{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T01:26:38Z","timestamp":1760145998009,"version":"build-2065373602"},"reference-count":57,"publisher":"MDPI AG","issue":"10","license":[{"start":{"date-parts":[[2024,9,25]],"date-time":"2024-09-25T00:00:00Z","timestamp":1727222400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"DAAD \u201cInternationale Forschungsaufenthalte f\u00fcr Informatikerinnen und Informatiker (IFI)\u201d"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>Seasonal influenza viruses continuously evolve via antigenic drift. This leads to recurring epidemics, globally significant mortality rates, and the need for annually updated vaccines. Co-occurring mutations in hemagglutinin (HA) and neuraminidase (NA) are suggested to have synergistic interactions where mutations can increase the chances of immune escape and viral fitness. Association rule mining was used to identify temporal relationships of co-occurring HA\u2013NA mutations of influenza virus A\/H3N2 and its role in antigenic evolution. A total of 64 clusters were found. These included well-known mutations responsible for antigenic drift, as well as previously undiscovered groups. A majority (41\/64) were associated with known antigenic sites, and 38\/64 involved mutations across both HA and NA. The emergence and disappearance of N-glycosylation sites in the pattern of N-X-[S\/T] were also identified, which are crucial post-translational processes to maintain protein stability and functional balance (e.g., emergence of NA:339ASP and disappearance of HA:187ASP). Our study offers an alternative approach to the existing mutual-information and phylogenetic methods used to identify co-occurring mutations, enabling faster processing of large amounts of data. Our approach can facilitate the prediction of critical mutations given their occurrence in a previous season, facilitating vaccine development for the next flu season and leading to better preparation for future pandemics.<\/jats:p>","DOI":"10.3390\/v16101515","type":"journal-article","created":{"date-parts":[[2024,9,25]],"date-time":"2024-09-25T04:01:24Z","timestamp":1727236884000},"page":"1515","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Evolutionary Insights from Association Rule Mining of Co-Occurring Mutations in Influenza Hemagglutinin and Neuraminidase"],"prefix":"10.3390","volume":"16","author":[{"given":"Valentina","family":"Galeone","sequence":"first","affiliation":[{"name":"Institute of Computer Science, Freie Universit\u00e4t Berlin, 14195 Berlin, Germany"},{"name":"Australian e-Health Research Centre, Commonwealth Scientific and Industrial Research Organisation, Sydney, NSW 2145, Australia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1865-1541","authenticated-orcid":false,"given":"Carol","family":"Lee","sequence":"additional","affiliation":[{"name":"Australian e-Health Research Centre, Commonwealth Scientific and Industrial Research Organisation, Sydney, NSW 2145, Australia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Michael T.","family":"Monaghan","sequence":"additional","affiliation":[{"name":"Institute of Biology, Freie Universit\u00e4t Berlin, 14195 Berlin, Germany"},{"name":"Leibniz Institute of Freshwater Ecology and Inland Fisheries (IGB), 12587 Berlin, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Denis C.","family":"Bauer","sequence":"additional","affiliation":[{"name":"Australian e-Health Research Centre, Commonwealth Scientific and Industrial Research Organisation, Sydney, NSW 2145, Australia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5688-5758","authenticated-orcid":false,"given":"Laurence O. W.","family":"Wilson","sequence":"additional","affiliation":[{"name":"Australian e-Health Research Centre, Commonwealth Scientific and Industrial Research Organisation, Sydney, NSW 2145, Australia"},{"name":"Department of Biomedical Sciences, Macquarie University, Sydney, NSW 2109, Australia"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2024,9,25]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1285","DOI":"10.1016\/S0140-6736(17)33293-2","article-title":"Estimates of global seasonal influenza-associated respiratory mortality: A modelling study","volume":"391","author":"Iuliano","year":"2018","journal-title":"Lancet"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"831","DOI":"10.1007\/s00705-018-3708-y","article-title":"The emerging influenza virus threat: Status and new prospects for its therapy and control","volume":"163","author":"Kumar","year":"2018","journal-title":"Arch. Virol."