{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,25]],"date-time":"2026-09-25T09:59:20Z","timestamp":1790330360556,"version":"4.1.0"},"reference-count":73,"publisher":"Springer Science and Business Media LLC","license":[{"start":{"date-parts":[[2026,9,25]],"date-time":"2026-09-25T00:00:00Z","timestamp":1790294400000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2026,9,25]],"date-time":"2026-09-25T00:00:00Z","timestamp":1790294400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"funder":[{"DOI":"10.13039\/501100001711","name":"Schweizerischer Nationalfonds zur F\u00f6rderung der Wissenschaftlichen Forschung","doi-asserted-by":"publisher","award":["Ambizione Fellowship PZ00P3_180085"],"award-info":[{"award-number":["Ambizione Fellowship PZ00P3_180085"]}],"id":[{"id":"10.13039\/501100001711","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001711","name":"Schweizerischer Nationalfonds zur F\u00f6rderung der Wissenschaftlichen Forschung","doi-asserted-by":"publisher","award":["Starting Grant TMSGI3_211369"],"award-info":[{"award-number":["Starting Grant TMSGI3_211369"]}],"id":[{"id":"10.13039\/501100001711","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001711","name":"Schweizerischer Nationalfonds zur F\u00f6rderung der Wissenschaftlichen Forschung","doi-asserted-by":"publisher","award":["NCCR AntiResist (grant number 180541)"],"award-info":[{"award-number":["NCCR AntiResist (grant number 180541)"]}],"id":[{"id":"10.13039\/501100001711","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001711","name":"Schweizerischer Nationalfonds zur F\u00f6rderung der Wissenschaftlichen Forschung","doi-asserted-by":"publisher","award":["NCCR AntiResist (grant number 180541)"],"award-info":[{"award-number":["NCCR AntiResist (grant number 180541)"]}],"id":[{"id":"10.13039\/501100001711","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001711","name":"Schweizerischer Nationalfonds zur F\u00f6rderung der Wissenschaftlichen Forschung","doi-asserted-by":"publisher","award":["NCCR AntiResist (grant number 180541)"],"award-info":[{"award-number":["NCCR AntiResist (grant number 180541)"]}],"id":[{"id":"10.13039\/501100001711","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Nat Microbiol"],"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>The arms race of bacteriophages and their bacterial hosts has inspired major breakthroughs in biotechnology and shaped phages as fierce predators with great clinical potential to fight multidrug-resistant bacterial pathogens. However, the large amount of genes of unknown function in phage genomes remains a major obstacle for the molecular understanding of phage\u2013host interactions. Here we present HIDEN-SEQ (hidden Acr-enabled transposon-insertion sequencing), a transposon-insertion sequencing method for phages that systematically links viral genes to selectable phenotypes. Using model phage T4, we show that HIDEN-SEQ readily reproduces the gene essentiality map established over decades of research. Our method is easily portable across diverse non-model phages and reveals conditionally essential genes in multiple bacterial hosts and growth conditions, including previously unknown antidefence factors that we matched to specific antiviral defences. We anticipate that HIDEN-SEQ will be leveraged to reveal functions of viral genes with direct relevance for microbial ecology, biotechnology and phage therapy.