Identification of microbiota biomarkers with orthologous gene annotation for type 2 diabetes
Type 2 diabetes (T2D) is a systematic chronic metabolic condition with abnormal sugar metabolism dysfunction, and its complications are the most harmful to human beings and may be life-threatening after long-term durations. Considering the high incidence and severity at late stage, researchers have...
- Autores:
-
Zhang, Yu-Hang
Guo, Wei
Tao, Zeng
ShiQi, Zhang
Chen, Lei
Gamarra, Margarita
Mansour, Romany F.
Escorcia-Gutierrez, Jose
Huang, Tao
Yu Dong, Cai
- Tipo de recurso:
- Article of journal
- Fecha de publicación:
- 2021
- Institución:
- Corporación Universidad de la Costa
- Repositorio:
- REDICUC - Repositorio CUC
- Idioma:
- eng
- OAI Identifier:
- oai:repositorio.cuc.edu.co:11323/8555
- Acceso en línea:
- https://hdl.handle.net/11323/8555
https://repositorio.cuc.edu.co/
- Palabra clave:
- type 2 diabetes
gut microbiome
machine learning
feature selection
support vector machine
microbiota biomarkers
- Rights
- openAccess
- License
- CC0 1.0 Universal
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dc.title.spa.fl_str_mv |
Identification of microbiota biomarkers with orthologous gene annotation for type 2 diabetes |
title |
Identification of microbiota biomarkers with orthologous gene annotation for type 2 diabetes |
spellingShingle |
Identification of microbiota biomarkers with orthologous gene annotation for type 2 diabetes type 2 diabetes gut microbiome machine learning feature selection support vector machine microbiota biomarkers |
title_short |
Identification of microbiota biomarkers with orthologous gene annotation for type 2 diabetes |
title_full |
Identification of microbiota biomarkers with orthologous gene annotation for type 2 diabetes |
title_fullStr |
Identification of microbiota biomarkers with orthologous gene annotation for type 2 diabetes |
title_full_unstemmed |
Identification of microbiota biomarkers with orthologous gene annotation for type 2 diabetes |
title_sort |
Identification of microbiota biomarkers with orthologous gene annotation for type 2 diabetes |
dc.creator.fl_str_mv |
Zhang, Yu-Hang Guo, Wei Tao, Zeng ShiQi, Zhang Chen, Lei Gamarra, Margarita Mansour, Romany F. Escorcia-Gutierrez, Jose Huang, Tao Yu Dong, Cai |
dc.contributor.author.spa.fl_str_mv |
Zhang, Yu-Hang Guo, Wei Tao, Zeng ShiQi, Zhang Chen, Lei Gamarra, Margarita Mansour, Romany F. Escorcia-Gutierrez, Jose Huang, Tao Yu Dong, Cai |
dc.subject.spa.fl_str_mv |
type 2 diabetes gut microbiome machine learning feature selection support vector machine microbiota biomarkers |
topic |
type 2 diabetes gut microbiome machine learning feature selection support vector machine microbiota biomarkers |
description |
Type 2 diabetes (T2D) is a systematic chronic metabolic condition with abnormal sugar metabolism dysfunction, and its complications are the most harmful to human beings and may be life-threatening after long-term durations. Considering the high incidence and severity at late stage, researchers have been focusing on the identification of specific biomarkers and potential drug targets for T2D at the genomic, epigenomic, and transcriptomic levels. Microbes participate in the pathogenesis of multiple metabolic diseases including diabetes. However, the related studies are still non-systematic and lack the functional exploration on identified microbes. To fill this gap between gut microbiome and diabetes study, we first introduced eggNOG database and KEGG ORTHOLOGY (KO) database for orthologous (protein/gene) annotation of microbiota. Two datasets with these annotations were employed, which were analyzed by multiple machine-learning models for identifying significant microbiota biomarkers of T2D. The powerful feature selection method, Max-Relevance and Min-Redundancy (mRMR), was first applied to the datasets, resulting in a feature list for each dataset. Then, the list was fed into the incremental feature selection (IFS), incorporating support vector machine (SVM) as the classification algorithm, to extract essential annotations and build efficient classifiers. This study not only revealed potential pathological factors for diabetes at the microbiome level but also provided us new candidates for drug development against diabetes. |
publishDate |
2021 |
dc.date.accessioned.none.fl_str_mv |
2021-08-19T15:34:31Z |
dc.date.available.none.fl_str_mv |
2021-08-19T15:34:31Z |
dc.date.issued.none.fl_str_mv |
2021-07-09 |
dc.type.spa.fl_str_mv |
Artículo de revista |
dc.type.coar.fl_str_mv |
