scholarly journals Rapid, sensitive, full genome sequencing of Severe Acute Respiratory Syndrome Virus Coronavirus 2 (SARS-CoV-2)

2020 ◽  
Author(s):  
Clinton R Paden ◽  
Ying Tao ◽  
Krista Queen ◽  
Jing Zhang ◽  
Yan Li ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Sanger Sequencing ◽  
Respiratory Syndrome Virus ◽  
Full Genome ◽  
Miseq Sequencing ◽  
Genomic Information ◽  
Rt Pcr ◽  
Full Genome Sequencing ◽  
High Quality ◽  
Global Pandemic

AbstractSARS-CoV-2 recently emerged, resulting a global pandemic. Rapid genomic information is critical to understanding transmission and pathogenesis. Here, we describe validated protocols for generating high-quality full-length genomes from primary samples. The first employs multiplex RT-PCR followed by MinION or MiSeq sequencing. The second uses singleplex, nested RT-PCR and Sanger sequencing.

scholarly journals Sequencing and analysis of complete genome of the attenuated Hanvet1.VN strain used for vaccine production against porcine reproductive and respiratory syndrome

2018 ◽  
Vol 16 (1) ◽  
pp. 51-57
Author(s):  
Nguyễn Thị Nga ◽  
Hà Thị Thu ◽  
Nguyễn Thị Hoa ◽  
Vũ Thị Hiền ◽  
Trần Thị Thu Hiền ◽  
...  
Keyword(s):  
North American ◽  
Sanger Sequencing ◽  
Respiratory Syndrome Virus ◽  
Whole Genome ◽  
Type Ii ◽  
Vaccine Production ◽  
Rt Pcr ◽  
North American Strain ◽  
American Strain ◽  
Vietnamese Strain

The porcine reproductive and respiratory syndrome virus (PRRSV) attenuated strain Hanvet1.VN has been developed by the Pharmaceutical and Veterinary Material J.S.C (HANVET) by passaging HY-2010 strain on MARC-145 cells for 80 passages and used for PRRS vaccine production. In this study, we sequenced and analyzed the whole genome of the attenuated Hanvet1.VN strain. The total RNA was extracted from the Hanvet1.VN strain, RT-PCR was used for amplification of 15 separate segments of the whole genome. The amplified segments were cloned into the pCR2.1 vector and sequenced by Sanger sequencing. The sequences were analyzed with BioEdit and DNA Star Software. The results showed that, GP5 of the Hanvet1.VN attenuated strain had 100% identity in amino acid (aa) sequences with one of the pathogenic Vietnamese strain isolated in Quang Nam Province and had 98% identity with that of the Chinese 07NM strain. However, the identity of aa sequence of the Hanvet1.VN GP5 was much lower in the comparison with GP5 of VR2332, and it was only 87%. The MP and NP proteins were highly conserved compared with pathogenic strains circulating in Vietnam (07QN) and China (07NM) (99-100%, respectively). The other eight proteins of the Hanvet1.VN strain showed changes from 1.2% in NP1a to 3.9% in GP2 compared with the 07QN strain. However, the aa identity of all Hanvet1.VN proteins were very low when compared with proteins of PRRSV type II strain (North American strain, VR2332), ranged from 86.25% to 97.7%. Our results showed that the Hanvet1.VN attenuated vaccine strain had protective immunogenicity similar to that strain circulating in Vietnam closely related to a strain from China but different from the type II North American strain VR2332. Hence, for importing PRRSV vaccine, especially from American or Europe Countries, antigenic compatibility of the PRRSV vaccine and strains circulating in Vietnam should be concerned in PRRSV vaccine production.

scholarly journals A fast and robust method for full genome sequencing of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) Type 1 and Type 2

2013 ◽  
Vol 193 (2) ◽  
pp. 697-705 ◽  
Author(s):  
Lise K. Kvisgaard ◽  
Charlotte K. Hjulsager ◽  
Ulrik Fahnøe ◽  
Solvej Ø. Breum ◽  
Tahar Ait-Ali ◽  
...  
Keyword(s):  
Genome Sequencing ◽  
Respiratory Syndrome Virus ◽  
Full Genome ◽  
Robust Method ◽  
Full Genome Sequencing

scholarly journals SARS-CoV-2 variants in Paraguay: Detection and surveillance with a readily modifiable, multiplex real-time RT-PCR

2021 ◽  
Author(s):  
Magaly Martinez ◽  
Phuong-Vi Nguyen ◽  
Maxwell Su ◽  
Fatima Cardozo ◽  
Adriana Valenzuela ◽  
...  
Keyword(s):  
Real Time ◽  
Sanger Sequencing ◽  
Lower Cost ◽  
Population Level ◽  
Nanopore Sequencing ◽  
Receptor Binding Domain ◽  
Rt Pcr ◽  
High Quality ◽  
Rna Detection ◽  
Detection Of Mutations

