Hybrid Assembly of vanA-Encoding Plasmids

Long-read MinION and paired-end short-read MiSeq FASTQ files were used for hybrid assemblies using Unicycler (v0.4.8).²² (link) The genetic organization of vanA-encoding plasmids from Irish isolates was determined following hybrid assembly and these were used as reference sequences for further analysis. MiSeq reads were mapped against reference plasmid sequences and percentage depth and breadth of coverage calculated using Burrows–Wheeler Aligner, SAMtools and BedTools.^23–25 (link) Alignment quality was assessed using Tablet.²⁶ (link) vanA-encoding genomes resolved by hybrid assembly were annotated using RAST v2.0 (http://rast.nmpdr.org/)²⁷ (link) and visualized using SnapGene (GSL Biotech; SnapGene.com">https://SnapGene.com). Irish VREfm and VSEfm sequence reads and sequences resolved by hybrid assembly have been deposited in GenBank under BioProject PRJNA734127. Danish VREfm sequence reads have been deposited in GenBank under BioProjects PRJNA573568, PRJNA686881, PRJNA691722, PRJNA702038 and PRJNA740173.

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Egan S.A., Kavanagh N.L., Shore A.C., Mollerup S., Samaniego Castruita J.A., O’Connell B., McManus B.A., Brennan G.I., Pinholt M., Westh H, & Coleman D.C. (2021). Genomic analysis of 600 vancomycin-resistant Enterococcus faecium reveals a high prevalence of ST80 and spread of similar vanA regions via IS1216E and plasmid transfer in diverse genetic lineages in Ireland. Journal of Antimicrobial Chemotherapy, 77(2), 320-330.

Publication 2021

SnapGene

Genomes Hybrid Plasmid Rast Sequences analysis Tablet

Corresponding Organization : Dublin Dental University Hospital

Other organizations : Hvidovre Hospital, St. James's Hospital

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Variable analysis

independent variables

Long-read MinION and paired-end short-read MiSeq FASTQ files

dependent variables

Genetic organization of vanA-encoding plasmids
Percentage depth and breadth of coverage of MiSeq reads mapped against reference plasmid sequences

control variables

Unicycler (v0.4.8) used for hybrid assemblies
Burrows–Wheeler Aligner, SAMtools and BedTools used for mapping and coverage calculations
Tablet used for assessing alignment quality
RAST v2.0 used for genome annotation
SnapGene used for visualization

Annotations

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