analysis IVW results showed that genus Eubacterium rectale group id.14374 (p = 0.043, OR = 0.805) and genus Eubacterium brachy group id.11296 (p = 0.036, OR = 0.904) had significant negative causal relationship with constipation (Supplementary Table 2) . However, the remaining nine genus Eubacterium did not have a significant causal relationship with constipation (Supplementary Table 2) . The scatter plot results of IVW algorithm showed negative causal relationship between Eubacterium rectale group id.14374/ genus Eubacterium brachy group id.11296 and constipation ( Fig. 1 a- 1 b, Supplementary Fig. 1) . It can be seen from the forest map of the IVW algorithm that the effect values of Eubacterium rectale group id.14374/ genus Eubacterium brachy group id.11296 on constipation were all less than 0, indicating that the two genus of Eubacterium were protective factors on the constipation ( Fig. 1 c- 1 d, Supplementary Fig. 2) . In addition, the samples were symmetrically distributed along both sides of the IVW line in the funnel plot, indicating that the MR analysis conformed to randomness ( Fig. 1 e- 1 f, Supplementary Fig. 3) . To evaluate the reliability of the UVMR analysis, we performed a sensitivity analysis. Heterogeneity results showed that there was no heterogeneity in genus Eubacterium rectale group id.14374 on constipation (Q pvalue = 0.439) and genus Eubacterium brachy group id.11296 on constipation (Q pvalue = 0.928) MR analysis ( Table 1 , Supplementary Table 3) . Horizontal pleiotropy test results showed that the P-value of the two exposure factors were all greater than 0.05 (genus Eubacterium rectale group id.14374 pvalue = 0.78, genus Eubacterium brachy group id.11296 pvalue = 0.45), indicated that there was no horizontal pleiotropy of the analysis results ( Table 2 , Supplementary Table 4) . The LOO test results of two exposure factors were on the left side of 0, indicating that there were no SNP with overly sensitive outcome ( Fig. 2 a- 2 b, Supplementary Fig. 4) . Table 1 The heterogeneity test of causality between genus Eubacterium rectale/genus Eubacterium brachy groups and constipation based on MR results outcome exposure method Q Q_df Q_pval Constipation genus Eubacterium rectale group MR Egger 2.578125498 2 0.276 Constipation genus Eubacterium rectale group Inverse variance weighted 2.708439662 3 0.439 Constipation genus Eubacterium brachy group MR Egger 0.658221819 4 0.956 Constipation genus Eubacterium brachy group Inverse variance weighted 1.364387018 5 0.928 MR mendelian randomization Table 2 The horizontal pleiotropy test of causality between genus Eubacterium rectale/genus Eubacterium brachy groups and constipation based on MR results outcome exposure egger_intercept se pval Constipation genus Eubacterium rectale group -0.009880127 0.031074472 0.780650712 Constipation genus Eubacterium brachy group 0.021345491 0.025401128 0.448015795 MR mendelian randomization, se standard error The genus Eubacterium rectale group id.14374 still was protective factors on constipation in MVMR analysis In MVMR analysis, genus Eubacterium rectale group id.14374 still had a significant negative causal effect on constipation (p = 0.007, OR = 0.80), indicating that its causal effect on constipation was more independent, while the causal effect of genus Eubacterium brachy group id.11296 on constipation was diluted (p = 0.151, OR = 0.94) ( Table 3 , Fig. 3 ) . Table 3 The MVMR results between between genus Eubacterium rectale/genus Eubacterium brachy groups and constipation exposure outcome b se pval or or_lci95 or_uci95 genus Eubacterium brachy group Constipation -0.062937802 0.043836734 0.151 0.939001876 0.861691639 1.023248322 genus Eubacterium rectale group Constipation -0.219443616 0.081435751 0.007 0.802965431 0.684506124 0.941925076 MR mendelian randomization, se, standard error, OR odds ratio, CI confidence interval Discussion In our study, a novel exploration into the causative link between the genus Eubacterium and constipation was undertaken, with 11 species within the Eubacillus genus being investigated as the exposure element, while constipation served as the observed effect. Our findings indicated that both the genus Eubacterium rectale group and the genus Eubacterium brachy group acted as protective elements against constipation in univariate MR analyses. Furthermore, in multivariate analysis, the genus Eubacterium rectale group continued to stand out as a significant protectiveguard. These insights lay a theoretical foundation for clinical investigations into the relationship between Eubacterium and constipation, offering diverse analytical perspectives. Pracimvot first introduced the term 'Eubacterium' in 1938 as a designation for a cluster of beneficial bacteria derived from human stool samples(Andreesen 1992 ). Commonly found within the mouths and gastrointestinal tracts of mammals, such as the stomachs of cud-chewing animals, the Eubacterium genus is also present in various environmental settings(Karcher et al. 2020 ). This genus plays a central role in the gut flora in humans and is extensively prevalent among various demographic groups across continents such as Africa, Australia, Europe, India, South America, Asia, and North America(Ayeni et al. 2018 ; Escobar et al. 2014 ; Gomez-Arango et al. 2018 ; Mueller et al. 2006 ). The Eubacterium rectale group and the Eubacterium brachy group represent specific bacterial groups within this genus, each with unique features and functions. Although both belong to the genus Eubacterium, they may differ in terms of bacterial composition, metabolic products, and other aspects. The genus Eubacterium rectale, a species within the human microbial population, is recognized for its capacity to generate butyrate(Lu et al. 2022 ), a short-chain fatty acid (SCFAs) with a crucial role in protectiveguarding the gut lining and reducing inflammation within the intestines(Biagi et al. 2013 ; Magrone and Jirillo 2013 ). As individuals age, there is a notable reduction in the variety of gut microbes, coinciding with a diminished production of butyrate by this specific genus, a factor linked to the onset of gastrointestinal disorders(Ling et al. 2022 ). Throughout the digestive process, an optimal level of Eubacterium rectale supports the uptake of indigestible elements like dietary fibers, while also generating butyrate, which protectiveguards intestinal health and assists in preventing constipation(Topping and Clifton 2001 ). Moreover, this genus is a member of the intestinal bacterial consortium that ferments fibers like cellulose and resistant starch. It contributes to the production of additional varieties of SCFAs, including acetic and propionic acids, which are essential for overall gut health(Mukherjee et al. 2020 ). SCFAs, known for their crucial role in the modulating intestinal inflammation, offer beneficial effects on constipation by enhancing intestinal