Objectives The objective of the current study was to develop a lower-cost and scalable protocol to identify and monitor SARS-CoV-2 variants in Paraguay by pairing real-time RT-PCR detection of spike mutations with amplicon Sanger sequencing and whole-genome Nanopore sequencing. Methods 201 acute-phase nasopharyngeal samples from SARS-CoV-2-positive individuals were tested with two rRT-PCRs: 1) N2RP assay to confirm SARS-CoV-2 RNA detection (CDC N2 target), and 2) the Spike SNP assay to detect mutations in the spike receptor binding domain. The assay was performed with probes to identify mutations associated with the following variants: alpha (501Y), beta/gamma (417variant/484K/501Y), delta (452R/478K), and lambda (452Q/490S). Results All samples were positive for SARS-CoV-2 in the N2RP assay (mean Ct, 20.8; SD 5.6); 198/201 (98.5%) tested positive in the Spike SNP assay. The most common genotype was 417variant/484K/501Y, detected in 102/198 samples (51.5%) and most consistent with P.1 lineage (gamma variant) in Paraguay. No mutations (K417 only) were found in 64/198 (32.3%); and K417/484K was identified in 22/198 (11.1%), consistent with P.2 (zeta). Seven samples (3.5%) tested positive for 452R without 478K, and one sample with genotype K417/501Y was confirmed as B.1.1.7 (alpha). Results were confirmed by Sanger sequencing in 181/181 samples (100%) with high-quality amplicon sequences, and variant calls were consistent with Nanopore sequencing in 29/29 samples. Conclusions The Spike SNP assay provides accurate detection of mutations associated with SARS-CoV-2 variants. This can be implemented in laboratories performing rRT-PCR to improve population-level surveillance for these mutations and inform the judicious use of scarce sequencing resources.

scholarly journals Massive Multiplexing Can Deliver a $1 Test for COVID-19

2020 ◽  
Author(s):  
Paul DN Hebert ◽  
Sean WJ Prosser ◽  
Natalia V Ivanova ◽  
Evgeny V Zakharov ◽  
Sujeevan Ratnasingham
Keyword(s):  
Large Scale ◽  
Low Cost ◽  
Rna Extraction ◽  
Respiratory Syndrome Virus ◽  
Major Advance ◽  
Rt Pcr ◽  
Screening Programs ◽  
Global Pandemic ◽  
Gene Coding ◽  
Large Scale Screening

ABSTRACTThe severe acute respiratory syndrome virus, SARS-CoV-2 (hereafter COVID-19), rapidly achieved global pandemic status, provoking large-scale screening programs in many nations. Their activation makes it imperative to identify methods that can deliver a diagnostic result at low cost. This paper describes an approach which employs sequence variation in the gene coding for its envelope protein as the basis for a scalable, inexpensive test for COVID-19. It achieves this by coupling a simple RNA extraction protocol with low-volume RT-PCR, followed by E-Gel screening and sequencing on high-throughput platforms to analyze 10,000 samples in a run. Slight modifications to the protocol could support screening programs for other known viruses and for viral discovery. Just as the $1,000 genome is transforming medicine, a $1 diagnostic test for viral and bacterial pathogens would represent a major advance for public health.

scholarly journals A simplified Sanger sequencing method for full genome sequencing of multiple subtypes of human influenza A viruses

2015 ◽  
Vol 68 ◽  
pp. 43-48 ◽  
Author(s):  
Yi-Mo Deng ◽  
Natalie Spirason ◽  
Pina Iannello ◽  
Lauren Jelley ◽  
Hilda Lau ◽  
...  
Keyword(s):  
Genome Sequencing ◽  
Influenza A ◽  
Sanger Sequencing ◽  
Full Genome ◽  
Human Influenza ◽  
Full Genome Sequencing ◽  
Influenza A Viruses ◽  
Sequencing Method

scholarly journals Rapid, Sensitive, Full-Genome Sequencing of Severe Acute Respiratory Syndrome Coronavirus 2

2020 ◽  
Vol 26 (10) ◽  
pp. 2401-2405 ◽  
Author(s):  
Clinton R. Paden ◽  
Ying Tao ◽  
Krista Queen ◽  
Jing Zhang ◽  
Yan Li ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Genome Sequencing ◽  
Full Genome ◽  
Full Genome Sequencing

scholarly journals Novel rapid identification of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) by real-time RT-PCR using BD Max Open System in Taiwan

