scholarly journals Genetic variability of SARS-CoV-2 in biological samples from patients in Moscow

2021 ◽  
Vol 97 (6) ◽  
pp. 511-517
Author(s):  
A. S. Speranskaya ◽  
V. V. Kaptelova ◽  
A. E. Samoilov ◽  
A. Yu. Bukharina ◽  
O. Yu. Shipulina ◽  
...  
Keyword(s):  
Genetic Variants ◽  
Reference Genome ◽  
Threshold Value ◽  
Nucleotide Position ◽  
Nucleotide Substitutions ◽  
Genomic Variants ◽  
Variable Position ◽  
Polymerase Chain ◽  
Autopsy Specimens ◽  
Total Coverage

Currently, a lot of attention is given to SARS-CoV-2 subpopulations and their coexistence with different genomic variants within the same patient. In this study, we performed next-generation whole-genome sequencing and assembly of viruses from samples representing swabs or autopsy specimens obtained from patients diagnosed with СOVID-19, which were initially confirmed by the real-time polymerase chain reaction (Ct = 10.4–19.8). Samples were prepared for sequencing by using the SCV-2000bp protocol. The obtained data were checked for presence of more than one SARS-CoV-2 genetic variants in a sample. Variants of nucleotide substitutions, coverage for each variant, and location of the variable position in the reference genome were detected with tools incorporated in the CLC Genomics Workbench program. In our search for variable nucleotide positions, we assumed that the sample had two genetic variants (not more); the threshold value ≥ 90% was set for probability of the identified variant. Variants represented by less than 20% of the reads in the total coverage were not taken into consideration. The obtained results showed that 5 samples had variability, i.e. they had several genetic variants of SARS-CoV-2. In 4 samples, both of the detected genomic variants differed only in one nucleotide position. The fifth sample demonstrated more substantial differences: a total of 3 variable positions and one three-nucleotide deletion. Our study shows that different genetic variants of SARS-CoV-2 can coexist within the same patient.

Paris I Dysfibrinogenemia: A Point Mutation in Intron 8 Results in Insertion of a 15 Amino Acid Sequence in the Fibrinogen γ-Chain

1993 ◽  
Vol 69 (03) ◽  
pp. 217-220 ◽  
Author(s):  
Jonathan B Rosenberg ◽  
Peter J Newman ◽  
Michael W Mosesson ◽  
Marie-Claude Guillin ◽  
David L Amrani
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Point Mutation ◽  
Genomic Dna ◽  
Comparative Sequence Analysis ◽  
Chain Gene ◽  
Molecular Defect ◽  
Nucleotide Position ◽  
Normal Individuals ◽  
Polymerase Chain

SummaryParis I dysfibrinogenemia results in the production of a fibrinogen molecule containing a functionally abnormal γ-chain. We determined the basis of the molecular defect using polymerase chain reaction (PCR) to amplify the γ-chain region of the Paris I subject’s genomic DNA. Comparative sequence analysis of cloned PCR segments of normal and Paris I genomic DNA revealed only an A→G point mutation occurring at nucleotide position 6588 within intron 8 of the Paris I γ-chain gene. We examined six normal individuals and found only normal sequence in this region, indicating that this change is not likely to represent a normal polymorphism. This nucleotide change leads to a 45 bp fragment being inserted between exons 8 and 9 in the mature γparis I chain mRNA, and encodes a 15 amino acid insert after γ350 [M-C-G-E-A-L-P-M-L-K-D-P-C-Y]. Alternative splicing of this region from intron 8 into the mature Paris I γ-chain mRNA also results after translation into a substitution of S for G at position γ351. Biochemical studies of 14C-iodoacetamide incorporation into disulfide-reduced Paris I and normal fibrinogen corroborated the molecular biologic predictions that two additional cysteine residues exist within the γpariS I chain. We conclude that the insertion of this amino acid sequence leads to a conformationallyaltered, and dysfunctional γ-chain in Paris I fibrinogen.

scholarly journals A Real-World Comparison of SARS-CoV-2 Rapid Antigen vs. Polymerase Chain Reaction Testing in Florida

2021 ◽  
Author(s):  
Lao-Tzu Allan-Blitz ◽  
Jeffrey D. Klausner
Keyword(s):  
Polymerase Chain Reaction ◽  
Real World ◽  
Threshold Value ◽  
Test Results ◽  
Chain Reaction ◽  
Cycle Threshold ◽  
Large Populations ◽  
Symptom Status ◽  
Polymerase Chain ◽  
Antigen Tests