http://purl.org/coar/resource_type/c_2df8fbb1 |
dc.type.coar.spa.fl_str_mv |
http://purl.org/coar/resource_type/c_6501 |
dc.type.content.spa.fl_str_mv |
Text |
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info:eu-repo/semantics/article |
dc.type.redcol.spa.fl_str_mv |
http://purl.org/redcol/resource_type/ART |
dc.type.version.spa.fl_str_mv |
info:eu-repo/semantics/acceptedVersion |
format |
http://purl.org/coar/resource_type/c_6501 |
status_str |
acceptedVersion |
dc.identifier.issn.spa.fl_str_mv |
1664-302X |
dc.identifier.uri.spa.fl_str_mv |
https://hdl.handle.net/11323/8555 |
dc.identifier.doi.spa.fl_str_mv |
10.3389/fmicb.2021.711244 |
dc.identifier.instname.spa.fl_str_mv |
Corporación Universidad de la Costa |
dc.identifier.reponame.spa.fl_str_mv |
REDICUC - Repositorio CUC |
dc.identifier.repourl.spa.fl_str_mv |
https://repositorio.cuc.edu.co/ |
identifier_str_mv |
1664-302X 10.3389/fmicb.2021.711244 Corporación Universidad de la Costa REDICUC - Repositorio CUC |
url |
https://hdl.handle.net/11323/8555 https://repositorio.cuc.edu.co/ |
dc.language.iso.none.fl_str_mv |
eng |
language |
eng |
dc.relation.references.spa.fl_str_mv |
Arthur, R., Rohrmann, S., Møller, H., Selvin, E., Dobs, A. S., Kanarek, N., et al. (2017). Pre-diabetes and serum sex steroid hormones among US men. Andrology 5, 49–57. doi: 10.1111/andr.12287 Bakris, G. L., Agarwal, R., Anker, S. D., Pitt, B., Ruilope, L. M., Rossing, P., et al. (2020). Effect of finerenone on chronic kidney disease outcomes in type 2 diabetes. N. Engl. J. Med. 383, 2219–2229. doi: 10.1056/nejmoa2025845 Beli, E., Prabakaran, S., Krishnan, P., Evans-Molina, C., and Grant, M. B. (2019). Loss of diurnal oscillatory rhythms in gut microbiota correlates with changes in circulating metabolites in type 2 diabetic db/db mice. Nutrients 11:2310. doi: 10.3390/nu11102310 Bullard, K. M., Cowie, C. C., Lessem, S. E., Saydah, S. H., Menke, A., Geiss, L. S., et al. (2018). Prevalence of diagnosed diabetes in adults by diabetes type—United States, 2016. Morb. Mortal. Wkly. Rep. 67:359. doi: 10.15585/mmwr.mm6712a2 Carrillo-Larco, R. M., Altez-Fernandez, C., Acevedo-Rodriguez, J. G., Ortiz-Acha, K., and Ugarte-Gil, C. (2019). Leptospirosis as a risk factor for chronic kidney disease: a systematic review of observational studies. PLoS Neglect. Trop. Dis. 13:e0007458. doi: 10.1371/journal.pntd.0007458 Chatterjee, S., Khunti, K., and Davies, M. J. (2017). Type 2 diabetes. Lancet 389, 2239–2251. Chen, L., Wang, S., Zhang, Y.-H., Li, J., Xing, Z.-H., Yang, J., et al. (2017). Identify key sequence features to improve CRISPR sgRNA efficacy. IEEE Access 5, 26582–26590. doi: 10.1109/access.2017.2775703 Chen, L., Zeng, T., Pan, X., Zhang, Y. H., Huang, T., and Cai, Y. D. (2019). Identifying methylation pattern and genes associated with breast cancer subtypes. Int. J. Mol. Sci. 20:4269. doi: 10.3390/ijms20174269 Cortes, C., and Vapnik, V. (1995). Support-vector networks. Mach. Learn. 20, 273–297. Deputy, N. P., Kim, S. Y., Conrey, E. J., and Bullard, K. M. (2018). Prevalence and changes in preexisting diabetes and gestational diabetes among women who had a live birth—United States, 2012–2016. Morb. Mortal. Wkly. Rep. 67:1201. doi: 10.15585/mmwr.mm6743a2 Farnsworth, C. W., Shehatou, C. T., Maynard, R., Nishitani, K., Kates, S. L., Zuscik, M. J., et al. (2015). A humoral immune defect distinguishes the response to Staphylococcus aureus infections in mice with obesity and type 2 diabetes from that in mice with type 1 diabetes. Infect. Immun. 83, 2264–2274. doi: 10.1128/iai.03074-14 Fischer, E., Günter, K., and Braun, V. (1989). Involvement of ExbB and TonB in transport across the outer membrane of Escherichia coli: phenotypic complementation of exb mutants by overexpressed tonB and physical stabilization of TonB by ExbB. J. Bacteriol. 171, 5127–5134. doi: 10.1128/jb.171.9.5127-5134.1989 Forslund, K., Hildebrand, F., Nielsen, T., Falony, G., Le Chatelier, E., Sunagawa, S., et al. (2015). Disentangling type 2 diabetes and metformin treatment signatures in the human gut microbiota. Nature 528, 262–266. Gan, Y.-H. (2013). Host susceptibility factors to bacterial infections in type 2 diabetes. PLoS Pathog. 9:e1003794. doi: 10.1371/journal.ppat.1003794 Gherasim, C., Lofgren, M., and Banerjee, R. (2013). Navigating the B12 road: assimilation, delivery, and disorders of cobalamin. J. Biol. Chem. 288, 13186–13193. doi: 10.1074/jbc.r113.458810 Goldstein, B. J. (2002). Insulin resistance as the core defect in type 2 diabetes mellitus. Am. J. Cardiol. 