<\/jats:p>","DOI":"10.1038\/s41564-026-02455-8","type":"journal-article","created":{"date-parts":[[2026,9,25]],"date-time":"2026-09-25T09:02:24Z","timestamp":1790326944000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Systematic mapping of bacteriophage gene essentiality with HIDEN-SEQ"],"prefix":"10.1038","author":[{"ORCID":"https:\/\/orcid.org\/0009-0002-1819-6155","authenticated-orcid":false,"given":"Dorentina","family":"Humolli","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0006-3760-8142","authenticated-orcid":false,"given":"Damien","family":"Piel","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0003-2698-3439","authenticated-orcid":false,"given":"Jessica","family":"Ransome","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kathrin","family":"Bausch","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4875-5368","authenticated-orcid":false,"given":"Sarah","family":"Tschudin-Sutter","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0009-3256-3987","authenticated-orcid":false,"given":"Monica","family":"Ortelli","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7288-1052","authenticated-orcid":false,"given":"Christoph","family":"Dehio","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3162-6634","authenticated-orcid":false,"given":"Jan-Willem","family":"Veening","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2106-1286","authenticated-orcid":false,"given":"Alexander","family":"Harms","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2026,9,25]]},"reference":[{"key":"2455_CR1","doi-asserted-by":"publisher","first-page":"356","DOI":"10.1038\/nature04160","volume":"437","author":"CA Suttle","year":"2005","unstructured":"Suttle, C. A. Viruses in the sea. Nature 437, 356\u2013361 (2005).","journal-title":"Nature"},{"key":"2455_CR2","doi-asserted-by":"publisher","first-page":"197","DOI":"10.1146\/annurev-virology-100114-054952","volume":"3","author":"AG Cobi\u00e1n G\u00fcemes","year":"2016","unstructured":"Cobi\u00e1n G\u00fcemes, A. G. et al. Viruses as winners in the game of life. Annu. Rev. Virol. 3, 197\u2013214 (2016).","journal-title":"Annu. Rev. Virol."},{"key":"2455_CR3","doi-asserted-by":"publisher","first-page":"801","DOI":"10.1038\/nrmicro1750","volume":"5","author":"CA Suttle","year":"2007","unstructured":"Suttle, C. A. Marine viruses\u2014major players in the global ecosystem. Nat. Rev. Microbiol. 5, 801\u2013812 (2007).","journal-title":"Nat. Rev. Microbiol."},{"key":"2455_CR4","doi-asserted-by":"publisher","first-page":"327","DOI":"10.1038\/s41586-019-1894-8","volume":"577","author":"HG Hampton","year":"2020","unstructured":"Hampton, H. G., Watson, B. N. J. & Fineran, P. C. The arms race between bacteria and their phage foes. Nature 577, 327\u2013336 (2020).","journal-title":"Nature"},{"key":"2455_CR5","doi-asserted-by":"publisher","first-page":"49","DOI":"10.1038\/s41579-021-00602-y","volume":"20","author":"A Chevallereau","year":"2022","unstructured":"Chevallereau, A., Pons, B. J., van Houte, S. & Westra, E. R. Interactions between bacterial and phage communities in natural environments. Nat. Rev. Microbiol. 20, 49\u201362 (2022).","journal-title":"Nat. Rev. Microbiol."},{"key":"2455_CR6","doi-asserted-by":"publisher","first-page":"777","DOI":"10.1038\/nrmicro3564","volume":"13","author":"GPC Salmond","year":"2015","unstructured":"Salmond, G. P. C. & Fineran, P. C. A century of the phage: past, present and future. Nat. Rev. Microbiol. 13, 777\u2013786 (2015).","journal-title":"Nat. Rev. Microbiol."},{"key":"2455_CR7","doi-asserted-by":"publisher","first-page":"17","DOI":"10.1016\/j.cell.2022.11.017","volume":"186","author":"SA Strathdee","year":"2023","unstructured":"Strathdee, S. A., Hatfull, G. F., Mutalik, V. K. & Schooley, R. T. Phage therapy: from biological mechanisms to future directions. Cell 186, 17\u201331 (2023).","journal-title":"Cell"},{"key":"2455_CR8","doi-asserted-by":"publisher","first-page":"8107","DOI":"10.1128\/JVI.01340-15","volume":"89","author":"GF Hatfull","year":"2015","unstructured":"Hatfull, G. F. Dark matter of the biosphere: the amazing world of bacteriophage diversity. J. Virol. 89, 8107\u20138110 (2015).","journal-title":"J. Virol."