scholarly journals Staphylococcus aureus viewed from the perspective of 40,000+ genomes

PeerJ ◽  
2018 ◽  
Vol 6 ◽  
pp. e5261 ◽  
Author(s):  
Robert A. Petit ◽  
Timothy D. Read
Keyword(s):  
Staphylococcus Aureus ◽  
De Novo ◽  
Wide Spectrum ◽  
Low Cost ◽  
Application Programming Interface ◽  
Relevant Information ◽  
Clinical Diagnostics ◽  
Nucleotide Polymorphisms ◽  
Analysis Pipeline ◽  
European Nucleotide Archive

Low-cost Illumina sequencing of clinically-important bacterial pathogens has generated thousands of publicly available genomic datasets. Analyzing these genomes and extracting relevant information for each pathogen and the associated clinical phenotypes requires not only resources and bioinformatic skills but organism-specific knowledge. In light of these issues, we created Staphopia, an analysis pipeline, database and application programming interface, focused on Staphylococcus aureus, a common colonizer of humans and a major antibiotic-resistant pathogen responsible for a wide spectrum of hospital and community-associated infections. Written in Python, Staphopia’s analysis pipeline consists of submodules running open-source tools. It accepts raw FASTQ reads as an input, which undergo quality control filtration, error correction and reduction to a maximum of approximately 100× chromosome coverage. This reduction significantly reduces total runtime without detrimentally affecting the results. The pipeline performs de novo assembly-based and mapping-based analysis. Automated gene calling and annotation is performed on the assembled contigs. Read-mapping is used to call variants (single nucleotide polymorphisms and insertion/deletions) against a reference S. aureus chromosome (N315, ST5). We ran the analysis pipeline on more than 43,000 S. aureus shotgun Illumina genome projects in the public European Nucleotide Archive database in November 2017. We found that only a quarter of known multi-locus sequence types (STs) were represented but the top 10 STs made up 70% of all genomes. methicillin-resistant S. aureus (MRSA) were 64% of all genomes. Using the Staphopia database we selected 380 high quality genomes deposited with good metadata, each from a different multi-locus ST, as a non-redundant diversity set for studying S. aureus evolution. In addition to answering basic science questions, Staphopia could serve as a potential platform for rapid clinical diagnostics of S. aureus isolates in the future. The system could also be adapted as a template for other organism-specific databases.

scholarly journals Staphylococcus aureus viewed from the perspective of 40,000+ genomes

2018 ◽  
Author(s):  
Robert A Petit III ◽  
Timothy D Read
Keyword(s):  
Staphylococcus Aureus ◽  
De Novo ◽  
Wide Spectrum ◽  
Low Cost ◽  
Application Programming Interface ◽  
Relevant Information ◽  
Clinical Diagnostics ◽  
Nucleotide Polymorphisms ◽  
Analysis Pipeline ◽  
Antibiotic Resistant

Low-cost Illumina sequencing of clinically-important bacterial pathogens has generated thousands of publicly available genomic datasets. Analyzing these genomes and extracting relevant information for each pathogen and the associated clinical phenotypes requires not only resources and bioinformatic skills but organism-specific knowledge. In light of these issues, we created Staphopia, an analysis pipeline, database and Application Programming Interface, focused on Staphylococcus aureus, a common colonizer of humans and a major antibiotic-resistant pathogen responsible for a wide spectrum of hospital and community-associated infections. Written in Python, Staphopia’s analysis pipeline consists of submodules running open-source tools. It accepts raw FASTQ reads as an input, which undergo quality control filtration, error correction and reduction to a maximum of approximately 100x chromosome coverage. This reduction significantly reduces total runtime without detrimentally affecting the results. The pipeline performs de novo assembly-based and mapping-based analysis. Automated gene calling and annotation is performed on the assembled contigs. Read-mapping is used to call variants (single nucleotide polymorphisms and insertion/deletions) against a reference S. aureus chromosome (Type strain, N315). We ran the analysis pipeline on more than 43,000 S. aureus shotgun Illumina genome projects in the public ENA database in November 2017. We found that only a quarter of known multi-locus sequence types (STs) were represented but the top ten STs made up 70% of all genomes. MRSA (methicillin resistant S. aureus) were 64% of all genomes. Using the Staphopia database we selected 380 high quality genomes deposited with good metadata, each from a different multi-locus sequence type, as a non-redundant diversity set for studying S. aureus evolution. In addition to answering basic science questions, Staphopia could serve as a potential platform for rapid clinical diagnostics of S. aureus isolates in the future. The system could also be adapted as a template for other organism-specific databases.

