Safety Evaluation and Whole Genome Sequencing of Aspergillus japonicas PJ01 Reveal Its Potential to Degrade Citrus Segments in Juice Processing

Aspergillus japonicas PJ01 (A. japonicas PJ01) is a strain isolated from the rotten branches. In previ-ous studies, it was shown that it can produce complex enzymes to degrade polysaccharide com-ponents. In this study, we evaluated the safety of its crude enzyme solution. Acute oral toxicity, subchronic toxicity, micronucleus and sperm malformation tests all validated the high biologi-cal safety for the crude enzymes. Secondly, we carried out the citrus segment degradation ex-periment of crude enzyme solution. Compared with the control group, the crude enzyme solu-tion of A. japonicas PJ01 can completely degrade the segments in 50 min, which provides the basis for enzymatic peeling during juice processing. The whole genome sequencing showed that the genome of A. japonicus PJ01 has a GC content of 51.37% with a size of 36204647 bp, and encoded 10070 genes. GO, COG, KEGG and CAZy databases were used in gene annotation analyses. Pathway enrichment showed many genes related to carbohydrate metabolism, rich in genes re-lated to pectinase, xylanase and carboxylcellulase. Therefore, the complex enzyme produced by A. japonicus PJ01 can be used in gizzard juice processing to achieve efficient enzymatic decapsu-lation.

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Evaluating coverage bias in next-generation sequencing of Escherichia coli

PLoS ONE ◽

10.1371/journal.pone.0253440 ◽

2021 ◽

Vol 16 (6) ◽

pp. e0253440

Author(s):

Samantha Gunasekera ◽

Sam Abraham ◽

Marc Stegger ◽

Stanley Pang ◽

Penghao Wang ◽

...

Keyword(s):

Escherichia Coli ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

De Novo ◽

Fragment Size ◽

Gc Content ◽

Main Concern ◽

Whole Genome ◽

Assembly Quality ◽

Coverage Bias

Whole-genome sequencing is essential to many facets of infectious disease research. However, technical limitations such as bias in coverage and tagmentation, and difficulties characterising genomic regions with extreme GC content have created significant obstacles in its use. Illumina has claimed that the recently released DNA Prep library preparation kit, formerly known as Nextera Flex, overcomes some of these limitations. This study aimed to assess bias in coverage, tagmentation, GC content, average fragment size distribution, and de novo assembly quality using both the Nextera XT and DNA Prep kits from Illumina. When performing whole-genome sequencing on Escherichia coli and where coverage bias is the main concern, the DNA Prep kit may provide higher quality results; though de novo assembly quality, tagmentation bias and GC content related bias are unlikely to improve. Based on these results, laboratories with existing workflows based on Nextera XT would see minor benefits in transitioning to the DNA Prep kit if they were primarily studying organisms with neutral GC content.

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Whole-Genome Sequencing of Mycobacterium tilburgii Strain MEPHI

Microbiology Resource Announcements ◽

10.1128/mra.00933-19 ◽

2019 ◽

Vol 8 (40) ◽

Author(s):

Jamal Saad ◽

Michel Drancourt ◽

Margaret M. Hannan ◽

Patrick J. Stapleton ◽

Simon Grandjean Lapierre

Keyword(s):

Whole Genome Sequencing ◽

Clinical Isolate ◽

Genome Sequencing ◽

Gc Content ◽

Whole Genome ◽

Content Type ◽

Phylogenetic Placement ◽

Mycobacterium Simiae

Mycobacterium tilburgii is a fastidious mycobacterium which has previously been reported to cause severe disseminated infections. Genome sequencing of the M. tilburgii MEPHI clinical isolate yielded 3.14 Mb, with 66.3% GC content, and confirmed phylogenetic placement within the Mycobacterium simiae complex.