},{"key":"2455_CR9","doi-asserted-by":"publisher","first-page":"e31955","DOI":"10.7554\/eLife.31955","volume":"7","author":"G Mahmoudabadi","year":"2018","unstructured":"Mahmoudabadi, G. & Phillips, R. A comprehensive and quantitative exploration of thousands of viral genomes. Elife 7, e31955 (2018).","journal-title":"Elife"},{"key":"2455_CR10","doi-asserted-by":"publisher","first-page":"86","DOI":"10.1128\/MMBR.67.1.86-156.2003","volume":"67","author":"ES Miller","year":"2003","unstructured":"Miller, E. S. et al. Bacteriophage T4 genome. Microbiol. Mol. Biol. Rev. 67, 86\u2013156 (2003).","journal-title":"Microbiol. Mol. Biol. Rev."},{"key":"2455_CR11","doi-asserted-by":"publisher","first-page":"298","DOI":"10.1016\/j.coviro.2011.06.009","volume":"1","author":"GF Hatfull","year":"2011","unstructured":"Hatfull, G. F. & Hendrix, R. W. Bacteriophages and their genomes. Curr. Opin. Virol. 1, 298\u2013303 (2011).","journal-title":"Curr. Opin. Virol."},{"key":"2455_CR12","doi-asserted-by":"publisher","first-page":"1838","DOI":"10.1016\/j.molcel.2025.04.010","volume":"85","author":"S Silas","year":"2025","unstructured":"Silas, S. et al. Activation of bacterial programmed cell death by phage inhibitors of host immunity. Mol. Cell 85, 1838\u20131851 (2025).","journal-title":"Mol. Cell"},{"key":"2455_CR13","doi-asserted-by":"publisher","first-page":"307","DOI":"10.1146\/annurev-virology-031413-085500","volume":"1","author":"RL Dy","year":"2014","unstructured":"Dy, R. L., Richter, C., Salmond, G. P. C. & Fineran, P. C. Remarkable mechanisms in microbes to resist phage infections. Annu. Rev. Virol. 1, 307\u2013331 (2014).","journal-title":"Annu. Rev. Virol."},{"key":"2455_CR14","doi-asserted-by":"publisher","first-page":"309","DOI":"10.1146\/annurev-biochem-011420-111224","volume":"89","author":"AR Davidson","year":"2020","unstructured":"Davidson, A. R. et al. Anti-CRISPRs: protein inhibitors of CRISPR\u2013Cas systems. Annu. Rev. Biochem. 89, 309\u2013332 (2020).","journal-title":"Annu. Rev. Biochem."},{"key":"2455_CR15","doi-asserted-by":"publisher","first-page":"e3002416","DOI":"10.1371\/journal.pbio.3002416","volume":"21","author":"D Piya","year":"2023","unstructured":"Piya, D. et al. Systematic and scalable genome-wide essentiality mapping to identify nonessential genes in phages. PLoS Biol. 21, e3002416 (2023).","journal-title":"PLoS Biol."},{"key":"2455_CR16","doi-asserted-by":"publisher","first-page":"694","DOI":"10.1038\/s41564-025-01935-7","volume":"10","author":"BA Adler","year":"2025","unstructured":"Adler, B. A. et al. CRISPRi-ART enables functional genomics of diverse bacteriophages using RNA-binding dCas13d. Nat. Microbiol. 10, 694\u2013709 (2025).","journal-title":"Nat. Microbiol."},{"key":"2455_CR17","doi-asserted-by":"publisher","first-page":"1363","DOI":"10.1016\/j.chom.2025.06.018","volume":"33","author":"J Chen","year":"2025","unstructured":"Chen, J. et al. Systematic, high-throughput characterization of bacteriophage gene essentiality on diverse hosts. Cell Host Microbe 33, 1363\u20131380 (2025).","journal-title":"Cell Host Microbe"},{"key":"2455_CR18","doi-asserted-by":"publisher","unstructured":"Ojima, S., et al. Systematic discovery of phage genes that inactivate bacterial immune systems. Preprint at bioRxiv https:\/\/doi.org\/10.1101\/2024.04.14.589459 (2024).","DOI":"10.1101\/2024.04.14.589459"},{"key":"2455_CR19","doi-asserted-by":"publisher","first-page":"E848","DOI":"10.1073\/pnas.1222538110","volume":"110","author":"WP Robins","year":"2013","unstructured":"Robins, W. P., Faruque, S. M. & Mekalanos, J. J. Coupling mutagenesis and parallel deep sequencing to probe essential residues in a genome or gene. Proc. Natl Acad. Sci. USA 110, E848\u2013E857 (2013).","journal-title":"Proc. Natl Acad. Sci. USA"},{"key":"2455_CR20","doi-asserted-by":"publisher","first-page":"1195","DOI":"10.1038\/s41586-025-09499-6","volume":"646","author":"M Gerovac","year":"2025","unstructured":"Gerovac, M. et al. Programmable antisense oligomers for phage functional genomics. Nature 646, 1195\u20131203 (2025).","journal-title":"Nature"},{"key":"2455_CR21","doi-asserted-by":"publisher","first-page":"767","DOI":"10.1038\/nmeth.1377","volume":"6","author":"T van Opijnen","year":"2009","unstructured":"van Opijnen, T., Bodi, K. L. & Camilli, A. Tn-seq: high-throughput parallel sequencing for fitness and genetic interaction studies in microorganisms. Nat. Methods 6, 767\u2013772 (2009).","journal-title":"Nat. Methods"},{"key":"2455_CR22","doi-asserted-by":"publisher","first-page":"110231","DOI":"10.1016\/j.jbc.2025.110231","volume":"301","author":"LM Nolan","year":"2025","unstructured":"Nolan, L. M., Webber, M. A. & Filloux, A. Throwing a spotlight on genomic dark matter: the power and potential of transposon-insertion sequencing. J. Biol. Chem. 301, 110231 (2025).","journal-title":"J. Biol. Chem."},{"key":"2455_CR23","doi-asserted-by":"publisher","first-page":"1405","DOI":"10.1128\/JB.187.4.1405-1414.2005","volume":"187","author":"S Kiljunen","year":"2005","unstructured":"Kiljunen, S., Vilen, H., Pajunen, M., Savilahti, H. & Skurnik, M. Nonessential genes of phage \u03c6YeO3-12 include genes involved in adaptation to growth on Yersinia enterocolitica serotype O:3. J. Bacteriol. 187, 1405 (2005).","journal-title":"J. Bacteriol."},{"key":"2455_CR24","doi-asserted-by":"publisher","first-page":"91","DOI":"10.1016\/0378-1119(92)90712-X","volume":"114","author":"D Kersulyte","year":"1992","unstructured":"Kersulyte, D., Rajendra Krishnan, B. & Berg, D. E. Nonrandom orientation of transposon Tn5supF insertions in phage lambda. Gene 114, 91\u201396 (1992).","journal-title":"Gene"},{"key":"2455_CR25","doi-asserted-by":"publisher","first-page":"1289","DOI":"10.1111\/j.1365-2958.1992.tb00850.x","volume":"6","author":"DL Woodworth","year":"1992","unstructured":"Woodworth, D. L. & Kreuzer, K. N. A system of transposon mutagenesis for bacteriophage T4. Mol. Microbiol. 6, 1289\u20131296 (1992).","journal-title":"Mol. Microbiol."},{"key":"2455_CR26","doi-asserted-by":"publisher","first-page":"119","DOI":"10.1038\/nrmicro.2015.7","volume":"14","author":"MC Chao","year":"2016","unstructured":"Chao, M. C., Abel, S., Davis, B. M. & Waldor, M. K. The design and analysis of transposon insertion sequencing experiments. Nat. Rev. Microbiol. 14, 119\u2013128 (2016).","journal-title":"Nat. Rev. Microbiol."},{"key":"2455_CR27","doi-asserted-by":"publisher","first-page":"54","DOI":"10.1126\/science.abb6151","volume":"369","author":"AJ Meeske","year":"2020","unstructured":"Meeske, A. J. et al. A phage-encoded anti-CRISPR enables complete evasion of type VI-A CRISPR\u2013Cas immunity. Science 369, 54\u201359 (2020).","journal-title":"Science"},{"key":"2455_CR28","doi-asserted-by":"publisher","first-page":"1956","DOI":"10.1038\/s41564-022-01243-4","volume":"7","author":"J Guan","year":"2022","unstructured":"Guan, J. et al. Bacteriophage genome engineering with CRISPR\u2013Cas13a. Nat. Microbiol. 7, 1956\u20131966 (2022).","journal-title":"Nat. Microbiol."},{"key":"2455_CR29","doi-asserted-by":"publisher","first-page":"aaf5573","DOI":"10.1126\/science.aaf5573","volume":"353","author":"OO Abudayyeh","year":"2016","unstructured":"Abudayyeh, O. O. et al. C2c2 is a single-component programmable RNA-guided RNA-targeting CRISPR effector. Science 353, aaf5573 (2016).","journal-title":"Science"},{"key":"2455_CR30","doi-asserted-by":"publisher","first-page":"1967","DOI":"10.1038\/s41564-022-01258-x","volume":"7","author":"BA Adler","year":"2022","unstructured":"Adler, B. A. et al. Broad-spectrum CRISPR\u2013Cas13a enables efficient phage genome editing. Nat. Microbiol. 7, 1967\u20131979 (2022).","journal-title":"Nat. Microbiol."