Hackflex library preparation enables low-cost metagenomic profiling DOI Creative Commons
Samantha L. Goldman, Jon G. Sanders, Daniel D. Sprockett

et al.

ISME Communications, Journal Year: 2024, Volume and Issue: 4(1)

Published: Jan. 1, 2024

Shotgun metagenomic sequencing provides valuable insights into microbial communities, but the high cost of library preparation with standard kits and protocols is a barrier for many. New methods such as Hackflex use diluted commercially available reagents to greatly reduce costs. However, these have not been systematically validated sequencing. Here, we evaluate performance by libraries from known mock communities well mouse fecal samples prepared Hackflex, Illumina DNA Prep, TruSeq methods. successfully recovered all members Zymo community, performing best concentrations <1 ng/μL. Furthermore, was able delineate microbiota individual inbred mice same breeding stock at facility, statistical modeling indicated that ID explained greater fraction variance in composition than did method. These results show suitable generating inventories bacterial through

Language: Английский

Quantifying bias introduced by sample collection in relative and absolute microbiome measurements DOI
Dylan G. Maghini, Mai Dvorak, Alex Dahlen

et al.

Nature Biotechnology, Journal Year: 2023, Volume and Issue: 42(2), P. 328 - 338

Published: April 27, 2023

Language: Английский

Citations

58
Unveiling microbial diversity: harnessing long-read sequencing technology DOI
Daniel Paiva Agustinho, Yilei Fu, Vipin K. Menon

et al.

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

Published: April 30, 2024

Language: Английский

Citations

29
Enhancing Clinical Utility: Utilization of International Standards and Guidelines for Metagenomic Sequencing in Infectious Disease Diagnosis DOI Open Access

Chau-Ming Kan,

Hin Fung Tsang,

Xiao M. Pei

et al.

International Journal of Molecular Sciences, Journal Year: 2024, Volume and Issue: 25(6), P. 3333 - 3333

Published: March 15, 2024

Metagenomic sequencing has emerged as a transformative tool in infectious disease diagnosis, offering comprehensive and unbiased approach to pathogen detection. Leveraging international standards guidelines is essential for ensuring the quality reliability of metagenomic clinical practice. This review explores implications application diagnosis. By adhering established standards, such those outlined by regulatory bodies expert consensus, healthcare providers can enhance accuracy utility sequencing. The integration into workflows streamline diagnostic processes, improve identification, optimize patient care. Strategies implementing these diagnosis using are discussed, highlighting importance standardized approaches advancing precision initiatives.

Language: Английский

Citations

6
Functional metaproteomics for enzyme discovery DOI

Marina Prisacar,

Lars I. Leichert

Methods in enzymology on CD-ROM/Methods in enzymology, Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 1, 2025

Language: Английский

Citations

0
Benchmarking of shotgun sequencing depth highlights strain-level limitations of metagenomic analysis DOI Creative Commons

Nicole Treichel,

Thomas C. A. Hitch, John Penders

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2025, Volume and Issue: unknown

Published: March 27, 2025

Abstract Shallow metagenomics promises taxonomic and functional insights into samples at an affordable price. To determine the depth of sequencing required for specific analysis, benchmarking is using defined communities. We used complex mixtures DNA from cultured gut bacteria analysed composition, strain-level resolution, profiles nine depths (0.1-10.0 Gb). Reference-based analysis provided accurate taxonomic, 0.5-1.0 Gb. In contrast, de-novo metagenome-assembled genome (MAG) reconstruction deep (>10 Furthermore, it was found that even high-quality MAGs were chimeric, with few (54.5 %) accurately representing original strains. Functionally, 2 Gb reliable pathway level, but sufficient proteome coverage only achieved 10 Library preparation host-DNA identified as confounders in shallow metagenomic analysis. This comprehensive mock communities provides guidance to increasing community scientists interested metagenomics, highlights limitations capturing diversity.

Language: Английский

Citations

0
Expressions of resistome is linked to the key functions and stability of active rumen microbiome DOI Creative Commons
Tao Ma, Rahat Zaheer, Tim A. McAllister

et al.

Animal Microbiome, Journal Year: 2022, Volume and Issue: 4(1)

Published: June 4, 2022

Abstract Background The resistome describes the array of antibiotic resistant genes (ARGs) present within a microbial community. Recent research has documented in rumen ruminants and revealed that type abundance ARGs could be affected by diet and/or treatment. However, most these studies only assessed using metagenomics, expression its biological function microbiome remains largely unexplored. Results We characterized pools (resistome) their activities 48 beef cattle belonging to three breeds (Angus, Charolais, Kinsella composite hybrid), shotgun metagenomics metatranscriptomics. Sixty (including 20 plasmid-associated) were expressed which accounted for about 30% total number (187) identified metagenomic datasets, with tetW mefA exhibiting highest level expression. In addition, bacterial hosts 17 identified. active was less diverse hybrid than Angus, however, did not differ among breeds. Although associated feed efficiency, positively correlated metabolic pathways ‘attenuation values’ (a measurement stability) microbiome, suggesting influences stability functionality microbiome. Moreover, Ruminococcus spp. , Prevotella ruminicola Muribaculaceae spp . Collinsella aerofaciens all as ARGs, possibly promoting dominance carbohydrate degraders Conclusions Findings from this study provide new insight into vivo, may inform strategies limit spread ubiquitously found broader environment without negatively impacting key functional outcomes

Language: Английский

Citations

12
Unveiling the microbial symphony: Next-Gen sequencing and bioinformatics insights into the human gut microbiome DOI Creative Commons

Keerti Maheshwari,

Pankaj Musyuni, Amitava Moulick

et al.

