scholarly journals Detection of Mutations Associated with Variants of Concern Via High Throughput Sequencing of SARS-CoV-2 Isolated from NYC Wastewater

2021 ◽  
Author(s):  
Davida S. Smyth ◽  
Monica Trujillo ◽  
Kristen Cheung ◽  
Anna Gao ◽  
Irene Hoxie ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Herd Immunity ◽  
Control Measures ◽  
Clinical Samples ◽  
Genetic Sequencing ◽  
Novel Variants ◽  
The World ◽  
Induced Immunity

ABSTRACTMonitoring SARS-CoV-2 genetic diversity is strongly indicated because diversifying selection may lead to the emergence of novel variants resistant to naturally acquired or vaccine-induced immunity. To date, most data on SARS-CoV-2 genetic diversity has come from the sequencing of clinical samples, but such studies may suffer limitations due to costs and throughput. Wastewater-based epidemiology may provide an alternative and complementary approach for monitoring communities for novel variants. Given that SARS-CoV-2 can infect the cells of the human gut and is found in high concentrations in feces, wastewater may be a valuable source of SARS-CoV-2 RNA, which can be deep sequenced to provide information on the circulating variants in a community. Here we describe a safe, affordable protocol for the sequencing of SARS-CoV-2 RNA using high-throughput Illumina sequencing technology. Our targeted sequencing approach revealed the presence of mutations associated with several Variants of Concern at appreciable frequencies. Our work demonstrates that wastewater-based SARS-CoV-2 sequencing can inform surveillance efforts monitoring the community spread of SARS-CoV-2 Variants of Concern and detect the appearance of novel emerging variants more cheaply, safely, and efficiently than the sequencing of individual clinical samples.IMPORTANCEThe SARS-CoV-2 pandemic has caused millions of deaths around the world as countries struggle to contain infections. The pandemic will not end until herd immunity is reached, that is, when most of the population has either recovered from SARS-CoV-2 infection or is vaccinated against SARS-CoV-2. However, the emergence of new SARS-CoV-2 variants of concern threatens to erase gains. Emerging new variants may re-infect persons who have recovered from COVID-19 or may evade vaccine-induced immunity. However, scaling up SARS-CoV-2 genetic sequencing to monitor Variants of Concern in communities around the world is challenging. Wastewater-based sequencing of SARS-CoV-2 RNA can be used to monitor the presence of emerging variants in large communities to enact control measures to minimize the spread of these variants. We describe here the identification of alleles associated with several variants of concern in wastewater obtained from NYC watersheds.

scholarly journals Tracking Cryptic SARS-CoV-2 Lineages Detected in NYC Wastewater

2021 ◽  
Author(s):  
Davida S Smyth ◽  
Monica Trujillo ◽  
Devon A Gregory ◽  
Kristen Cheung ◽  
Anna Gao ◽  
...  
Keyword(s):  
New York ◽  
New York City ◽  
High Throughput Sequencing ◽  
Protein Gene ◽  
Wastewater Treatment Plants ◽  
Clinical Samples ◽  
Direct Inference ◽  
Animal Reservoir ◽  
Novel Variants ◽  
Induced Immunity

Tracking SARS-CoV-2 genetic diversity is strongly indicated because diversifying selection may lead to the emergence of novel variants resistant to naturally acquired or vaccine-induced immunity. To monitor New York City (NYC) for the presence of novel variants, we amplified regions of the SARS-CoV-2 Spike protein gene from RNA acquired from all 14 NYC wastewater treatment plants (WWTPs) and ascertained the diversity of lineages from these samples using high throughput sequencing. Here we report the detection and increasing frequencies of novel SARS-CoV-2 lineages not recognized in GISAIDs EpiCoV database. These lineages contain mutations rarely observed in clinical samples, including Q493K, Q498Y, H519N and T572N. Many of these mutations were found to expand the tropism of SARS-CoV-2 pseudoviruses by allowing infection of cells expressing the human, mouse, or rat ACE2 receptor. In addition, pseudoviruses containing the Spike amino acid sequence of these lineages were found to be resistant to many different classes of receptor binding domain (RBD) binding neutralizing monoclonal antibodies. We offer several hypotheses for the anomalous presence of these mutations, including the possibility of a non-human animal reservoir. Although wastewater sampling cannot provide direct inference of SARS-CoV-2 clinical sequences, our research revealed several lineages that could be relevant to public health and they would not have been discovered if not for wastewater surveillance.

