新冠病毒分析
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https://www.frontiersin.org/articles/10.3389/fmicb.2021.665041/full
A Novel SARS-CoV-2 Viral Sequence Bioinformatic Pipeline Has Found Genetic Evidence That the Viral 3′ Untranslated Region (UTR) Is Evolving and Generating Increased Viral DiversityNext-Generation Sequencing and FASTA Dataset Processing
To process next generation sequencing datasets, we employed our pipeline (SARS-CoV-2_freebayes) consisting of a bash/UNIX script that runs several programs in sequential order. We processed imputed list of SRA accessions with sra-tools7, generating compressed FASTQ files per sequencing, automatically trimmed with fastp tool (Chen et al., 2018). Then, we aligned each trimmed fastq file against a provided reference genome (Wuhan-Hu-1, GenBank Accession: MN908947.3) using Minimap2 splice-aware aligner in preset mode -ax sr (Li, 2018). We sorted and indexed the resulting BAM files by using Samtools (Li et al., 2009) and performed variant calling on every sorted BAM file, obtaining major frequency viral variants per genome in VCF format using the Freebayes variant calling program, as frequency-based pooled caller (−F 0.49)8 (Garrison and Marth, 2012). Then, we used Jacquard program9 in the python environment (Sanner, 1999) to merge every VCF file containing variants associated to each bam file into a single VCF file, containing aggregated variants from all genomes. In the resulting merged VCF file, we recalculated viral frequencies using several UNIX tools (Kernighan and Morgan, 1982), in combination with vcflib10. We used the variants per genome logfile “logfile_variants_SRA_freebayes” to construct Figure 1B using GraphPad Prism 8 software11. We processed GISAID FASTA genomes in a similar manner. We preprocess a single GISAID genome collection with SeqKit (Shen et al., 2016) to decompose a single FASTA file into individual FASTA files, each file containing a single genome. Then, we aligned every FASTA genome against SARS-CoV-2 reference genome (NC_045512.2) using Minimap2 aligner with preset -ax asm5 (Li, 2018) and performed variant calling on each BAM file using Freebayes variant caller with –min-alternate-count 1 (C 1) option (see text footnote 8), outputting variants in VCF format. With these operations, we obtained major frequency viral variants in VCF format from each FASTA genome. Then, we aggregated variants into a single VCF file, as described with Jacquard. We constructed Figure 1B graph by using variants per genome logfile, reported in the output file “logfile_variants_GISAID_freebayes” and imputed into the GraphPad Prism 8 software. We filtered out highly homoplasic sites from merged variant calls, as already reported to be frequent in SARS-CoV-2 sequencing see: https://virological.org/t/issues-with-sars-cov-2-sequencing-data/473. All these computational analyses are described here: https://github.com/cfarkas/SARS-CoV-2-freebayes (case examples I and II, respectively). -
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https://www.medrxiv.org/content/10.1101/2020.03.30.20048108v3
中国 美国等CDC用的试剂盒对比 -
https://sci-hub.st/10.3390/cimb43020061
Next-Generation Sequencing (NGS) in COVID-19: A Tool for
SARS-CoV-2 Diagnosis, Monitoring New Strains and
Phylodynamic Modeling in Molecular Epidemiology
1.文章提到了两种新冠的检测方法
Molecular-Based Testing Methods 即 RT-PCR
Serological and Immunological-Based Testing Methods
抗体只能检测这个之前有没有得过新冠 而且不是很准确 因为有的人就算被感染过也可能没有抗体
2.NGS检测病原的优势
一个是不用培养 而且不要做针对性的假设 一股脑全给测出来 但是这种方法的成本肯定没有panel低
Using NGS technology for the diagnosis of infectious diseases offers an unbiased approach detecting pathogens that does not rely on culturing or the need for clinical hypotheses.
While standard testing procedures require clinicians to identify possible explanations for a patient’s symptoms and employ tests aimed at those specific pathogens, NGS testing can reveal the presence of all types of microorganisms present in a sample, including bacteria,viruses, fungi, and parasites.
另外一个优势就是病人可能同时 感染了多种病毒 例如新冠 流感病毒 可能同时感染 这对后期的治疗 病因都有影响 这个优势其实就是上面说的第二点 NGS 可以同时测出来多种病原 -
https://gitlab.com/uit-sfb/cbf
https://covid19.sfb.uit.no/
新冠的数据库 包括其源码
他们开发了一个CBF框架 可以管理这些数据 -
InterARTIC: an interactive web application for whole-genome nanopore sequencing analysis of SARS-CoV-2 and other viruses
https://github.com/Psy-Fer/interARTIC/ -
poreCov-An Easy to Use, Fast, and Robust Workflow for SARS-CoV-2 Genome Reconstruction via Nanopore Sequencing
https://www.frontiersin.org/articles/10.3389/fgene.2021.711437/full
https://github.com/replikation/poreCov -
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https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7400123/
Mutational Frequencies of SARS-CoV-2 Genome during the Beginning Months of the Outbreak in USA
Mutation frequency was calculated by taking the ratio of the number of total nucleotide mutations and the number of genome sequences in each week. -