integrity and regulating immune responses(Chen et al. 2015 ). More specifically, SCFAs positively influence the expression levels of junctional proteins (such as claudin-1 and occludin), thereby increasing the density of intercellular junctions(Wang et al. 2012 ; Willemsen et al. 2003 ). This process effectively protectiveguards against the infiltration of harmful substances into intestinal tissues. Additionally, SCFAs activate G protein-coupled receptors (GPCRs), a group of proteins responsible for transmitting vital cellular signals. This activation process directly impacts the activity and functionality of immune cells, thereby contributing to immune response modulation(Corrêa-Oliveira et al. 2016 ). Eubacterium rectale plays a role in alleviating constipation by enhancing the movement of the intestines. It has been documented that SCFAs can influence the movement within the intestines regardless of pH levels. The time it takes for substances to move through the intestines may decrease when propionate and butyrate solutions are introduced into the colon(Cherbut et al. 1998 ; Tazoe et al. 2008 ). Moreover, introducing mixtures that contain SCFAs has been shown to boost motility in the ileum more effectively than either air or saline solutions(Jouët et al. 2013 ). In conclusion, Eubacterium rectale helps manage constipation by preserving intestinal mucosal integrity, modulating gut immunity, reducing inflammation, and enhancing intestinal motility. Current understanding of the metabolic processes and roles of Eubacterium brachy remains limited. Nonetheless, research indicates that it might share metabolic activities with Eubacterium rectale, including the synthesis of short-chain fatty acids(Mukherjee et al. 2020 ). Additionally, several studies have linked Eubacterium brachy to gastrointestinal well-being and inflammation-related oral conditions(Li et al. 2023 ; Moutsopoulos et al. 2015 ; Vincent et al. 1986 ), though the precise pathways involved remain to be elucidated. Recent research demonstrates a decline in Eubacterium brachy population following nine weeks on a diet rich in fats(Pessoa et al. 2023 ), yet the connection between such a diet, Eubacterium brachy levels, and constipation remains undetermined. Moreover, a reduction in Eubacterium brachy has been noted in cases of inflammatory bowel disease(Li et al. 2023 ), which implies that Eubacterium brachy may exert a beneficial effect on intestinal health. Significantly, the modulation of constipation by Eubacterium brachy appears to be linked to the production of its metabolites known as SCFAs. These compounds are potentially crucial for their roles in regulating the immune system, managing inflammation, and protectiveguarding the gut's integrity. However, it is still uncertain whether other factors or different metabolites are involved in this regulatory mechanism, an area that requires further exploration. Considering the co-presence of both the Eubacterium rectale group and the Eubacterium brachy group in the human intestines, we conducted additional multivariate MR analyses with both groups as exposure variables and constipation as the outcome. While the univariate MR analysis provided evidence of causality, it is important and valid to perform multivariate MR analysis, as it can potentially yield more evidence(Chen et al. 2022 ; Zhou et al. 2021 ). The findings of our study revealed that the Eubacterium rectale group remained a significant protective factor in the multivariate MR analysis, whereas the significance of the Eubacterium brachy group decreased. This suggests that the Eubacterium brachy group may have a lesser causal role in constipation compared to the Eubacterium rectale group. It is also possible that these two groups interact in the development of constipation, emphasizing the need for further research in this area. Undoubtedly, the Eubacterium rectale group has been identified as a protective factor against constipation in both univariate and multivariate MR analyses. This consistent finding is expected to establish a new theoretical basis for the development of targeted interventions that utilize this genus to alleviate constipation symptoms. Conclusion Our investigation faced certain inherent constraints. Despite our examination of the prevalent occurrence of Eubacterium across continents such as Africa, Australia, Europe, India, South America, Asia, and North America, MR analyses were constrained to European population data, thereby omitting data from other ethnic groups and necessitating a more circumspect approach when applying findings to diverse populations. Additionally, the goal of MR analyses in gauging the life-long influence of Eubacterium rectale and Eubacterium brachy groups raised the potential of age-related changes in the genetic instruments, potentially impacting the results' interpretability and accuracy. Nevertheless, our study's clinical significance is underlined by the identification of a causal link between the Eubacterium rectale group and constipation, laying a theoretical foundation for future clinical applications. Our future research will aim to expand the sample size, further investigate the interactions between different bacterial species, and incorporate other research methodologies to validate and reinforce the findings of this study. Declarations Competing Interests The authors have no relevant financial or non-financial interests to disclose. Data Availability The datasets analysed during the current study are available from the corresponding author on reasonable request. Ethics statement The data utilized in our analysis were derived from publicly available sources and were obtained, ensuring compliance with the principles of the declaration of Helsinki and approval by the relevant ethics committees. Furthermore, our findings were presented in alignment with the guidelines outlined in the Mendelian Randomization STrengthening the Reporting of OB servational studies using Genetic Epidemiology (MR STROBE) framework. References Andreesen JR (1992) The genus Eubacterium. 