90, 3–10. doi: 10.1016/s0002-9149(02)02553-5 Górski, A., Międzybrodzki, R., Weber-Da̧browska, B., Fortuna, W., Letkiewicz, S., Rogóż, P., et al. (2016). Phage therapy: combating infections with potential for evolving from merely a treatment for complications to targeting diseases. Front. Microbiol. 7:1515. doi: 10.3389/fmicb.2016.01515 Gurung, M., Li, Z., You, H., Rodrigues, R., Jump, D. B., Morgun, A., et al. (2020). Role of gut microbiota in type 2 diabetes pathophysiology. EBioMedicine 51:102590. doi: 10.1016/j.ebiom.2019.11.051 He, S., Guo, F., Zou, Q., and Ding, H. (2020). MRMD2.0: a python tool for machine learning with feature ranking and reduction. Curr. Bioinform. 15, 1213–1221. doi: 10.2174/1574893615999200503030350 Jia, Y., Zhao, R., and Chen, L. (2020). Similarity-based machine learning model for predicting the metabolic pathways of compounds. IEEE Access 8, 130687–130696. doi: 10.1109/access.2020.3009439 Kahn, S. E., Hull, R. L., and Utzschneider, K. M. (2006). Mechanisms linking obesity to insulin resistance and type 2 diabetes. Nature 444, 840–846. doi: 10.1038/nature05482 Kibirige, D., and Mwebaze, R. (2013). Vitamin B12 deficiency among patients with diabetes mellitus: is routine screening and supplementation justified? J. Diabetes Metab. Disord. 12:17. Kodera, T., Smirnov, S. V., Samsonova, N. N., Kozlov, Y. I., Koyama, R., Hibi, M., et al. (2009). A novel L-isoleucine hydroxylating enzyme, L-isoleucine dioxygenase from Bacillus thuringiensis, produces (2S, 3R, 4S)-4-hydroxyisoleucine. Biochem. Biophys. Res. Commun. 390, 506–510. doi: 10.1016/j.bbrc.2009.09.126 Kohavi, R. (1995). “A study of cross-validation and bootstrap for accuracy estimation and model selection,” in Proceedings of the International joint Conference on Artificial Intelligence, (London: Lawrence Erlbaum Associates Ltd), 1137–1145. Lai, Y.-R., Chiu, W.-C., Huang, C.-C., Tsai, N.-W., Wang, H.-C., Lin, W.-C., et al. (2019). HbA1C variability is strongly associated with the severity of peripheral neuropathy in patients with type 2 diabetes. Front. Neurosci. 13:90. doi: 10.3389/fnins.2019.00090 Li, T., Xu, X., Xu, Y., Jin, P., Chen, J., Shi, Y., et al. (2019). PPARG polymorphisms are associated with unexplained mild vision loss in patients with type 2 diabetes mellitus. J. Ophthalmol. 2019:5284867. Liang, H., Chen, L., Zhao, X., and Zhang, X. (2020). Prediction of drug side effects with a refined negative sample selection strategy. Comput. Math. Methods Med. 2020:1573543. Liu, C., Feng, X., Li, Q., Wang, Y., Li, Q., and Hua, M. (2016). Adiponectin, TNF-α and inflammatory cytokines and risk of type 2 diabetes: a systematic review and meta-analysis. Cytokine 86, 100–109. doi: 10.1016/j.cyto.2016.06.028 Liu, H., Hu, B., Chen, L., and Lu, L. (2021). Identifying protein subcellular location with embedding features learned from networks. Curr. Proteom. [Epub ahead of print]. Liu, H. A., and Setiono, R. (1998). Incremental feature selection. Appl. Intellig. 9, 217–230. Ma, Y., You, X., Mai, G., Tokuyasu, T., and Liu, C. (2018). A human gut phage catalog correlates the gut phageome with type 2 diabetes. Microbiome 6:24. Maes, M., Kubera, M., Leunis, J. C., Berk, M., Geffard, M., and Bosmans, E. (2013). In depression, bacterial translocation may drive inflammatory responses, oxidative and nitrosative stress (O&NS), and autoimmune responses directed against O&NS-damaged neoepitopes. Acta Psychiatr. Scand. 127, 344–354. doi: 10.1111/j.1600-0447.2012.01908.x Mao, X., Cai, T., Olyarchuk, J. G., and Wei, L. (2005). Automated genome annotation and pathway identification using the KEGG Orthology (KO) as a controlled vocabulary. Bioinformatics 21, 3787–3793. doi: 10.1093/bioinformatics/bti430 Matthews, B. (1975). Comparison of the predicted and observed secondary structure of T4 phage lysozyme. Biochim. Biophys. Acta Protein Struct. 405, 442–451. doi: 10.1016/0005-2795(75)90109-9 Pan, X., Li, H., Zeng, T., Li, Z., Chen, L., Huang, T., et al. (2021). Identification of protein subcellular localization with network and functional embeddings. Front. Genet. 11:626500. doi: 10.3389/fgene.2020.626500 Peng, H., Long, F., and Ding, C. (2005). Feature selection based on mutual information: criteria of max-dependency, max-relevance, and min-redundancy. IEEE Trans. Pattern Anal. Mach. Intellig. 27, 1226–1238. doi: 10.1109/tpami.2005.159 J. Platt (ed.) (1998a). Fast Training of Support Vector Machines Using Sequential Minimal Optimization. Cambridge, MA: MIT Press. Platt, J. (1998b). Sequential Minimal Optimizaton: A Fast Algorithm for Training Support Vector Machines. Technical Report MSR-TR-98–14. Redmond: Microsoft Corporation. Powell, S., Forslund, K., Szklarczyk, D., Trachana, K., Roth, A., Huerta-Cepas, J., et al. (2014). eggNOG v4. 