PeerJ ◽  
2020 ◽  
Vol 8 ◽  
pp. e9318 ◽  
Author(s):  
Cherng-Lih Perng ◽  
Ming-Jr Jian ◽  
Chih-Kai Chang ◽  
Jung-Chung Lin ◽  
Kuo-Ming Yeh ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Real Time ◽  
Turnaround Time ◽  
Rapid Identification ◽  
Clinical Samples ◽  
Respiratory Syndrome Virus ◽  
Acid Extraction ◽  
Prevention Measures ◽  
Rt Pcr ◽  
African Region

Coronavirus disease 2019 has become a worldwide pandemic. By April 7, 2020, approximately 1,279,722 confirmed cases were reported worldwide including those in Asia, European Region, African Region and Region of the Americas. Rapid and accurate detection of Severe Acute Respiratory Syndrome Virus 2 (SARS-CoV-2) is critical for patient care and implementing public health measures to control the spread of infection. In this study, we developed and validated a rapid total nucleic acid extraction method based on real‐time RT-PCR for reliable, high‐throughput identification of SARS-CoV-2 using the BD MAX platform. For clinical validation, 300 throat swab and 100 sputum clinical samples were examined by both the BD MAX platform and in-house real-time RT-PCR methods, which showed 100% concordant results. This BD MAX protocol is fully automated and the turnaround time from sample to results is approximately 2.5 h for 24 samples compared to 4.8 h by in-house real-time RT-PCR. Our developed BD MAX RT-PCR assay can accurately identify SARS-CoV-2 infection and shorten the turnaround time to increase the effectiveness of control and prevention measures for this emerging infectious disease.

scholarly journals Insights from Genomes and Genetic Epidemiology of SARS-CoV-2 isolates from the state of Andhra Pradesh

2021 ◽  
Author(s):  
Pallavali Roja Rani ◽  
Mohamed Imran ◽  
J. Vijaya Lakshmi ◽  
Bani Jolly ◽  
S. Afsar ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Genetic Epidemiology ◽  
Andhra Pradesh ◽  
The State ◽  
Rt Pcr ◽  
High Quality ◽  
Epidemiological Analysis ◽  
The Third ◽  
Global Pandemic ◽  
Large Numbers

AbstractCoronavirus disease (COVID-19) emerged from a city in China and has now spread as a global pandemic affecting millions of individuals. The causative agent, SARS-CoV-2 is being extensively studied in terms of its genetic epidemiology using genomic approaches. Andhra Pradesh is one of the major states of India with the third-largest number of COVID-19 cases with limited understanding of its genetic epidemiology. In this study, we have sequenced 293 SARS-CoV-2 genome isolates from Andhra Pradesh with a mean coverage of 13,324X. We identified 564 high-quality SARS-CoV-2 variants, out of which 15 are novel. A total of 18 variants mapped to RT-PCR primer/probe sites, and 4 variants are known to be associated with an increase in infectivity. Phylogenetic analysis of the genomes revealed the circulating SARS-CoV-2 in Andhra Pradesh majorly clustered under the clade A2a (94%), while 6% fall under the I/A3i clade, a clade previously defined to be present in large numbers in India. To the best of our knowledge, this is the most comprehensive genetic epidemiological analysis performed for the state of Andhra Pradesh.

The relevance of dietetic approach of Ayurveda from the perspective of COVID-19 pandemic

2020 ◽  
Vol 11 (SPL1) ◽  
pp. 748-752
Author(s):  
Swapnali Khabade ◽  
Bharat Rathi ◽  
Renu Rathi
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Healthy Person ◽  
World Health ◽  
Physical And Mental Health ◽  
Leading Role ◽  
Global Pandemic ◽  
The People ◽  
The World ◽  
Health Organization ◽  
The Individual

A novel, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), causes severe acute respiratory syndrome and spread globally from Wuhan, China. In March 2020 the World Health Organization declared the SARS-Cov-2 virus as a COVID- 19, a global pandemic. This pandemic happened to be followed by some restrictions, and specially lockdown playing the leading role for the people to get disassociated with their personal and social schedules. And now the food is the most necessary thing to take care of. It seems the new challenge for the individual is self-isolation to maintain themselves on the health basis and fight against the pandemic situation by boosting their immunity. Food organised by proper diet may maintain the physical and mental health of the individual. Ayurveda aims to promote and preserve the health, strength and the longevity of the healthy person and to cure the disease by properly channelling with and without Ahara. In Ayurveda, diet (Ahara) is considered as one of the critical pillars of life, and Langhana plays an important role too. This article will review the relevance of dietetic approach described in Ayurveda with and without food (Asthavidhi visheshaytana & Lanhgan) during COVID-19 like a pandemic.

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