},{"key":"2455_CR31","doi-asserted-by":"publisher","first-page":"241","DOI":"10.1038\/s41586-019-1257-5","volume":"570","author":"AJ Meeske","year":"2019","unstructured":"Meeske, A. J., Nakandakari-Higa, S. & Marraffini, L. A. Cas13-induced cellular dormancy prevents the rise of CRISPR-resistant bacteriophage. Nature 570, 241\u2013245 (2019).","journal-title":"Nature"},{"key":"2455_CR32","doi-asserted-by":"publisher","unstructured":"Chan, A. et al. Bacteriophage genome-wide transposon mutagenesis. Cell https:\/\/doi.org\/10.1016\/j.cell.2026.06.030 (2026).","DOI":"10.1016\/j.cell.2026.06.030"},{"key":"2455_CR33","doi-asserted-by":"publisher","first-page":"1235","DOI":"10.1261\/rna.591807","volume":"13","author":"LK Wang","year":"2007","unstructured":"Wang, L. K., Nandakumar, J., Schwer, B. & Shuman, S. The C-terminal domain of T4 RNA ligase 1 confers specificity for tRNA repair. RNA 13, 1235 (2007).","journal-title":"RNA"},{"key":"2455_CR34","doi-asserted-by":"publisher","first-page":"669","DOI":"10.1111\/j.1365-2958.2012.07975.x","volume":"83","author":"Y Otsuka","year":"2012","unstructured":"Otsuka, Y. & Yonesaki, T. Dmd of bacteriophage T4 functions as an antitoxin against Escherichia coli LsoA and RnlA toxins. Mol. Microbiol. 83, 669\u2013681 (2012).","journal-title":"Mol. Microbiol."},{"key":"2455_CR35","doi-asserted-by":"publisher","first-page":"522","DOI":"10.1038\/s41586-022-04716-y","volume":"605","author":"SJ Hobbs","year":"2022","unstructured":"Hobbs, S. J. et al. Phage anti-CBASS and anti-Pycsar nucleases subvert bacterial immunity. Nature 605, 522\u2013526 (2022).","journal-title":"Nature"},{"key":"2455_CR36","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-025-56887-7","volume":"16","author":"A Johannesman","year":"2025","unstructured":"Johannesman, A., Awasthi, L. C., Carlson, N. & LeRoux, M. Phages carry orphan antitoxin-like enzymes to neutralize the DarTG1 toxin\u2013antitoxin defense system. Nat. Commun. 16, 1598 (2025).","journal-title":"Nat. Commun."},{"key":"2455_CR37","doi-asserted-by":"publisher","first-page":"1109","DOI":"10.1016\/j.molcel.2016.11.014","volume":"64","author":"G Jankevicius","year":"2016","unstructured":"Jankevicius, G., Ariza, A., Ahel, M. & Ahel, I. The toxin\u2013antitoxin system DarTG catalyzes reversible ADP-ribosylation of DNA. Mol. Cell 64, 1109\u20131116 (2016).","journal-title":"Mol. Cell"},{"key":"2455_CR38","doi-asserted-by":"publisher","first-page":"780","DOI":"10.1016\/j.str.2023.04.008","volume":"31","author":"A Deep","year":"2023","unstructured":"Deep, A. et al. Structural insights into DarT toxin neutralization by cognate DarG antitoxin: ssDNA mimicry by DarG C-terminal domain keeps the DarT toxin inhibited. Structure 31, 780\u2013789 (2023).","journal-title":"Structure"},{"key":"2455_CR39","doi-asserted-by":"publisher","first-page":"117219","DOI":"10.1016\/j.celrep.2026.117219","volume":"45","author":"CJ Morgan","year":"2026","unstructured":"Morgan, C. J. et al. The phage nucleus synergizes with an anti-defense protein to resist bacterial immunity. Cell Rep. 45, 117219 (2026).","journal-title":"Cell Rep."},{"key":"2455_CR40","doi-asserted-by":"publisher","first-page":"42","DOI":"10.1128\/jvi.51.1.42-46.1984","volume":"51","author":"NJ Olson","year":"1984","unstructured":"Olson, N. J. & Marchin, G. L. Valyl-tRNA synthetase modification-dependent restriction of bacteriophage T4. J. Virol. 51, 42\u201346 (1984).","journal-title":"J. Virol."},{"key":"2455_CR41","doi-asserted-by":"publisher","first-page":"e3001424","DOI":"10.1371\/journal.pbio.3001424","volume":"19","author":"E Maffei","year":"2021","unstructured":"Maffei, E. et al. Systematic exploration of Escherichia coli phage\u2013host interactions with the BASEL phage collection. PLoS Biol. 19, e3001424 (2021).","journal-title":"PLoS Biol."},{"key":"2455_CR42","doi-asserted-by":"publisher","DOI":"10.1186\/1743-422X-10-76","volume":"10","author":"AM Kropinski","year":"2013","unstructured":"Kropinski, A. M. et al. The host-range, genomics and proteomics of Escherichia coli O157:H7 bacteriophage rV5. Virol. J. 10, 76 (2013).","journal-title":"Virol. J."