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Cell Metab 22: 320-31. doi: 10.1016/j.cmet.2015.07.001 Sumida K, Molnar MZ, Potukuchi PK, Thomas F, Lu JL, Yamagata K, Kalantar-Zadeh K, Kovesdy CP (2019) Constipation and risk of death and cardiovascular events. Atherosclerosis 281: 114-120. doi: 10.1016/j.atherosclerosis.2018.12.021 Tazoe H, Otomo Y, Kaji I, Tanaka R, Karaki SI, Kuwahara A (2008) Roles of short-chain fatty acids receptors, GPR41 and GPR43 on colonic functions. J Physiol Pharmacol 59 Suppl 2: 251-62. Topping DL, Clifton PM (2001) Short-chain fatty acids and human colonic function: roles of resistant starch and nonstarch polysaccharides. Physiol Rev 81: 1031-64. doi: 10.1152/physrev.2001.81.3.1031 Vincent JW, Falkler WA, Jr., Suzuki JB (1986) Systemic antibody response of clinically characterized patients with antigens of Eubacterium brachy initially and following periodontal therapy. J Periodontol 57: 625-31. doi: 10.1902/jop.1986.57.10.625 Wang HB, Wang PY, Wang X, Wan YL, Liu YC (2012) Butyrate enhances intestinal epithelial barrier function via up-regulation of tight junction protein Claudin-1 transcription. Dig Dis Sci 57: 3126-35. doi: 10.1007/s10620-012-2259-4 Wang P, Shen X, Wang Y, Jia X (2023) Association between constipation and major depression in adult Americans: evidence from NHANES 2005-2010. Front Psychiatry 14: 1152435. doi: 10.3389/fpsyt.2023.1152435 Willemsen LE, Koetsier MA, van Deventer SJ, van Tol EA (2003) Short chain fatty acids stimulate epithelial mucin 2 expression through differential effects on prostaglandin E(1) and E(2) production by intestinal myofibroblasts. Gut 52: 1442-7. doi: 10.1136/gut.52.10.1442 Wu M, Zheng W, Song X, Bao B, Wang Y, Ramanan D, Yang D, Liu R, Macbeth J, Do E, Andrade W, Yang T, Cho H, Gazzaniga F, Ilves M, Coronado D, Thompson C, Hang S, Chiu I, Moffitt J, Hsiao A, Mekalanos J, Benoist C, Kasper D (2024) Gut complement induced by the microbiota combats pathogens and spares commensals. Cell. doi: 10.1016/j.cell.2023.12.036 Xu J, Zhang S, Tian Y, Si H, Zeng Y, Wu Y, Liu Y, Li M, Sun K, Wu L, Shen B (2022) Genetic Causal Association between Iron Status and Osteoarthritis: A Two-Sample Mendelian Randomization. Nutrients 14. doi: 10.3390/nu14183683 Yan XY, Yao JP, Li YQ, Xiao XJ, Yang WQ, Chen SJ, Tang TC, Yang YQ, Qu L, Hou YJ, Chen M, Li Y (2023) Effects of acupuncture on gut microbiota and short-chain fatty acids in patients with functional constipation: a randomized placebo-controlled trial. Front Pharmacol 14: 1223742. doi: 10.3389/fphar.2023.1223742 Zhou W, Liu G, Hung RJ, Haycock PC, Aldrich MC, Andrew AS, Arnold SM, Bickeböller H, Bojesen SE, Brennan P, Brunnström H, Melander O, Caporaso NE, Landi MT, Chen C, Goodman GE, Christiani DC, Cox A, Field JK, Johansson M, Kiemeney LA, Lam S, Lazarus P, Le Marchand L, Rennert G, Risch A, Schabath MB, Shete SS, Tardón A, Zienolddiny S, Shen H, Amos CI (2021) Causal relationships between body mass index, smoking and lung cancer: Univariable and multivariable Mendelian randomization. Int J Cancer 148: 1077-1086. doi: 10.1002/ijc.33292 Additional Declarations No competing interests reported. Supplementary Files SupplementaryFig.1.tif SupplementaryFig.2.tif SupplementaryFig.3.tif SupplementaryFig.4.tif SupplementaryTable1.xlsx SupplementaryTable2.xlsx SupplementaryTable3.xlsx SupplementaryTable4.xlsx SupplementaryInformationsandLegends.docx Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4482347","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":317841877,"identity":"e2fb1998-0470-45f3-8cea-9aa6e3a4850e","order_by":0,"name":"Dingwen Fan","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAxElEQVRIiWNgGAWjYBACeWb+DwcSKmyY+dkbiNRi2M5geODDmTR2yZ4DxFpznsH44My2Q/wGNxKI1MHYzJBwmOfMAWmDm4833mCosYkmqIWdmeHAYZ6KO8aSt9OKLRiOpeU2ELaFsQFoy7Nkvts5ZhJANmEtDIeZGQ7zth2ub7h5hmgtbAxA7x9mFrjBQ6QWw2YeBlAgM0v2AP2SQIxf5PnPMH+AROXhjTc+1NgQ4TAkYCCRQIpyiBZSdYyCUTAKRsHIAAAWUURh1k5WvAAAAABJRU5ErkJggg==","orcid":"","institution":"Hospital of Chengdu University of Traditional Chinese Medicine","correspondingAuthor":true,"prefix":"","firstName":"Dingwen","middleName":"","lastName":"Fan","suffix":""},{"id":317841878,"identity":"67a78a6e-b73d-490b-b50a-5db70d171afc","order_by":1,"name":"Liangliang Zhang","email":"","orcid":"","institution":"Hospital of Chengdu University of Traditional Chinese Medicine","correspondingAuthor":false,"prefix":"","firstName":"Liangliang","middleName":"","lastName":"Zhang","suffix":""},{"id":317841879,"identity":"aa65de58-0082-43da-85fb-55b5586d9979","order_by":2,"name":"Hongchang Liu","email":"","orcid":"","institution":"Hospital of Chengdu University of Traditional Chinese Medicine","correspondingAuthor":false,"prefix":"","firstName":"Hongchang","middleName":"","lastName":"Liu","suffix":""},{"id":317841880,"identity":"b7721eb0-9191-44a0-ac0d-a6dfac9573a7","order_by":3,"name":"Dan Zhu","email":"","orcid":"","institution":"Chengdu Fifth People's Hospital","correspondingAuthor":false,"prefix":"","firstName":"Dan","middleName":"","lastName":"Zhu","suffix":""}],"badges":[],"createdAt":"2024-05-27 05:08:25","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4482347/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4482347/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":58924845,"identity":"377d6191-5e49-49f6-a23b-32d7bdda5b62","added_by":"auto","created_at":"2024-06-24 07:59:22","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":366709,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eUVMR analyses of the relationship genus Eubacterium rectale/genus Eubacterium brachy groups and constipation \u003c/strong\u003ea. Scatterplot plot of causal association analysis between Eubacterium rectale genus group and constipation. b. Scatterplot plot of causal association analysis between Eubacterium brachy group and constipation. c. The forest plot of causal association analysis between Eubacterium rectale genus group and constipation. d. The forest plot of causal association analysis between Eubacterium brachy group and constipation. e. Funnel plot of Eubacterium rectale genus group and constipation. f. Funnel plot of Eubacterium brachy group and constipation.\u003c/p\u003e","description":"","filename":"figure1.png","url":"https://assets-eu.researchsquare.com/files/rs-4482347/v1/65600296d3804b0548b38f5e.png"},{"id":58924846,"identity":"82ff748c-dbed-445f-81fd-b96bc72fb895","added_by":"auto","created_at":"2024-06-24 07:59:22","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":153848,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eUVMR LOO sensitivity analyses for genus Eubacterium rectale/genus Eubacterium brachy groups and constipation \u003c/strong\u003ea. UVMR LOO sensitivity analysis for genus Eubacterium rectale group and constipation. b. UVMR LOO sensitivity analysis for genus Eubacterium brachy group and constipation.