Unravelling the efficacy of Enterococcus faecalis: A statistical approach to select cost effective medium

Unravelling the efficacy of Enterococcus faecalis: A statistical approach to select cost effective medium | 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 Article Unravelling the efficacy of Enterococcus faecalis: A statistical approach to select cost effective medium Mangala Lakshmi Ragavan, Hemalatha S This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6180079/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 The human gut microbiota comprises millions of microbes that confer various health benefits. Maintaining a healthy gut microbiome necessitates the consumption of appropriate dietary supplements. This study endeavours to identify an optimal dietary supplement for fostering the growth of Enterococcus faecalis (isolated from human origin) and Lactobacillus acidophilus (MTCC 10307). E. faecalis displays notable tolerance under simulated gastrointestinal conditions (87%), akin to L. acidophilus (88%). Initially, a screening of diverse dietary materials (including millets and legumes) was conducted using the Plackett-Burman Design (PBD) method. Among the 11 materials assessed, foxtail millet, barnyard millet, finger millet, and cowpea demonstrated significant effects on the growth rate of probiotics. However, further optimization of conditions is necessary to augment the growth of these probiotics. Subsequently, the Box-Behnken Design (BBD) was employed to derive the ANOVA for a quadratic model utilizing Design Expert software. The growth rate was positively influenced by variables such as (A) foxtail millet, (B) barnyard millet, (C) finger millet, and (D) cowpea, either individually or in combinations of two variables (AB, AC, BC, BD, CD). Particularly noteworthy was the enhancement of E. faecalis growth by approximately 92.78% with the supplementation of foxtail and barnyard millets at a concentration of 100 mg/ml. Moreover, growth kinetics indicated that increased microbial growth resulted in a reduction of substrate (foxtail millet) concentration. The specific growth rate of 1.26 Log CFU/ml was observed in E. faecalis monoculture, with foxtail consumption accounting for 95% of this growth. However, co-cultures of E. faecalis and L. acidophilus led to an inhibition of up to 84% in the growth rate of S. typhimurium . Consequently, millet-based media showcased a superior capacity to enhance probiotic growth compared to MRS media. Thus, foxtail millets emerge as a compelling dietary supplement for augmenting the human gut microbiome. Biological sciences/Biotechnology Biological sciences/Microbiology Probiotics GIT tolerance Millets Legumes Response surface methodology Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 Full Text Additional Declarations No competing interests reported. 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-6180079","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":445564540,"identity":"69d758a6-c35b-4743-9b00-aa5a510d41c4","order_by":0,"name":"Mangala Lakshmi Ragavan","email":"","orcid":"","institution":"BS Abdur Rahman Crescent Institute of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Mangala","middleName":"Lakshmi","lastName":"Ragavan","suffix":""},{"id":445564541,"identity":"f4a7b0e9-bbe7-4cea-837c-3ac145b12a2e","order_by":1,"name":"Hemalatha S","email":"data:image/png;base64,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","orcid":"","institution":"BS Abdur Rahman Crescent Institute of Science and Technology","correspondingAuthor":true,"prefix":"","firstName":"Hemalatha","middleName":"","lastName":"S","suffix":""}],"badges":[],"createdAt":"2025-03-07 17:23:22","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6180079/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6180079/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":81141807,"identity":"90f7a647-98a5-4556-955b-7445454a24fa","added_by":"auto","created_at":"2025-04-22 16:55:26","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":124877,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003eE. faecalis\u003c/em\u003eexhibitexcellent probiotic properties\u003cstrong\u003e(a)\u003c/strong\u003eGrowth analysis of microbes, \u003cstrong\u003e(b)\u003c/strong\u003e Survival of microbes under simulated GIT conditions\u003c/p\u003e","description":"","filename":"Figure1.png","url":"https://assets-eu.researchsquare.com/files/rs-6180079/v1/17d781704f1fa87cb17afa40.png"},{"id":81141809,"identity":"64a4e6dd-e273-4968-a84f-63cc5c34fc0e","added_by":"auto","created_at":"2025-04-22 16:55:26","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":130440,"visible":true,"origin":"","legend":"\u003cp\u003eCell surface hydrophobicity index