scholarly journals Current Perspectives on High-Throughput Sequencing (HTS) for Adventitious Virus Detection: Upstream Sample Processing and Library Preparation

Viruses ◽  
2018 ◽  
Vol 10 (10) ◽  
pp. 566 ◽  
Author(s):  
Siemon Ng ◽  
Cassandra Braxton ◽  
Marc Eloit ◽  
Szi Feng ◽  
Romain Fragnoud ◽  
...  
Keyword(s):  
Sample Preparation ◽  
Nucleic Acids ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Virus Detection ◽  
Extraction Methods ◽  
Control Measures ◽  
Acid Extraction ◽  
Library Preparation ◽  
Sample Processing

A key step for broad viral detection using high-throughput sequencing (HTS) is optimizing the sample preparation strategy for extracting viral-specific nucleic acids since viral genomes are diverse: They can be single-stranded or double-stranded RNA or DNA, and can vary from a few thousand bases to over millions of bases, which might introduce biases during nucleic acid extraction. In addition, viral particles can be enveloped or non-enveloped with variable resistance to pre-treatment, which may influence their susceptibility to extraction procedures. Since the identity of the potential adventitious agents is unknown prior to their detection, efficient sample preparation should be unbiased toward all different viral types in order to maximize the probability of detecting any potential adventitious viruses using HTS. Furthermore, the quality assessment of each step for sample processing is also a critical but challenging aspect. This paper presents our current perspectives for optimizing upstream sample processing and library preparation as part of the discussion in the Advanced Virus Detection Technologies Interest group (AVDTIG). The topics include: Use of nuclease treatment to enrich for encapsidated nucleic acids, techniques for amplifying low amounts of virus nucleic acids, selection of different extraction methods, relevant controls, the use of spike recovery experiments, and quality control measures during library preparation.

scholarly journals High-Throughput Sequencing Reveals Further Diversity of Little Cherry Virus 1 with Implications for Diagnostics

Viruses ◽  
2018 ◽  
Vol 10 (7) ◽  
pp. 385 ◽  
Author(s):  
Asimina Katsiani ◽  
Varvara Maliogka ◽  
Nikolaos Katis ◽  
Laurence Svanella-Dumas ◽  
Antonio Olmos ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Sour Cherry ◽  
Prunus Species ◽  
Cherry Tree ◽  
High Genetic Diversity ◽  
Viral Genomes ◽  
Virus Diversity ◽  
First Time

Little cherry virus 1 (LChV1, Velarivirus, Closteroviridae) is a widespread pathogen of sweet or sour cherry and other Prunus species, which exhibits high genetic diversity and lacks a putative efficient transmission vector. Thus far, four distinct phylogenetic clusters of LChV1 have been described, including isolates from different Prunus species. The recent application of high throughput sequencing (HTS) technologies in fruit tree virology has facilitated the acquisition of new viral genomes and the study of virus diversity. In the present work, several new LChV1 isolates from different countries were fully sequenced using different HTS approaches. Our results reveal the presence of further genetic diversity within the LChV1 species. Interestingly, mixed infections of the same sweet cherry tree with different LChV1 variants were identified for the first time. Taken together, the high intra-host and intra-species diversities of LChV1 might affect its pathogenicity and have clear implications for its accurate diagnostics.

scholarly journals Detection Of Genomic Variants Of SARS-CoV-2 Circulating In Wastewater By High-Throughput Sequencing

