Unveiling microbial diversity: harnessing long-read sequencing technology

Nature Methods, Journal Year: 2024, Volume and Issue: 21(6), P. 954 - 966

Published: April 30, 2024

Language: Английский

---

Recommendations for the introduction of metagenomic high-throughput sequencing in clinical virology, part I: Wet lab procedure

Journal of Clinical Virology, Journal Year: 2020, Volume and Issue: 134, P. 104691 - 104691

Published: Nov. 18, 2020

Metagenomic high-throughput sequencing (mHTS) is a hypothesis-free, universal pathogen detection technique for determination of the DNA/RNA sequences in variety sample types and infectious syndromes. mHTS still its early stages translating into clinical application. To support development, implementation standardization procedures virus diagnostics, European Society Clinical Virology (ESCV) Network on Next-Generation Sequencing (ENNGS) has been established. The aim ENNGS to bring together professionals involved viral diagnostics share methodologies experiences, develop application recommendations. This manuscript aims provide practical recommendations wet lab necessary give development validation laboratory methods, including quality assurance, control assessment protocols.

Language: Английский

---

Recommendations for the introduction of metagenomic next-generation sequencing in clinical virology, part II: bioinformatic analysis and reporting

Journal of Clinical Virology, Journal Year: 2021, Volume and Issue: 138, P. 104812 - 104812

Published: March 26, 2021

Metagenomic next-generation sequencing (mNGS) is an untargeted technique for determination of microbial DNA/RNA sequences in a variety sample types from patients with infectious syndromes. mNGS still its early stages broader translation into clinical applications. To further support the development, implementation, optimization and standardization procedures virus diagnostics, European Society Clinical Virology (ESCV) Network on Next-Generation Sequencing (ENNGS) has been established. The aim ENNGS to bring together professionals involved viral diagnostics share methodologies experiences, develop application guidelines. Following publication Recommendations introduction virology, part I: wet lab procedure this journal, current manuscript aims provide practical recommendations bioinformatic analysis data reporting results clinicians.

Language: Английский

---

BugSeq: a highly accurate cloud platform for long-read metagenomic analyses

BMC Bioinformatics, Journal Year: 2021, Volume and Issue: 22(1)

Published: March 25, 2021

Abstract Background As the use of nanopore sequencing for metagenomic analysis increases, tools capable performing long-read taxonomic classification (ie. determining composition a sample) in fast and accurate manner are needed. Existing were either designed short-read data (eg. Centrifuge), take days to analyse modern sequencer outputs MetaMaps) or suffer from suboptimal accuracy CDKAM). Additionally, all require command line expertise do not scale cloud. Results We present BugSeq, novel, highly classifier reads. evaluate BugSeq on simulated data, mock microbial communities real clinical samples. On ZymoBIOMICS Even Log communities, (F1 = 0.95 at species level) offers better read than MetaMaps 0.89–0.94) fraction time. significantly improves Centrifuge 0.79–0.93) CDKAM 0.91–0.94) while offering competitive run times. When applied 41 samples patients with lower respiratory tract infections, produces greater concordance microbiological culture qPCR compared “What’s In My Pot” analysis. Conclusion is deployed cloud easy scalable analyses. freely available non-commercial https://bugseq.com/free .

Language: Английский

---

The human “contaminome”: bacterial, viral, and computational contamination in whole genome sequences from 1000 families

Scientific Reports, Journal Year: 2022, Volume and Issue: 12(1)

Published: June 14, 2022

Abstract The unmapped readspace of whole genome sequencing data tends to be large but is often ignored. We posit that it contains valuable signals both human infection and contamination. Using poorly aligned reads from sequences (WGS) over 1000 families nearly 5000 individuals, we present insights into common viral, bacterial, computational contamination plague studies. several notable results: (1) In addition known contaminants such as Epstein-Barr virus phiX, blood lymphocyte cell lines contain many other contaminants, likely originating storage, prep, pipelines. (2) Sequencing plate biological sample source a strongly influence profile. And, (3) Y-chromosome fragments not on the reference commonly mismap bacterial genomes. Both experiment-derived prominent in next-generation data. Such can compromise results WGS well metagenomics studies, standard protocols for identifying removing should developed ensure fidelity sequencing-based

Language: Английский

---

The Skin Microbiome: Current Techniques, Challenges, and Future Directions

Microorganisms, Journal Year: 2023, Volume and Issue: 11(5), P. 1222 - 1222

Published: May 6, 2023

Skin acts as a barrier that promotes the colonization of bacteria, fungi, archaea, and viruses whose membership function may differ depending on various specialized niches or micro-environments skin. The group microorganisms inhabiting skin, also known skin microbiome, offers protection against pathogens while actively interacting with host's immune system. Some members microbiome can act opportunistic pathogens. is influenced by factors such site, birth mode, genetics, environment, products, conditions. association(s) health disease has (have) been identified characterized via culture-dependent culture-independent methods. Culture-independent methods (such high-throughput sequencing), in particular, have expanded our understanding microbiome's role maintaining promoting disease. However, intrinsic challenges associated low microbial biomass high host content samples hindered advancements field. In addition, limitations current collection extraction biases derived from sample preparation analysis significantly results conclusions many studies. Therefore, present review discusses technical processing samples, advantages disadvantages sequencing approaches, potential future areas focus for