of microbes\u003c/p\u003e","description":"","filename":"Figure2.png","url":"https://assets-eu.researchsquare.com/files/rs-6180079/v1/b0993f227e8e8c6587323b3c.png"},{"id":81142367,"identity":"658ee367-472d-4324-87bb-32b5813a436e","added_by":"auto","created_at":"2025-04-22 17:03:26","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":113244,"visible":true,"origin":"","legend":"\u003cp\u003eAdhesion properties of \u003cem\u003eE. Faecalis\u003c/em\u003eand \u003cem\u003eL. acidophilus\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003e(a) Auto-aggregation ability and (b) Co-aggregation ability along with pathogens\u003c/p\u003e","description":"","filename":"Figure3.png","url":"https://assets-eu.researchsquare.com/files/rs-6180079/v1/827d21a91c9d1950feedbe05.png"},{"id":81143307,"identity":"d87cf7c7-5eb4-44a9-9211-f53a4b113089","added_by":"auto","created_at":"2025-04-22 17:19:26","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":2501265,"visible":true,"origin":"","legend":"\u003cp\u003eAntimicrobial activity of \u003cem\u003eL. acidophilus\u003c/em\u003e(LC) and \u003cem\u003eE. Faecalis\u003c/em\u003e(EF) against (a) \u003cem\u003eS. typhimurium\u003c/em\u003e, (b) \u003cem\u003eS. aureus\u003c/em\u003e(c) \u003cem\u003eK. pneumoniae\u003c/em\u003e, (d) \u003cem\u003eS. boydii\u003c/em\u003e, and (e) \u003cem\u003eE. coli\u003c/em\u003e\u003c/p\u003e","description":"","filename":"Figure4.png","url":"https://assets-eu.researchsquare.com/files/rs-6180079/v1/8105d1bcc820066a0e05f6d6.png"},{"id":81143306,"identity":"2e363279-1885-430e-a4aa-753743959ebb","added_by":"auto","created_at":"2025-04-22 17:19:26","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":72766,"visible":true,"origin":"","legend":"\u003cp\u003ePlackett-Burman design to screen efficient dietary materials (a) Pareto chart indicate the effect of variables, (b) correlation between actual and predicted values on growth rate of \u003cem\u003eE. faecalis\u003c/em\u003e\u003c/p\u003e","description":"","filename":"Figure5.png","url":"https://assets-eu.researchsquare.com/files/rs-6180079/v1/4a5b6a8449ee0b6912e39b9c.png"},{"id":81141818,"identity":"b5a4cdfb-638d-40a3-a195-ec80c7c5331a","added_by":"auto","created_at":"2025-04-22 16:55:26","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":132815,"visible":true,"origin":"","legend":"\u003cp\u003eProcess optimization to enhance the growth of \u003cem\u003eE. faecalis \u003c/em\u003eusing response surface methodology, 2D and 3D plots shows effect of interactions (a) foxtail and barnyard millet (b) foxtail and finger millet and (c) foxtail millet and cow pea\u003c/p\u003e","description":"","filename":"Figure6.png","url":"https://assets-eu.researchsquare.com/files/rs-6180079/v1/79c6a72586a7d6a90f92b756.png"},{"id":81142369,"identity":"03459349-840f-422f-bc77-20e6e8b1735b","added_by":"auto","created_at":"2025-04-22 17:03:26","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":162507,"visible":true,"origin":"","legend":"\u003cp\u003eProcess optimization to enhance the growth of \u003cem\u003eE. faecalis \u003c/em\u003eusing response surface methodology, 2D and 3D plots shows effects of interactions (a) barnyard millet and finger millet (b) barnyard millet and cow pea and (c) cow pea and finger millet\u003c/p\u003e","description":"","filename":"Figure7.png","url":"https://assets-eu.researchsquare.com/files/rs-6180079/v1/bded5fd82c5673df75b14e70.png"},{"id":81142373,"identity":"cea562b4-93f9-49cf-b6ac-8a4797975b93","added_by":"auto","created_at":"2025-04-22 17:03:26","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":75592,"visible":true,"origin":"","legend":"\u003cp\u003eStatistical analysis for optimized parameters (a) Normal plot of Residuals, (b) Actual vs predicted and (c) Box-Cox plot for power transforms\u003c/p\u003e","description":"","filename":"Figure8.png","url":"https://assets-eu.researchsquare.com/files/rs-6180079/v1/215f1d1f76fe39714ea1f0a7.png"},{"id":81141822,"identity":"a048c5d4-78b7-427b-b11e-b10ead89f264","added_by":"auto","created_at":"2025-04-22 16:55:26","extension":"png","order_by":9,"title":"Figure 9","display":"","copyAsset":false,"role":"figure","size":79083,"visible":true,"origin":"","legend":"\u003cp\u003eGrowth kinetics of \u003cem\u003eE. faecalis \u003c/em\u003e(a) specific growth rate at 12th h, (b) biomass conc. vs substrate conc.