There is currently a lack of experimental evidence for horizontal gene transfer (HGT) mechanisms in the human gut microbiota. The aim of this study was therefore to experimentally determine the HGT potential in the microbiota of a healthy preterm infant twin pair and to evaluate the global occurrence of the mobilized elements.
Stool samples were collected. Both shotgun metagenome sequencing and bacterial culturing were done for the same samples. A range of experimental conditions were used to test DNA transfer for the cultured isolates. Searches for global distribution of transferable elements were done for the ~120,000 metagenomic samples in the Sequence Read Archive (SRA) database.
DNA transfer experiments demonstrated frequent transmission of an ESBL encoding IncI1 plasmid, a high copy number ColEI plasmid, and bacteriophage P1. Both IncI1 and ColE1 were abundant in the stool samples. In vitro competition experiments showed that transconjugants containing IncI1 plasmids outcompeted the recipient strain in the absence of antibiotic selection. The SRA searches indicated a global distribution of the mobilizable elements, with chicken identified as a possible reservoir for the IncI1 ESBL encoding plasmid.
Our results experimentally support a major horizontal transmission and persistence potential of the preterm infant gut microbiota mobilome involving genes encoding ESBL.
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The following annotated sequences were deposited in the NCBI database. IncFIB from L-II was deposited with accession number MH422552, IncI1 from L-II with accession number MH422553, P1 bacteriophage from L-II with accession number MH445381, and P1 from transconjugant 2 (L-II) with accession number MH445380. Sequencing treads for the shotgun sequence data for the isolated bacterial strains and the transconjugants are deposited in Sequence Read Archive (SRA) with accession number SRP148649.
Tacconelli, E. & Magrini, N. Global Priority List of Antibiotic-resistant Bacteria to Guide Research, Discovery, and Development of New Antibiotics (World Health Organization, Geneva, 2017).
Logan, L. K. & Weinstein, R. A. The epidemiology of carbapenem-resistant enterobacteriaceae: the impact and evolution of a global menace. J. Infect. Dis. 215, S28–S36 (2017).
Brito, I. L. et al. Mobile genes in the human microbiome are structured from global to individual scales. Nature 535, 435−439 (2016).
Ravi, A. et al. Association of the gut microbiota mobilome with hospital location and birth weight in preterm infants. Pediatr. Res. 82, 829–838 (2017).
Morrow, A. L. et al. Early microbial and metabolomic signatures predict later onset of necrotizing enterocolitis in preterm infants. Microbiome 1, 13 (2013).
Westerbeek, E. A. et al. The intestinal bacterial colonisation in preterm infants: a review of the literature. Clin. Nutr. 25, 361–368 (2006).
Ward, D. V. et al. Metagenomic sequencing with strain-level resolution implicates uropathogenic E. coli in necrotizing enterocolitis and mortality in preterm infants. Cell Rep. 14, 2912–2924 (2016).
Wang, Y. et al. 16S rRNA gene-based analysis of fecal microbiota from preterm infants with and without necrotizing enterocolitis. ISME J. 3, 944–954 (2009).
Fernandez-Lopez, R., Redondo, S., Garcillan-Barcia, M. P. & de la Cruz, F. Towards a taxonomy of conjugative plasmids. Curr. Opin. Microbiol 38, 106–113 (2017).
Smillie, C., Garcillan-Barcia, M. P., Francia, M. V., Rocha, E. P. & de la Cruz, F. Mobility of plasmids. Microbiol. Mol. Biol. Rev. 74, 434–452 (2010).
Kovalevskaya, N. P. Mobile gene cassettes and integrons. Mol. Biol. 36, 196–201 (2002).
Rumnieks, J. & Tars, K. Diversity of pili-specific bacteriophages: genome sequence of IncM plasmid-dependent RNA phage M. BMC Microbiol. 12, 277 (2012).
Carattoli, A. Resistance plasmid families in Enterobacteriaceae. Antimicrob. Agents Chemother. 53, 2227–2238 (2009).
Thomas, C. M. Paradigms of plasmid organization. Mol. Microbiol. 37, 485–491 (2000).
Bauer, A. W., Kirby, W. M., Sherris, J. C. & Turck, M. Antibiotic susceptibility testing by a standardized single disk method. Am. J. Clin. Pathol. 45, 493–496 (1966).
Matuschek, E., Brown, D. F. J. & Kahlmeter, G. Development of the EUCAST disk diffusion antimicrobial susceptibility testing method and its implementation in routine microbiology laboratories. Clin. Microbiol. Infect. 20, O255–O266 (2014).
Zaman, M. A., Pasha, M. H. & Akhter, M. Plasmid curing of Escherichia coli cells with ethidium bromide, sodium dodecyl sulfate and acridine orange. Banglad. J. Microbiol. 27, 28–31 (2010).
Levi, K., Rynge, M., Abeysinghe, E. A., & Edwards, R. Searching the sequence read archive using Jetstream and Wrangler. in PEARC '18: Proceedings of the Practice and Experience on Advanced Research Computing, Pittsburgh, PA, USA, July 22–26 (ACM, New York, 2018).
Torres, P. J., Edwards, R. A. & McNair, K. A. PARTIE: a partition engine to separate metagenomic and amplicon projects in the Sequence Read Archive. Bioinformatics 33, 2389–2391 (2017).
