Targeted IS-element sequencing uncovers transposition dynamics during selective pressure in enterococci

Abstract

Insertion sequences (IS) are simple transposons implicated in the genome evolution of diverse pathogenic bacterial species. Enterococci have emerged as important human intestinal pathogens with newly adapted virulence potential and antibiotic resistance. These genetic features arose in tandem with large-scale genome evolution mediated by mobile elements. Pathoadaptation in enterococci is thought to be mediated in part by the IS element IS256 through gene inactivation and recombination events. However, the regulation of IS256 and the mechanisms controlling its activation are not well understood. Here, we adapt an IS256-specfic deep sequencing method to describe how chronic lytic phage infection drives a widespread expansion of IS256 in E. faecalis and how antibiotic exposure is associated with IS256 expansion in both E. faecalis and E. faecium during a clinical human infection. We show through comparative genomics that IS256 is primarily found in hospital-adapted enterococcal isolates. IS256 transposase gene expression analyses reveal that IS256 mobility is regulated at the transcriptional and translational levels in E. faecalis, indicating tight control of IS256 activation in the absence of selective pressure. Our findings reveal that stressors such as phages and antibiotic exposure drives rapid genome-scale transposition in the enterococci. IS256 expansion can therefore explain how selective pressures mediate evolution of the enterococcal genome, ultimately leading to the emergence of dominant nosocomial lineages that threaten human health.

A metagenomics pipeline reveals insertion sequence-driven evolution of the microbiota

“…In addition to metabolic and MGE system targeting by IS elements, another common iORF was ermF, a macrolide resistance gene that acts by methylating the 23S rRNA subunit to inhibit protein translation (78). We previously found that gentamicin resistance operons are frequently interrupted by IS elements during enterococcal infection (19). We presume that in the absence of antibiotic pressure, bacteria can dampen their effort toward maintaining high-level antibiotic resistance at the population level to favor their fitness.…”

“…Antibiotic exposure enriches IS insertions in E. faecium during a chronic human infection and antibiotics can stimulate IS element activity in Staphylococcus aureus (19,87). In each of these scenarios this activity was linked to the IS element IS256, which is strongly associated with pathogenic and hospital-associated bacteria (18).…”

“…Following the growth of these transformants, we isolated their genomic DNA and performed IS-Seq, a technique that enriches for ISOSDB412 amplicons during Illumina DNA sequencing library construction and read mapping to identify specific ISOSDB412 insertion locations (19).…”

“…Genomic DNA was extracted from 1.5 mL cultures of Bacteroides strains using the Zymobionics DNA Miniprep kit (Zymo Research, Irvine, CA) and used as input for IS-Seq as described previously 19 . Briefly, NGS libraries were produced using the Illumina DNA Prep Kit (Illumina, San Diego, CA) and amplified using ISOSDB412 IS-Seq Step1 and p7 primers for 13 cycles using Q5 Master Mix (New England Biolabs, Ipswich, MA).…”

“…This analysis revealed that Bacteroides caccae gained and lost numerous IS insertions over the course of sampling (20). Finally, we previously utilized a targeted IS element sequencing approach to identify novel insertions of the IS element IS256 in intestinal Enterococcus faecium populations isolated from an individual undergoing multiple antibiotic treatment (19). Specific antibiotics treatment were associated with increased IS256 abundances and enrichment of IS256 insertions in genes related to antibiotic resistance and virulence, suggesting that antibiotic treatment drives IS-mediated pathoadaptation in the human intestine.…”