},{"key":"ref_3","doi-asserted-by":"crossref","unstructured":"Shao, W., Li, X., Goraya, M.U., Wang, S., and Chen, J.-L. (2017). Evolution of Influenza A Virus by Mutation and Re-Assortment. Int. J. Mol. Sci., 18.","DOI":"10.3390\/ijms18081650"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"47","DOI":"10.1038\/nrmicro.2017.118","article-title":"The evolution of seasonal influenza viruses","volume":"16","author":"Petrova","year":"2018","journal-title":"Nat. Rev. Microbiol."},{"key":"ref_5","doi-asserted-by":"crossref","unstructured":"Kosik, I., and Yewdell, J.W. (2019). Influenza Hemagglutinin and Neuraminidase: Yin\u2013Yang Proteins Coevolving to Thwart Immunity. Viruses, 11.","DOI":"10.3390\/v11040346"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"26","DOI":"10.32607\/20758251-2009-1-2-26-32","article-title":"Influenza virus neuraminidase: Structure and function","volume":"1","author":"Shtyrya","year":"2009","journal-title":"Acta Naturae"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1385","DOI":"10.3201\/eid1008.040107","article-title":"Predicting Antigenic Variants of Influenza A\/H3N2 Viruses","volume":"10","author":"Lee","year":"2004","journal-title":"Emerg. Infect. Dis."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"1153","DOI":"10.1016\/j.bbamem.2013.10.004","article-title":"Receptor binding and pH stability\u2014How influenza A virus hemagglutinin affects host-specific virus infection","volume":"1838","author":"Mair","year":"2014","journal-title":"Biochim. Biophys. Acta."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"825","DOI":"10.1126\/science.1135165","article-title":"Immunization by avian H5 influenza hemagglutinin mutants with altered receptor binding specificity","volume":"317","author":"Yang","year":"2007","journal-title":"Science"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"149","DOI":"10.1186\/s12985-019-1258-x","article-title":"Influenza A virus hemagglutinin mutations associated with use of neuraminidase inhibitors correlate with decreased inhibition by anti-influenza antibodies","volume":"16","author":"lyushina","year":"2019","journal-title":"Virol. J."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"e01439-21","DOI":"10.1128\/spectrum.01439-21","article-title":"Identification of Hemagglutinin Mutations Caused by Neuraminidase Antibody Pressure","volume":"9","author":"Wang","year":"2021","journal-title":"Microbiol. Spectr."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"6283","DOI":"10.1073\/pnas.0701396104","article-title":"Simultaneous amino acid substitutions at antigenic sites drive influenza A hemagglutinin evolution","volume":"104","author":"Shih","year":"2007","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"vead037","DOI":"10.1093\/ve\/vead037","article-title":"Mutual information networks reveal evolutionary relationships within the influenza A virus polymerase","volume":"9","author":"Arcos","year":"2023","journal-title":"Virus Evol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"20","DOI":"10.1016\/j.compbiolchem.2012.06.004","article-title":"A novel empirical mutual information approach to identify co-evolving amino acid positions of influenza A viruses","volume":"39","author":"Gong","year":"2012","journal-title":"Comput. Biol. Chem."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"2309","DOI":"10.1093\/bioinformatics\/btp423","article-title":"Using a mutual information-based site transition network to map the genetic evolution of influenza A\/H3N2 virus","volume":"25","author":"Xia","year":"2009","journal-title":"Bioinformatics"},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Chen, H., Zhou, X., Zheng, J., and Kwoh, C.