DNA methylation is an important mechanism involved in bacteria limiting foreign DNA acquisition, maintenance of mobile genetic elements, DNA mismatch repair, and gene expression. Changes in DNA methylation pattern are observed in bacteria under stress conditions, including exposure to antimicrobial compounds. These changes can result in transient and fast-appearing adaptive antibiotic resistance (AdR) phenotypes, e.g., strain overexpressing efflux pumps. DNA methylation can be related to DNA mutation rate, because it is involved in DNA mismatch repair systems and because methylated bases are well-known mutational hotspots. The AdR process can be the first important step in the selection of antibiotic-resistant strains, allowing the survival of the bacterial population until more efficient resistant mutants emerge. Epigenetic modifications can be investigated by third-generation sequencing platforms that allow us to simultaneously detect all the methylated bases along with the DNA sequencing. In this scenario, this sequencing technology enables the study of epigenetic modifications in link with antibiotic resistance and will help to investigate the relationship between methylation and mutation in the development of stable mechanisms of resistance.

Biomedical Center Faculty of Medicine Charles University Pilsen Czechia

Department of Microbiology Faculty of Medicine and University Hospital in Pilsen Charles University Pilsen Czechia

Romeo ed Enrica Invernizzi Pediatric Research Center Department of Biomedical and Clinical Sciences University of Milan Milan Italy

Romeo ed Enrica Invernizzi Pediatric Research Center Department of Bioscience University of Milan Milan Italy

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Acharya S., Foster P. L., Brooks P., Fishel R. (2003). The coordinated functions of the E. coli MutS and MutL proteins in mismatch repair. Mol. Cell 12, 233–246. 10.1016/S1097-2765(03)00219-3 PubMed DOI

Adam M., Murali B., Glenn N. O., Potter S. S. (2008). Epigenetic inheritance based evolution of antibiotic resistance in bacteria. BMC Evol. Biol. 8, 52. 10.1186/1471-2148-8-52 PubMed DOI PMC

Adhikari S., Curtis P. D. (2016). DNA methyltransferases and epigenetic regulation in bacteria. FEMS Microbiol. Rev. 40, 575–591. 10.1093/femsre/fuw023 PubMed DOI

Aminov R. I. (2010). A brief history of the antibiotic era: lessons learned and challenges for the future. Front. Microbiol. 1, 134. 10.3389/fmicb.2010.00134 PubMed DOI PMC

Andersson D. I., Levin B. R. (1999). The biological cost of antibiotic resistance. Curr. Opin. Microbiol. 2, 489–493. 10.1016/S1369-5274(99)00005-3 PubMed DOI

Ardui S., Ameur A., Vermeesch J. R., Hestand M. S. (2018). Single molecule real-time (SMRT) sequencing comes of age: applications and utilities for medical diagnostics. Nucleic Acids Res. 46, 2159–2168. 10.1093/nar/gky066 PubMed DOI PMC

Barclay M. L., Begg E. J., Chambers S. T. (1992). Adaptive resistance following single doses of gentamicin in a dynamic in vitro model. Antimicrob. Agents Chemother. 36, 1951–1957. 10.1128/AAC.36.9.1951 PubMed DOI PMC

Beaulaurier J., Schadt E. E., Fang G. (2019). Deciphering bacterial epigenomes using modern sequencing technologies. Nat. Rev. Genet. 20, 157–172. 10.1038/s41576-018-0081-3 PubMed DOI PMC

Beissinger M., Buchner J. (1998). How chaperones fold proteins. Biol. Chem. 379, 245–259. PubMed

Blow M. J., Clark T. A., Daum C. G., Deutschbauer A. M., Fomenkov A., Fries R., et al. . (2016). The epigenomic landscape of prokaryotes. PLoS Genet. 12, e1005854. 10.1371/journal.pgen.1005854 PubMed DOI PMC

Branton D., Deamer D. W., Marziali A., Bayley H., Benner S. A., Butler T., et al. . (2008). The potential and challenges of nanopore sequencing. Nat. Biotechnol. 26, 1146–1153. 10.1038/nbt.1495 PubMed DOI PMC

Calmann M. A., Marinus M. G. (2003). Regulated expression of the Escherichia coli dam gene. J. Bacteriol. 185, 5012–5014. 10.1128/JB.185.16.5012-5014.2003 PubMed DOI PMC

