Chromosome and Plasmids of the Tick-Borne Relapsing Fever Agent
                    Borrelia hermsii

The zoonotic pathogen Borrelia hermsii bears its multiple paralogous genes for variable antigens on several linear plasmids. Application of combined long-read and short-read next-generation sequencing provided complete sequences for antigen-encoding plasmids as well as other linear and circular plasmids and the linear chromosome of the genome.

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Filling the gap of short-read next generation sequencing in PGD by long-read approach

10.26226/morressier.5af300b2738ab10027aa98ef ◽

2018 ◽

Author(s):

Dona Ngar Yin Ho

Keyword(s):

Next Generation Sequencing ◽

Next Generation ◽

Short Read ◽

Long Read ◽

Generation Sequencing

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Potential of whole-genome sequencing-based pharmacogenetic profiling

Pharmacogenomics ◽

10.2217/pgs-2020-0155 ◽

2021 ◽

Author(s):

Sylvan Manuel Caspar ◽

Timo Schneider ◽

Patricia Stoll ◽

Janine Meienberg ◽

Gabor Matyas

Keyword(s):

Next Generation Sequencing ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Whole Genome ◽

Next Generation ◽

Short Read ◽

Long Read ◽

Improving Patient Care ◽

Gene Diagnostics ◽

Generation Sequencing

Pharmacogenetics represents a major driver of precision medicine, promising individualized drug selection and dosing. Traditionally, pharmacogenetic profiling has been performed using targeted genotyping that focuses on common/known variants. Recently, whole-genome sequencing (WGS) is emerging as a more comprehensive short-read next-generation sequencing approach, enabling both gene diagnostics and pharmacogenetic profiling, including rare/novel variants, in a single assay. Using the example of the pharmacogene CYP2D6, we demonstrate the potential of WGS-based pharmacogenetic profiling as well as emphasize the limitations of short-read next-generation sequencing. In the near future, we envision a shift toward long-read sequencing as the predominant method for gene diagnostics and pharmacogenetic profiling, providing unprecedented data quality and improving patient care.

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Improved variant discovery through local re-alignment of short-read next-generation sequencing data using SRMA

Genome Biology ◽

10.1186/gb-2010-11-10-r99 ◽

2010 ◽

Vol 11 (10) ◽

Cited By ~ 53

Author(s):

Nils Homer ◽

Stanley F Nelson

Keyword(s):

Next Generation Sequencing ◽

Next Generation Sequencing Data ◽

Next Generation ◽

Sequencing Data ◽

Short Read ◽

Variant Discovery ◽

Generation Sequencing

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An improved experimental pipeline for preparing circular ssDNA viruses for next-generation sequencing

10.1101/2020.09.30.321224 ◽

2020 ◽

Cited By ~ 1

Author(s):

Catherine D. Aimone ◽

J. Steen Hoyer ◽

Anna E. Dye ◽

David O. Deppong ◽

Siobain Duffy ◽

...

Keyword(s):

Next Generation Sequencing ◽

Rolling Circle Amplification ◽

Next Generation ◽

Viral Dna ◽

Single Stranded Dna ◽

Dna Viruses ◽

Short Read ◽

Rolling Circle ◽

Optimized Protocol ◽

Generation Sequencing

AbstractWe present an optimized protocol for enhanced amplification and enrichment of viral DNA for Next Generation Sequencing of begomovirus genomes. The rapid ability of these viruses to evolve threatens many crops and underscores the importance of using next generation sequencing efficiently to detect and understand the diversity of these viruses. We combined enhanced rolling circle amplification (RCA) with EquiPhi29 polymerase and size selection to generate a cost-effective, short-read sequencing method. This optimized protocol produced short-read sequencing with at least 50% of the reads mapping to the viral reference genome. We provide other insights into common misconceptions about RCA and lessons we have learned from sequencing single-stranded DNA viruses. Our protocol can be used to examine viral DNA as it moves through the entire pathosystem from host to vector, providing valuable information for viral DNA population studies, and would likely work well with other CRESS DNA viruses.HighlightsProtocol for short-read, high throughput sequencing of single-stranded DNA viruses using random primersComparison of the sequencing of total DNA versus size-selected DNAComparison of phi29 and Equiphi29 DNA polymerases for rolling circle amplification of viral single-stranded DNA genomes

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PERHAPS: Paired-End short Reads-based HAPlotyping from next-generation Sequencing data

Briefings in Bioinformatics ◽

10.1093/bib/bbaa320 ◽

2020 ◽

Author(s):

Jie Huang ◽

Stefano Pallotti ◽

Qianling Zhou ◽

Marcus Kleber ◽

Xiaomeng Xin ◽

...

