PMID- 31508405
OWN - NLM
STAT- PubMed-not-MEDLINE
LR  - 20201001
IS  - 2296-2565 (Print)
IS  - 2296-2565 (Electronic)
IS  - 2296-2565 (Linking)
VI  - 7
DP  - 2019
TI  - Evaluation of Rapid Library Preparation Protocols for Whole Genome Sequencing 
      Based Outbreak Investigation.
PG  - 241
LID - 10.3389/fpubh.2019.00241 [doi]
LID - 241
AB  - Whole genome sequencing (WGS) has become the new gold standard for bacterial 
      outbreak investigation, due to the high resolution available for typing. While 
      sequencing is currently predominantly performed on Illumina devices, the 
      preceding library preparation can be performed using various protocols. Enzymatic 
      fragmentation library preparation protocols are fast, have minimal hands-on time, 
      and work with small quantities of DNA. The aim of our study was to compare three 
      library preparation protocols for molecular typing: Nextera XT (Illumina); 
      Nextera Flex (Illumina); and QIAseq FX (Qiagen). We selected 12 ATCC strains from 
      human Gram-positive and Gram-negative pathogens with %G+C-content ranging from 
      27% (Fusobacterium nucleatum) to 73% (Micrococcus luteus), each having a high 
      quality complete genome assembly available, to allow in-depth analysis of the 
      resulting Illumina sequence data quality. Additionally, we selected isolates from 
      previously analyzed cases of vancomycin-resistant Enterococcus faecium (VRE) (n = 
      7) and a local outbreak of Klebsiella aerogenes (n = 5). The number of protocol 
      steps and time required were compared, in order to test the suitability for 
      routine laboratory work. Data analyses were performed with standard tools 
      commonly used in outbreak situations: Ridom SeqSphere+ for cgMLST; CLC genomics 
      workbench for SNP analysis; and open source programs. Nextera Flex and QIAseq FX 
      were found to be less sensitive than Nextera XT to variable %G+C-content, 
      resulting in an almost uniform distribution of read-depth. Therefore, low 
      coverage regions are reduced to a minimum resulting in a more complete 
      representation of the genome. Thus, with these two protocols, more alleles were 
      detected in the cgMLST analysis, producing a higher resolution of closely related 
      isolates. Furthermore, they result in a more complete representation of accessory 
      genes. In particular, the high data quality and relative simplicity of the 
      workflow of Nextera Flex stood out in this comparison. This thorough comparison 
      within an ISO/IEC 17025 accredited environment will be of interest to those 
      aiming to optimize their clinical microbiological genome sequencing.
FAU - Seth-Smith, Helena M B
AU  - Seth-Smith HMB
AD  - Division of Clinical Bacteriology and Mycology, University Hospital Basel, Basel, 
      Switzerland.
AD  - Applied Microbiology Research, Department of Biomedicine, University of Basel, 
      Basel, Switzerland.
AD  - DBM Bioinformatics Core Facility, SIB Swiss Institute of Bioinformatics, Basel, 
      Switzerland.
FAU - Bonfiglio, Ferdinando
AU  - Bonfiglio F
AD  - Applied Microbiology Research, Department of Biomedicine, University of Basel, 
      Basel, Switzerland.
AD  - Personalized Health Basel, University of Basel, Basel, Switzerland.
FAU - Cuenod, Aline
AU  - Cuenod A
AD  - Division of Clinical Bacteriology and Mycology, University Hospital Basel, Basel, 
      Switzerland.
AD  - Applied Microbiology Research, Department of Biomedicine, University of Basel, 
      Basel, Switzerland.
FAU - Reist, Josiane
AU  - Reist J
AD  - Applied Microbiology Research, Department of Biomedicine, University of Basel, 
      Basel, Switzerland.
FAU - Egli, Adrian
AU  - Egli A
AD  - Division of Clinical Bacteriology and Mycology, University Hospital Basel, Basel, 
      Switzerland.
AD  - Applied Microbiology Research, Department of Biomedicine, University of Basel, 
      Basel, Switzerland.
FAU - Wuthrich, Daniel
AU  - Wuthrich D
AD  - Division of Clinical Bacteriology and Mycology, University Hospital Basel, Basel, 
      Switzerland.
AD  - Applied Microbiology Research, Department of Biomedicine, University of Basel, 
      Basel, Switzerland.
AD  - DBM Bioinformatics Core Facility, SIB Swiss Institute of Bioinformatics, Basel, 
      Switzerland.
LA  - eng
PT  - Journal Article
DEP - 20190827
PL  - Switzerland
TA  - Front Public Health
JT  - Frontiers in public health
JID - 101616579
PMC - PMC6719548
OTO - NOTNLM
OT  - Illumina
OT  - NGS
OT  - bacteria
OT  - comparison
OT  - library
OT  - next generation sequencing
OT  - prokaryotes
OT  - whole genome sequencing
EDAT- 2019/09/12 06:00
MHDA- 2019/09/12 06:01
PMCR- 2019/08/27
CRDT- 2019/09/12 06:00
PHST- 2019/04/09 00:00 [received]
PHST- 2019/08/12 00:00 [accepted]
PHST- 2019/09/12 06:00 [entrez]
PHST- 2019/09/12 06:00 [pubmed]
PHST- 2019/09/12 06:01 [medline]
PHST- 2019/08/27 00:00 [pmc-release]
AID - 10.3389/fpubh.2019.00241 [doi]
PST - epublish
SO  - Front Public Health. 2019 Aug 27;7:241. doi: 10.3389/fpubh.2019.00241. 
      eCollection 2019.