\u003c/p\u003e","description":"","filename":"figure2.png","url":"https://assets-eu.researchsquare.com/files/rs-4482347/v1/dfa996184cd3e0be2329ee88.png"},{"id":58924844,"identity":"7921b554-6f25-40bd-a573-b32ae6db7372","added_by":"auto","created_at":"2024-06-24 07:59:22","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":53678,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eMVMR analyses of the relationship genus Eubacterium rectale/genus Eubacterium brachy groups and constipation\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"figure3.png","url":"https://assets-eu.researchsquare.com/files/rs-4482347/v1/336bd5c3db977ab5dcbcc6db.png"},{"id":59115506,"identity":"e7c81434-59dd-497e-8b9c-2a5288300d33","added_by":"auto","created_at":"2024-06-26 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encountered in the realm of digestive issues and is characterized by challenges in stool evacuation, discomfort during bowel movements, or the need for marked exertion to eliminate feces(Bharucha \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2007\u003c/span\u003e; Rao \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e2009\u003c/span\u003e; Rao and Singh \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). The global prevalence of constipation is approximately 14%, with the incidence varying across different demographics(Black and Ford \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Specifically, the incidence rates in children and the elderly (aged 60 and over) are about 9.5% and 33.5%, respectively(Forootan et al. \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Koppen et al. \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). While constipation is not inherently life-threatening, chronic constipation can substantially degrade the quality of life(Bharucha and Wald \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Furthermore, constipation markedly elevates the risk of coronary heart disease and ischemic stroke and increases all-cause mortality among the elderly, thus imposing a significant health burden on this age group(Sumida et al. \u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Substantial current evidence suggests that the gut microbiome, bile secretion, lifestyle, and behavioral psychology are all factors influencing constipation.(Cha et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Erhardt et al. \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Neves et al. \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Wang et al. \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Central to intestinal well-being and digestion is the gut microbiome\u0026mdash;a rich ecosystem of microorganisms such as bacteria, fungi, and viruses that inhabit the human gastrointestinal tract(Wu et al. \u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). In the past few years, there has been a growing focus on investigating the correlation between gut microorganisms and illnesses(Cui et al. \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2024\u003c/span\u003e; Gilbert et al. \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Harnett et al. \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Kong et al. \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2023\u003c/span\u003e); however, substantial knowledge regarding the direct impact of gut microbes on constipation is still lacking(Oliphant and Allen-Vercoe \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Quigley \u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e2019\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe Eubacterium genus, comprising prevalent anaerobic bacteria within the gastrointestinal tracts of both humans and animals(Mukherjee et al. \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2020\u003c/span\u003e), is essential for the metabolic processing of carbohydrates(Pessoa et al. \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e2023\u003c/span\u003e), amino acids(Jin et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2023\u003c/span\u003e), and fatty acids(Yan et al. \u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e2023\u003c/span\u003e), as well as for normal digestive processes(Gomes et al. \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Crucial to the integrity of intestinal health is the presence of an appropriate population of Eubacterium in the gut's microbial ecosystem. Studies have revealed a clear association between a diminished presence or complete lack of Eubacterium strains and the development of inflammatory bowel conditions and constipation(El-Salhy \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Hong and Rhee \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Rendeli et al. \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). In theory, Eubacterium could influence constipation through various mechanisms. Initially, Eubacterium strains can be involved in carbohydrate metabolism and generate short-chain fatty acids, which are crucial for preserving the health of the intestinal lining and enhancing the movement of the intestines(Shoaie et al. \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). Additionally, these strains of Eubacterium may collaborate with other intestinal microbes to maintain a healthy microbial balance in the gut and affect the activities of the intestinal immune system(Hiippala et al. \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Magrone and Jirillo \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). Nevertheless, even though there is research backing the link between Eubacterium and constipation(Galkin et al. \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Odamaki et al. \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e2016\u003c/span\u003e), the gut microbial environment is intricate and affected by various elements, such as diet, lifestyle, and genetics. Consequently, the exact nature of the cause-and-effect relationship between Eubacterium and constipation remains uncertain.\u003c/p\u003e \u003cp\u003eMendelian randomization (MR) represents a technique for identifying causal links between exposure factors and outcomes(Burgess and Thompson \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). MR analyses are contingent upon three pivotal assumptions. The first is the relevance assumption, signifying that the chosen instrumental variables (IVs) should have a robust correlation with the exposure of interest, such as a specific biomarker or behavior. Next, the independence assumption dictates that the IVs need to be free from any confounding factors. Lastly, the exclusion-restriction assumption demands that the influence of the instrumental variable on the disease outcome should only be mediated through its effect on the exposure, and not via alternative pathways(Burgess et al. \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). This method capitalizes on the naturally random distribution of genetic variations, which are unconnected to ethnicity, gender, or status at birth, to offer a robust analysis of clinical intervention efficacy while reducing the risks of reverse causality and confounding variables(Burgess et al. \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2012\u003c/span\u003e; Richmond and Davey Smith \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Our study applied MR with the intention of clarifying the causative link between Eubacterium and constipation, in order to provide a theoretical basis for the improvement of clinical practices.