scholarly journals Staphylococcus aureus viewed from the perspective of 40,000+ genomes

2018 ◽  
Author(s):  
Robert A Petit III ◽  
Timothy D Read
Keyword(s):  
Staphylococcus Aureus ◽  
De Novo ◽  
Wide Spectrum ◽  
Low Cost ◽  
Application Programming Interface ◽  
Relevant Information ◽  
Clinical Diagnostics ◽  
Nucleotide Polymorphisms ◽  
Analysis Pipeline ◽  
Antibiotic Resistant

Low-cost Illumina sequencing of clinically-important bacterial pathogens has generated thousands of publicly available genomic datasets. Analyzing these genomes and extracting relevant information for each pathogen and the associated clinical phenotypes requires not only resources and bioinformatic skills but organism-specific knowledge. In light of these issues, we created Staphopia, an analysis pipeline, database and Application Programming Interface, focused on Staphylococcus aureus, a common colonizer of humans and a major antibiotic-resistant pathogen responsible for a wide spectrum of hospital and community-associated infections. Written in Python, Staphopia’s analysis pipeline consists of submodules running open-source tools. It accepts raw FASTQ reads as an input, which undergo quality control filtration, error correction and reduction to a maximum of approximately 100x chromosome coverage. This reduction significantly reduces total runtime without detrimentally affecting the results. The pipeline performs de novo assembly-based and mapping-based analysis. Automated gene calling and annotation is performed on the assembled contigs. Read-mapping is used to call variants (single nucleotide polymorphisms and insertion/deletions) against a reference S. aureus chromosome (Type strain, N315). We ran the analysis pipeline on more than 43,000 S. aureus shotgun Illumina genome projects in the public ENA database in November 2017. We found that only a quarter of known multi-locus sequence types (STs) were represented but the top ten STs made up 70% of all genomes. MRSA (methicillin resistant S. aureus) were 64% of all genomes. Using the Staphopia database we selected 380 high quality genomes deposited with good metadata, each from a different multi-locus sequence type, as a non-redundant diversity set for studying S. aureus evolution. In addition to answering basic science questions, Staphopia could serve as a potential platform for rapid clinical diagnostics of S. aureus isolates in the future. The system could also be adapted as a template for other organism-specific databases.

scholarly journals Staphylococcus aureus viewed from the perspective of 40,000+ genomes

2018 ◽  
Author(s):  
Robert A Petit III ◽  
Timothy D Read
Keyword(s):  
Staphylococcus Aureus ◽  
De Novo ◽  
Wide Spectrum ◽  
Low Cost ◽  
Application Programming Interface ◽  
Relevant Information ◽  
Clinical Diagnostics ◽  
Nucleotide Polymorphisms ◽  
Analysis Pipeline ◽  
Antibiotic Resistant

Low-cost Illumina sequencing of clinically-important bacterial pathogens has generated thousands of publicly available genomic datasets. Analyzing these genomes and extracting relevant information for each pathogen and the associated clinical phenotypes requires not only resources and bioinformatic skills but organism-specific knowledge. In light of these issues, we created Staphopia, an analysis pipeline, database and Application Programming Interface, focused on Staphylococcus aureus, a common colonizer of humans and a major antibiotic-resistant pathogen responsible for a wide spectrum of hospital and community-associated infections. Written in Python, Staphopia’s analysis pipeline consists of submodules running open-source tools. It accepts raw FASTQ reads as an input, which undergo quality control filtration, error correction and reduction to a maximum of approximately 100x chromosome coverage. This reduction significantly reduces total runtime without detrimentally affecting the results. The pipeline performs de novo assembly-based and mapping-based analysis. Automated gene calling and annotation is performed on the assembled contigs. Read-mapping is used to call variants (single nucleotide polymorphisms and insertion/deletions) against a reference S. aureus chromosome (Type strain, N315). We ran the analysis pipeline on more than 43,000 S. aureus shotgun Illumina genome projects in the public ENA database in November 2017. We found that only a quarter of known multi-locus sequence types (STs) were represented but the top ten STs made up 70% of all genomes. MRSA (methicillin resistant S. aureus) were 64% of all genomes. Using the Staphopia database we selected 380 high quality genomes deposited with good metadata, each from a different multi-locus sequence type, as a non-redundant diversity set for studying S. aureus evolution. In addition to answering basic science questions, Staphopia could serve as a potential platform for rapid clinical diagnostics of S. aureus isolates in the future. The system could also be adapted as a template for other organism-specific databases.

