A Metagenomic Nanopore Sequence Analysis Combined with Conventional Screening and Spectroscopic Methods for Deciphering the Antimicrobial Metabolites Produced by Alcaligenes faecalis Soil Isolate MZ921504

Eltokhy, Mohamed A; Saad, Bishoy T; Eltayeb, Wafaa N; El-Ansary, Mona R; Aboshanab, Khaled M.; Ashour, M. S. E.

doi:10.3390/antibiotics10111382

Cited by 4 publications

(5 citation statements)

References 40 publications

(71 reference statements)

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“…However, the use of NGS technologies in clinical diagnostics necessitates a large initial investment in the sequencer, which is a barrier for local research institutions in underdeveloped nations, as well as small research institutes and hospitals. MinION, the first commercially available sequencer based on Nanopore technology, might be a viable alternative [ 43 , 44 ]. MinION has previously been utilized effectively to identify mutations in TP53 and ABL1 genes in CLL and CML patients [ 45 , 46 , 47 , 48 ], respectively.…”

Section: Discussionmentioning

confidence: 99%

Evaluation of the Available Variant Calling Tools for Oxford Nanopore Sequencing in Breast Cancer

Helal¹,

Saad²,

Saad³

et al. 2022

Genes

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The goal of biomarker testing, in the field of personalized medicine, is to guide treatments to achieve the best possible results for each patient. The accurate and reliable identification of everyone’s genome variants is essential for the success of clinical genomics, employing third-generation sequencing. Different variant calling techniques have been used and recommended by both Oxford Nanopore Technologies (ONT) and Nanopore communities. A thorough examination of the variant callers might give critical guidance for third-generation sequencing-based clinical genomics. In this study, two reference genome sample datasets (NA12878) and (NA24385) and the set of high-confidence variant calls provided by the Genome in a Bottle (GIAB) were used to allow the evaluation of the performance of six variant calling tools, including Human-SNP-wf, Clair3, Clair, NanoCaller, Longshot, and Medaka, as an integral step in the in-house variant detection workflow. Out of the six variant callers understudy, Clair3 and Human-SNP-wf that has Clair3 incorporated into it achieved the highest performance rates in comparison to the other variant callers. Evaluation of the results for the tool was expressed in terms of Precision, Recall, and F1-score using Hap.py tools for the comparison. In conclusion, our findings give important insights for identifying accurate variants from third-generation sequencing of personal genomes using different variant detection tools available for long-read sequencing.

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Section: Discussionmentioning

confidence: 99%

Evaluation of the Available Variant Calling Tools for Oxford Nanopore Sequencing in Breast Cancer

Helal¹,

Saad²,

Saad³

et al. 2022

Genes

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“…4.0 to guarantee there is enough pure genomic material before the sequencing run, 400 ng/7 μL (55 ng/μL), as mentioned by Oxford nanopore manual. Metagenomics were performed at HITS Solutions Co (Bioinformatics Department, Cairo, Egypt, accessed on 5 December 2021) [ 15 ].…”

Section: Methodsmentioning

confidence: 99%

“…Before loading on the flow cell, a total of 34 μL of sequencing Buffer and 25.5 μL of loading Beads were added to 12 μL of the DNA libraries and 4.5 μL nuclease free water. After that, priming and loading onto FLO-MIN106 flow cell was performed [ 15 ].…”

Section: Methodsmentioning

confidence: 99%

“…A centrifuge was used to classify sequencing reads to a taxonomic identifier, as previously reported [ 60 ]. Results were visualized using re-centrifuged [ 15 ].…”

Section: Methodsmentioning

confidence: 99%

“…Metagenomics analysis identifies all DNA present in a given community [ 13 ]. Metagenomics made it possible to discover antimicrobials from natural sources [ 14 ] as bioinformatics analysis and systematic characterization made it possible to explore the hidden biosynthetic gene clusters [ 15 ].…”

Section: Introductionmentioning

confidence: 99%

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Exploring the Nature of the Antimicrobial Metabolites Produced by Paenibacillus ehimensis Soil Isolate MZ921932 Using a Metagenomic Nanopore Sequencing Coupled with LC-Mass Analysis