\u003c/p\u003e"},{"header":"Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eSource of data and study analysis\u003c/h2\u003e \u003cp\u003eThe Integrative Epidemiology Unit (IEU) Open Genome-Wide Association Study (GWAS) database (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://gwas.mrcieu.ac.uk/\u003c/span\u003e\u003cspan address=\"https://gwas.mrcieu.ac.uk/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) was used to search for genus Eubacterium and constipation, and the traitID and GWAS data were obtained. The 11 species of genus Eubacillus was the exposure factor, constipation was the outcome. The datasets of 11 species of genus Eubacillus shown in \u003cb\u003eSupplementary Table\u0026nbsp;1\u003c/b\u003e, the ebi-a-GCST90017003 of constipation included 5,708,381 single nucleotide polymorphisms (SNPs) from 14,306 samples.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003eUnivariable Mendelian randomization (UVMR) analysis\u003c/h2\u003e \u003cp\u003eThe extract_instruments in TwoSampleMR package (Hemani et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2018\u003c/span\u003e) was used to read exposure factors and filter instrument variables (IVs) with p\u0026thinsp;\u0026lt;\u0026thinsp;5*10\u0026thinsp;\u0026minus;\u0026thinsp;6. The linkage disequilibrium analysis (LDA) were performed to ensure independence (clump\u0026thinsp;=\u0026thinsp;TRUE, r2\u0026thinsp;=\u0026thinsp;0.001 and kb\u0026thinsp;=\u0026thinsp;100). The mr function of TwoSampleMR package combined with five algorithms (MR Egger (Bowden et al. \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2015\u003c/span\u003e), Weighted median(Bowden et al. \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2016\u003c/span\u003e), Inverse variance weighted (IVW)(Burgess et al. \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2015\u003c/span\u003e), Simple mode, Weighted mode(Hartwig et al. \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2017\u003c/span\u003e)) to carry out UVMR analysis. Results were mainly referred to IVW. The foundational assumptions of this MR analysis were based on three key principles: a strong correlation between the IVs and the exposure, the independence of IVs from potential confounders, and the exclusive influence of IVs on the outcome through the exposure without involvement in alternative pathways.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003eMultivariable-MR (MVMR) analysis\u003c/h2\u003e \u003cp\u003eThe input data in MVMR analysis were outcome and exposure factors that were significantly associated with constipation in UVMR analysis. The SNPs significantly associated with exposure factors were searched by the TwoSampleMR package \u0026ldquo;mv extract exposures\u0026rdquo; (screening criteria: p\u0026thinsp;=\u0026thinsp;5e-08, clump\u0026thinsp;=\u0026thinsp;TRUE, r2\u0026thinsp;=\u0026thinsp;0.001 and kb\u0026thinsp;=\u0026thinsp;10000). The TwoSampleMR package \u0026ldquo;extract outcome data\u0026rdquo; was used to read exposure factors and screen SNPs (screening criteria: proxies\u0026thinsp;=\u0026thinsp;TRUE; rsq\u0026thinsp;=\u0026thinsp;0.8). Finally, MVMR was performed through the TwoSampleMR package function mv_harmonise_data\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003eSensitivity analysis\u003c/h2\u003e \u003cp\u003eTo assess the reliability of MR analysis, sensitivity analysis were performed. In the heterogeneity test, Q pvalue greater than 0.05 indicated that there was no heterogeneity. In pleiotropy test, p value greater than 0.05 indicated that there was no horizontal pleiotropy. Leave-one-out (LOO) sensitivity test was mainly to judge the impact of the remaining SNPs on the outcome after removing the SNP one by one. If the result changed significantly, the outcome was sensitive to the SNP(Xu et al. \u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e2022\u003c/span\u003e) .\u003c/p\u003e \u003c/div\u003e"},{"header":"Result","content":"\u003cp\u003e \u003cb\u003eEubacterium rectale group id.14374/Eubacterium rectale group id.14374 had a significant causal relationship with constipation inUVMR analysis\u003c/b\u003e \u003c/p\u003e \u003cp\u003eIVW results showed that genus Eubacterium rectale group id.14374 (p\u0026thinsp;=\u0026thinsp;0.043, OR\u0026thinsp;=\u0026thinsp;0.805) and genus Eubacterium brachy group id.11296 (p\u0026thinsp;=\u0026thinsp;0.036, OR\u0026thinsp;=\u0026thinsp;0.904) had significant negative causal relationship with constipation \u003cb\u003e(Supplementary Table\u0026nbsp;2)\u003c/b\u003e. However, the remaining nine genus Eubacterium did not have a significant causal relationship with constipation \u003cb\u003e(Supplementary Table\u0026nbsp;2)\u003c/b\u003e. The scatter plot results of IVW algorithm showed negative causal relationship between Eubacterium rectale group id.14374/ genus Eubacterium brachy group id.11296 and constipation \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea-\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb, \u003cb\u003eSupplementary Fig.\u0026nbsp;1)\u003c/b\u003e. It can be seen from the forest map of the IVW algorithm that the effect values of Eubacterium rectale group id.14374/ genus Eubacterium brachy group id.11296 on constipation were all less than 0, indicating that the two genus of Eubacterium were protective factors on the constipation \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ec-\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ed, \u003cb\u003eSupplementary Fig.\u0026nbsp;2)\u003c/b\u003e. In addition, the samples were symmetrically distributed along both sides of the IVW line in the funnel plot, indicating that the MR analysis conformed to randomness \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ee-\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ef, \u003cb\u003eSupplementary Fig.\u0026nbsp;3)\u003c/b\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eTo evaluate the reliability of the UVMR analysis, we performed a sensitivity analysis. Heterogeneity results showed that there was no heterogeneity in genus Eubacterium rectale group id.14374 on constipation (Q pvalue\u0026thinsp;=\u0026thinsp;0.439) and genus Eubacterium brachy group id.11296 on constipation (Q pvalue\u0026thinsp;=\u0026thinsp;0.928) MR analysis \u003cb\u003e(\u003c/b\u003eTable\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e, \u003cb\u003eSupplementary Table\u0026nbsp;3)\u003c/b\u003e. Horizontal pleiotropy test results showed that the P-value of the two exposure factors were all greater than 0.05 (genus Eubacterium rectale group id.14374 pvalue\u0026thinsp;=\u0026thinsp;0.78, genus Eubacterium brachy group id.11296 pvalue\u0026thinsp;=\u0026thinsp;0.45), indicated that there was no horizontal pleiotropy of the analysis results \u003cb\u003e(\u003c/b\u003eTable\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e, \u003cb\u003eSupplementary Table\u0026nbsp;4)\u003c/b\u003e. The LOO test results of two exposure factors were on the left side of 0, indicating that there were no SNP with overly sensitive outcome \u003cb\u003e(\u003c/b\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ea-\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eb, \u003cb\u003eSupplementary Fig.