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Whole Genome Sequencing Assembly and Annotation of Halobacillus Trueperi S61 Isolated From the Qarhan Salt Lake

10.21203/rs.3.rs-1188620/v1 ◽

2022 ◽

Author(s):

Wei Li ◽

Shuo Shen ◽

Jian Wang

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Protein Function ◽

Salt Lake ◽

Gc Content ◽

Whole Genome ◽

Biological Processes ◽

Circular Chromosome ◽

Advanced Analysis ◽

Qarhan Salt Lake

Abstract Background: Halophilic microbial as prospective resources of biotechnology due to the advantages of flexible survivability. Qarhan Salt Lake is the second largest Salt Lake in the world which contains rich-unique extremophiles and deserved in-depth exploration. Results: Present study first time isolated novel strain Halobacillus trueperi S61 from Qarhan Salt Lake and performed whole-genome sequencing through combined third-generation PacBio and second-generation Illumina technology. The whole genome of Halobacillus trueperi S61 identified 57549 total reads and consists a complete circular chromosome of 4047887 bp with 43.86% GC content without gaps. Total number of 139 non-coding RNA (included 86 tRNA, 30 rRNA and 23 sRNA), 16 gene islands with 260275 bp and two prophages (with 82682 length) were predicted. In addition, the whole genome of Halobacillus trueperi S61 summarized basic annotation for 3982 protein-coding genes, 3980, 3667, 2998 and 2303 unigenes were annotated with Nr, Swissport, KOG and KEGG database. Combined with advanced analysis, 561 carbohydrate enzymes and 4416 pathogen host interactions related genes were identified. The protein function of Halobacillus trueperi S61 was mainly focus on biological processes, and the protein function was mainly distributed in gene transcription and amino acids, and carbohydrates metabolism. Conclusions: The complete whole genome sequence assembly and annotation of novel strain Halobacillus trueperi S61 isolated from Qarhan Salt Lake mainly focus on protein biological processes and antibiotic resistance, provides a potential resource for biotechnology.

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Validation and Implementation of CLIA-Compliant Whole Genome Sequencing (WGS) in Public Health Laboratory

10.1101/107003 ◽

2017 ◽

Author(s):

Varvara K. Kozyreva ◽

Chau-Linda Truong ◽

Alexander L. Greninger ◽

John Crandall ◽

Rituparna Mukhopadhyay ◽

...

Keyword(s):

Public Health ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Gc Content ◽

End Users ◽

Whole Genome ◽

Base Calling ◽

Specificity And Sensitivity ◽

Wide Range ◽

Performance Specifications

AbstractBackgroundPublic health microbiology laboratories (PHL) are at the cusp of unprecedented improvements in pathogen identification, antibiotic resistance detection, and outbreak investigation by using whole genome sequencing (WGS). However, considerable challenges remain due to the lack of common standards.Objectives1) Establish the performance specifications of WGS applications used in PHL to conform with CLIA (Clinical Laboratory Improvements Act) guidelines for laboratory developed tests (LDT), 2) Develop quality assurance (QA) and quality control (QC) measures, 3) Establish reporting language for end users with or without WGS expertise, 4) Create a validation set of microorganisms to be used for future validations of WGS platforms and multi-laboratory comparisons and, 5) Create modular templates for the validation of different sequencing platforms.MethodsMiSeq Sequencer and Illumina chemistry (Illumina, Inc.) were used to generate genomes for 34 bacterial isolates with genome sizes from 1.8 to 4.7 Mb and wide range of GC content (32.1%-66.1%). A customized CLCbio Genomics Workbench - shell script bioinformatics pipeline was used for the data analysis.ResultsWe developed a validation panel comprising ten Enterobacteriaceae isolates, five gram-positive cocci, five gram-negative non-fermenting species, nine Mycobacterium tuberculosis, and five miscellaneous bacteria; the set represented typical workflow in the PHL. The accuracy of MiSeq platform for individual base calling was >99.9% with similar results shown for reproducibility/repeatability of genome-wide base calling. The accuracy of phylogenetic analysis was 100%. The specificity and sensitivity inferred from MLST and genotyping tests were 100%. A test report format was developed for the end users with and without WGS knowledge.ConclusionWGS was validated for routine use in PHL according to CLIA guidelines for LDTs. The validation panel, sequencing analytics, and raw sequences will be available for future multi-laboratory comparisons of WGS in PHL. Additionally, the WGS performance specifications and modular validation template are likely to be adaptable for the validation of other platforms and reagents kits.

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Non-invasive detection of urothelial carcinoma (UC) by cost-effective low-coverage whole genome sequencing from urine exfoliated cells DNA.