scholarly journals Rapid low-cost assembly of the Drosophila melanogaster reference genome using low-coverage, long-read sequencing

2018 ◽  
Author(s):  
Edwin A. Solares ◽  
Mahul Chakraborty ◽  
Danny E. Miller ◽  
Shannon Kalsow ◽  
Kate Hall ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Genetic Variation ◽  
Large Scale ◽  
Reference Genome ◽  
De Novo ◽  
Low Cost ◽  
Nucleotide Polymorphisms ◽  
Structural Variants ◽  
High Coverage ◽  
Reference Assembly

ABSTRACTAccurate and comprehensive characterization of genetic variation is essential for deciphering the genetic basis of diseases and other phenotypes. A vast amount of genetic variation stems from large-scale sequence changes arising from the duplication, deletion, inversion, and translocation of sequences. In the past 10 years, high-throughput short reads have greatly expanded our ability to assay sequence variation due to single nucleotide polymorphisms. However, a recent de novo assembly of a second Drosophila melanogaster reference genome has revealed that short read genotyping methods miss hundreds of structural variants, including those affecting phenotypes. While genomes assembled using high-coverage long reads can achieve high levels of contiguity and completeness, concerns about cost, errors, and low yield have limited widespread adoption of such sequencing approaches. Here we resequenced the reference strain of D. melanogaster (ISO1) on a single Oxford Nanopore MinION flow cell run for 24 hours. Using only reads longer than 1 kb or with at least 30x coverage, we assembled a highly contiguous de novo genome. The addition of inexpensive paired reads and subsequent scaffolding using an optical map technology achieved an assembly with completeness and contiguity comparable to the D. melanogaster reference assembly. Comparison of our assembly to the reference assembly of ISO1 uncovered a number of structural variants (SVs), including novel LTR transposable element insertions and duplications affecting genes with developmental, behavioral, and metabolic functions. Collectively, these SVs provide a snapshot of the dynamics of genome evolution. Furthermore, our assembly and comparison to the D. melanogaster reference genome demonstrates that high-quality de novo assembly of reference genomes and comprehensive variant discovery using such assemblies are now possible by a single lab for under $1,000 (USD).

scholarly journals The recent evolutionary rescue of a staple crop depended on over half a century of global germplasm exchange

2021 ◽  
Author(s):  
Kebede T Muleta ◽  
Terry Felderhoff ◽  
Noah Winans ◽  
Rachel Walstead ◽  
Jean Rigaud Charles ◽  
...  
Keyword(s):  
Population Genomics ◽  
De Novo ◽  
Low Cost ◽  
Nucleotide Polymorphisms ◽  
Sequencing Data ◽  
Melanaphis Sacchari ◽  
Germplasm Exchange ◽  
Breeding Programs ◽  
A Genome ◽  
Evolutionary Rescue

Rapid environmental change can lead to extinction of populations or evolutionary rescue via genetic adaptation. In the past several years, smallholder and commercial cultivation of sorghum (Sorghum bicolor), a global cereal and forage crop, has been threatened by a global outbreak of an aggressive new biotype of sugarcane aphid (SCA; Melanaphis sacchari). Here we characterized genomic signatures of adaptation in a Haitian sorghum breeding population, which had been recently founded from admixed global germplasm, extensively intercrossed, and subjected to intense selection under SCA infestation. We conducted evolutionary population genomics analyses of 296 post-selection Haitian lines compared to 767 global accessions at 159,683 single nucleotide polymorphisms. Despite intense selection, the Haitian population retains high nucleotide diversity through much of the genome due to diverse founders and an intercrossing strategy. A genome-wide fixation (FST) scan and geographic analyses suggests that adaptation to SCA in the Haiti is conferred by a globally-rare East African allele of RMES1, which has also spread to breeding programs in Africa, Asia, and the Americas. De novo genome sequencing data for SCA resistant and susceptible lines revealed putative causative variants at RMES1. Convenient low-cost markers were developed from the RMES1 selective sweep and successfully predicted resistance in independent U.S. x African breeding lines and eight U.S. commercial and public breeding programs, demonstrating the global relevance of the findings. Together, the findings highlight the potential of evolutionary genomics to develop adaptive trait breeding technology and the value of global germplasm exchange to facilitate evolutionary rescue.