\u0026nbsp;4)\u003c/b\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eThe heterogeneity test of causality between genus Eubacterium rectale/genus Eubacterium brachy groups and constipation based on MR results\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"6\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eoutcome\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eexposure\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003emethod\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eQ\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eQ_df\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eQ_pval\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eConstipation\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003egenus Eubacterium rectale group\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMR Egger\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e2.578125498\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.276\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eConstipation\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003egenus Eubacterium rectale group\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eInverse variance weighted\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e2.708439662\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.439\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eConstipation\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003egenus Eubacterium brachy group\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMR Egger\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.658221819\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.956\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eConstipation\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003egenus Eubacterium brachy group\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eInverse variance weighted\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e1.364387018\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.928\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"6\"\u003eMR mendelian randomization\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eThe horizontal pleiotropy test of causality between genus Eubacterium rectale/genus Eubacterium brachy groups and constipation based on MR results\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eoutcome\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eexposure\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eegger_intercept\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003ese\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003epval\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eConstipation\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003egenus Eubacterium rectale group\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e-0.009880127\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.031074472\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.780650712\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eConstipation\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003egenus Eubacterium brachy group\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.021345491\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.025401128\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.448015795\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"5\"\u003eMR mendelian randomization, se standard error\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eThe genus Eubacterium rectale group id.14374 still was protective factors on constipation in MVMR analysis\u003c/h2\u003e \u003cp\u003eIn MVMR analysis, genus Eubacterium rectale group id.14374 still had a significant negative causal effect on constipation (p\u0026thinsp;=\u0026thinsp;0.007, OR\u0026thinsp;=\u0026thinsp;0.80), indicating that its causal effect on constipation was more independent, while the causal effect of genus Eubacterium brachy group id.11296 on constipation was diluted (p\u0026thinsp;=\u0026thinsp;0.151, OR\u0026thinsp;=\u0026thinsp;0.94) \u003cb\u003e(\u003c/b\u003eTable\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e, Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e\u003cb\u003e)\u003c/b\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eThe MVMR results between between genus Eubacterium rectale/genus Eubacterium brachy groups and constipation\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"8\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eexposure\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eoutcome\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eb\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003ese\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003epval\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eor\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eor_lci95\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eor_uci95\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003egenus Eubacterium brachy group\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eConstipation\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e-0.062937802\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.043836734\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.151\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.939001876\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.861691639\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e1.023248322\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003egenus Eubacterium rectale group\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eConstipation\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e-0.219443616\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.081435751\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.007\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e \u003cp\u003e0.802965431\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e \u003cp\u003e0.684506124\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e \u003cp\u003e0.941925076\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eMR mendelian randomization, se, standard error, OR odds ratio, CI confidence interval\u003c/p\u003e \u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eIn our study, a novel exploration into the causative link between the genus Eubacterium and constipation was undertaken, with 11 species within the Eubacillus genus being investigated as the exposure element, while constipation served as the observed effect. Our findings indicated that both the genus Eubacterium rectale group and the genus Eubacterium brachy group acted as protective elements against constipation in univariate MR analyses. Furthermore, in multivariate analysis, the genus Eubacterium rectale group continued to stand out as a significant protectiveguard. These insights lay a theoretical foundation for clinical investigations into the relationship between Eubacterium and constipation, offering diverse analytical perspectives.