-K. (2016). Rules of co-occurring mutations characterize the antigenic evolution of human influenza A\/H3N2, A\/H1N1 and B viruses. BMC Med. Genom., 9.","DOI":"10.1186\/s12920-016-0230-5"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"78","DOI":"10.1016\/j.procs.2016.05.180","article-title":"Market Basket Analysis: Identify the Changing Trends of Market Data Using Association Rule Mining","volume":"85","author":"Kaur","year":"2016","journal-title":"Procedia Comput. Sci."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"181","DOI":"10.1016\/j.jbi.2015.07.018","article-title":"Knowledge discovery and sequence-based prediction of pandemic influenza using an integrated classification and association rule mining (CBA) algorithm","volume":"57","author":"Kargarfard","year":"2015","journal-title":"J. Biomed. Inform."},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Kargarfard, F., Sami, A., Mohammadi-Dehcheshmeh, M., and Ebrahimie, E. (2016). Novel approach for identification of influenza virus host range and zoonotic transmissible sequences by determination of host-related associative positions in viral genome segments. BMC Genom., 17.","DOI":"10.1186\/s12864-016-3250-9"},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Tandan, M., Acharya, Y., Pokharel, S., and Timilsina, M. (2021). Discovering symptom patterns of COVID-19 patients using association rule mining. Comput. Biol. Med., 131.","DOI":"10.1016\/j.compbiomed.2021.104249"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"109","DOI":"10.1016\/j.mib.2015.06.015","article-title":"Viral evolution: Beyond drift and shift","volume":"26","author":"Greenbaum","year":"2015","journal-title":"Curr. Opin. Microbiol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"73","DOI":"10.2174\/1574893611666161123142425","article-title":"A Weighted Association Rule Mining Method for Predicting HCV-Human Protein Interactions","volume":"13","author":"Indhumathy","year":"2018","journal-title":"Curr. Bioinform."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"428","DOI":"10.1109\/TCBB.2009.6","article-title":"Data mining on DNA sequences of hepatitis B virus","volume":"8","author":"Leung","year":"2011","journal-title":"IEEE\/ACM Trans. Comput. Biol. Bioinform."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"106411","DOI":"10.1016\/j.compag.2021.106411","article-title":"Multidrug resistance analysis method for pathogens of cow mastitis based on weighted-association rule mining and similarity comparison","volume":"190","author":"Liang","year":"2021","journal-title":"Comput. Electron. Agric."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"100595","DOI":"10.1016\/j.imu.2021.100595","article-title":"Identification of cancer related genes using feature selection and association rule mining","volume":"24","author":"Gakii","year":"2021","journal-title":"Inform. Med. Unlocked."},{"key":"ref_26","first-page":"1","article-title":"Mining frequent patterns without candidate generation","volume":"29","author":"Han","year":"2000","journal-title":"Data Min. Knowl. Discov."},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Zaki, M.J., and Gouda, K. (2003, January 24\u201327). Fast vertical mining using diffsets. Proceedings of the Ninth ACM SIGKDD International Conference on Knowledge Discovery and Data Mining, New York, NY, USA.","DOI":"10.1145\/956750.956788"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"1178122X1878832","DOI":"10.1177\/1178122X18788328","article-title":"Comparative Co-Evolution Analysis Between the HA and NA Genes of Influenza A Virus","volume":"9","author":"Jang","year":"2018","journal-title":"Virology"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"veab090","DOI":"10.1093\/ve\/veab090","article-title":"Spatial and temporal coevolution of N2 neuraminidase and H1 and H3 hemagglutinin genes of influenza A virus in US swine","volume":"7","author":"Zeller","year":"2021","journal-title":"Virus Evol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"D678","DOI":"10.1093\/nar\/gkac1003","article-title":"Introducing the Bacterial and Viral Bioinformatics Resource Center (BV-BRC): A resource combining PATRIC, IRD and ViPR","volume":"51","author":"Olson","year":"2023","journal-title":"Nucleic Acids Res."