Language: Английский

---

Evaluation of the Impact of Concentration and Extraction Methods on the Targeted Sequencing of Human Viruses from Wastewater

Environmental Science & Technology, Journal Year: 2024, Volume and Issue: 58(19), P. 8239 - 8250

Published: May 1, 2024

Sequencing human viruses in wastewater is challenging due to their low abundance compared the total microbial background. This study impact of four virus concentration/extraction methods (Innovaprep, Nanotrap, Promega, and Solids extraction) on probe-capture enrichment for followed by sequencing. Different yielded distinct profiles. Innovaprep ultrafiltration (following solids removal) had highest sequencing sensitivity richness, resulting successful assembly several near-complete genomes. However, it was less sensitive detecting SARS-CoV-2 digital polymerase chain reaction (dPCR) Promega Nanotrap. Across all preparation methods, astroviruses polyomaviruses were most highly abundant viruses, rare. These findings suggest that success can be increased using reduce nontarget nucleic acids extract, though absolute concentration extracted acid, as indicated Qubit, targeted dPCR, may not directly related performance. Further, broadly panels capture viral diversity but risks losing signals specific low-abundance viruses. Overall, this highlights importance aligning wet lab bioinformatic with goals when employing from wastewater.

Language: Английский

---

Deep viral blood metagenomics reveals extensive anellovirus diversity in healthy humans

Scientific Reports, Journal Year: 2021, Volume and Issue: 11(1)

Published: March 25, 2021

Abstract Human blood metagenomics has revealed the presence of different types viruses in apparently healthy subjects. By far, anelloviruses constitute viral family that is more frequently found human blood, although amplification biases and contaminations pose a major challenge this field. To investigate further, we subjected pooled plasma samples from 120 donors Spain to high-speed centrifugation, RNA DNA extraction, random amplification, massive parallel sequencing. Our results confirm extensive such samples, which represented nearly 97% total sequence reads obtained. We assembled 114 genomes belonging family, revealing remarkable diversity. Phylogenetic analysis ORF1 suggested 28 potentially novel anellovirus species, 24 were validated by Sanger sequencing discard artifacts. These findings underscore importance implementing efficient purification procedures enrich fraction as an essential step virome studies question pathological role anelloviruses.

Language: Английский

---

Metagenomic nanopore sequencing for exploring the nature of antimicrobial metabolites of Bacillus haynesii

AMB Express, Journal Year: 2024, Volume and Issue: 14(1)

Published: May 4, 2024

Abstract Multidrug-resistant (MDR) pathogens are a rising global health worry that imposes an urgent need for the discovery of novel antibiotics particularly those natural origin. In this context, we aimed to use metagenomic nanopore sequence analysis soil microbiota coupled with conventional phenotypic screening and genomic identifying antimicrobial metabolites produced by promising isolate(s). study, whole metagenome sample(s) was performed using MinION™ (Oxford Nanopore Technologies). Aligning sequences probable secondary metabolite gene clusters were extracted analyzed antiSMASH version 2 DeepBGC. Results showed most abundant taxa Bifidobacterium, Burkholderia , Nocardiaceae (99.21%, followed Sphingomonadaceae (82.03%) B. haynesii (34%). Phenotypic respective samples has resulted in Bacillus isolate exhibited broad-spectrum antibacterial activities against various MDR pathogens. It identified microscopical, cultural, molecular methods as (B.) MZ922052. The revealed conservation seven biosynthetic namely, siderophore lichenicidin VK21-A1/A2 (95% identity), lichenysin (100%), fengycin (53%), terpenes bacteriocin Lasso peptide (95%) bacillibactin (53%). conclusion, helped identify MZ922052 harboring metabolites. This is first report bacteriocin, lichenysin,

Language: Английский

---

Improved diagnosis of viral encephalitis in adult and pediatric hematological patients using viral metagenomics

Journal of Clinical Virology, Journal Year: 2020, Volume and Issue: 130, P. 104566 - 104566

Published: July 31, 2020

Metagenomic sequencing is a powerful technique that enables detection of the full spectrum pathogens present in any specimen single test. Hence, metagenomics increasingly being applied for viruses clinical cases with suspected infections unknown etiology and large number relevant potential causes. This typically case patients presenting encephalitis, particular when immunity impaired by underlying disorders. In this study, viral has been to cohort hematological encephalitis origin. Because loads cerebrospinal fluid are generally low, technical performance metagenomic protocol enrichment capture probes targeting all known vertebrate sequences was studied. Subsequently, optimized Viral increased sequence read count on samples 100 - 10.000 fold, compared unenriched sequencing. five out 41 (12%) virus detected which had not current routine diagnostics. BK polyomavirus, hepatitis E virus, human herpes virus-6 Epstein Barr were identified unbiased approach. study demonstrated origin may benefit from early testing as step

Language: Английский

---