\u003c/p\u003e \u003cp\u003ePracimvot first introduced the term 'Eubacterium' in 1938 as a designation for a cluster of beneficial bacteria derived from human stool samples(Andreesen \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1992\u003c/span\u003e). Commonly found within the mouths and gastrointestinal tracts of mammals, such as the stomachs of cud-chewing animals, the Eubacterium genus is also present in various environmental settings(Karcher et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). This genus plays a central role in the gut flora in humans and is extensively prevalent among various demographic groups across continents such as Africa, Australia, Europe, India, South America, Asia, and North America(Ayeni et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Escobar et al. \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Gomez-Arango et al. \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Mueller et al. \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2006\u003c/span\u003e). The Eubacterium rectale group and the Eubacterium brachy group represent specific bacterial groups within this genus, each with unique features and functions. Although both belong to the genus Eubacterium, they may differ in terms of bacterial composition, metabolic products, and other aspects.\u003c/p\u003e \u003cp\u003eThe genus Eubacterium rectale, a species within the human microbial population, is recognized for its capacity to generate butyrate(Lu et al. \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2022\u003c/span\u003e), a short-chain fatty acid (SCFAs) with a crucial role in protectiveguarding the gut lining and reducing inflammation within the intestines(Biagi et al. \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2013\u003c/span\u003e; Magrone and Jirillo \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). As individuals age, there is a notable reduction in the variety of gut microbes, coinciding with a diminished production of butyrate by this specific genus, a factor linked to the onset of gastrointestinal disorders(Ling et al. \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Throughout the digestive process, an optimal level of Eubacterium rectale supports the uptake of indigestible elements like dietary fibers, while also generating butyrate, which protectiveguards intestinal health and assists in preventing constipation(Topping and Clifton \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e2001\u003c/span\u003e). Moreover, this genus is a member of the intestinal bacterial consortium that ferments fibers like cellulose and resistant starch. It contributes to the production of additional varieties of SCFAs, including acetic and propionic acids, which are essential for overall gut health(Mukherjee et al. \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). SCFAs, known for their crucial role in the modulating intestinal inflammation, offer beneficial effects on constipation by enhancing intestinal integrity and regulating immune responses(Chen et al. \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). More specifically, SCFAs positively influence the expression levels of junctional proteins (such as claudin-1 and occludin), thereby increasing the density of intercellular junctions(Wang et al. \u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e2012\u003c/span\u003e; Willemsen et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2003\u003c/span\u003e). This process effectively protectiveguards against the infiltration of harmful substances into intestinal tissues. Additionally, SCFAs activate G protein-coupled receptors (GPCRs), a group of proteins responsible for transmitting vital cellular signals. This activation process directly impacts the activity and functionality of immune cells, thereby contributing to immune response modulation(Corr\u0026ecirc;a-Oliveira et al. \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). Eubacterium rectale plays a role in alleviating constipation by enhancing the movement of the intestines. It has been documented that SCFAs can influence the movement within the intestines regardless of pH levels. The time it takes for substances to move through the intestines may decrease when propionate and butyrate solutions are introduced into the colon(Cherbut et al. \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e1998\u003c/span\u003e; Tazoe et al. \u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e2008\u003c/span\u003e). Moreover, introducing mixtures that contain SCFAs has been shown to boost motility in the ileum more effectively than either air or saline solutions(Jou\u0026euml;t et al. \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). In conclusion, Eubacterium rectale helps manage constipation by preserving intestinal mucosal integrity, modulating gut immunity, reducing inflammation, and enhancing intestinal motility.\u003c/p\u003e \u003cp\u003eCurrent understanding of the metabolic processes and roles of Eubacterium brachy remains limited. Nonetheless, research indicates that it might share metabolic activities with Eubacterium rectale, including the synthesis of short-chain fatty acids(Mukherjee et al. \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Additionally, several studies have linked Eubacterium brachy to gastrointestinal well-being and inflammation-related oral conditions(Li et al. \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Moutsopoulos et al. \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Vincent et al. \u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e1986\u003c/span\u003e), though the precise pathways involved remain to be elucidated. Recent research demonstrates a decline in Eubacterium brachy population following nine weeks on a diet rich in fats(Pessoa et al. \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e2023\u003c/span\u003e), yet the connection between such a diet, Eubacterium brachy levels, and constipation remains undetermined. Moreover, a reduction in Eubacterium brachy has been noted in cases of inflammatory bowel disease(Li et al. \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2023\u003c/span\u003e), which implies that Eubacterium brachy may exert a beneficial effect on intestinal health. Significantly, the modulation of constipation by Eubacterium brachy appears to be linked to the production of its metabolites known as SCFAs. These compounds are potentially crucial for their roles in regulating the immune system, managing inflammation, and protectiveguarding the gut's integrity. However, it is still uncertain whether other factors or different metabolites are involved in this regulatory mechanism, an area that requires further exploration.