Cao B., Chen C., DeMott M. S., Cheng Q., Clark T. A., Xiong X., et al. . (2014). Genomic mapping of phosphorothioates reveals partial modification of short consensus sequences. Nat. Commun. 5, 1–13. 10.1038/ncomms4951 PubMed DOI PMC

Carvalho A., Mazel D., Baharoglu Z. (2021). Deficiency in cytosine DNA methylation leads to high chaperonin expression and tolerance to aminoglycosides in Vibrio cholerae. PLoS Genet. 17, e1009748. 10.1371/journal.pgen.1009748 PubMed DOI PMC

Casadesús J., Low D. (2006). Epigenetic gene regulation in the bacterial world. Microbiol. Mol. Biol. Rev. 70, 830–856. 10.1128/MMBR.00016-06 PubMed DOI PMC

Chen C., Wang L., Chen S., Wu X., Gu M., Chen X., et al. . (2017). Convergence of DNA methylation and phosphorothioation epigenetics in bacterial genomes. Proc. Natl. Acad. Sci. USA. 114, 4501–4506. 10.1073/pnas.1702450114 PubMed DOI PMC

Chen L., Li H., Chen T., Yu L., Guo H., Chen Y., et al. . (2018). Genome-wide DNA methylation and transcriptome changes in Mycobacterium tuberculosis with rifampicin and isoniazid resistance. Int. J. Clin. Exp. Pathol. 11, 3036–3045. PubMed PMC

Cheng X. (1995). Structure and function of DNA methyltransferases. Annu. Rev. Biophys. Biomol. Struct. 24, 293–318. 10.1146/annurev.bb.24.060195.001453 PubMed DOI

Cherry J. L. (2018). Methylation-induced hypermutation in natural populations of bacteria. J. Bacteriol. 200, e00371–e00418. 10.1128/JB.00371-18 PubMed DOI PMC

Cherry J. L. (2021). Extreme c-to-a hypermutation at a site of cytosine-N4 methylation. mBio. 12, e00172–21. 10.1128/mBio.00172-21 PubMed DOI PMC

Ciuffreda L., Rodríguez-Pérez H., Flores C. (2021). Nanopore sequencing and its application to the study of microbial communities. Comput. Struct. Biotechnol. J. 19, 1497–1511. 10.1016/j.csbj.2021.02.020 PubMed DOI PMC

Drotschmann K., Aronshtam A., Fritz H. J., Marinus M. G. (1998). The Escherichia coli MutL protein stimulates binding of Vsr and MutS to heteroduplex DNA. Nucleic Acids Res. 26, 948–953. 10.1093/nar/26.4.948 PubMed DOI PMC

Dunn D. B., Smith J. D. (1955). Occurrence of a new base in the deoxyribonucleic acid of a strain of Bacterium coli. Nature 175, 336–337. 10.1038/175336a0 PubMed DOI

Dwyer D. J., Belenky P. A., Yang J. H., MacDonald I. C., Martell J. D., Takahashi N., et al. . (2014). Antibiotics induce redox-related physiological alterations as part of their lethality. Proc. Natl. Acad. Sci. USA. 111, E2100–E2109. 10.1073/pnas.1401876111 PubMed DOI PMC

Ebbensgaard A. E., Løbner-Olesen A., Frimodt-Møller J. (2020). The role of efflux pumps in the transition from low-level to clinical antibiotic resistance. Antibiotics 9, 855. 10.3390/antibiotics9120855 PubMed DOI PMC

Eid J., Fehr A., Gray J., Luong K., Lyle J., Otto G., et al. . (2009). Real-time DNA sequencing from single polymerase molecules. Science 323, 133–138. 10.1126/science.1162986 PubMed DOI

El Meouche I., Dunlop M. J. (2018). Heterogeneity in efflux pump expression predisposes antibiotic-resistant cells to mutation. Science 362, 686–690. 10.1126/science.aar7981 PubMed DOI PMC

El'Garch F., Jeannot K., Hocquet D., Llanes-Barakat C., Plésiat P. (2007). Cumulative effects of several nonenzymatic mechanisms on the resistance of Pseudomonas aeruginosa to aminoglycosides. Antimicrob. Agents Chemother. 51, 1016–1021. 10.1128/AAC.00704-06 PubMed DOI PMC

Fernández L., Hancock R. E. W. (2013). Adaptive and mutational resistance: Role of porins and efflux pumps in drug resistance. Clin. Microbiol. Rev. 26, 163–163. 10.1128/CMR.00094-12 PubMed DOI PMC