Background The reported sensitivity of rapid, antigen-based diagnostics for SARS-CoV-2 infection varies. Few studies have evaluated rapid antigen tests in real-world settings or among large populations. Methods Beginning October 2020, Florida offered individuals presenting for SARS-CoV-2 testing polymerase chain reaction (PCR) testing if they tested positive by the Abbott BinaxNOW TM COVID-19 Ag Card, were symptomatic, or required or requested PCR testing. We compared test results among individuals who received both types of tests at four publicly-accessible testing sites across Florida. We calculated the positive percent agreement (PPA) between the two test types by symptom status. Subsequently, we evaluated the PPA among individuals regardless of symptoms with lower cycle threshold values (<30). Results Overall, 18,457 individuals were tested via both methods, of which 3,153 (17.1%) were positive by PCR. The PPA for the Abbott BinaxNOW TM COVID-19 Ag Card using the PCR comparator was 49.2% (95% CI 47.4%-50.9%). That performance was moderately improved among symptomatic individuals (51.9%; 95% CI 49.7%-54.0%). When restricted to positive PCR tests with a cycle threshold value <30, regardless of symptom status, the PPA was 75.3% (95% CI 72.8%-77.6%). Conclusion The PPA of the Abbott BinaxNOW TM COVID-19 Ag Card with PCR was lower than among previous reports. Our findings may reflect the performance of the BinaxNOW TM antigen test in real-world settings.

scholarly journals Identification of Genetic Variants of Human Papillomavirus in a Group of Mexican HIV/AIDS Patients and Their Possible Association with Cervical Cancer

2021 ◽  
Vol 70 (4) ◽  
pp. 501-509
Author(s):  
FELIPE ORTIZ-GUTIÉRREZ ◽  
LILIA SÁNCHEZ-MINUTTI ◽  
JOSÉ F. MARTÍNEZ-HERRERA ◽  
INDIANA D. TORRES-ESCOBAR ◽  
ELIAS B. PEZZAT-SAID ◽  
...  
Keyword(s):  
Cervical Cancer ◽  
Human Papillomavirus ◽  
Genetic Variants ◽  
Direct Relationship ◽  
Low Risk ◽  
Hiv Positive ◽  
Hiv Positive Women ◽  
Immunodeficiency Virus ◽  
Regional Population ◽  
Polymerase Chain

Infections caused by the human immunodeficiency virus (HIV) and human papillomavirus (HPV) cause thousands of deaths worldwide each year. So far, there has been no consensus on whether there is a direct relationship between the incidence of neoplasms and the immunosuppression caused by HIV that could help understand if coinfection increases the likelihood of cervical cancer. The objective of the study was to identify the presence of genetic variants of HPV in a group of HIV-positive women and their possible association with cervical cancer. Cervical samples were taken from HIV-positive patients for cytological analysis to identify the HPV genotype by polymerase chain reaction (PCR) and sequencing. The most preva¬lent L1 capsid protein mutations in the HPV genotype were ana¬lyzed in silico. Various types of HPV were identified, both high-risk (HR) and low-risk (LR). The most prevalent genotype was HPV51. Analysis of the L1 gene sequences of HPV51 isolates showed nucleo¬tide variations. Of the samples analyzed in Puebla, Mexico, HPV51 had the highest incidence (17.5%, 7/40). Different mutations, which could be used as population markers, were detected in this area, and they have not been reported in the L1 databases for HPV51 in Mexico. Genotypes 6, 14, 86, 87, 89, and 91, not detected or reported in samples from patients with HPV in Mexico, were also identified. Data from the population analyzed suggest no direct relationship between HIV immunosuppression and cervical cancer, regardless of the high- or low-risk HPV genotype. Furthermore, it is possible to develop regional population markers for the detection of HPV based on the mutations that occur in the sequence of nucleotides analyzed.

scholarly journals New sensitive method for the detection of the A3243G mutation of human mitochondrial deoxyribonucleic acid in diabetes mellitus patients by ligation-mediated polymerase chain reaction

1998 ◽  
Vol 44 (10) ◽  
pp. 2088-2093 ◽  
Author(s):  
Michiyo Urata ◽  
Machiko Wakiyama ◽  
Masanori Iwase ◽  
Makoto Yoneda ◽  
Sachiko Kinoshita ◽  
...  
Keyword(s):  
Diabetes Mellitus ◽  
Polymerase Chain Reaction ◽  
Conventional Method ◽  
Peripheral Blood ◽  
Blood Cells ◽  
Nucleotide Position ◽  
Peripheral Blood Cells ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Sensitive Method