Journal of Clinical Oncology ◽

10.1200/jco.2020.38.15_suppl.1552 ◽

2020 ◽

Vol 38 (15_suppl) ◽

pp. 1552-1552

Author(s):

Shuxiong Zeng ◽

Dongxing Nai ◽

Yingdie Ye ◽

Jiatao Ji ◽

Zunlin Zhou ◽

...

Keyword(s):

Urothelial Carcinoma ◽

Whole Genome Sequencing ◽

Cancer Patients ◽

Genome Sequencing ◽

Chromosomal Aberrations ◽

Cost Effective ◽

Urine Samples ◽

Control Group ◽

Whole Genome ◽

Exfoliated Cells

1552 Background: Urothelial carcinoma (UC) is a malignancy with frequent chromosomal aberrations. The FISH assays were more sensitive as compared to cytology tests. Here we investigated cost-effective whole genome sequencing technology, which is able to detect all chromosomal aberrations for UC diagnoses. Methods: UC patients and control group are prospectively recruited in trial NCT03998371. First-morning-voided urine were freshly collected before TURBT or cystectomy. Urine Exfoliated Cells DNA was analyzed by illumina HiSeq X10, followed by genotyping by bioinformatics workflow UCAD. Results: 195 individuals were prospectively recruited. 121 UC patients and 67 non-tumor diseases were included in this study. 7 other malignancies as confirmed by pathological testing were excluded. Frequent chromosome copy number changes were found in cancer patients as compared non-tumor controls, including chromosome 3 gain, 17 gain, 7 gain and 9p loss used in FISH assays were found. In addition to that, chr9q loss, 8q gain, 5q loss, 17p loss, 11p loss, 1q gain, 8p loss, 10q loss, 6q loss, 4q loss and 11q loss were also frequent in cancer patients (AUC > 0.65). Metacentric chromosomes showed better AUC compared to acrocentric and telocentric chromosomes (P = 1.7e-03). A novel diagnosis model UCAD was built by incorporating all the chromosomal changes. The model reached performance of AUC = 0.933. At the optimal cutoff |Z| > = 3.16, the sensitivity, specificity and accuracy were 84.7%, 97.9% and 89.0% respectively. The prediction positivity was found correlated with urine microscopy visible epithelial cells (P = 0.00069), tumor invasiveness (Ta/Tis vs the other, P = 0.0048) and tumor grade (P = 0.0030), but not microscopy RBC/WBC findings, urine culture findings, smoke and drinking history. The UCAD model outperformed cytology tests by predicting all 16-cytology positive and 12 cytology negative tumors with comparable specificity. The model found 75.0% more tumors. And UCAD identified more upper urinary tract cancer (P = 0.012) and smaller tumors ( < 3cm, P = 5.9e-04). The adding of cytology to UCAD did not improve diagnosing sensitivity and specificity. UCAD reproduce the diagnoses among morning - void urine, morning, afternoon urine samples with correlation coefficient R2> 0.98. All the urine samples showed high concordances with matched tumor samples (R2> 0.85). Conclusions: UCAD could be a high specific, robust UC diagnoses method with improved sensitivity as compared to cytology tests. Clinical trial information: NCT03998371.

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Characterization of multidrug-resistant Acinetobacter baumannii strain ATCC BAA1605 using whole-genome sequencing

BMC Research Notes ◽

10.1186/s13104-021-05493-z ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Kah Ern Ten ◽

Muhammad Zarul Hanifah Md Zoqratt ◽

Qasim Ayub ◽

Hock Siew Tan

Keyword(s):