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"W276","DOI":"10.1093\/nar\/gkac240","article-title":"Search and sequence analysis tools services from EMBL-EBI in 2022","volume":"50","author":"Madeira","year":"2022","journal-title":"Nucleic Acids Res."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"403","DOI":"10.1016\/S0022-2836(05)80360-2","article-title":"Basic local alignment search tool","volume":"215","author":"Altschul","year":"1990","journal-title":"J. Mol. Biol."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"638","DOI":"10.21105\/joss.00638","article-title":"MLxtend: Providing machine learning and data science utilities and extensions to Python\u2019s scientific computing stack","volume":"3","author":"Raschka","year":"2018","journal-title":"J. Open Source Softw."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"487","DOI":"10.1016\/0022-2836(91)80069-7","article-title":"Refined atomic structures of N9 subtype influenza virus neuraminidase and escape mutants","volume":"221","author":"Tulip","year":"1991","journal-title":"J. Mol. Biol."},{"key":"ref_35","unstructured":"Zhang, T. (2000, January 18\u201320). Association Rules. Proceedings of the 4th Pacific-Asia Conference on Knowledge Discovery and Data Mining, Current Issues and New Applications (PADKK \u201900), Kyoto, Japan."},{"key":"ref_36","doi-asserted-by":"crossref","unstructured":"Hagberg, A.A., Schult, D.A., and Swart, P.J. (2008, January 19\u201324). Exploring network structure, dynamics, and function using NetworkX. Proceedings of the 7th Python in Science Conference SciPy2008, Pasadena, CA, USA.","DOI":"10.25080\/TCWV9851"},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Perrone, G., Unpingco, J., and Lu, H. (2020). Network visualizations with Pyvis and VisJS. arXiv.","DOI":"10.25080\/Majora-342d178e-008"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"268","DOI":"10.1093\/molbev\/msu300","article-title":"IQ-TREE: A fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies","volume":"32","author":"Nguyen","year":"2015","journal-title":"Mol. Biol. Evol."},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Dang, C.C., Le, Q.S., Gascuel, O., and Le, V.S. (2010). FLU, an amino acid substitution model for influenza proteins. BMC Ecol. Evol., 10.","DOI":"10.1186\/1471-2148-10-99"},{"key":"ref_40","first-page":"976","article-title":"Substitutions Near the Receptor Binding Site Determine Major Antigenic Change During Influenza Virus Evolution","volume":"342","author":"Koel","year":"2013","journal-title":"IScience"},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"Tria, F., Pompei, S., and Loreto, V. (2013). Dynamically correlated mutations drive human Influenza A evolution. Sci. Rep., 3.","DOI":"10.1038\/srep02705"},{"key":"ref_42","doi-asserted-by":"crossref","unstructured":"Dou, D., Revol, R., \u00d6stbye, H., Wang, H., and Daniels, R. (2018). Influenza A Virus Cell Entry, Replication, Virion Assembly and Movement. Front. immunol., 9.","DOI":"10.3389\/fimmu.2018.01581"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"1363","DOI":"10.1016\/j.chom.2022.09.003","article-title":"The evolutionary potential of influenza A virus hemagglutinin is highly constrained by epistatic interactions with neuraminidase","volume":"30","author":"Liu","year":"2022","journal-title":"Cell Host Microbe"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"795","DOI":"10.1586\/14787210.4.5.795","article-title":"Resistance to anti-influenza drugs: Adamantanes and neuraminidase inhibitors","volume":"4","author":"Hurt","year":"2006","journal-title":"Expert Rev. Anti-infect. Ther."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"1175","DOI":"10.1016\/S0140-6736(05)67338-2","article-title":"Incidence of adamantane resistance among influenza A (H3N2) viruses isolated worldwide from 1994 to 2005: A cause for concern","volume":"366","author":"Bright","year":"2005","journal-title":"Lancet"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"523","DOI":"10.1086\/318537","article-title":"Selection of Influenza Virus Mutants in Experimentally Infected Volunteers Treated with Oseltamivir","volume":"183","author":"Gubareva","year":"2005","journal-title":"J. Infect. Dis."