0: nested orthology inference across 3686 organisms. Nucleic Acids Res. 42, D231–D239. Sanahuja, J., Alonso, N., Diez, J., Ortega, E., Rubinat, E., Traveset, A., et al. (2016). Increased burden of cerebral small vessel disease in patients with type 2 diabetes and retinopathy. Diabetes Care 39, 1614–1620. doi: 10.2337/dc15-2671 Schlienger, J.-L. (2013). Type 2 diabetes complications. Presse Med. 42, 839–848. Suzuki, Y., Nishijima, S., Furuta, Y., Yoshimura, J., Suda, W., Oshima, K., et al. (2019). Long-read metagenomic exploration of extrachromosomal mobile genetic elements in the human gut. Microbiome 7:119. Tahir, M., and Idris, A. (2020). MD-LBP: an efficient computational model for protein subcellular localization from hela cell lines using SVM. Curr. Bioinform. 15, 204–211. doi: 10.2174/1574893614666190723120716 Tanaka, A., Shima, K., Fukuda, M., Tahara, Y., Yamamoto, Y., and Kumahara, Y. (1989). Tubular dysfunction in the early stage of diabetic nephropathy. Med. J. Osaka Univ. 38, 57–63. Teh, S.-H., You, R.-I., Yang, Y.-C., Hsu, C. Y., and Pang, C.-Y. (2020). A cohort study: the association between autoimmune disorders and leptospirosis. Sci. Rep. 10:3276. Tomaszewski, J. E., Brooks, J. S. J., Hicks, D., and Livolsi, V. A. (1992). Diabetic mastopathy: a distinctive clinicopathologic entity. Hum. Pathol. 23, 780–786. doi: 10.1016/0046-8177(92)90348-7 Vergès, B., Rouland, A., Baillot-Rudoni, S., Brindisi, M. C., Duvillard, L., Simoneau, I., et al. (2021). Increased body fat mass reduces the association between fructosamine and glycated hemoglobin in obese type 2 diabetes patients. J. Diabetes Investig. 12, 619–624. doi: 10.1111/jdi.13383 Wang, X., Xu, X., and Xia, Y. (2017). Further analysis reveals new gut microbiome markers of type 2 diabetes mellitus. Antonie Van Leeuwenhoek 110, 445–453. doi: 10.1007/s10482-016-0805-3 Witten, I. H., and Frank, E. (eds) (2005). Data Mining:Practical Machine Learning Tools and Techniques. San Francisco: Kaufmann. Wu, H., Esteve, E., Tremaroli, V., Khan, M. T., Caesar, R., Mannerås-Holm, L., et al. (2017). Metformin alters the gut microbiome of individuals with treatment-naive type 2 diabetes, contributing to the therapeutic effects of the drug. Nat. Med. 23:850. doi: 10.1038/nm.4345 Yan, A., Issar, T., Tummanapalli, S., Markoulli, M., Kwai, N., Poynten, A., et al. (2020). Relationship between corneal confocal microscopy and markers of peripheral nerve structure and function in Type 2 diabetes. Diabet. Med. 37, 326–334. doi: 10.1111/dme.13952 Zafar, M. I., and Gao, F. (2016). 4-hydroxyisoleucine: a potential new treatment for type 2 diabetes mellitus. BioDrugs 30, 255–262. doi: 10.1007/s40259-016-0177-2 Zhang, F., Wang, M., Yang, J., Xu, Q., Liang, C., Chen, B., et al. (2019). Response of gut microbiota in type 2 diabetes to hypoglycemic agents. Endocrine 66, 485–493. doi: 10.1007/s12020-019-02041-5 Zhang, S., Pan, X., Zeng, T., Guo, W., Gan, Z., Zhang, Y. H., et al. (2019). Copy number variation pattern for discriminating MACROD2 states of colorectal cancer subtypes. Front. Bioeng. Biotechnol. 7:407. doi: 10.3389/fbioe.2019.00407 Zhang, S., Zeng, T., Hu, B., Zhang, Y. H., Feng, K., Chen, L., et al. (2020). Discriminating origin tissues of tumor cell lines by methylation signatures and Dys-methylated rules. Front. Bioeng. Biotechnol. 8:507. doi: 10.3389/fbioe.2020.00507 Zhang, Y. H., Li, H., Zeng, T., Chen, L., Li, Z., Huang, T., et al. (2021a). Identifying transcriptomic signatures and rules for SARS-CoV-2 infection. Front. Cell Dev. Biol. 8:627302. doi: 10.3389/fcell.2020.627302 Zhang, Y.-H., Zeng, T., Chen, L., Huang, T., and Cai, Y.-D. (2021b). Detecting the multiomics signatures of factor-specific inflammatory effects on airway smooth muscles. Front. Genet. 11:599970. doi: 10.3389/fgene.2020.599970 Zhang, Y.-H., Zeng, T., Chen, L., Huang, T., and Cai, Y.-D. (2021c). Determining protein–protein functional associations by functional rules based on gene ontology and KEGG pathway. Biochim. Biophys. Acta Proteins Proteom. 1869:140621. doi: 10.1016/j.bbapap.2021.140621 Zhao, X., Chen, L., and Lu, J. (2018). A similarity-based method for prediction of drug side effects with heterogeneous information. Math. Biosci. 306, 136–144. doi: 10.1016/j.mbs.2018.09.010 Zheng, Y., Ley, S. H., and Hu, F. B. (2018). Global aetiology and epidemiology of type 2 diabetes mellitus and its complications. Nat. Rev. Endocrinol. 