},{"key":"2455_CR43","doi-asserted-by":"publisher","first-page":"113","DOI":"10.1038\/s41579-019-0278-2","volume":"18","author":"A Bernheim","year":"2020","unstructured":"Bernheim, A. & Sorek, R. The pan-immune system of bacteria: antiviral defence as a community resource. Nat. Rev. Microbiol. 18, 113\u2013119 (2020).","journal-title":"Nat. Rev. Microbiol."},{"key":"2455_CR44","doi-asserted-by":"publisher","first-page":"gkae1171","DOI":"10.1093\/nar\/gkae1171","volume":"53","author":"F Tesson","year":"2025","unstructured":"Tesson, F. et al. Exploring the diversity of anti-defense systems across prokaryotes, phages and mobile genetic elements. Nucleic Acids Res. 53, gkae1171 (2025).","journal-title":"Nucleic Acids Res."},{"key":"2455_CR45","doi-asserted-by":"publisher","first-page":"1212","DOI":"10.1016\/j.tim.2024.05.005","volume":"32","author":"K Murtazalieva","year":"2024","unstructured":"Murtazalieva, K., Mu, A., Petrovskaya, A. & Finn, R. D. The growing repertoire of phage anti-defence systems. Trends Microbiol. 32, 1212\u20131228 (2024).","journal-title":"Trends Microbiol."},{"key":"2455_CR46","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-022-30269-9","volume":"13","author":"F Tesson","year":"2022","unstructured":"Tesson, F. et al. Systematic and quantitative view of the antiviral arsenal of prokaryotes. Nat. Commun. 13, 2561 (2022).","journal-title":"Nat. Commun."},{"key":"2455_CR47","first-page":"253","volume":"4","author":"T Koch","year":"2018","unstructured":"Koch, T., Raudonikiene, A., Wilkens, K. & R\u00fcger, W. Over expression, purification, and characterization of the ADP-ribosyltransferase (gpAlt) of bacteriophage T4: ADP-ribosylation of E. coli RNA polymerase modulates T4 \u2018early\u2019 transcription. Gene Expr. 4, 253 (2018).","journal-title":"Gene Expr."},{"key":"2455_CR48","doi-asserted-by":"publisher","first-page":"188","DOI":"10.1111\/mmi.13225","volume":"99","author":"AM Alawneh","year":"2016","unstructured":"Alawneh, A. M., Qi, D., Yonesaki, T. & Otsuka, Y. An ADP-ribosyltransferase Alt of bacteriophage T4 negatively regulates the Escherichia coli MazF toxin of a toxin\u2013antitoxin module. Mol. Microbiol. 99, 188\u2013198 (2016).","journal-title":"Mol. Microbiol."},{"key":"2455_CR49","doi-asserted-by":"publisher","first-page":"779","DOI":"10.1016\/j.jmb.2006.11.049","volume":"366","author":"CL Bair","year":"2007","unstructured":"Bair, C. L., Rifat, D. & Black, L. W. Exclusion of glucosyl-hydroxymethylcytosine DNA containing bacteriophages is overcome by the injected protein inhibitor IPI*. J. Mol. Biol. 366, 779\u2013789 (2007).","journal-title":"J. Mol. Biol."},{"key":"2455_CR50","doi-asserted-by":"publisher","first-page":"4181","DOI":"10.1002\/j.1460-2075.1994.tb06736.x","volume":"13","author":"F Repoila","year":"1994","unstructured":"Repoila, F., T\u00e9tart, F., Bouet, J. Y. & Krisch, H. M. Genomic polymorphism in the T-even bacteriophages. EMBO J. 13, 4181 (1994).","journal-title":"EMBO J."},{"key":"2455_CR51","unstructured":"Kyte, N., Fuchs, M., Smith, L. M. & Fineran, P. C. Defining the essential genome of diverse phages with phage Tn-seq. Nat. Microbiol. (in the press)."},{"key":"2455_CR52","doi-asserted-by":"publisher","first-page":"23","DOI":"10.1007\/978-1-0716-1621-5_2","volume":"2357","author":"E Maffei","year":"2021","unstructured":"Maffei, E., Fino, C. & Harms, A. Antibiotic tolerance and persistence studied throughout bacterial growth phases. Methods Mol. Biol. 2357, 23\u201340 (2021).","journal-title":"Methods Mol. Biol."},{"key":"2455_CR53","doi-asserted-by":"publisher","first-page":"1028","DOI":"10.1038\/s41564-022-01153-5","volume":"7","author":"M LeRoux","year":"2022","unstructured":"LeRoux, M. et al. The DarTG toxin\u2013antitoxin system provides phage defence by ADP-ribosylating viral DNA. Nat. Microbiol. 7, 1028\u20131040 (2022).","journal-title":"Nat. Microbiol."