An accessible, efficient and global approach for the large-scale sequencing of bacterial genomes

Perez-Sepulveda, Blanca; Heavens, Darren; Pulford, Caisey V; Predeus, Alexander V.; Low, Ross; Webster, Hermione J.; Dykes, Gregory F.; Schudoma, Christian; Rowe, Will; Lipscombe, James; Watkins, Christopher B.; Kumwenda, Benjamin; Shearer, Neil; Costigan, Karl; Baker, Kate S.; Feasey, Nicholas; Hinton, Jay C. D.; Hall, Neil

doi:10.1186/s13059-021-02536-3

Cited by 36 publications

(34 citation statements)

References 53 publications

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“…Second phase: Determination of class 1 integrons by WGS . Genomic DNA extraction, library preparation and WGS for 225 clinical S. Typhimurium isolates were performed as part of the 10KSG consortium and were described previously (Perez-Sepulveda et al ., 2020). For 9 clinical isolates the WGS was made with Miseq Illumina platform.…”

Section: Methodsmentioning

confidence: 99%

“…De Novo assembly were performed with CLC Genomics Workbench (Qiagen) version 8.0.1. “Rapid Annotation using Subsystem Technology, RAST” version 2.0 and SEED Viewer (http://rast.nmpdr.org/seedviewer.cgi) were used for annotation of genomes and with the pipeline available from https://github.com/apredeus/10k_genomes (Perez-Sepulveda et al ., 2020).…”

Section: Methodsmentioning

confidence: 99%

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Class 1 integrons in clinical and swine industry isolates of Salmonella Typhimurium from Colombia, dating 1997 to 2017

Flórez-Delgado

Ubillus

Perez-Sepulveda

et al. 2022

Preprint

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BackgroundSalmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium) has been linked to outbreaks of foodborne gastroenteritis disease, and the emergence of antimicrobial-resistant clones. In Colombia, laboratory surveillance of Salmonella spp. between 1997-2018 revealed that S. Typhimurium was the most ubiquitous serovar (27.57% of all Salmonella isolates), with increasing levels of antimicrobial resistance to several families of antibiotics. Hypothesis. Resistant isolates of S. Typhimurium recovered from clinical and swine samples carry class 1 integrons that are linked to antimicrobial resistance genes. Aim. Identify class 1 integrons, and investigate their association with other mobile genetic elements, and their relationship to the antimicrobial resistance of Colombian S. Typhimurium isolates. Methods. In this study, 392 clinical isolates of S. Typhimurium were analyzed, of which 237 were obtained from blood culture, 155 from non-invasive sources and 50 isolates from swine of which 32 were obtained from a slaughterhouse, 10 from a point of sale and 8 from cutting plant. Class 1 integrons and plasmid incompatibility groups were analyzed by PCR and Whole Genome Sequencing (WGS), and the region surrounding of the integrons identified by WGS. The phylogenetic relationship was established by MLST and SNP analysis. Results. Overall, 39.03% (153/392) of the clinical isolates and 22% (11/50) of the swine S. Typhimurium isolates carried complete class 1 integrons. Twelve types of gene cassette arrays were identified, including dfr7-aac-blaOXA-2 (Int1-Col1) as the most frequent in clinical isolates (75.2% = 115/153). Clinical and swine isolates that carried class 1 integrons were resistant to up to 5 and up to 3 antimicrobial families, respectively. The Int1-Col1 integron was most prevalent in stool isolates and was associated with Tn21. The most common plasmid incompatibility group was IncA/C. Conclusions. The widespread presence of the integron (IntI1-Col1) in Colombia since 1997 was striking. We speculate that the relationship between integrons, source and mobile elements favors the dispersion of antibiotic resistance determinants in Colombian S. Typhimurium isolates.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Class 1 integrons in clinical and swine industry isolates of Salmonella Typhimurium from Colombia, dating 1997 to 2017

Flórez-Delgado

Ubillus

Perez-Sepulveda

et al. 2022

Preprint

Self Cite

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“…With continual improvements in sequencing length, accuracy, convenience, and affordability, Illumina platform sequencers have emerged as a dominant force in short-read sequencing. For example, a recent large-scale global study of more than 10,000 Salmonella genomes was achieved with Illumina sequencing, where the total cost of consumables (DNA extraction and genome sequence generation—excluding staff time) was under USD 10 per sample [ 17 ]. However, the limitations due to read length, now typically 75–300 bp, and the requirement for template sequence amplification prior to sequencing do still apply.…”