Whole Genome Sequencing ◽

Acinetobacter Baumannii ◽

Genome Sequencing ◽

Gc Content ◽

Multidrug Resistant ◽

Genomic Islands ◽

Whole Genome ◽

Strain Atcc ◽

Circular Chromosome ◽

Genome Data

Abstract Objective The nosocomial pathogen, Acinetobacter baumannii, has acquired clinical significance due to its ability to persist in hospital settings and survive antibiotic treatment, which eventually resulted in the rapid spread of this bacterium with antimicrobial resistance (AMR) phenotypes. This study used a multidrug-resistant A. baumannii (strain ATCC BAA1605) as a model to study the genomic features of this pathogen. Results One circular chromosome and one circular plasmid were discovered in the complete genome of A. baumannii ATCC BAA1605 using whole-genome sequencing. The chromosome is 4,039,171 bp long with a GC content of 39.24%. Many AMR genes, which confer resistance to major classes of antibiotics (beta-lactams, aminoglycosides, tetracycline, sulphonamides), were found on the chromosome. Two genomic islands were predicted on the chromosome, one of which (Genomic Island 1) contains a cluster of AMR genes and mobile elements, suggesting the possibility of horizontal gene transfer. A subtype I-F CRISPR-Cas system was also identified on the chromosome of A. baumannii ATCC BAA1605. This study provides valuable genome data that can be used as a reference for future studies on A. baumannii. The genome of A. baumannii ATCC BAA1605 has been deposited at GenBank under accession no. CP058625 and CP058626.

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Whole-genome sequencing and gene annotation of Georgian grape cultivars

Acta Horticulturae ◽

10.17660/actahortic.2019.1248.41 ◽

2019 ◽

pp. 287-294

Author(s):

V. Tabidze ◽

I. Pipia ◽

M. Gogniashvili ◽

N. Kunelauri ◽

T. Beridze

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Gene Annotation ◽

Whole Genome ◽

Grape Cultivars

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Retrospective evaluation of whole exome and genome mutation calls in 746 cancer samples

Nature Communications ◽

10.1038/s41467-020-18151-y ◽

2020 ◽

Vol 11 (1) ◽

Author(s):

Matthew H. Bailey ◽

◽

William U. Meyerson ◽

Lewis Jonathan Dursi ◽

Liang-Bo Wang ◽

...

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Gc Content ◽

Cancer Genome ◽

The Cancer Genome Atlas ◽

Whole Genome Sequencing Data ◽

Whole Genome ◽

Sequencing Data ◽

Whole Exome ◽

Cancer Genome Atlas

AbstractThe Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) curated consensus somatic mutation calls using whole exome sequencing (WES) and whole genome sequencing (WGS), respectively. Here, as part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium, which aggregated whole genome sequencing data from 2,658 cancers across 38 tumour types, we compare WES and WGS side-by-side from 746 TCGA samples, finding that ~80% of mutations overlap in covered exonic regions. We estimate that low variant allele fraction (VAF < 15%) and clonal heterogeneity contribute up to 68% of private WGS mutations and 71% of private WES mutations. We observe that ~30% of private WGS mutations trace to mutations identified by a single variant caller in WES consensus efforts. WGS captures both ~50% more variation in exonic regions and un-observed mutations in loci with variable GC-content. Together, our analysis highlights technological divergences between two reproducible somatic variant detection efforts.

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Squamate Chromosome Size and GC Content Assessed by Flow Karyotyping

Cytogenetic and Genome Research ◽

10.1159/000497265 ◽

2019 ◽

Vol 157 (1-2) ◽

pp. 46-52 ◽

Cited By ~ 1

Author(s):

Fumio Kasai ◽

Patricia C.M. O'Brien ◽

Malcolm A. Ferguson-Smith

Keyword(s):

Gene Mapping ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Chromosome Painting ◽

Karyotype Evolution ◽

Gc Content ◽

Whole Genome ◽

Chromosome Size ◽

The Relationship ◽

Reptilian Species

Chromosome homologies in reptiles have been investigated extensively by gene mapping and chromosome painting. Relative chromosome size can be estimated roughly from conventional karyotypes, but chromosome GC content cannot be evaluated by any of these approaches. However, GC content can be obtained by whole-genome sequencing, although complete data are available only for a limited number of reptilian species. Chromosomes can be characterized by size and GC content in bivariate flow karyotypes, in which the distribution of peaks represents the differences. We have analysed flow karyotypes from 9 representative squamate species and show chromosome profiles for each species based on the relationship between size and GC content. Our results reveal that the GC content of macrochromosomes is invariable in the 9 species. A higher GC content was found in microchromosomes, similar to profiles previously determined in crocodile, turtle, and chicken. The findings suggest that karyotype evolution in reptiles is characterized by unique features of chromosome GC content.

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