14:88. doi: 10.1038/nrendo.2017.151 Zhou, J.-P., Chen, L., Wang, T., and Liu, M. (2020). iATC-FRAKEL: a simple multi-label web-server for recognizing anatomical therapeutic chemical classes of drugs with their fingerprints only. Bioinformatics 36, 3568–3569. doi: 10.1093/bioinformatics/btaa166 Zhu, Y., Hu, B., Chen, L., and Dai, Q. (2021). iMPTCE-Hnetwork: a multi-label classifier for identifying metabolic pathway types of chemicals and enzymes with a heterogeneous network. Comput. Math. Methods Med. 2021:66 83051. Zou, Q., Qu, K., Luo, Y., Yin, D., Ju, Y., and Tang, H. (2018). Predicting diabetes mellitus with machine learning techniques. Front. Genet. 9:515. doi: 10.3389/fgene.2018.00515 |
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Zhang, Yu-HangGuo, WeiTao, ZengShiQi, ZhangChen, LeiGamarra, MargaritaMansour, Romany F.Escorcia-Gutierrez, JoseHuang, TaoYu Dong, Cai2021-08-19T15:34:31Z2021-08-19T15:34:31Z2021-07-091664-302Xhttps://hdl.handle.net/11323/855510.3389/fmicb.2021.711244Corporación Universidad de la CostaREDICUC - Repositorio CUChttps://repositorio.cuc.edu.co/Type 2 diabetes (T2D) is a systematic chronic metabolic condition with abnormal sugar metabolism dysfunction, and its complications are the most harmful to human beings and may be life-threatening after long-term durations. Considering the high incidence and severity at late stage, researchers have been focusing on the identification of specific biomarkers and potential drug targets for T2D at the genomic, epigenomic, and transcriptomic levels. Microbes participate in the pathogenesis of multiple metabolic diseases including diabetes. However, the related studies are still non-systematic and lack the functional exploration on identified microbes. To fill this gap between gut microbiome and diabetes study, we first introduced eggNOG database and KEGG ORTHOLOGY (KO) database for orthologous (protein/gene) annotation of microbiota. Two datasets with these annotations were employed, which were analyzed by multiple machine-learning models for identifying significant microbiota biomarkers of T2D. The powerful feature selection method, Max-Relevance and Min-Redundancy (mRMR), was first applied to the datasets, resulting in a feature list for each dataset. Then, the list was fed into the incremental feature selection (IFS), incorporating support vector machine (SVM) as the classification algorithm, to extract essential annotations and build efficient classifiers. This study not only revealed potential pathological factors for diabetes at the microbiome level but also provided us new candidates for drug development against diabetes.Zhang, Yu-Hang-will be generated-orcid-0000-0003-3825-0796-600Guo, Wei-will be generated-orcid-0000-0001-5445-7872-600Tao, ZengShiQi, ZhangChen, Lei-will be generated-orcid-0000-0001-6652-610X-600Gamarra, Margarita-will be generated-orcid-0000-0003-1834-2984-600Mansour, Romany F.-will be generated-orcid-0000-0002-4290-0387-600Escorcia-Gutierrez, Jose-will be generated-orcid-0000-0003-0518-3187-600Huang, Tao-will be generated-orcid-0000-0002-1888-7389-600Yu Dong, Caiapplication/pdfengCC0 1.0 Universalhttp://creativecommons.org/publicdomain/zero/1.0/info:eu-repo/semantics/openAccesshttp://purl.org/coar/access_right/c_abf2FRONTIERS IN MICROBIOLOGYhttps://www.frontiersin.org/articles/10.3389/fmicb.2021.711244/fulltype 2 diabetesgut microbiomemachine learningfeature selectionsupport vector machinemicrobiota biomarkersIdentification of microbiota biomarkers with orthologous gene annotation for type 2 diabetesArtículo de revistahttp://purl.org/coar/resource_type/c_6501http://purl.org/coar/resource_type/c_2df8fbb1Textinfo:eu-repo/semantics/articlehttp://purl.org/redcol/resource_type/ARTinfo:eu-repo/semantics/acceptedVersionArthur, R., Rohrmann, S., Møller, H., Selvin, E., Dobs, A. S., Kanarek, N., et al. (2017). Pre-diabetes and serum sex steroid hormones among US men. Andrology 5, 49–57. doi: 10.1111/andr.12287Bakris, G. L., Agarwal, R., Anker, S. D., Pitt, B., Ruilope, L. M., Rossing, P., et al. (2020). Effect of finerenone on chronic kidney disease outcomes in type 2 diabetes. N. Engl. J. Med. 383, 2219–2229. doi: 10.1056/nejmoa2025845Beli, E., Prabakaran, S., Krishnan, P., Evans-Molina, C., and Grant, M. B. (2019). Loss of diurnal oscillatory rhythms in gut microbiota correlates with changes in circulating metabolites in type 2 diabetic db/db mice. Nutrients 11:2310. doi: 10.3390/nu11102310Bullard, K. M., Cowie, C. C., Lessem, S. E., Saydah, S. H., Menke, A., Geiss, L. S., et al. (2018). Prevalence of diagnosed diabetes in adults by diabetes type—United States, 2016. Morb. Mortal. Wkly. Rep. 67:359. doi: 10.15585/mmwr.mm6712a2Carrillo-Larco, R. M., Altez-Fernandez, C., Acevedo-Rodriguez, J. G., Ortiz-Acha, K., and Ugarte-Gil, C. (2019). Leptospirosis as a risk factor for chronic kidney disease: a systematic review of observational studies. PLoS Neglect. Trop. Dis. 13:e0007458. doi: 10.1371/journal.pntd.0007458Chatterjee, S., Khunti, K., and Davies, M. J. (2017). Type 2 diabetes. Lancet 389, 2239–2251.Chen, L., Wang, S., Zhang, Y.