Langmead, B. & Salzberg, S. L. Fast gapped-read alignment with Bowtie 2. Nat. Methods 9, 357–359 (2012).
Towns, J., Cockerill, T., Dahan, M. et al. XSEDE: accelerating scientific discovery. Comput. Sci. Eng. 16, 62–74 (2014).
Lambrecht, E., Van Meervenne, E. & Boon, N. et al. Characterization of cefotaxime- and ciprofloxacin-resistant commensal Escherichia coli originating from belgian farm animals indicates high antibiotic resistance transfer rates. Microb. Drug Resist. 24, 707–717 (2018).
Ravi, A., Avershina, E. & Foley, S. L. et al. The commensal infant gut meta-mobilome as a potential reservoir for persistent multidrug resistance integrons. Sci. Rep. 5, 15317 (2015).
Ravi, A., Valdes-Varela, L., Gueimonde, M. & Rudi, K. Transmission and persistence of IncF conjugative plasmids in the gut microbiota of full-term infants. FEMS Microbiol. Ecol. 94, fix158 (2018).
Smillie, C. S. et al. Ecology drives a global network of gene exchange connecting the human microbiome. Nature 480, 241–244 (2011).
Leverstein-van Hall, M. A., Dierikx, C. M., Cohen Stuart, J. et al. Dutch patients, retail chicken meat and poultry share the same ESBL genes, plasmids and strains. Clin. Microbiol. Infect. 17, 873–880 (2011).
Hughes, D. & Andersson, D. I. Persistence of antibiotic resistance in bacterial populations. FEMS Microbiol. Rev. 35, 901–911 (2011).
Sidjabat, H. E. et al. Expansive spread of IncI1 plasmids carrying blaCMY-2 amongst Escherichia coli. Int. J. Antimicrob. Agents 44, 203–208 (2014).
Riccobono, E. et al. Characterization of IncI1 sequence type 71 epidemic plasmid lineage responsible for the recent dissemination of CTX-M-65 extended-spectrum beta-lactamase in the Bolivian Chaco region. Antimicrob. Agents Chemother. 59, 5340–5347 (2015).
Madec, J. Y., Haenni, M., Metayer, V., Saras, E. & Nicolas-Chanoine, M. H. High prevalence of the animal-associated bla CTX-M-1 IncI1/ST3 plasmid in human Escherichia coli isolates. Antimicrob. Agents Chemother. 59, 5860–5861 (2015).
Madec, J. Y. Sequence type 48 Escherichia coli carrying the blaCTX-M-1 IncI1/ST3 plasmid in drinking water in France. Antimicrob. Agents Chemother. 60, 6430–6432 (2016).
Norizuki, C. et al. Specific blaCTX-M-8/IncI1 plasmid transfer among genetically diverse Escherichia coli isolates between humans and chickens. Antimicrob. Agents Chemother. 61, e00663-17 (2017).
Lopatkin, A. J. et al. Persistence and reversal of plasmid-mediated antibiotic resistance. Nat. Commun. 8, 1689 (2017).
Liebert, C. A., Hall, R. M. & Summers, A. O. Transposon Tn21, flagship of the floating genome. Microbiol. Mol. Biol. Rev. 63, 507–522 (1999).
Shintani, M., Sanchez, Z. K. & Kimbara, K. Genomics of microbial plasmids: classification and identification based on replication and transfer systems and host taxonomy. Front. Microbiol. 6, 242 (2015).
Gaimster, H. & Summers, D. Plasmids in the driving seat: the regulatory RNA Rcd gives plasmid ColE1 control over division and growth of its E. coli host. Plasmid 78, 59–64 (2015).
Yarmolinsky, M. B. Bacteriophage P1 in retrospect and in prospect. J. Bacteriol. 186, 7025–7028 (2004).
Yang, L. et al. Characterization of a P1-like bacteriophage carrying CTX-M-27 in Salmonella spp. resistant to third generation cephalosporins isolated from pork in China. Sci. Rep. 7, 40710 (2017).
Park, H. et al. The success of fecal microbial transplantation in Clostridium difficile infection correlates with bacteriophage relative abundance in the donor: a retrospective cohort study. Gut Microbes. (2019). https://doi.org/10.1080/19490976.2019.1586037.
Babicki, S., Arndt, D. & Marcu, A. et al. Heatmapper: web-enabled heat mapping for all. Nucleic Acids Res. 44, W147–W153 (2016).
We thank Norwegian University of Life Sciences for the financial support, and the Spanish Ministry of Science and Universities for the grant AGL2015-70487-P. Travels and stays in Spain and Norway for this study were supported by EEA Coordinated Mobility of Researchers NILS Science and Sustainability Project 017-CM-01-2013.
M.V., M.C.C., A.R., and G.P.-M. collected the clinical material and isolated the bacteria. M.H. did the main transmission experiments. I.L.A., A.R., and J.L. contributed with the sequencing and sequence interpretation. M.S., S.L.F., and D.B.D. contributed with the phenotypic characterization of the strains. K.R. wrote the paper with input from all the coauthors.
The authors declare no competing interests.
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Hagbø, M., Ravi, A., Angell, I. et al. Experimental support for multidrug resistance transfer potential in the preterm infant gut microbiota. Pediatr Res 88, 57–65 (2020). https://doi.org/10.1038/s41390-019-0491-8
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