Health Sciences Review, Journal Year: 2024, Volume and Issue: 11, P. 100173 - 100173

Published: April 4, 2024

Hundreds of bacteria, viruses, eukaryotes, and archaea comprise the human intestinal microbiota, which constitutes a complex ecology. Recent years have established significance gastrointestinal microbiome in study biological microorganisms. Diverse techniques for cultural extraction analysis make it difficult to cultivate gut bacteria species. To determine causative microorganism, is necessary investigate microbes bacteria. Novel efficacious remedies illnesses are required. Bioinformatics next-generation sequencing can assist enormous quantities sequenced data bacterial investigations, thereby expanding sequencing. However, advancements technology broadened scope bioinformatics identification analysis. With such as metagenomic, 16S rRNA, meta-transcriptomic sequencing, experimental regarding immunological response genetic manipulation seems be helpful identifying diseases. Despite significant progress over past two decades, there still lack understanding about ecology diseases treatments. The present review highlights how microbial gene information help manage issues.

Language: Английский

Citations

2
Next Generation Sequencing in Metagenomics and Metatranscriptomics DOI

Ajay Kumar,

Anuj Yadav

Published: Jan. 1, 2024

This study explores metagenomics and metatranscriptomics, delving into the intricacies of understanding complex microbial communities through Next-Generation Sequencing (NGS) technologies. Metagenomics includes analyzing genetic material directly extracted from environmental samples, offering insights community composition, diversity, functional potential. In contrast, metatranscriptomics provides a dynamic snapshot gene expression patterns, revealing how these actively respond to their environments. We meticulously explore stages gathering extracting DNA/RNA, preparing libraries, utilizing sequencing platforms, conducting data analysis, showcasing comprehensive journey raw samples meaningful biological insights. Through lens we elucidate significance quality control, read alignment, quantification, annotation. Moreover, accentuate broad applications disciplines, ranging studies host-microbiome interactions biotechnological advancements. The effective integration metagenomic metatranscriptomic enables researchers unlock complementary insights, providing multidimensional view functions. modern NGS has revolutionized research. NGS's ability generate vast amounts DNA RNA sequence enabled at an unprecedented depth. advancement transcended limitations traditional culture-based methods, allowing for unculturable microorganisms intricate relationships within ecosystems. brought precision scalability by structures, potential functions present in samples. real-time expression, shedding light on microbes' active roles By bridging with activity, offers holistic function, adapt, changing conditions. conclusion, technologies stand as cornerstone research, propelling fields new frontiers discovery. Their offer detailed dynamics, is revolutionizing our comprehension realm. far-reaching implications, spanning diverse such ecology human health.

Language: Английский

Citations

2
Enhancing insights into diseases through horizontal gene transfer event detection from gut microbiome DOI Creative Commons
Shuai Wang, Yiqi Jiang,

Lijia Che

et al.

Nucleic Acids Research, Journal Year: 2024, Volume and Issue: 52(14), P. e61 - e61

Published: June 17, 2024

Horizontal gene transfer (HGT) phenomena pervade the gut microbiome and significantly impact human health. Yet, no current method can accurately identify complete HGT events, including transferred sequence associated deletion insertion breakpoints from shotgun metagenomic data. Here, we develop LocalHGT, which facilitates reliable swift detection of events data, delivering an accuracy 99.4%-verified by Nanopore data-across 200 samples, achieving average F1 score 0.99 on 100 simulated LocalHGT enables a systematic characterization within across 2098 revealing that multiple recipient genome sites become targets sequence, microhomology is enriched in breakpoint junctions (P-value = 3.3e-58), HGTs function as host-specific fingerprints indicated higher similarity intra-personal temporal samples than inter-personal 4.3e-303). Crucially, showed potential contributions to colorectal cancer (CRC) acute diarrhoea, evidenced enrichment butyrate metabolism pathway 3.8e-17) shigellosis 5.9e-13) respective HGTs. Furthermore, differential demonstrated promise biomarkers for predicting various diseases. Integrating into CRC prediction model achieved AUC 0.87.

Language: Английский

Citations

2
Evaluating supervised and unsupervised background noise correction in human gut microbiome data DOI Creative Commons
Leah Briscoe, Brunilda Balliu, Sriram Sankararaman

et al.

PLoS Computational Biology, Journal Year: 2022, Volume and Issue: 18(2), P. e1009838 - e1009838

Published: Feb. 7, 2022

The ability to predict human phenotypes and identify biomarkers of disease from metagenomic data is crucial for the development therapeutics microbiome-associated diseases. However, commonly affected by technical variables unrelated phenotype interest, such as sequencing protocol, which can make it difficult find disease. Supervised methods correct background noise, originally designed gene expression RNA-seq data, are applied microbiome but may be limited because they cannot account unmeasured sources variation. Unsupervised approaches address this issue, current ill-equipped deal with unique aspects compositional, highly skewed, sparse. We perform a comparative analysis different denoising transformations in combination supervised correction well an unsupervised principal component approach that presently used other domains has not been date. comparable reducing false discovery approaches, added benefit needing know variation apriori. prediction tasks, appears only improve when contribute majority variance data. As new larger datasets become increasingly available, noise will essential generating reproducible analyses.

Language: Английский

Citations

11