Analysis of the ARTIC version 3 and version 4 SARS-CoV-2 primers and their impact on the detection of the G142D amino acid substitution in the spike protein

The ARTIC Network provides a common resource of PCR primer sequences and recommendations for amplifying SARS-CoV-2 genomes. The initial tiling strategy was developed with the reference genome Wuhan-01, and subsequent iterations have addressed areas of low amplification and sequence drop out. Recently, a new version (V4) was released, based on new variant genome sequences, in response to the realization that some V3 primers were located in regions with key mutations. Herein, we compare the performance of the ARTIC V3 and V4 primer sets with a matched set of 663 SARS-CoV-2 clinical samples sequenced with an Illumina NovaSeq 6000 instrument. We observe general improvements in sequencing depth and quality, and improved resolution of the SNP causing the D950N variation in the spike protein. Importantly, we also find nearly universal presence of spike protein substitution G142D in Delta-lineage samples. Due to the prior release and widespread use of the ARTIC V3 primers during the initial surge of the Delta variant, it is likely that the G142D amino acid substitution is substantially underrepresented among early Delta variant genomes deposited in public repositories. In addition to the improved performance of the ARTIC V4 primer set, this study also illustrates the importance of the primer scheme in downstream analyses.

Download Full-text

Analysis of the ARTIC Version 3 and Version 4 SARS-CoV-2 Primers and Their Impact on the Detection of the G142D Amino Acid Substitution in the Spike Protein

Microbiology Spectrum ◽

10.1128/spectrum.01803-21 ◽

2021 ◽

Author(s):

James J. Davis ◽

S. Wesley Long ◽

Paul A. Christensen ◽

Randall J. Olsen ◽

Robert Olson ◽

...

Keyword(s):

Amino Acid ◽

Amino Acid Substitution ◽

Spike Protein ◽

Clinical Samples

ARTIC Network primers are commonly used by laboratories worldwide to amplify and sequence SARS-CoV-2 present in clinical samples. As new variants have evolved and spread, it was found that the V3 primer set poorly amplified several key mutations.

Download Full-text

Evaluation of a new variant in the aggrecan gene potentially associated with chondrodysplastic dwarfism in Miniature horses

Scientific Reports ◽

10.1038/s41598-020-72192-3 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Danilo Giorgi Abranches de Andrade ◽

Roberta Martins Basso ◽

Angelo José Magro ◽

Renée Laufer-Amorim ◽

Alexandre Secorun Borges ◽

...

Keyword(s):

Genetic Testing ◽

Amino Acid ◽

Amino Acid Substitution ◽

Autosomal Recessive ◽

Autosomal Recessive Disorder ◽

Dwarf Phenotype ◽

Horse Genome ◽

Complex Interactions ◽

New Variant ◽

Exon 11

Abstract Chondrodysplastic dwarfism in Miniature horses is an autosomal recessive disorder previously associated with four mutations (D1, D2, D3*, and D4) in the aggrecan (ACAN) gene. The aim of this study was to identify additional variants in the candidate ACAN gene associated with chondrodysplastic dwarfism in Miniature horses. Fifteen dwarf Miniature horses were found to possess only one of the dwarfism-causing variants, and two possessed none of the variants. The ACAN exons (EquCab3.0) of seven dwarf Miniature horses were sequenced. A missense SNP in coding exon 11 (g.95271115A > T, c.6465A > T—RefSeq XM_005602799.2), which resulted in the amino acid substitution p.Leu2155Phe (RefSeq XP_005602856.2), was initially associated with the dwarf phenotype. The variant was tested and found present in 14 dwarf foals as well as one parent of each, and both parents of a dwarf possessing two copies. Genetic testing of 347 phenotypically normal Miniature horses demonstrated that none had more than one of the dwarf alleles or c.6465A > T. However, a study of large breeds revealed the presence of c.6465A > T, which was present in homozygosis in two Mangalarga Marchador horses. We suggest that c.6465A > T as a marker of disequilibrium or complex interactions in the Miniature horse genome could contribute to the associated dwarfism.

Download Full-text

Congenital Dyserythropoietic Anemia Type 1: A New Variant Pathogenic CDAN1 Mutation

Blood ◽

10.1182/blood.v126.23.4551.4551 ◽

2015 ◽

Vol 126 (23) ◽

pp. 4551-4551 ◽

Cited By ~ 1

Author(s):

Jenny McDaniel ◽

Stuart Cramer

Keyword(s):

