Nextseq 1000 2000 p2 reagent

A total of 70 unique CRE clinical isolates archived at the clinical microbiology laboratory of Fujita Health University Hospital were used (Table 2). Each isolate was collected from a different patient except for two non-CPE, CRE isolates (Escherichia coli and Serratia marcescens), which were collected from the same patient. The whole genome of all isolates, except for one Klebsiella pneumoniae CPE isolate, were sequenced by MiSeq or NextSeq 2000 instruments using the NexteraXT DNA library preparation kit or QIAseq FX DNA library kit (Qiagen, Hulsterweg, Netherlands) and MiSeq reagent kit v3 600 cycles or NextSeq 1000/2000 P2 reagents 200 cycles (Illumina, San Diego, CA). Sequence reads were assembled using SPAdes v3.13.1. The remaining isolate was sequenced by a MinION device using the ligation sequencing kit SQK-LSK109 (Oxford Nanopore Technologies, Oxford, UK). Sequence reads were assembled using Flye v2.7. Carbapenemase genes were detected in 51 of the 70 isolates and included bla_IMP-1 (n = 48), bla_OXA-48 (1), bla_NDM-5 (1), and bla_IMI-2 (1). The remaining 19 isolates were non-CPE, CRE isolates. Antimicrobial susceptibility testing was conducted on a Vitek 2 instrument using the AST-N229 card (bioMérieux).

Sequencing was performed on the NextSeq 2000 platform (Illumina Inc, USA) using NextSeq 1000/2000 P2 Reagents (100 Cycles) v3. PhiX Control v3 library (Illumina Inc, USA) was spiked in at a concentration of 1% to enable troubleshooting in the event of run failure. Sequencing was single‐end 1 × 75.