-H., Li, J., Xing, Z.-H., Yang, J., et al. (2017). Identify key sequence features to improve CRISPR sgRNA efficacy. IEEE Access 5, 26582–26590. doi: 10.1109/access.2017.2775703Chen, L., Zeng, T., Pan, X., Zhang, Y. H., Huang, T., and Cai, Y. D. (2019). Identifying methylation pattern and genes associated with breast cancer subtypes. Int. J. Mol. Sci. 20:4269. doi: 10.3390/ijms20174269Cortes, C., and Vapnik, V. (1995). Support-vector networks. Mach. Learn. 20, 273–297.Deputy, N. P., Kim, S. Y., Conrey, E. J., and Bullard, K. M. (2018). Prevalence and changes in preexisting diabetes and gestational diabetes among women who had a live birth—United States, 2012–2016. Morb. Mortal. Wkly. Rep. 67:1201. doi: 10.15585/mmwr.mm6743a2Farnsworth, C. W., Shehatou, C. T., Maynard, R., Nishitani, K., Kates, S. L., Zuscik, M. J., et al. (2015). A humoral immune defect distinguishes the response to Staphylococcus aureus infections in mice with obesity and type 2 diabetes from that in mice with type 1 diabetes. Infect. Immun. 83, 2264–2274. doi: 10.1128/iai.03074-14Fischer, E., Günter, K., and Braun, V. (1989). Involvement of ExbB and TonB in transport across the outer membrane of Escherichia coli: phenotypic complementation of exb mutants by overexpressed tonB and physical stabilization of TonB by ExbB. J. Bacteriol. 171, 5127–5134. doi: 10.1128/jb.171.9.5127-5134.1989Forslund, K., Hildebrand, F., Nielsen, T., Falony, G., Le Chatelier, E., Sunagawa, S., et al. (2015). Disentangling type 2 diabetes and metformin treatment signatures in the human gut microbiota. Nature 528, 262–266.Gan, Y.-H. (2013). Host susceptibility factors to bacterial infections in type 2 diabetes. PLoS Pathog. 9:e1003794. doi: 10.1371/journal.ppat.1003794Gherasim, C., Lofgren, M., and Banerjee, R. (2013). Navigating the B12 road: assimilation, delivery, and disorders of cobalamin. J. Biol. Chem. 288, 13186–13193. doi: 10.1074/jbc.r113.458810Goldstein, B. J. (2002). Insulin resistance as the core defect in type 2 diabetes mellitus. Am. J. Cardiol. 90, 3–10. doi: 10.1016/s0002-9149(02)02553-5Górski, A., Międzybrodzki, R., Weber-Da̧browska, B., Fortuna, W., Letkiewicz, S., Rogóż, P., et al. (2016). Phage therapy: combating infections with potential for evolving from merely a treatment for complications to targeting diseases. Front. Microbiol. 7:1515. doi: 10.3389/fmicb.2016.01515Gurung, M., Li, Z., You, H., Rodrigues, R., Jump, D. B., Morgun, A., et al. (2020). Role of gut microbiota in type 2 diabetes pathophysiology. EBioMedicine 51:102590. doi: 10.1016/j.ebiom.2019.11.051He, S., Guo, F., Zou, Q., and Ding, H. (2020). MRMD2.0: a python tool for machine learning with feature ranking and reduction. Curr. Bioinform. 15, 1213–1221. doi: 10.2174/1574893615999200503030350Jia, Y., Zhao, R., and Chen, L. (2020). Similarity-based machine learning model for predicting the metabolic pathways of compounds. IEEE Access 8, 130687–130696. doi: 10.1109/access.2020.3009439Kahn, S. E., Hull, R. L., and Utzschneider, K. M. (2006). Mechanisms linking obesity to insulin resistance and type 2 diabetes. Nature 444, 840–846. doi: 10.1038/nature05482Kibirige, D., and Mwebaze, R. (2013). Vitamin B12 deficiency among patients with diabetes mellitus: is routine screening and supplementation justified? J. Diabetes Metab. Disord. 12:17.Kodera, T., Smirnov, S. V., Samsonova, N. N., Kozlov, Y. I., Koyama, R., Hibi, M., et al. (2009). A novel L-isoleucine hydroxylating enzyme, L-isoleucine dioxygenase from Bacillus thuringiensis, produces (2S, 3R, 4S)-4-hydroxyisoleucine. Biochem. Biophys. Res. Commun. 390, 506–510. doi: 10.1016/j.bbrc.2009.09.126Kohavi, R. (1995). “A study of cross-validation and bootstrap for accuracy estimation and model selection,” in Proceedings of the International joint Conference on Artificial Intelligence, (London: Lawrence Erlbaum Associates Ltd), 1137–1145.Lai, Y.-R., Chiu, W.-C., Huang, C.