Bone Marrow ◽

Amino Acid ◽

Amino Acid Substitution ◽

Sequence Variant ◽

Type I ◽

Red Cell ◽

Female Fetus ◽

Congenital Dyserythropoietic Anemia ◽

Bone Marrow Evaluation ◽

New Variant

Abstract Introduction: Congenital dyserythropoietic anemia (CDA) is a rare autosomal recessive condition that results in dyserythropoiesis and iron overload. Bone marrow evaluation in these patients demonstrates distinctive abnormalities in red cell precursors including multinucleated erythroblasts and chromatin bridges. Dysmorphisms such as distal limb abnormalities, skin pigmentation defects, vertebral malformations, and short stature may also be present. Mutations in the CDAN1/codanin-1 gene underlie the majority of CDA type I cases. We describe a previously unreported pathogenic variant. Case report: The female fetus of a gravida 2 para 1 woman was noted to have hepatomegaly, cardiomegaly, and elevated middle cerebral artery velocity concerning for severe intrauterine anemia at approximately 20 weeks gestation. The parents had one previous pregnancy that resulted in a stillborn male infant with hydrops fetalis and club foot. Given concern for anemia with this pregnancy, percutaneous umbilical cord blood sampling was performed demonstrating a fetal hematocrit of 9%. This severe fetal anemia was initially thought to be secondary to ABO incompatibility as mother was O- with a positive antibody screen and infant was A+. The fetus required four intrauterine transfusions and was delivered at 34 weeks 6 days via cesarean section due to history of previous cesarean. Exam revealed a phenotypically normal female with APGARs of 9/9 and a birth weight of 2130 grams. Following delivery the infant had serial blood counts performed over a 6-week period, which noted a steady decrease in her hematocrit to 16%. A transfusion was given, and a bone marrow evaluation revealed a cellular marrow with red cell hyperplasia and dyserythropoiesis. Next generation sequencing through PreventionGenetics was performed revealing a novel CDAN1 alteration. Results: The patient was found to be heterozygous for two mutations in trans in the CDAN1 gene: c.2072dupT and c.2093A>T. Discussion: We present a patient with severe fetal and infant transfusion dependent anemia who has two novel CDAN1 gene variants not previously described. The sequence variant c.2072dupT is predicted to result in a frameshift and premature protein termination and is expected to be pathogenic. The second variant c.2093A>T is predicted to result in an amino acid substitution (p.Glu698Val). Another amino acid substitution (p.Glu698Lys) was previously reported as pathogenic in an individual with congenital dyserythropoietic anemia. Based upon the clinical picture, morphologic characteristics, and genetic findings, we have concluded that this presentation is consistent with a diagnosis of CDA type I. Through the utilization of improved diagnostic techniques we continue to gain knowledge regarding the molecular underpinnings of CDA type I. Disclosures No relevant conflicts of interest to declare.

Download Full-text

AN AMINO ACID SUBSTITUTION MATRIX FOR PROTEIN CONFORMATION IDENTIFICATION

Journal of Bioinformatics and Computational Biology ◽

10.1142/s0219720006002156 ◽

2006 ◽

Vol 04 (03) ◽

pp. 769-782 ◽

Cited By ~ 3

Author(s):

XIN LIU ◽

WEI-MOU ZHENG

Keyword(s):

Amino Acid ◽

Amino Acid Substitution ◽

Substitution Matrix ◽

Structure Alignment ◽

Amino Acid Substitution Matrix ◽

Substitution Matrices ◽

The Matrix ◽

Structure Relationship ◽

Improved Performance ◽

Amino Acid Substitution Matrices

Amino acid substitution matrices play an essential role in protein sequence alignment, a fundamental task in bioinformatics. Most widely used matrices, such as PAM matrices derived from homologous sequences and BLOSUM matrices derived from aligned segments of PROSITE, did not integrate conformation information in their construction. There are a few structure-based matrices, which are derived from limited data of structure alignment. Using databases PDB_SELECT and DSSP, we create a database of sequence-conformation blocks which explicitly represent sequence-structure relationship. Members in a block are identical in conformation and are highly similar in sequence. From this block database, we derive a conformation-specific amino acid substitution matrix CBSM60. The matrix shows an improved performance in conformational segment search and homolog detection.

Download Full-text

A single amino acid substitution (R441A) in the receptor-binding domain of SARS coronavirus spike protein disrupts the antigenic structure and binding activity

Biochemical and Biophysical Research Communications ◽

10.1016/j.bbrc.2006.03.139 ◽

2006 ◽

Vol 344 (1) ◽

pp. 106-113 ◽

Cited By ~ 13

Author(s):

Yuxian He ◽

Jingjing Li ◽

Shibo Jiang

Keyword(s):

Amino Acid ◽

Receptor Binding ◽

Amino Acid Substitution ◽

Binding Activity ◽

Single Amino Acid Substitution ◽

Spike Protein ◽

Single Amino Acid ◽

Antigenic Structure ◽

Receptor Binding Domain ◽

Sars Coronavirus

Download Full-text

New Variant of CTX-M-Type Extended-Spectrum β-Lactamases, CTX-M-71, with a Gly238Cys Substitution in a Klebsiella pneumoniae Isolate from Bulgaria

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00461-09 ◽

2009 ◽

Vol 53 (10) ◽

pp. 4518-4521 ◽

Cited By ~ 8

Author(s):

Ines Schneider ◽

Anne Marie Queenan ◽

Rumyana Markovska ◽

Boyka Markova ◽

Emma Keuleyan ◽

...