\u003c/p\u003e","description":"","filename":"Figure9.png","url":"https://assets-eu.researchsquare.com/files/rs-6180079/v1/4aff5dacf40fbf8000ad8b7b.png"},{"id":81143040,"identity":"33460a7f-3ade-49b8-8200-b004d6b29a9e","added_by":"auto","created_at":"2025-04-22 17:11:26","extension":"png","order_by":10,"title":"Figure 10","display":"","copyAsset":false,"role":"figure","size":62949,"visible":true,"origin":"","legend":"\u003cp\u003eGrowth kinetics for co-culture studies to evaluate the efficiency of probiotic growth rate over pathogens(S – \u003cem\u003eS. typhimurium\u003c/em\u003e; ES – \u003cem\u003eE. faecalis\u003c/em\u003e + \u003cem\u003eS. typhimurium\u003c/em\u003e; LS – \u003cem\u003eL. acidophilus\u003c/em\u003e + \u003cem\u003eS. typhimurium\u003c/em\u003e; ELS - \u003cem\u003eE. faecalis\u003c/em\u003e + \u003cem\u003eL. acidophilus\u003c/em\u003e + \u003cem\u003eS. typhimurium\u003c/em\u003e)\u003c/p\u003e","description":"","filename":"Figure10.png","url":"https://assets-eu.researchsquare.com/files/rs-6180079/v1/7e4c3c916e5c6d07a205ab92.png"},{"id":87973825,"identity":"d31662e7-2e6d-4a0c-ac57-5497ea9214d1","added_by":"auto","created_at":"2025-07-31 03:46:45","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1464584,"visible":true,"origin":"","legend":"","description":"","filename":"RevisedManuscriptLakshmi19324.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6180079/v1_covered_fee09695-644b-4a2b-873d-aca8796f59a6.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Unravelling the efficacy of Enterococcus faecalis: A statistical approach to select cost effective medium","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"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":"Probiotics, GIT tolerance, Millets, Legumes, Response surface methodology","lastPublishedDoi":"10.21203/rs.3.rs-6180079/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6180079/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThe human gut microbiota comprises millions of microbes that confer various health benefits. Maintaining a healthy gut microbiome necessitates the consumption of appropriate dietary supplements. This study endeavours to identify an optimal dietary supplement for fostering the growth of \u003cem\u003eEnterococcus faecalis\u003c/em\u003e (isolated from human origin) and \u003cem\u003eLactobacillus acidophilus\u003c/em\u003e (MTCC 10307). \u003cem\u003eE. faecalis\u003c/em\u003e displays notable tolerance under simulated gastrointestinal conditions (87%), akin to \u003cem\u003eL. acidophilus\u003c/em\u003e(88%). Initially, a screening of diverse dietary materials (including millets and legumes) was conducted using the Plackett-Burman Design (PBD) method. Among the 11 materials assessed, foxtail millet, barnyard millet, finger millet, and cowpea demonstrated significant effects on the growth rate of probiotics. However, further optimization of conditions is necessary to augment the growth of these probiotics. Subsequently, the Box-Behnken Design (BBD) was employed to derive the ANOVA for a quadratic model utilizing Design Expert software. The growth rate was positively influenced by variables such as (A) foxtail millet, (B) barnyard millet, (C) finger millet, and (D) cowpea, either individually or in combinations of two variables (AB, AC, BC, BD, CD). Particularly noteworthy was the enhancement of \u003cem\u003eE. faecalis \u003c/em\u003egrowth by approximately 92.78% with the supplementation of foxtail and barnyard millets at a concentration of 100 mg/ml. Moreover, growth kinetics indicated that increased microbial growth resulted in a reduction of substrate (foxtail millet) concentration. The specific growth rate of 1.26 Log CFU/ml was observed in \u003cem\u003eE. faecalis\u003c/em\u003e monoculture, with foxtail consumption accounting for 95% of this growth. However, co-cultures of \u003cem\u003eE. faecalis\u003c/em\u003e and \u003cem\u003eL. acidophilus\u003c/em\u003e led to an inhibition of up to 84% in the growth rate of \u003cem\u003eS. typhimurium\u003c/em\u003e. Consequently, millet-based media showcased a superior capacity to enhance probiotic growth compared to MRS media. Thus, foxtail millets emerge as a compelling dietary supplement for augmenting the human gut microbiome.\u003c/p\u003e","manuscriptTitle":"Unravelling the efficacy of Enterococcus faecalis: A statistical approach to select cost effective medium","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-04-22 16:55:21","doi":"10.21203/rs.3.rs-6180079/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":"a2a623a1-431d-461b-9f5f-109e36133943","owner":[],"postedDate":"April 22nd, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":47430492,"name":"Biological sciences/Biotechnology"},{"id":47430493,"name":"Biological sciences/Microbiology"}],"tags":[],"updatedAt":"2025-07-31T03:38:32+00:00","versionOfRecord":[],"versionCreatedAt":"2025-04-22 16:55:21","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6180079","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6180079","identity":"rs-6180079","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}