Abstract An adenine-to-guanine mutation at nucleotide position (np) 3243 in the mitochondrial tRNALeu(UUR) gene is closely associated with various clinical phenotypes of diabetes mellitus. Because the mutation creates a new restriction site for the restriction enzyme ApaI, the mutation is usually detected and quantified by ApaI cleavage of the PCR products including np 3243. The sensitivity of the conventional method is, however, 5–10% heteroplasmy. The percentage of heteroplasmy of the mutation is usually highest in the affected tissues and is much lower in peripheral blood cells, which are used most frequently for the analysis. The sensitivity of the conventional method, however, is not sufficient to detect the mutation from peripheral blood cells. Utilizing ligation-mediated polymerase chain reaction, we have developed a feasible and sensitive method to detect 0.01% heteroplasmy of the 3243 mutation in peripheral leukocytes.

Identification of Heterozygous and Homozygous Individuals with the Novel RYR1 Mutation Cys35Arg in a Large Kindred

1997 ◽  
Vol 86 (3) ◽  
pp. 620-626 ◽  
Author(s):  
P. J. Lynch ◽  
R. Krivosic-Horber ◽  
H. Reyford ◽  
N. Monnier ◽  
K. Quane ◽  
...  
Keyword(s):  
Linkage Analysis ◽  
Malignant Hyperthermia ◽  
Nucleotide Position ◽  
The Novel ◽  
Autosomal Dominant Disorder ◽  
Large Pedigree ◽  
Malignant Hyperthermia Susceptible ◽  
Polymerase Chain ◽  
Clear Differentiation ◽  
Muscle Biopsies

Background Malignant hyperthermia (MH) is a potentially fatal, often autosomal dominant, disorder of skeletal muscle and is triggered in susceptible people by all commonly used inhalational anesthetics. In this article, the authors describe a malignant hyperthermia susceptible (MHS) kindred in which both parents of the proband are MHS and are first-degree cousins. Haplotype analysis in this kindred with chromosome 19 linked markers revealed that the proband and another sibling were homozygous for the affected RYR1 allele. Methods Eighteen members of this large pedigree were investigated, with a clinical examination for signs of a myopathy, a caffeine halothane contracture test, a histo-enzymologic study on the muscle biopsies, and linkage analysis on genomic DNA isolated from family blood samples. RYR1 cDNA was amplified by polymerase chain reaction and was cloned and sequenced, facilitating mutation detection. Results Linkage analysis demonstrated linkage between RYR1-linked markers and MH susceptibility in this family. DNA sequencing identified a T to C transition at nucleotide position 103, resulting in the substitution of an arginine for cysteine 35, representing the most N-terminal mutation reported to date in the RYR1 gene. This mutation segregates fully with the MHS trait, generating a lod score of 4.65 in favor of linkage to MHS at a recombination frequency of 0.0. Conclusions The proband in this kindred is the first reported homozygote to have presented with an MH episode. The homozygotes in this pedigree do not have an overt myopathy. The sensitivity of muscle samples to caffeine clearly distinguished the two homozygotes from other heterozygous-susceptible individuals. No clear differentiation was observed with the halothane contracture results.

The Associations of Genetic Variants in E-cadherin Gene With Clinical Outcome of Epithelial Ovarian Cancer

2016 ◽  
Vol 26 (9) ◽  
pp. 1601-1607 ◽  
Author(s):  
Wang Juan ◽  
Kang Shan ◽  
Wang Na ◽  
Zhou Rong-Miao ◽  
Li Yan
Keyword(s):  
Ovarian Cancer ◽  
Epithelial Ovarian Cancer ◽  
Genetic Variants ◽  
Metastatic Cancer ◽  
Progression Free Survival ◽  
Northern China ◽  
Free Survival ◽  
Increased Risk ◽  
Polymerase Chain ◽  
E Cadherin

ObjectiveThe E-cadherin protein plays major roles in tumor progression, invasion, and metastasis. Polymorphisms located in the E-cadherin gene (CDH1) may contribute to increased risks of specific cancers. In this study, we evaluated the associations between genetic variants inCDH1and the clinical outcomes of patients with epithelial ovarian cancer (EOC).Materials and MethodsWe assessed the−160C/Aand−347G/GApolymorphisms in the promoter region, as well as the3′-UTR +54C/Tpolymorphism of E-cadherin, in 257 patients with EOC by ligase detection reaction–polymerase chain reaction.ResultsMultivariate analysis showed that patients with EOC with theCDH1 −347GA/GAgenotype had shorter progression-free survival and overall survival (hazard ratio [HR], 2.16; 95% confidence interval [CI], 1.06–4.40 and HR, 2.06; 95% CI, 1.01–4.19, respectively) compared to those carrying the G/G genotype. Likewise, the patients with theCDH1 −160A/Agenotype had a shorter progression-free survival than those with the C/C genotype (HR, 4.12; 95% CI, 1.43–111.88). No significant association was detected between theCDH1 3′-UTR +54C/Tpolymorphism and survival of the patients with EOC.ConclusionsTheCDH1 −347GA/GAand−160A/Agenotypes may be prognostic markers that can help to identify patients at increased risk of invasive/metastatic cancer in northern China.