Foster P. L. (2005). Stress responses and genetic variation in bacteria. Mutation Res. Fund. Mol. Mech. Mutagenesis 569, 3–11. 10.1016/j.mrfmmm.2004.07.017 PubMed DOI PMC

George A. M., Levy S. B. (1983). Amplifiable resistance to tetracycline, chloramphenicol, and other antibiotics in Escherichia coli: involvement of a non-plasmid-determined efflux of tetracycline. J. Bacteriol. 155, 531–540. 10.1128/jb.155.2.531-540.1983 PubMed DOI PMC

Ghosh D., Veeraraghavan B., Elangovan R., Vivekanandan P. (2020). Antibiotic resistance and epigenetics: more to it than meets the eye. Antimicrob. Agents Chemother. 64, e02225–e02239. 10.1128/AAC.02225-19 PubMed DOI PMC

Goldberg A. D., Allis C. D., Bernstein E. (2007). Epigenetics: a landscape takes shape. Cell 128, 635–638. 10.1016/j.cell.2007.02.006 PubMed DOI

Goltermann L., Sarusie M. V., Bentin T. (2015). Chaperonin GroEL/GroES over-expression promotes aminoglycoside resistance and reduces drug susceptibilities in escherichia coli following exposure to sublethal aminoglycoside doses. Front. Microbiol. 6, 1572. 10.3389/fmicb.2015.01572 PubMed DOI PMC

Hughes L., Roberts W., Johnson D. (2021). The impact of DNA adenine methyltransferase knockout on the development of triclosan resistance and antibiotic cross-resistance in. Access Microbiol 3, acmi000178. 10.1099/acmi.0.000178 PubMed DOI PMC

Jensen T. Ø., Tellgren-Roth C., Redl S., Maury J., Jacobsen S. A. B., Pedersen L. E., et al. . (2019). Genome-wide systematic identification of methyltransferase recognition and modification patterns. Nat. Commun. 10, 3311. 10.1038/s41467-019-11179-9 PubMed DOI PMC

Jian H., Xu G., Yi Y., Hao Y., Wang Y., Xiong L., et al. . (2021). The origin and impeded dissemination of the DNA phosphorothioation system in prokaryotes. Nat. Commun. 12, 6382. 10.1038/s41467-021-26636-7 PubMed DOI PMC

Kang S., Lee H., Han J. S., Hwang D. S. (1999). Interaction of SeqA and Dam methylase on the hemimethylated origin of Escherichia coli chromosomal DNA replication. J. Biol. Chem. 274, 11463–11468. 10.1074/jbc.274.17.11463 PubMed DOI

Kapoor G., Saigal S., Elongavan A. (2017). Action and resistance mechanisms of antibiotics: a guide for clinicians. J. Anaesthesiol. Clin. Pharmacol. 33, 300–305. 10.4103/joacp.JOACP_349_15 PubMed DOI PMC

Kellner S., DeMott M. S., Cheng C. P., Russell B. S., Cao B., You D., et al. . (2017). Oxidation of phosphorothioate DNA modifications leads to lethal genomic instability. Nat. Chem. Biol. 13, 888–894. 10.1038/nchembio.2407 PubMed DOI PMC

Kulakauskas S., Lubys A., Ehrlich S. D. (1995). DNA restriction-modification systems mediate plasmid maintenance. J. Bacteriol. 177, 3451–3454. 10.1128/jb.177.12.3451-3454.1995 PubMed DOI PMC

Lambert P. A. (2005). Bacterial resistance to antibiotics: modified target sites. Adv. Drug Deliv. Rev. 57, 1471–1485. 10.1016/j.addr.2005.04.003 PubMed DOI

Li G.-M. (2008). Mechanisms and functions of DNA mismatch repair. Cell Res. 18, 85–98. 10.1038/cr.2007.115 PubMed DOI

Lieb M., Rehmat S., Bhagwat A. S. (2001). Interaction of MutS and Vsr: some dominant-negative mutS mutations that disable methyladenine-directed mismatch repair are active in very-short-patch repair. J. Bacteriol. 183, 6487–6490. 10.1128/JB.183.21.6487-6490.2001 PubMed DOI PMC

Marinus M. G. (2012). DNA mismatch repair. EcoSal Plus. 5, 10.1128/ecosalplus.7.2.5 PubMed DOI PMC