Section: Short-read Sequencing Technologies Have Enabled the Systemat...mentioning

confidence: 99%

“…The relative strengths and weaknesses of the second- and third-generation sequencing technologies mean that one platform, or the other or even a hybrid approach, may be more suitable to a certain research project. Studies requiring whole-genome sequencing (WGS) of a large number of related strains can benefit from the accuracy and cost-effectiveness of short-read sequencing [ 17 , 24 ], but can additionally include a long-read platform to gap-close if the aim is to also produce more complete genome sequences or analyse the plasmid content [ 25 ]. According to the metadata that we mined from PATRIC (March 2022; Figure 1 ), the most frequently used platform when sequencing a bacterial pathogen with only one technology is Illumina (~91% of sequenced strains), followed by PacBio (5%).…”

Section: The Promise Of Long-read Sequencing Technologies and Hybrid ...mentioning

confidence: 99%

The Notable Achievements and the Prospects of Bacterial Pathogen Genomics

2022

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Throughout the entirety of human history, bacterial pathogens have played an important role and even shaped the fate of civilizations. The application of genomics within the last 27 years has radically changed the way we understand the biology and evolution of these pathogens. In this review, we discuss how the short- (Illumina) and long-read (PacBio, Oxford Nanopore) sequencing technologies have shaped the discipline of bacterial pathogen genomics, in terms of fundamental research (i.e., evolution of pathogenicity), forensics, food safety, and routine clinical microbiology. We have mined and discuss some of the most prominent data/bioinformatics resources such as NCBI pathogens, PATRIC, and Pathogenwatch. Based on this mining, we present some of the most popular sequencing technologies, hybrid approaches, assemblers, and annotation pipelines. A small number of bacterial pathogens are of very high importance, and we also present the wealth of the genomic data for these species (i.e., which ones they are, the number of antimicrobial resistance genes per genome, the number of virulence factors). Finally, we discuss how this discipline will probably be transformed in the near future, especially by transitioning into metagenome-assembled genomes (MAGs), thanks to long-read sequencing.

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“…The majority of isolates (n=67) were processed at the University of Liverpool (UK) and sequenced at the Earlham Institute (UK). DNA extraction and sequencing were carried out using an optimized method for large-scale sequencing [30], including the bespoke LITE (Low Input, Transposase Enabled) pipeline for library construction, and Illumina HiSeq sequencing technology. The remaining isolates (n=33) were processed and sequenced at the MRCG.…”

Section: Dna Extraction and Whole Genome Sequencingmentioning

confidence: 99%

Genomic diversity and antimicrobial resistance among non-typhoidal Salmonella associated with human disease in The Gambia

et al. 2022

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Non-typhoidal Salmonella associated with multidrug resistance cause invasive disease in sub-Saharan Africa. Specific lineages of serovars Typhimurium and Enteritidis have been implicated. Here we characterized the genomic diversity of 100 clinical non-typhoidal Salmonella collected from 93 patients in 2001 from the eastern, and in 2006–2018 from the western regions of The Gambia respectively. A total of 93 isolates (64 invasive, 23 gastroenteritis and six other sites) representing a single infection episode were phenotypically tested for antimicrobial susceptibility using the Kirby–Bauer disc diffusion technique. Whole genome sequencing of 100 isolates was performed using Illumina, and the reads were assembled and analysed using SPAdes. The Salmonella in Silico Typing Resource (SISTR) was used for serotyping. SNP differences among the 93 isolates were determined using Roary, and phylogenetic analysis was performed in the context of 495 African strains from the European Nucleotide Archive. Salmonella serovars Typhimurium (26/64; 30.6 %) and Enteritidis (13/64; 20.3 %) were associated with invasive disease, whilst other serovars were mainly responsible for gastroenteritis (17/23; 73.9 %). The presence of three major serovar Enteritidis clades was confirmed, including the invasive West African clade, which made up more than half (11/16; 68.8 %) of the genomes. Multidrug resistance was confined among the serovar Enteritidis West African clade. The presence of this epidemic virulent clade has potential for spread of resistance and thus important implications for systematic patient management. Surveillance and epidemiological investigations to inform control are warranted.

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An accessible, efficient and global approach for the large-scale sequencing of bacterial genomes

Cited by 36 publications

References 53 publications

Class 1 integrons in clinical and swine industry isolates of Salmonella Typhimurium from Colombia, dating 1997 to 2017

Class 1 integrons in clinical and swine industry isolates of Salmonella Typhimurium from Colombia, dating 1997 to 2017

The Notable Achievements and the Prospects of Bacterial Pathogen Genomics

Genomic diversity and antimicrobial resistance among non-typhoidal Salmonella associated with human disease in The Gambia

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