scholarly journals Mutations in a membrane permease or hpt lead to 6-thioguanine resistance in Staphylococcus aureus

2021 ◽  
Author(s):  
Denny Chin ◽  
Mariya I. Goncheva ◽  
Ronald S. Flannagan ◽  
David E. Heinrichs
Keyword(s):  
Staphylococcus Aureus ◽  
De Novo ◽  
Toxin Production ◽  
Lesion Size ◽  
Missense Mutations ◽  
Nucleotide Polymorphisms ◽  
Coagulase Negative Staphylococci ◽  
Evolution Of Resistance ◽  
High Concentrations

We recently discovered that 6-thioguanine (6-TG) is an anti-virulence compound that is produced by a number of coagulase negative staphylococci. In Staphylococcus aureus , it inhibits de novo purine biosynthesis and ribosomal protein expression, thus inhibiting growth and abrogating toxin production. Mechanisms by which S. aureus may develop resistance to this compound are currently unknown. Here, we show that 6-TG-resistant S. aureus mutants emerge spontaneously when the bacteria are subjected to high concentrations of 6-TG in vitro . Whole genome sequencing of these mutants revealed frameshift and missense mutations in a xanthine-uracil permease family protein ( stgP : s ix t hio g uanine p ermease) and single nucleotide polymorphisms in hypoxanthine phosphoribosyltransferase ( hpt ). These mutations engender S. aureus the ability to resist both the growth inhibitory and toxin down regulation effects of 6-TG. While prophylactic administration of 6-TG ameliorates necrotic lesions in subcutaneous infection of mice with MRSA strain USA300-LAC, the drug did not reduce lesion size formed by the 6-TG resistant strains. These findings identify mechanisms of 6-TG resistance and this information can be leveraged to inform strategies to slow the evolution of resistance.

scholarly journals Rapid and Sensitive Detection of miRNA Based on AC Electrokinetic Capacitive Sensing for Point-of-Care Applications

Sensors ◽  
2021 ◽  
Vol 21 (12) ◽  
pp. 3985
Author(s):  
Nan Wan ◽  
Yu Jiang ◽  
Jiamei Huang ◽  
Rania Oueslati ◽  
Shigetoshi Eda ◽  
...  
Keyword(s):  
Limit Of Detection ◽  
Point Of Care ◽  
Low Cost ◽  
Clinical Diagnostics ◽  
Detection Methods ◽  
Capacitive Sensing ◽  
Application Potential ◽  
Mirna Detection ◽  
Mirna 25 ◽  
Low Sensitivity

A sensitive and efficient method for microRNAs (miRNAs) detection is strongly desired by clinicians and, in recent years, the search for such a method has drawn much attention. There has been significant interest in using miRNA as biomarkers for multiple diseases and conditions in clinical diagnostics. Presently, most miRNA detection methods suffer from drawbacks, e.g., low sensitivity, long assay time, expensive equipment, trained personnel, or unsuitability for point-of-care. New methodologies are needed to overcome these limitations to allow rapid, sensitive, low-cost, easy-to-use, and portable methods for miRNA detection at the point of care. In this work, to overcome these shortcomings, we integrated capacitive sensing and alternating current electrokinetic effects to detect specific miRNA-16b molecules, as a model, with the limit of detection reaching 1.0 femto molar (fM) levels. The specificity of the sensor was verified by testing miRNA-25, which has the same length as miRNA-16b. The sensor we developed demonstrated significant improvements in sensitivity, response time and cost over other miRNA detection methods, and has application potential at point-of-care.

scholarly journals De Novo SNP Discovery and Genotyping of Iranian Pimpinella Species Using ddRAD Sequencing

Agronomy ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 1342
Author(s):  
Shaghayegh Mehravi ◽  
Gholam Ali Ranjbar ◽  
Ghader Mirzaghaderi ◽  
Anita Alice Severn-Ellis ◽  
Armin Scheben ◽  
...  
Keyword(s):  
De Novo ◽  
Genetic Relationships ◽  
Nucleotide Polymorphisms ◽  
High Quality ◽  
Genomic Resources ◽  
High Quality Snps ◽  
The Family ◽  
Double Digestion ◽  
Flanking Sequences ◽  
Downstream Analysis