Keyword(s):

Amino Acid ◽

Klebsiella Pneumoniae ◽

Amino Acid Substitution ◽

Hydrolytic Activity ◽

Extended Spectrum ◽

New Variant

ABSTRACT A single K lebsiella pneumoniae strain isolated in a Bulgarian hospital was found to produce CTX-M-71, a new CTX-M variant characterized by one amino acid substitution from glycine to cysteine at position 238 in comparison to CTX-M-15. This exchange decreased the hydrolytic activity of the β-lactamase for cefotaxime, ceftazidime, and cefepime.

Download Full-text

Evidence for the dependence of the SARS-Cov-2 Delta high diffusivity on the associated N:G215C nucleocapsid mutation

10.21203/rs.3.rs-846225/v1 ◽

2021 ◽

Author(s):

Vito Marchitelli ◽

Claudia Troise ◽

Antonio Parisi ◽

Angelica Bianco ◽

Laura Del Sambro ◽

...

Keyword(s):

Amino Acid ◽

Genome Sequencing ◽

Virus Genome ◽

Relative Increase ◽

Amino Acid Sequences ◽

Spike Protein ◽

The United Kingdom ◽

New Variant ◽

Adaptive Effect ◽

Selective Environment

Abstract The continuous evolution of the SARS-CoV-2 virus genome and the consequent substitutions observed in the amino acid sequences, can induce significant changes in parameters such as diffusivity and pathogenicity, and causes constant concern regarding the efficacy of vaccines against the new variants in circulation. In recent months there has been an increase in the number of infections first in India, and more recently in the United Kingdom. The genome sequencing of the samples showed that this increase coincided with the emergence of a new variant, B.1.617.2, also known as VOC Delta. This variant is rapidly becoming dominant in several countries, causing increasing concern for its extreme diffusivity and its ability to often overcome the vaccines. The high diffusivity of this variant is normally ascribed to the Spike protein mutations. However, we will show here that it is rather due to the nucleocapsid substitution N:G215C. This is made clear by comparing, in the genomic sequences available on the GISAID database, the relative increase of the Delta variant with and without the associated N:G215C substitution. Once the extreme diffusivity of the Delta variant with associated nucleocapsid aminoacidic substitution is evidenced, we tentatively explain it as possibly due to the adaptive effect of the highly selective environment in Countries with high levels of vaccination.

Download Full-text

The lethal triad: SARS-CoV-2 Spike, ACE2 and TMPRSS2. Mutations in host and pathogen may affect the course of pandemic

10.1101/2021.01.12.426365 ◽

2021 ◽

Author(s):

Matteo Calcagnile ◽

Patricia Forgez ◽

Marco Alifano ◽

Pietro Alifano

Keyword(s):

Amino Acid ◽

Receptor Binding ◽

Virus Transmission ◽

Threat Assessment ◽

Spike Protein ◽

Binding Motif ◽

Lethal Triad ◽

New Variant ◽

South East England ◽

And Control

AbstractVariants of SARS-CoV-2 have been identified rapidly after the beginning of pandemic. One of them, involving the spike protein and called D614G, represents a substantial percentage of currently isolated strains. While research on this variant was ongoing worldwide, on December 20th 2020 the European Centre for Disease Prevention and Control reported a Threat Assessment Brief describing the emergence of a new variant of SARS-CoV-2, named B.1.1.7, harboring multiple mutations mostly affecting the Spike protein. This viral variant has been recently associated with a rapid increase in COVID-19 cases in South East England, with alarming implications for future virus transmission rates. Specifically, of the nine amino acid replacements that characterize the Spike in the emerging variant, four are found in the region between the Fusion Peptide and the RBD domain (namely the already known D614G, together with A570D, P681H, T716I), and one, N501Y, is found in the Spike Receptor Binding Domain – Receptor Binding Motif (RBD-RBM). In this study, by using in silico biology, we provide evidence that these amino acid replacements have dramatic effects on the interactions between SARS-CoV-2 Spike and the host ACE2 receptor or TMPRSS2, the protease that induces the fusogenic activity of Spike. Mostly, we show that these effects are strongly dependent on ACE2 and TMPRSS2 polymorphism, suggesting that dynamics of pandemics are strongly influenced not only by virus variation but also by host genetic background.

Download Full-text