Marinus M. G., Casadesus J. (2009). Roles of DNA adenine methylation in host-pathogen interactions: mismatch repair, transcriptional regulation, and more. FEMS Microbiol. Rev. 33, 488–503. 10.1111/j.1574-6976.2008.00159.x PubMed DOI PMC

Messer W., Noyer-Weidner M. (1988). Timing and targeting: the biological functions of Dam methylation in E. coli. Cell 54, 735–737. 10.1016/S0092-8674(88)90911-7 PubMed DOI

Moore L. D., Le T., Fan G. (2013). DNA methylation and its basic function. Neuropsychopharmacology 38, 23–38. 10.1038/npp.2012.112 PubMed DOI PMC

Motta S. S., Cluzel P., Aldana M. (2015). Adaptive resistance in bacteria requires epigenetic inheritance, genetic noise, and cost of efflux pumps. PLoS ONE 10, e0118464. 10.1371/journal.pone.0118464 PubMed DOI PMC

Mruk I., Kobayashi I. (2014). To be or not to be: regulation of restriction–modification systems and other toxin–antitoxin systems. Nucleic Acids Res. 42, 70–86. 10.1093/nar/gkt711 PubMed DOI PMC

Nature E. (2013). The antibiotic alarm. Nature 495, 141. 10.1038/495141a PubMed DOI

Nye T. M., Jacob K. M., Holley E. K., Nevarez J. M., Dawid S., Simmons L. A., et al. . (2019). DNA methylation from a Type I restriction modification system influences gene expression and virulence in Streptococcus pyogenes. PLoS Pathog. 15, e1007841. 10.1371/journal.ppat.1007841 PubMed DOI PMC

Oliveira P. H., Fang G. (2021). Conserved DNA Methyltransferases: A Window into Fundamental Mechanisms of Epigenetic Regulation in Bacteria. Trends Microbiol. 29, 28–40. 10.1016/j.tim.2020.04.007 PubMed DOI PMC

Olofsson S. K., Cars O. (2007). Optimizing drug exposure to minimize selection of antibiotic resistance. Clin. Infect. Dis. 45, S129–36. 10.1086/519256 PubMed DOI

Oshima T., Wada C., Kawagoe Y., Ara T., Maeda M., Masuda Y., et al. . (2002). Genome-wide analysis of deoxyadenosine methyltransferase-mediated control of gene expression in Escherichia coli. Mol. Microbiol. 45, 673–695. 10.1046/j.1365-2958.2002.03037.x PubMed DOI

Palmer B. R., Marinus M. G. (1994). The dam and dcm strains of Escherichia coli–a review. Gene 143, 1–12. 10.1016/0378-1119(94)90597-5 PubMed DOI

Payelleville A., Brillard J. (2021). Novel Identification of Bacterial Epigenetic Regulations Would Benefit From a Better Exploitation of Methylomic Data. Front. Microbiol. 12, 685670. 10.3389/fmicb.2021.685670 PubMed DOI PMC

Peterson S. N., Reich N. O. (2006). GATC flanking sequences regulate Dam activity: evidence for how Dam specificity may influence pap expression. J. Mol. Biol. 355, 459–472. 10.1016/j.jmb.2005.11.003 PubMed DOI

Ramirez M. S., Tolmasky M. E. (2010). Aminoglycoside modifying enzymes. Drug Resist. Updat. 13, 151–171. 10.1016/j.drup.2010.08.003 PubMed DOI PMC

Sánchez-Romero M. A., Casadesús J. (2014). Contribution of phenotypic heterogeneity to adaptive antibiotic resistance. Proc. Natl. Acad. Sci. USA. 111, 355–360. 10.1073/pnas.1316084111 PubMed DOI PMC

Sánchez-Romero M. A., Casadesús J. (2020). The bacterial epigenome. Nat. Rev. Microbiol. 18, 7–20. 10.1038/s41579-019-0286-2 PubMed DOI

Sánchez-Romero M. A., Casadesús J. (2021). Waddington's landscapes in the bacterial world. Front. Microbiol. 12, 1167. 10.3389/fmicb.2021.685080 PubMed DOI PMC

Sandoval-Motta S., Aldana M. (2016). Adaptive resistance to antibiotics in bacteria: a systems biology perspective. Wiley Interdiscip. Rev. Syst. Biol. Med. 8, 253–267. 10.1002/wsbm.1335 PubMed DOI