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"270","DOI":"10.1016\/j.virol.2009.03.026","article-title":"The Origin and Global Emergence of Adamantane Resistant A\/H3N2 Influenza Viruses","volume":"388","author":"Nelson","year":"2009","journal-title":"Virology"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"174","DOI":"10.1089\/vim.2017.0141","article-title":"Influenza Virus: Dealing with a Drifting and Shifting Pathogen","volume":"31","author":"Kim","year":"2018","journal-title":"Viral Immunol."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"2216","DOI":"10.1038\/s41564-019-0522-6","article-title":"The neuraminidase of A(H3N2) influenza viruses circulating since 2016 is antigenically distinct from the A\/Hong Kong\/4801\/2014 vaccine strain","volume":"4","author":"Wan","year":"2019","journal-title":"Nat. Microbiol."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"2348","DOI":"10.1074\/mcp.R119.001491","article-title":"Why Glycosylation Matters in Building a Better Flu Vaccine","volume":"18","author":"Chang","year":"2019","journal-title":"Mol. Cell. Proteom."},{"key":"ref_51","doi-asserted-by":"crossref","unstructured":"Dudin, G.A., Aziz, I.M., Alzayed, R.M., Ahmed, A., Hussain, T., Somily, A.M., Alsaadi, M.M., and Almajhdi, F.N. (2023). Genetic Diversity and Evolutionary Kinetics of Influenza A Virus H3N2 Subtypes Circulating in Riyadh, Saudi Arabia. Vaccines, 11.","DOI":"10.3390\/vaccines11030702"},{"key":"ref_52","doi-asserted-by":"crossref","unstructured":"Phyu, W.W., Saito, R., Kyaw, Y., Lin, N., Win, S.M.K., Win, N.C., Ja, L.D., Htwe, K.T.Z., Aung, T.Z., and Tin, H.H. (2023). Evolutionary Dynamics of Whole-Genome Influenza A\/H3N2 Viruses Isolated in Myanmar from 2015 to 2019. Viruses, 14.","DOI":"10.3390\/v14112414"},{"key":"ref_53","doi-asserted-by":"crossref","unstructured":"Boonnak, K., Mansanguan, C., Schuerch, D., Boonyuen, U., Lerdsamran, H., Jiamsomboon, K., Sae Wang, F., Huntrup, A., Prasertsopon, J., and Kosoltanapiwat, N. (2021). Molecular Characterization of Seasonal Influenza A and B from Hospitalized Patients in Thailand in 2018\u20132019. Viruses, 13.","DOI":"10.3390\/v13060977"},{"key":"ref_54","doi-asserted-by":"crossref","unstructured":"Ge, J., Lin, X., Guo, J., Liu, L., Li, Z., Lan, Y., Liu, L., Guo, J., Lu, J., and Huang, W. (2022). The Antibody Response Against Neuraminidase in Human Influenza A (H3N2) Virus Infections During 2018\/2019 Flu Season: Focusing on the Epitopes of 329-N-Glycosylation and E344 in N2. Front. Microbiol., 13.","DOI":"10.3389\/fmicb.2022.845088"},{"key":"ref_55","doi-asserted-by":"crossref","unstructured":"Simon, B., Pichon, M., Valette, M., Burfin, G., Richard, M., Lina, B., and Josset, L. (2019). Whole Genome Sequencing of A(H3N2) Influenza Viruses Reveals Variants Associated with Severity during the 2016\u20132017 Season. Viruses, 11.","DOI":"10.3390\/v11020108"},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"6015","DOI":"10.1128\/JVI.74.13.6015-6020.2000","article-title":"Balanced hemagglutinin and neuraminidase activities are critical for efficient replication of influenza A virus","volume":"74","author":"Mitnaul","year":"2000","journal-title":"J. Virol."},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"583","DOI":"10.1038\/s41586-021-03819-2","article-title":"Highly accurate protein structure prediction with AlphaFold","volume":"596","author":"Jumper","year":"2021","journal-title":"Nature"}],"container-title":["Viruses"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1999-4915\/16\/10\/1515\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T16:02:20Z","timestamp":1760112140000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1999-4915\/16\/10\/1515"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,9,25]]},"references-count":57,"journal-issue":{"issue":"10","published-online":{"date-parts":[[2024,10]]}},"alternative-id":["v16101515"],"URL":"https:\/\/doi.org\/10.3390\/v16101515","relation":{},"ISSN":["1999-4915"],"issn-type":[{"type":"electronic","value":"1999-4915"}],"subject":[],"published":{"date-parts":[[2024,9,25]]}}}