},{"key":"2455_CR54","doi-asserted-by":"publisher","first-page":"69","DOI":"10.1007\/978-1-60327-164-6_7","volume":"501","author":"AM Kropinski","year":"2009","unstructured":"Kropinski, A. M., Mazzocco, A., Waddell, T. E., Lingohr, E. & Johnson, R. P. Enumeration of bacteriophages by double agar overlay plaque assay. Methods Mol. Biol. 501, 69\u201376 (2009).","journal-title":"Methods Mol. Biol."},{"key":"2455_CR55","doi-asserted-by":"publisher","first-page":"343","DOI":"10.1038\/nmeth.1318","volume":"6","author":"DG Gibson","year":"2009","unstructured":"Gibson, D. G. et al. Enzymatic assembly of DNA molecules up to several hundred kilobases. Nat. Methods 6, 343\u2013345 (2009).","journal-title":"Nat. Methods"},{"key":"2455_CR56","doi-asserted-by":"publisher","DOI":"10.1186\/1472-6750-8-91","volume":"8","author":"H Liu","year":"2008","unstructured":"Liu, H. & Naismith, J. H. An efficient one-step site-directed deletion, insertion, single and multiple-site plasmid mutagenesis protocol. BMC Biotechnol. 8, 91 (2008).","journal-title":"BMC Biotechnol."},{"key":"2455_CR57","doi-asserted-by":"publisher","unstructured":"Taveneau, C., et al. De novo design of potent CRISPR\u2013Cas13 inhibitors. Nat. Chem. Biol. https:\/\/doi.org\/10.1038\/s41589-025-02136-3 (2026).","DOI":"10.1038\/s41589-025-02136-3"},{"key":"2455_CR58","doi-asserted-by":"publisher","first-page":"1116","DOI":"10.1038\/nmeth.2681","volume":"10","author":"KM Esvelt","year":"2013","unstructured":"Esvelt, K. M. et al. Orthogonal Cas9 proteins for RNA-guided gene regulation and editing. Nat. Methods 10, 1116\u20131121 (2013).","journal-title":"Nat. Methods"},{"key":"2455_CR59","doi-asserted-by":"publisher","first-page":"184","DOI":"10.1038\/nbt.3437","volume":"34","author":"JG Doench","year":"2016","unstructured":"Doench, J. G. et al. Optimized sgRNA design to maximize activity and minimize off-target effects of CRISPR\u2013Cas9. Nat. Biotechnol. 34, 184\u2013191 (2016).","journal-title":"Nat. Biotechnol."},{"key":"2455_CR60","doi-asserted-by":"publisher","DOI":"10.1186\/s13100-020-0200-5","volume":"11","author":"M Tellier","year":"2020","unstructured":"Tellier, M. & Chalmers, R. Compensating for over-production inhibition of the Hsmar1 transposon in Escherichia coli using a series of constitutive promoters. Mobile DNA 11, 5 (2020).","journal-title":"Mobile DNA"},{"key":"2455_CR61","doi-asserted-by":"publisher","first-page":"2006.0008","DOI":"10.1038\/msb4100050","volume":"2","author":"T Baba","year":"2006","unstructured":"Baba, T. et al. Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection. Mol. Syst. Biol. 2, 2006.0008 (2006).","journal-title":"Mol. Syst. Biol."},{"key":"2455_CR62","unstructured":"Andrews, S. FastQC: a quality control tool for high throughput sequence data. Babraham Bioinformatics https:\/\/www.bioinformatics.babraham.ac.uk\/projects\/fastqc\/ (2010)."},{"key":"2455_CR63","doi-asserted-by":"publisher","DOI":"10.1002\/imt2.107","volume":"2","author":"S Chen","year":"2023","unstructured":"Chen, S. Ultrafast one-pass FASTQ data preprocessing, quality control, and deduplication using fastp. iMeta 2, e107 (2023).","journal-title":"iMeta"},{"key":"2455_CR64","doi-asserted-by":"publisher","first-page":"e191","DOI":"10.1002\/imt2.191","volume":"3","author":"W Shen","year":"2024","unstructured":"Shen, W., Sipos, B. & Zhao, L. SeqKit2: a Swiss army knife for sequence and alignment processing. iMeta 3, e191 (2024).","journal-title":"iMeta"},{"key":"2455_CR65","doi-asserted-by":"publisher","first-page":"e1004401","DOI":"10.1371\/journal.pcbi.1004401","volume":"11","author":"MA DeJesus","year":"2015","unstructured":"DeJesus, M. A., Ambadipudi, C., Baker, R., Sassetti, C. & Ioerger T. R. TRANSIT - A software tool for Himar1 TnSeq analysis. PLoS Comput. Biol. 11, e1004401 (2015).","journal-title":"PLoS Comput. Biol."