Sater M. R. A., Lamelas A., Wang G., Clark T. A., Röltgen K., Mane S., et al. . (2015). DNA methylation assessed by SMRT sequencing is linked to mutations in Neisseria meningitidis Isolates. PLoS ONE 10, e0144612. 10.1371/journal.pone.0144612 PubMed DOI PMC

Schaenzer A. J., Wright G. D. (2020). Antibiotic resistance by enzymatic modification of antibiotic targets. Trends Mol. Med. 26, 768–782. 10.1016/j.molmed.2020.05.001 PubMed DOI

Schlagman S. L., Hattman S., Marinus M. G. (1986). Direct role of the Escherichia coli dam DNA methyltransferase in methylation-directed mismatch repair. J. Bacteriol. 165, 896–900. 10.1128/jb.165.3.896-900.1986 PubMed DOI PMC

Spadar A., Perdigão J., Phelan J., Charleston J., Modesto A., Elias R., et al. . (2021). Methylation analysis of Klebsiella pneumoniae from Portuguese hospitals. Sci. Rep. 11, 6491. 10.1038/s41598-021-85724-2 PubMed DOI PMC

Tong T., Chen S., Wang L., Tang Y., Ryu J. Y., Jiang S., et al. . (2018). Occurrence, evolution, and functions of DNA phosphorothioate epigenetics in bacteria. Proc. Natl. Acad. Sci. USA. 115, E2988–E2996. 10.1073/pnas.1721916115 PubMed DOI PMC

Toprak E., Veres A., Michel J.-B., Chait R., Hartl D. L., Kishony R. (2011). Evolutionary paths to antibiotic resistance under dynamically sustained drug selection. Nat. Genet. 44, 101–105. 10.1038/ng.1034 PubMed DOI PMC

Unterholzner S. J., Poppenberger B., Rozhon W. (2013). Toxin-antitoxin systems: Biology, identification, and application. Mob. Genet. Elements 3, e26219. 10.4161/mge.26219 PubMed DOI PMC

Vasu K., Nagaraja V. (2013). Diverse functions of restriction-modification systems in addition to cellular defense. Microbiol. Mol. Biol. Rev. 77, 53–72. 10.1128/MMBR.00044-12 PubMed DOI PMC

Ventola C. L. (2015). The antibiotic resistance crisis: part 1: causes and threats. Pharmacy Therapeutics 40, 277–283. PubMed PMC

Waksman S. A. (1947). What is an antibiotic or an antibiotic substance? Mycologia 39, 565–569. 10.1080/00275514.1947.12017635 PubMed DOI

Wang L., Chen S., Xu T., Taghizadeh K., Wishnok J. S., Zhou X., et al. . (2007). Phosphorothioation of DNA in bacteria by dnd genes. Nat. Chem. Biol. 3, 709–710. 10.1038/nchembio.2007.39 PubMed DOI

Wang R., Lou J., Li J. (2019). A mobile restriction modification system consisting of methylases on the IncA/C plasmid. Mob. DNA 10, 26. 10.1186/s13100-019-0168-1 PubMed DOI PMC

Webber M. A., Piddock L. J. V. (2003). The importance of efflux pumps in bacterial antibiotic resistance. J. Antimicrob. Chemother. 51, 9–11. 10.1093/jac/dkg050 PubMed DOI

Wilson D. N. (2014). Ribosome-targeting antibiotics and mechanisms of bacterial resistance. Nat. Rev. Microbiol. 12, 35–48. 10.1038/nrmicro3155 PubMed DOI

Windels E. M., Van den Bergh B., Michiels J. (2020). Bacteria under antibiotic attack: Different strategies for evolutionary adaptation. PLoS Pathog. 16, e1008431. 10.1371/journal.ppat.1008431 PubMed DOI PMC

Wu X., Cao B., Aquino P., Chiu T.-P., Chen C., Jiang S., et al. . (2020). Epigenetic competition reveals density-dependent regulation and target site plasticity of phosphorothioate epigenetics in bacteria. Proc. Natl. Acad. Sci. USA. 117, 14322–14330. 10.1073/pnas.2002933117 PubMed DOI PMC

Yee R., Dien Bard J., Simner P. J. (2021). The genotype-to-phenotype dilemma: how should laboratories approach discordant susceptibility results? J. Clin. Microbiol. 59, e00138– e00220. 10.1128/JCM.00138-20 PubMed DOI PMC