-C., Tsai, N.-W., Wang, H.-C., Lin, W.-C., et al. (2019). HbA1C variability is strongly associated with the severity of peripheral neuropathy in patients with type 2 diabetes. Front. Neurosci. 13:90. doi: 10.3389/fnins.2019.00090Li, T., Xu, X., Xu, Y., Jin, P., Chen, J., Shi, Y., et al. (2019). PPARG polymorphisms are associated with unexplained mild vision loss in patients with type 2 diabetes mellitus. J. Ophthalmol. 2019:5284867.Liang, H., Chen, L., Zhao, X., and Zhang, X. (2020). Prediction of drug side effects with a refined negative sample selection strategy. Comput. Math. Methods Med. 2020:1573543.Liu, C., Feng, X., Li, Q., Wang, Y., Li, Q., and Hua, M. (2016). Adiponectin, TNF-α and inflammatory cytokines and risk of type 2 diabetes: a systematic review and meta-analysis. Cytokine 86, 100–109. doi: 10.1016/j.cyto.2016.06.028Liu, H., Hu, B., Chen, L., and Lu, L. (2021). Identifying protein subcellular location with embedding features learned from networks. Curr. Proteom. [Epub ahead of print].Liu, H. A., and Setiono, R. (1998). Incremental feature selection. Appl. Intellig. 9, 217–230.Ma, Y., You, X., Mai, G., Tokuyasu, T., and Liu, C. (2018). A human gut phage catalog correlates the gut phageome with type 2 diabetes. Microbiome 6:24.Maes, M., Kubera, M., Leunis, J. C., Berk, M., Geffard, M., and Bosmans, E. (2013). In depression, bacterial translocation may drive inflammatory responses, oxidative and nitrosative stress (O&NS), and autoimmune responses directed against O&NS-damaged neoepitopes. Acta Psychiatr. Scand. 127, 344–354. doi: 10.1111/j.1600-0447.2012.01908.xMao, X., Cai, T., Olyarchuk, J. G., and Wei, L. (2005). Automated genome annotation and pathway identification using the KEGG Orthology (KO) as a controlled vocabulary. Bioinformatics 21, 3787–3793. doi: 10.1093/bioinformatics/bti430Matthews, B. (1975). Comparison of the predicted and observed secondary structure of T4 phage lysozyme. Biochim. Biophys. Acta Protein Struct. 405, 442–451. doi: 10.1016/0005-2795(75)90109-9Pan, X., Li, H., Zeng, T., Li, Z., Chen, L., Huang, T., et al. (2021). Identification of protein subcellular localization with network and functional embeddings. Front. Genet. 11:626500. doi: 10.3389/fgene.2020.626500Peng, H., Long, F., and Ding, C. (2005). Feature selection based on mutual information: criteria of max-dependency, max-relevance, and min-redundancy. IEEE Trans. Pattern Anal. Mach. Intellig. 27, 1226–1238. doi: 10.1109/tpami.2005.159J. Platt (ed.) (1998a). Fast Training of Support Vector Machines Using Sequential Minimal Optimization. Cambridge, MA: MIT Press.Platt, J. (1998b). Sequential Minimal Optimizaton: A Fast Algorithm for Training Support Vector Machines. Technical Report MSR-TR-98–14. Redmond: Microsoft Corporation.Powell, S., Forslund, K., Szklarczyk, D., Trachana, K., Roth, A., Huerta-Cepas, J., et al. (2014). eggNOG v4. 0: nested orthology inference across 3686 organisms. Nucleic Acids Res. 42, D231–D239.Sanahuja, J., Alonso, N., Diez, J., Ortega, E., Rubinat, E., Traveset, A., et al. (2016). Increased burden of cerebral small vessel disease in patients with type 2 diabetes and retinopathy. Diabetes Care 39, 1614–1620. doi: 10.2337/dc15-2671Schlienger, J.-L. (2013). Type 2 diabetes complications. Presse Med. 42, 839–848.Suzuki, Y., Nishijima, S., Furuta, Y., Yoshimura, J., Suda, W., Oshima, K., et al. (2019). Long-read metagenomic exploration of extrachromosomal mobile genetic elements in the human gut. Microbiome 7:119.Tahir, M., and Idris, A. (2020). MD-LBP: an efficient computational model for protein subcellular localization from hela cell lines using SVM. Curr. Bioinform. 15, 204–211. doi: 10.2174/1574893614666190723120716Tanaka, A., Shima, K., Fukuda, M., Tahara, Y., Yamamoto, Y., and Kumahara, Y. (1989). Tubular dysfunction in the early stage of diabetic nephropathy. Med. J. Osaka Univ. 38, 57–63.Teh, S.-H., You, R.-I., Yang, Y.-C., Hsu, C. Y., and Pang, C.-Y. (2020). A cohort study: the association between autoimmune disorders and leptospirosis. Sci. Rep. 10:3276.Tomaszewski, J. E., Brooks, J. S. J., Hicks, D., and Livolsi, V. A. (1992). Diabetic mastopathy: a distinctive clinicopathologic entity. Hum. Pathol. 23, 780–786. doi: 10.1016/0046-8177(92)90348-7Vergès, B., Rouland, A., Baillot-Rudoni, S., Brindisi, M. C., Duvillard, L., Simoneau, I., et al. (2021). Increased body fat mass reduces the association between fructosamine and glycated hemoglobin in obese type 2 diabetes patients. J. Diabetes Investig. 