\u003c/p\u003e \u003cp\u003eConsidering the co-presence of both the Eubacterium rectale group and the Eubacterium brachy group in the human intestines, we conducted additional multivariate MR analyses with both groups as exposure variables and constipation as the outcome. While the univariate MR analysis provided evidence of causality, it is important and valid to perform multivariate MR analysis, as it can potentially yield more evidence(Chen et al. \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2022\u003c/span\u003e; Zhou et al. \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). The findings of our study revealed that the Eubacterium rectale group remained a significant protective factor in the multivariate MR analysis, whereas the significance of the Eubacterium brachy group decreased. This suggests that the Eubacterium brachy group may have a lesser causal role in constipation compared to the Eubacterium rectale group. It is also possible that these two groups interact in the development of constipation, emphasizing the need for further research in this area. Undoubtedly, the Eubacterium rectale group has been identified as a protective factor against constipation in both univariate and multivariate MR analyses. This consistent finding is expected to establish a new theoretical basis for the development of targeted interventions that utilize this genus to alleviate constipation symptoms.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eOur investigation faced certain inherent constraints. Despite our examination of the prevalent occurrence of Eubacterium across continents such as Africa, Australia, Europe, India, South America, Asia, and North America, MR analyses were constrained to European population data, thereby omitting data from other ethnic groups and necessitating a more circumspect approach when applying findings to diverse populations. Additionally, the goal of MR analyses in gauging the life-long influence of Eubacterium rectale and Eubacterium brachy groups raised the potential of age-related changes in the genetic instruments, potentially impacting the results' interpretability and accuracy. Nevertheless, our study's clinical significance is underlined by the identification of a causal link between the Eubacterium rectale group and constipation, laying a theoretical foundation for future clinical applications. Our future research will aim to expand the sample size, further investigate the interactions between different bacterial species, and incorporate other research methodologies to validate and reinforce the findings of this study.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eCompeting Interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors have no relevant financial or non-financial interests to disclose.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData Availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe datasets analysed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe data utilized in our analysis were derived from publicly available sources and were obtained, ensuring compliance with the principles of the declaration of Helsinki and approval by the relevant ethics committees. Furthermore, our findings were presented in alignment with the guidelines outlined in the Mendelian Randomization STrengthening the Reporting of OB servational studies using Genetic Epidemiology (MR STROBE) framework.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eAndreesen JR (1992) The genus Eubacterium. Prokaryotes: 1914-1924. \u003c/li\u003e\n\u003cli\u003eAyeni FA, Biagi E, Rampelli S, Fiori J, Soverini M, Audu HJ, Cristino S, Caporali L, Schnorr SL, Carelli V, Brigidi P, Candela M, Turroni S (2018) Infant and Adult Gut Microbiome and Metabolome in Rural Bassa and Urban Settlers from Nigeria. Cell Rep 23: 3056-3067. doi: 10.1016/j.celrep.2018.05.018\u003c/li\u003e\n\u003cli\u003eBharucha AE (2007) Constipation. Best Pract Res Clin Gastroenterol 21: 709-31. doi: 10.1016/j.bpg.2007.07.001\u003c/li\u003e\n\u003cli\u003eBharucha AE, Wald A (2019) Chronic Constipation. 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Int J Cancer 148: 1077-1086. doi: 10.1002/ijc.33292\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-4482347/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4482347/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eBackground: \u003c/strong\u003eConstipation may be related to an imbalance in flora imbalance. In this study, univariate and multivariate Mendelian randomization (MR) studies were used to further understand the causal relationship between different members of genus \u0026nbsp;Eubacteriumand constipation.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods: \u003c/strong\u003eThe constipation and genus Eubacterium datasets were obtained from the Integrative Epidemiology Unit (IEU) Open Genome-Wide Association Study (GWAS) database. The causal effects between constipation and genus Eubacterium were analyzed using MR-Egger, Weighted median, Simple mode, Weighted mode and Inverse variance weighted (IVW). In univariate MR analysis (UVMR), genus Eubacillus was the exposure factor, constipation was the outcome. In addition, the reliability of UVMR analysis was assessed by sensitivity analyses (heterogeneity tests, horizontal pleiotropy tests and leave-one-out (LOO) analysis). Finally, the MVMR analysis was performed.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults: \u003c/strong\u003eIn the UVMR analysis, genus Eubacterium rectale group id.14374 (p=0.043, OR=0.805) and genus Eubacterium brachy group id.11296 (p=0.036, OR=0.904) were all protective factors on constipation. The results of sensitivity analysis showed that UVMR analysis was reliable. In the MVMR analysis, genus Eubacterium rectalegroup was still a significant protective factor (p=0.007, OR=0.80), while genus Eubacterium brachy group was not a significant protective factor for constipation (p=0.151, OR=0.94) .\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusion: \u003c/strong\u003eThe results of study supported that genus Eubacterium rectale group had a significant causal relationship with constipation, and the increase of genus Eubacterium rectale group could reduce the risk of constipation.\u003c/p\u003e","manuscriptTitle":"The causal relationship between Eubacterium and constipation: a Mendelian randomization study","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-06-24 07:59:18","doi":"10.21203/rs.3.rs-4482347/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"96eed624-5f36-4e2f-bca6-829af0ab777d","owner":[],"postedDate":"June 24th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2024-06-26T13:48:14+00:00","versionOfRecord":[],"versionCreatedAt":"2024-06-24 07:59:18","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-4482347","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4482347","identity":"rs-4482347","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}