Journal: mBio

Article Title: Metagenomic Next-Generation Sequencing of Nasopharyngeal Specimens Collected from Confirmed and Suspect COVID-19 Patients

doi: 10.1128/mBio.01969-20

Figure Lengend Snippet: Bacterial diversity analysis of metagenomic sequencing results. (a, b, c) Alpha diversity analysis of metagenomic sequencing results. SARS-CoV-2 positivity was determined by LDT-RT-PCR. (a, b) Shannon diversity plot and Chao diversity plot of SARS-CoV-2-negative and -positive samples at the species level. (c) Shannon diversity plot of SARS-CoV-2-negative samples and SARS-CoV-2-positive samples at different periods post-onset of symptoms. (d, e, f) Beta diversity principal-coordinate analysis of metagenomic sequencing results at the species level. (d) Bray-Curtis analysis of bacterial community composition diversity between SARS-CoV-2-negative and SARS-CoV-2-positive samples. (e) Bray-Curtis analysis of bacterial community composition diversity grouped by PCR and sequencing positivity. (f) Bray-Curtis analysis of bacterial diversity in different disease severity groups. (a, b) Wilcoxon rank sum tests were performed between positive and negative SARS-CoV-2 groups for the Shannon diversity index ( P = 0.0097) and Chao diversity ( P = 0.0082). (c) Wilcoxon rank sum tests were performed between disease onset groups (no significance). (d, f) PERMANOVA tests were performed on Bray-Curtis distance matrices for SARS-CoV-2-positive and -negative groups ( P = 0.027) (d) and groups of disease severity ( P = 0.022) (f). (e) Pairwise PERMANOVA tests were performed on Bray-Curtis distance matrices between groups defined by SARS-CoV-2 positivity by RT-PCR and sequencing. RT-PCR+/sequencing+ (PCR+/Seq+) versus RT-PCR–/sequencing– (PCR–/Seq–) ( P = 0.007); PC1 and -2, principal components 1 and 2. The severity index was defined on a scale of 1 to 4, as follows: 4, not admitted; 3, admitted; 2, intensive care unit; and 1, required ventilator.

Article Snippet: Here, using direct Oxford Nanopore long-read third-generation metatranscriptomic and metagenomic sequencing of NP swab specimens from 50 patients under investigation for COVID-19, we detected SARS-CoV-2 sequences by applying the CosmosID bioinformatics platform.

Techniques: Sequencing, Reverse Transcription Polymerase Chain Reaction

Journal: Pharmacogenomics

Article Title: Sequencing the CYP2D6 gene: from variant allele discovery to clinical pharmacogenetic testing

doi: 10.2217/pgs-2017-0033

Figure Lengend Snippet: The hive plot edges display sequence similarity between CYP2D6-CYP2D7, CYP2D7-CYP2D8 and CYP2D8-CYP2D6 (clockwise from top). Three principle axes (0, 120 and 240°) of the hive plots represent the nucleotide composition of the multiple sequence alignment for the indicated gene: (A) exonic sequences (intronic sequences shown in black), (B) intronic sequences (exonic sequences shown in black) and (C) exonic and intronic sequences.

Article Snippet: The difficulties with pseudogenes and CNVs that are inherent to short-read CYP2D6 sequencing prompted the recent development of third-generation long-read CYP2D6 sequencing using Oxford Nanopore and PacBio.

Techniques: Sequencing

Journal: Pharmacogenomics

Article Title: Sequencing the CYP2D6 gene: from variant allele discovery to clinical pharmacogenetic testing

doi: 10.2217/pgs-2017-0033

Figure Lengend Snippet: Median read coverage of ExAC exomes; base pair resolution of three functional CYP2D6 transcript isoforms; and base pair resolution of the intersection with 'reliable genome' intervals.

Article Snippet: The difficulties with pseudogenes and CNVs that are inherent to short-read CYP2D6 sequencing prompted the recent development of third-generation long-read CYP2D6 sequencing using Oxford Nanopore and PacBio.

Techniques: Functional Assay

Journal: Pharmacogenomics

Article Title: Sequencing the CYP2D6 gene: from variant allele discovery to clinical pharmacogenetic testing

doi: 10.2217/pgs-2017-0033

Figure Lengend Snippet: Commercially available CYP2D6 genotyping and sequencing tests.

Article Snippet: The difficulties with pseudogenes and CNVs that are inherent to short-read CYP2D6 sequencing prompted the recent development of third-generation long-read CYP2D6 sequencing using Oxford Nanopore and PacBio.

Techniques: Sequencing, Luminex

Journal: Pharmacogenomics

Article Title: Sequencing the CYP2D6 gene: from variant allele discovery to clinical pharmacogenetic testing

doi: 10.2217/pgs-2017-0033

Figure Lengend Snippet: Results for NA12878 (CYP2D6*3/*4) are displayed from top to bottom panels for WGS from the 1000 Genomes Project, in-house WGS, WES, targeted capture with the PGRNseq platform, targeted PacBio CYP2D6 sequencing, and ALEC-corrected targeted PacBio CYP2D6 sequencing. Of note, discrepant and skewed allele frequencies in several loci from the WGS data indicate potential read misalignment errors. Moreover, the common CYP2D6 capture strategies (e.g., WES, PGRNseq) coupled with short-read Illumina sequencing result in significant read assignment to the CYP2D7 and CYP2D8 pseudogenes. These reads indicate a lack of specificity for CYP2D6 by these target enrichment approaches and/or informatic errors related to read misalignment. Targeted PacBio sequencing results in CYP2D6-specific sequencing and no misalignment to CYP2D7 or CYP2D8, but random errors throughout the sequencing reads are characteristic to this technology. These random errors can be minimized by circular consensus sequencing read analysis; however, further correction prior to variant calling can also be accomplished by available informatics tools (e.g., Amplicon Long-read Error Correction [ALEC]).

Article Snippet: The difficulties with pseudogenes and CNVs that are inherent to short-read CYP2D6 sequencing prompted the recent development of third-generation long-read CYP2D6 sequencing using Oxford Nanopore and PacBio.

Techniques: Sequencing, Illumina Sequencing, PacBio Sequencing, Variant Assay, Amplification

Journal: Pharmacogenomics

Article Title: Sequencing the CYP2D6 gene: from variant allele discovery to clinical pharmacogenetic testing

doi: 10.2217/pgs-2017-0033

Figure Lengend Snippet: Example variant CYP2D6 alleles and the ‘activity score’ framework. †

Article Snippet: The difficulties with pseudogenes and CNVs that are inherent to short-read CYP2D6 sequencing prompted the recent development of third-generation long-read CYP2D6 sequencing using Oxford Nanopore and PacBio.

Techniques: Variant Assay, Activity Assay