12, 619–624. doi: 10.1111/jdi.13383Wang, X., Xu, X., and Xia, Y. (2017). Further analysis reveals new gut microbiome markers of type 2 diabetes mellitus. Antonie Van Leeuwenhoek 110, 445–453. doi: 10.1007/s10482-016-0805-3Witten, I. H., and Frank, E. (eds) (2005). Data Mining:Practical Machine Learning Tools and Techniques. San Francisco: Kaufmann.Wu, H., Esteve, E., Tremaroli, V., Khan, M. T., Caesar, R., Mannerås-Holm, L., et al. (2017). Metformin alters the gut microbiome of individuals with treatment-naive type 2 diabetes, contributing to the therapeutic effects of the drug. Nat. Med. 23:850. doi: 10.1038/nm.4345Yan, A., Issar, T., Tummanapalli, S., Markoulli, M., Kwai, N., Poynten, A., et al. (2020). Relationship between corneal confocal microscopy and markers of peripheral nerve structure and function in Type 2 diabetes. Diabet. Med. 37, 326–334. doi: 10.1111/dme.13952Zafar, M. I., and Gao, F. (2016). 4-hydroxyisoleucine: a potential new treatment for type 2 diabetes mellitus. BioDrugs 30, 255–262. doi: 10.1007/s40259-016-0177-2Zhang, F., Wang, M., Yang, J., Xu, Q., Liang, C., Chen, B., et al. (2019). Response of gut microbiota in type 2 diabetes to hypoglycemic agents. Endocrine 66, 485–493. doi: 10.1007/s12020-019-02041-5Zhang, S., Pan, X., Zeng, T., Guo, W., Gan, Z., Zhang, Y. H., et al. (2019). Copy number variation pattern for discriminating MACROD2 states of colorectal cancer subtypes. Front. Bioeng. Biotechnol. 7:407. doi: 10.3389/fbioe.2019.00407Zhang, S., Zeng, T., Hu, B., Zhang, Y. H., Feng, K., Chen, L., et al. (2020). Discriminating origin tissues of tumor cell lines by methylation signatures and Dys-methylated rules. Front. Bioeng. Biotechnol. 8:507. doi: 10.3389/fbioe.2020.00507Zhang, Y. H., Li, H., Zeng, T., Chen, L., Li, Z., Huang, T., et al. (2021a). Identifying transcriptomic signatures and rules for SARS-CoV-2 infection. Front. Cell Dev. Biol. 8:627302. doi: 10.3389/fcell.2020.627302Zhang, Y.-H., Zeng, T., Chen, L., Huang, T., and Cai, Y.-D. (2021b). Detecting the multiomics signatures of factor-specific inflammatory effects on airway smooth muscles. Front. Genet. 11:599970. doi: 10.3389/fgene.2020.599970Zhang, Y.-H., Zeng, T., Chen, L., Huang, T., and Cai, Y.-D. (2021c). Determining protein–protein functional associations by functional rules based on gene ontology and KEGG pathway. Biochim. Biophys. Acta Proteins Proteom. 1869:140621. doi: 10.1016/j.bbapap.2021.140621Zhao, X., Chen, L., and Lu, J. (2018). A similarity-based method for prediction of drug side effects with heterogeneous information. Math. Biosci. 306, 136–144. doi: 10.1016/j.mbs.2018.09.010Zheng, Y., Ley, S. H., and Hu, F. B. (2018). Global aetiology and epidemiology of type 2 diabetes mellitus and its complications. Nat. Rev. Endocrinol. 14:88. doi: 10.1038/nrendo.2017.151Zhou, J.-P., Chen, L., Wang, T., and Liu, M. (2020). iATC-FRAKEL: a simple multi-label web-server for recognizing anatomical therapeutic chemical classes of drugs with their fingerprints only. Bioinformatics 36, 3568–3569. doi: 10.1093/bioinformatics/btaa166Zhu, Y., Hu, B., Chen, L., and Dai, Q. (2021). iMPTCE-Hnetwork: a multi-label classifier for identifying metabolic pathway types of chemicals and enzymes with a heterogeneous network. Comput. Math. Methods Med. 2021:66 83051.Zou, Q., Qu, K., Luo, Y., Yin, D., Ju, Y., and Tang, H. (2018). Predicting diabetes mellitus with machine learning techniques. Front. Genet. 9:515. doi: 10.3389/fgene.2018.00515PublicationORIGINALIdentification of Microbiota Biomarkers With Orthologous Gene Annotation for Type 2 Diabetes.pdfIdentification of Microbiota Biomarkers With Orthologous Gene Annotation for Type 2 Diabetes.pdfapplication/pdf6001426https://repositorio.cuc.edu.co/bitstreams/b34954bf-07ef-4125-b834-dd21caabeb67/download7ad9975e04318c91386617c52cfa3bf5MD51ORIGINALIdentification of Microbiota Biomarkers With Orthologous Gene Annotation for Type 2 Diabetes.pdfIdentification of Microbiota Biomarkers With Orthologous Gene Annotation for Type 2 Diabetes.pdfapplication/pdf6001426https://repositorio.cuc.edu.co/bitstreams/ad68b673-5619-4a46-a1c6-8a5673c3ef85/download7ad9975e04318c91386617c52cfa3bf5MD51CC-LICENSElicense_rdflicense_rdfapplication/rdf+xml; charset=utf-8701https://repositorio.cuc.edu.co/bitstreams/8d50e287-43cf-4957-b70b-cdfd5a7a4f0e/download42fd4ad1e89814f5e4a476b409eb708cMD52LICENSElicense.txtlicense.txttext/plain; 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Universalopen.accesshttps://repositorio.cuc.edu.coRepositorio de la Universidad de la Costa 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