The species of Pimpinella, one of the largest genera of the family Apiaceae, are traditionally cultivated for medicinal purposes. In this study, high-throughput double digest restriction-site associated DNA sequencing technology (ddRAD-seq) was used to identify single nucleotide polymorphisms (SNPs) in eight Pimpinella species from Iran. After double-digestion with the enzymes HpyCH4IV and HinfI, a total of 334,702,966 paired-end reads were de novo assembled into 1,270,791 loci with an average of 28.8 reads per locus. After stringent filtering, 2440 high-quality SNPs were identified for downstream analysis. Analysis of genetic relationships and population structure, based on these retained SNPs, indicated the presence of three major groups. Gene ontology and pathway analysis were determined by using comparison SNP-associated flanking sequences with a public non-redundant database. Due to the lack of genomic resources in this genus, our present study is the first report to provide high-quality SNPs in Pimpinella based on a de novo analysis pipeline using ddRAD-seq. This data will enhance the molecular knowledge of the genus Pimpinella and will provide an important source of information for breeders and the research community to enhance breeding programs and support the management of Pimpinella genomic resources.

scholarly journals Characterization of metabolic responses, genetic variations, and microsatellite instability in ammonia-stressed CHO cells grown in fed-batch cultures

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Dylan G. Chitwood ◽  
Qinghua Wang ◽  
Kathryn Elliott ◽  
Aiyana Bullock ◽  
Dwon Jordana ◽  
...  
Keyword(s):  
Microsatellite Instability ◽  
Microsatellite Loci ◽  
Cho Cells ◽  
Genome Stability ◽  
De Novo ◽  
Genome Instability ◽  
Chinese Hamster ◽  
Functional Genes ◽  
Nucleotide Polymorphisms ◽  
De Novo Mutations

Abstract Background As bioprocess intensification has increased over the last 30 years, yields from mammalian cell processes have increased from 10’s of milligrams to over 10’s of grams per liter. Most of these gains in productivity can be attributed to increasing cell densities within bioreactors. As such, strategies have been developed to minimize accumulation of metabolic wastes, such as lactate and ammonia. Unfortunately, neither cell growth nor biopharmaceutical production can occur without some waste metabolite accumulation. Inevitably, metabolic waste accumulation leads to decline and termination of the culture. While it is understood that the accumulation of these unwanted compounds imparts a suboptimal culture environment, little is known about the genotoxic properties of these compounds that may lead to global genome instability. In this study, we examined the effects of high and moderate extracellular ammonia on the physiology and genomic integrity of Chinese hamster ovary (CHO) cells. Results Through whole genome sequencing, we discovered 2394 variant sites within functional genes comprised of both single nucleotide polymorphisms and insertion/deletion mutations as a result of ammonia stress with high or moderate impact on functional genes. Furthermore, several of these de novo mutations were found in genes whose functions are to maintain genome stability, such as Tp53, Tnfsf11, Brca1, as well as Nfkb1. Furthermore, we characterized microsatellite content of the cultures using the CriGri-PICR Chinese hamster genome assembly and discovered an abundance of microsatellite loci that are not replicated faithfully in the ammonia-stressed cultures. Unfaithful replication of these loci is a signature of microsatellite instability. With rigorous filtering, we found 124 candidate microsatellite loci that may be suitable for further investigation to determine whether these loci may be reliable biomarkers to predict genome instability in CHO cultures. Conclusion This study advances our knowledge with regards to the effects of ammonia accumulation on CHO cell culture performance by identifying ammonia-sensitive genes linked to genome stability and lays the foundation for the development of a new diagnostic tool for assessing genome stability.

A pragmatic eLCR for an ultrasensitive detection of methicillin-resistant Staphylococcus aureus in joint synovial fluid: superior to qPCR

The Analyst ◽  
2021 ◽  
Author(s):  
Jinyuan Chen ◽  
Hongxiang Wei ◽  
Xinyu Fang ◽  
Yuanqing Cai ◽  
Zhenzhen Zhang ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Synovial Fluid ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
Detection Method ◽  
Low Cost ◽  
Ultrasensitive Detection ◽  
Gene Detection ◽  
Methicillin Resistant ◽  
Cost Identification

A pragmatic electrochemical mecA gene detection method for a rapid, accurate and low-cost identification of MRSA in the joint synovial fluid of PJI patients.

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