Keyword(s):

Next Generation Sequencing ◽

Snp Array ◽

Simple Approach ◽

Next Generation Sequencing Data ◽

Next Generation ◽

Sequencing Data ◽

Short Read ◽

Array Data ◽

Short Reads ◽

Generation Sequencing

Abstract The identification of rare haplotypes may greatly expand our knowledge in the genetic architecture of both complex and monogenic traits. To this aim, we developed PERHAPS (Paired-End short Reads-based HAPlotyping from next-generation Sequencing data), a new and simple approach to directly call haplotypes from short-read, paired-end Next Generation Sequencing (NGS) data. To benchmark this method, we considered the APOE classic polymorphism (*1/*2/*3/*4), since it represents one of the best examples of functional polymorphism arising from the haplotype combination of two Single Nucleotide Polymorphisms (SNPs). We leveraged the big Whole Exome Sequencing (WES) and SNP-array data obtained from the multi-ethnic UK BioBank (UKBB, N=48,855). By applying PERHAPS, based on piecing together the paired-end reads according to their FASTQ-labels, we extracted the haplotype data, along with their frequencies and the individual diplotype. Concordance rates between WES directly called diplotypes and the ones generated through statistical pre-phasing and imputation of SNP-array data are extremely high (>99%), either when stratifying the sample by SNP-array genotyping batch or self-reported ethnic group. Hardy-Weinberg Equilibrium tests and the comparison of obtained haplotype frequencies with the ones available from the 1000 Genome Project further supported the reliability of PERHAPS. Notably, we were able to determine the existence of the rare APOE*1 haplotype in two unrelated African subjects from UKBB, supporting its presence at appreciable frequency (approximatively 0.5%) in the African Yoruba population. Despite acknowledging some technical shortcomings, PERHAPS represents a novel and simple approach that will partly overcome the limitations in direct haplotype calling from short read-based sequencing.

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Whole genome sequencing of Borrelia miyamotoi isolate Izh-4: reference for a complex bacterial genome

BMC Genomics ◽

10.1186/s12864-019-6388-4 ◽

2020 ◽

Vol 21 (1) ◽

Cited By ~ 4

Author(s):

Konstantin V. Kuleshov ◽

Gabriele Margos ◽

Volker Fingerle ◽

Joris Koetsveld ◽

Irina A. Goptar ◽

...

Keyword(s):

Single Molecule ◽

Bacterial Genome ◽

Borrelia Miyamotoi ◽

Whole Genome ◽

Relapsing Fever ◽

Linear Chromosome ◽

The North ◽

Long Read ◽

Variable Major Proteins ◽

Tick Vectors

Abstract Background The genus Borrelia comprises spirochaetal bacteria maintained in natural transmission cycles by tick vectors and vertebrate reservoir hosts. The main groups are represented by a species complex including the causative agents of Lyme borreliosis and relapsing fever group Borrelia. Borrelia miyamotoi belongs to the relapsing fever group of spirochetes and forms distinct populations in North America, Asia, and Europe. As all Borrelia species B. miyamotoi possess an unusual and complex genome consisting of a linear chromosome and a number of linear and circular plasmids. The species is considered an emerging human pathogen and an increasing number of human cases are being described in the Northern hemisphere. The aim of this study was to produce a high quality reference genome that will facilitate future studies into genetic differences between different populations and the genome plasticity of B. miyamotoi. Results We used multiple available sequencing methods, including Pacific Bioscience single-molecule real-time technology (SMRT) and Oxford Nanopore technology (ONT) supplemented with highly accurate Illumina sequences, to explore the suitability for whole genome assembly of the Russian B. miyamotoi isolate, Izh-4. Plasmids were typed according to their potential plasmid partitioning genes (PF32, 49, 50, 57/62). Comparing and combining results of both long-read (SMRT and ONT) and short-read methods (Illumina), we determined that the genome of the isolate Izh-4 consisted of one linear chromosome, 12 linear and two circular plasmids. Whilst the majority of plasmids had corresponding contigs in the Asian B. miyamotoi isolate FR64b, there were only four that matched plasmids of the North American isolate CT13–2396, indicating differences between B. miyamotoi populations. Several plasmids, e.g. lp41, lp29, lp23, and lp24, were found to carry variable major proteins. Amongst those were variable large proteins (Vlp) subtype Vlp-α, Vlp-γ, Vlp-δ and also Vlp-β. Phylogenetic analysis of common plasmids types showed the uniqueness in Russian/Asian isolates of B. miyamotoi compared to other isolates. Conclusions We here describe the genome of a Russian B. miyamotoi clinical isolate, providing a solid basis for future comparative genomics of B. miyamotoi isolates. This will be a great impetus for further basic, molecular and epidemiological research on this emerging tick-borne pathogen.

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