2021 ◽  
Author(s):  
Alba Pérez-Cataluña ◽  
Álvaro Chiner-Oms ◽  
Enric Cuevas-Ferrando ◽  
Azahara Díaz-Reolid ◽  
Irene Falcó ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Community Level ◽  
Low Frequencies ◽  
Spike Gene ◽  
Genomic Variants ◽  
Clinical Databases ◽  
Novel Variants ◽  
First Time

The use of SARS-CoV-2 metagenomics in wastewater can allow the detection of variants circulating at community level. After comparing with clinical databases, we identified three novel variants in the spike gene, and six new variants in the spike detected for the first time in Spain. We finally support the hypothesis that this approach allows the identification of unknown SARS-CoV-2 variants or detected at only low frequencies in clinical genomes.Abstract Figure

scholarly journals Learning Sparse Log-Ratios for High-Throughput Sequencing Data

2021 ◽  
Author(s):  
Elliott Gordon-Rodriguez ◽  
Thomas P. Quinn ◽  
John P. Cunningham
Keyword(s):  
High Throughput ◽  
Latent Variables ◽  
High Throughput Sequencing ◽  
Compositional Data ◽  
Predictive Accuracy ◽  
Learning Algorithm ◽  
Sequencing Data ◽  
Genetic Sequencing ◽  
Wide Range ◽  
Benchmark Datasets

AbstractThe automatic discovery of interpretable features that are associated to an outcome of interest is a central goal of bioinformatics. In the context of high-throughput genetic sequencing data, and Compositional Data more generally, an important class of features are the log-ratios between subsets of the input variables. However, the space of these log-ratios grows combinatorially with the dimension of the input, and as a result, existing learning algorithms do not scale to increasingly common high-dimensional datasets. Building on recent literature on continuous relaxations of discrete latent variables, we design a novel learning algorithm that identifies sparse log-ratios several orders of magnitude faster than competing methods. As well as dramatically reducing runtime, our method outperforms its competitors in terms of sparsity and predictive accuracy, as measured across a wide range of benchmark datasets.

scholarly journals A cell-free DNA metagenomic sequencing assay that integrates the damage response to infection

2019 ◽  
Author(s):  
Alexandre Pellan Cheng ◽  
Philip Burnham ◽  
John Richard Lee ◽  
Matthew Pellan Cheng ◽  
Manikkam Suthanthiran ◽  
...  
Keyword(s):  
Urinary Tract ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Viral Infections ◽  
Clinical Samples ◽  
Metagenomic Sequencing ◽  
Kidney Transplant Recipients ◽  
Bladder Tissue ◽  
Cell Free Dna ◽  
Free Dna

ABSTRACTHigh-throughput metagenomic sequencing offers an unbiased approach to identify pathogens in clinical samples. Conventional metagenomic sequencing however does not integrate information about the host, which is often critical to distinguish infection from infectious disease, and to assess the severity of disease. Here, we explore the utility of high-throughput sequencing of cell-free DNA after bisulfite conversion to map the tissue and cell types of origin of host-derived cell-free DNA, and to profile the bacterial and viral metagenome. We applied this assay to 51 urinary cfDNA isolates collected from a cohort of kidney transplant recipients with and without bacterial and viral infection of the urinary tract. We find that the cell and tissue types of origin of urinary cell-free DNA can be derived from its genome-wide profile of methylation marks, and strongly depend on infection status. We find evidence of kidney and bladder tissue damage due to viral and bacterial infection, respectively, and of the recruitment of neutrophils to the urinary tract during infection. Through direct comparison to conventional metagenomic sequencing as well as clinical tests of infection, we find this assay accurately captures the bacterial and viral composition of the sample. The assay presented here is straightforward to implement, offers a systems view into bacterial and viral infections of the urinary tract, and can find future use as a tool for the differential diagnosis of infections.

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