scholarly journals Chemical Mutagenesis of Dengue Virus Type 4 Yields Mutant Viruses Which Are Temperature Sensitive in Vero Cells or Human Liver Cells and Attenuated in Mice

2001 ◽  
Vol 75 (20) ◽  
pp. 9731-9740 ◽  
Author(s):  
Joseph E. Blaney ◽  
Daniel H. Johnson ◽  
Cai-Yen Firestone ◽  
Christopher T. Hanson ◽  
Brian R. Murphy ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Virus Type ◽  
Human Liver ◽  
Vero Cells ◽  
Chemical Mutagenesis ◽  
Temperature Sensitive ◽  
Nucleotide Position ◽  
Nucleotide Substitutions ◽  
Ns3 Protein ◽  
Old Mice

ABSTRACT A recombinant live attenuated dengue virus type 4 (DEN4) vaccine candidate, 2AΔ30, was found previously to be generally well tolerated in humans, but a rash and an elevation of liver enzymes in the serum occurred in some vaccinees. 2AΔ30, a non-temperature-sensitive (non-ts) virus, contains a 30-nucleotide deletion (Δ30) in the 3′ untranslated region (UTR) of the viral genome. In the present study, chemical mutagenesis of DEN4 was utilized to generate attenuating mutations which may be useful in further attenuation of the 2AΔ30 candidate vaccine. Wild-type DEN4 2A virus was grown in Vero cells in the presence of 5-fluorouracil, and a panel of 1,248 clones were isolated. Twenty ts mutant viruses were identified that were ts in both simian Vero and human liver HuH-7 cells (n = 13) or only in HuH-7 cells (n = 7). Each of the 20 ts mutant viruses possessed an attenuation phenotype, as indicated by restricted replication in the brains of 7-day-old mice. The complete nucleotide sequence of the 20 ts mutant viruses identified nucleotide substitutions in structural and nonstructural genes as well as in the 5′ and 3′ UTRs, with more than one change occurring, in general, per mutant virus. A ts mutation in the NS3 protein (nucleotide position 4995) was introduced into a recombinant DEN4 virus possessing the Δ30 deletion, thereby creating rDEN4Δ30-4995, a recombinant virus which is ts and more attenuated than rDEN4Δ30 virus in the brains of mice. We are assembling a menu of attenuating mutations that should be useful in generating satisfactorily attenuated recombinant dengue vaccine viruses and in increasing our understanding of the pathogenesis of dengue virus.

Polymorphisms in the goat β-lactoglobulin gene

2005 ◽  
Vol 72 (3) ◽  
pp. 379-384 ◽  
Author(s):  
Maria Ballester ◽  
Armand Sánchez ◽  
Josep M Folch
Keyword(s):  
Genetic Variants ◽  
Promoter Region ◽  
Point Mutations ◽  
Proximal Promoter ◽  
Coding Region ◽  
Nucleotide Substitutions ◽  
Single Nucleotide ◽  
Β Lactoglobulin ◽  
Pyrosequencing Technology ◽  
Frequent Haplotype

β-lactoglobulin polymorphisms have been reported in the milk of different goat breeds, although no genetic variants affecting the protein have been characterized. In the present study, we amplified and sequenced the proximal promoter and the first six exons containing the entire coding region for the β-lactoglobulin gene in eleven goat breeds from Spain, France, Italy, Switzerland, Senegal and Asia to identify genetic variants. Fifteen polymorphisms were detected, nine in the promoter region and six in the exons of the β-lactoglobulin gene. All polymorphisms were single nucleotide substitutions with the exception of one deletion/insertion in the promoter region. The polymorphisms in the coding region did not produce any amino acid change. In addition, pyrosequencing technology was used to genotype four polymorphisms in the promoter region in 200 goats belonging to eleven breeds. Differences in allelic frequencies for these polymorphisms between breeds are described and a specific polymorphism for the Italian populations was identified. Finally, the analysis of association between these four promoter point mutations was investigated resulting in five haplotypes, GCGC being the most frequent haplotype in all breeds analysed.

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