},{"key":"2455_CR66","doi-asserted-by":"publisher","unstructured":"Li, H. Aligning sequence reads, clone sequences and assembly contigs with BWA-MEM. Preprint at https:\/\/doi.org\/10.48550\/arXiv.1303.3997 (2013).","DOI":"10.48550\/arXiv.1303.3997"},{"key":"2455_CR67","doi-asserted-by":"publisher","first-page":"2114","DOI":"10.1093\/bioinformatics\/btu170","volume":"30","author":"AM Bolger","year":"2014","unstructured":"Bolger, A. M., Lohse, M. & Usadel, B. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 30, 2114\u20132120 (2014).","journal-title":"Bioinformatics"},{"key":"2455_CR68","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pcbi.1005595","volume":"13","author":"RR Wick","year":"2017","unstructured":"Wick, R. R., Judd, L. M., Gorrie, C. L. & Holt, K. E. Unicycler: resolving bacterial genome assemblies from short and long sequencing reads. PLoS Comput. Biol. 13, e1005595 (2017).","journal-title":"PLoS Comput. Biol."},{"key":"2455_CR69","doi-asserted-by":"publisher","first-page":"e3003063","DOI":"10.1371\/journal.pbio.3003063","volume":"23","author":"D Humolli","year":"2025","unstructured":"Humolli, D. et al. Completing the BASEL phage collection to unlock hidden diversity for systematic exploration of phage\u2013host interactions. PLoS Biol. 23, e3003063 (2025).","journal-title":"PLoS Biol."},{"key":"2455_CR70","doi-asserted-by":"publisher","first-page":"2237","DOI":"10.1016\/j.jmb.2017.12.007","volume":"430","author":"L Zimmermann","year":"2018","unstructured":"Zimmermann, L. et al. A completely reimplemented MPI bioinformatics toolkit with a new HHpred server at its core. J. Mol. Biol. 430, 2237\u20132243 (2018).","journal-title":"J. Mol. Biol."},{"key":"2455_CR71","doi-asserted-by":"publisher","first-page":"e4519","DOI":"10.1002\/pro.4519","volume":"32","author":"L Holm","year":"2023","unstructured":"Holm, L., Laiho, A., T\u00f6r\u00f6nen, P. & Salgado, M. DALI shines a light on remote homologs: one hundred discoveries. Protein Sci. 32, e4519 (2023).","journal-title":"Protein Sci."},{"key":"2455_CR72","doi-asserted-by":"publisher","first-page":"679","DOI":"10.1038\/s41592-022-01488-1","volume":"19","author":"M Mirdita","year":"2022","unstructured":"Mirdita, M. et al. ColabFold: making protein folding accessible to all. Nat. Methods 19, 679\u2013682 (2022).","journal-title":"Nat. Methods"},{"key":"2455_CR73","doi-asserted-by":"publisher","unstructured":"Humolli, D. & Harms, A. Systematic mapping of bacteriophage gene essentiality with HIDEN\u2011SEQ. Zenodo https:\/\/doi.org\/10.5281\/zenodo.20179451 (2026).","DOI":"10.5281\/zenodo.20179451"}],"container-title":["Nature Microbiology"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.nature.com\/articles\/s41564-026-02455-8.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/www.nature.com\/articles\/s41564-026-02455-8","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/www.nature.com\/articles\/s41564-026-02455-8.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,9,25]],"date-time":"2026-09-25T09:02:27Z","timestamp":1790326947000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.nature.com\/articles\/s41564-026-02455-8"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,9,25]]},"references-count":73,"alternative-id":["2455"],"URL":"https:\/\/doi.org\/10.1038\/s41564-026-02455-8","relation":{},"ISSN":["2058-5276"],"issn-type":[{"value":"2058-5276","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,9,25]]},"assertion":[{"value":"2 December 2025","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"14 July 2026","order":2,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"25 September 2026","order":3,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"J.-W.V. is a scientific advisory board member at i-Seq Biotechnology. The other authors declare no competing interests.","order":1,"name":"Ethics","label":"Competing interests","group":{"name":"EthicsHeading","label":"Ethics"}}]}}