Eltokhy

Saad²,

Eltayeb

et al. 2021

Antibiotics

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The continuous emergence of multidrug-resistant (MDR) pathogens poses a global threat to public health. Accordingly, global efforts are continuously conducted to find new approaches to infection control by rapidly discovering antibiotics, particularly those that retain activities against MDR pathogens. In this study, metagenomic nanopore sequence analysis coupled with spectroscopic methods has been conducted for rapid exploring of the various active metabolites produced by Paenibacillus ehimensis soil isolate. Preliminary soil screening resulted in selection of a Gram-positive isolate identified via 16S ribosomal RNA gene sequencing as Paenibacillus ehimensis MZ921932. The isolate showed a broad range of activity against MDR Gram-positive, Gram-negative, and Candida spp. A metagenomics sequence analysis of the soil sample harboring Paenibacillus ehimensis isolate MZ921932 (NCBI GenBank accession PRJNA785410) revealed the presence of conserved biosynthetic gene clusters of petrobactin, tridecaptin, locillomycin (β-lactone), polymyxin, and macrobrevin (polyketides). The liquid chromatography/mass (LC/MS) analysis of the Paenibacillus ehimensis metabolites confirmed the presence of petrobactin, locillomycin, and macrobrevin. In conclusion, Paenibacillus ehimensis isolate MZ921932 is a promising rich source for broad spectrum antimicrobial metabolites. The metagenomic nanopore sequence analysis was a rapid, easy, and efficient method for the preliminary detection of the nature of the expected active metabolites. LC/MS spectral analysis was employed for further confirmation of the nature of the respective active metabolites.

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Benchmarking long-read aligners and SV callers for structural variation detection in Oxford nanopore sequencing data

Helal,

Saad,

Saad

et al. 2024

Sci Rep

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Structural variants (SVs) are one of the significant types of DNA mutations and are typically defined as larger-than-50-bp genomic alterations that include insertions, deletions, duplications, inversions, and translocations. These modifications can profoundly impact the phenotypic characteristics and contribute to disorders like cancer, response to treatment, and infections. Four long-read aligners and five SV callers have been evaluated using three Oxford Nanopore NGS human genome datasets in terms of precision, recall, and F1-score statistical metrics, depth of coverage, and speed of analysis. The best SV caller regarding recall, precision, and F1-score when matched with different aligners at different coverage levels tend to vary depending on the dataset and the specific SV types being analyzed. However, based on our findings, Sniffles and CuteSV tend to perform well across different aligners and coverage levels, followed by SVIM, PBSV, and SVDSS in the last place. The CuteSV caller has the highest average F1-score (82.51%) and recall (78.50%), and Sniffles has the highest average precision value (94.33%). Minimap2 as an aligner and Sniffles as an SV caller act as a strong base for the pipeline of SV calling because of their high speed and reasonable accomplishment. PBSV has a lower average F1-score, precision, and recall and may generate more false positives and overlook some actual SVs. Our results are valuable in the comprehensive evaluation of popular SV callers and aligners as they provide insight into the performance of several long-read aligners and SV callers and serve as a reference for researchers in selecting the most suitable tools for SV detection.

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A Metagenomic Nanopore Sequence Analysis Combined with Conventional Screening and Spectroscopic Methods for Deciphering the Antimicrobial Metabolites Produced by Alcaligenes faecalis Soil Isolate MZ921504

Cited by 4 publications

References 40 publications

Evaluation of the Available Variant Calling Tools for Oxford Nanopore Sequencing in Breast Cancer

Evaluation of the Available Variant Calling Tools for Oxford Nanopore Sequencing in Breast Cancer

Exploring the Nature of the Antimicrobial Metabolites Produced by Paenibacillus ehimensis Soil Isolate MZ921932 Using a Metagenomic Nanopore Sequencing Coupled with LC-Mass Analysis

Benchmarking long-read aligners and SV callers for structural variation detection in Oxford nanopore sequencing data

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