Biology subjects

Sherman, D.

Publications and source records attributed to Sherman, D..

3 recordsLinked to original sources

bioRxiv2025

Accurate MAG reconstruction from complex soil microbiome through combined short- and HiFi long-reads metagenomics

BackgroundAdvances in high-fidelity long-read (HiFi-LR) sequencing technologies have opened new opportunities to explore the microbial genomic diversity of complex environments, such as soils. While short-read (SR) sequencing has enabled broad insights at the gene level, the limited read length constrains the reconstruction of complete genomes. HiFi-LRs, in contrast, improve assembly continuity and completeness, supporting higher-resolution taxonomic and functional annotation. However, the cost and relatively low throughput of HiFi-LR sequencing can limit genome recovery--particularly at the binning stage, where coverage depth is critical. In this study, we assess the benefit of combining HiFi-LR and SR sequencing for genome-resolved characterization of a soil microbiome. ResultsWe generated metagenomic data for a tunnel-cultivated soil sample using high coverage Illumina SRs as well as a combination of two HiFi-LR sequencing platforms (PacBio Sequel II and PacBio Revio). We found that assemblies generated from pooled HiFi-LR data alone exhibited higher completeness compared to those from ultra-deep SR data. Incorporating SR-derived coverage information for the binning of HiFi-LR contigs further increased both the number and quality of recovered metagenome-assembled genomes (MAGs), with a 24% increase in MAG recovery (313 vs. 252) and lower contamination levels (116 vs. 132 contaminated bins; mean 7.09 vs. 8.07), compared to using HiFi-LR data alone. This approach enabled the recovery of 61 additional MAGs, including 67% of low-abundance and taxonomically diverse lineages such as Archaea, representing 36 novel lineages. ConclusionOur results demonstrate that integrating HiFi-LR and SR sequencing markedly enhances genome recovery and binning accuracy in a highly diverse environment such as soil. The hybrid approach employed leverages the strengths of both technologies, leading to more contiguous assemblies and enabling the recovery of a broader range of genomes, including low-abundance and taxonomically diverse taxa. While factors such as sequencing depth, cost, and DNA quality remain important considerations, our study provides practical guidance for designing future soil metagenomics projects and underscores the value of adopting long-read technologies for more comprehensive characterization of complex microbial communities.

genomics↗

bioRxiv2020

ODELAM: Rapid sequence-independent detection of drug resistance in clinical isolates of Mycobacterium tuberculosis

Antimicrobial-resistant Mycobacterium tuberculosis (Mtb) causes over 200,000 deaths each year. Current assays of antimicrobial resistance need knowledge of mutations that confer drug resistance, or long periods of culture time to test growth under drug pressure. We present ODELAM (One-cell Doubling Evaluation of Living Arrays of Mycobacterium), a time-lapse microscopy-based method that observes individual cells growing into microcolonies. ODELAM enables rapid quantitative measures of growth kinetics in as little as 30 hours under a wide variety of environmental conditions. We show the utility of ODELAM by identifying ofloxacin resistance in clinical isolates of Mtb and benchmark its performance with standard MIC assays. In one clinical isolate, ODELAM identified ofloxacin heteroresistance and identifies the presence of drug resistant colony forming units (CFU) at 1 per 1000 CFUs in as little as 48 hours. ODELAM is a powerful new tool that can rapidly evaluate Mtb drug resistance in a laboratory setting.

microbiology↗

bioRxiv2019

Phase variation in Mycobacterium tuberculosis glpK produces transiently heritable drug tolerance

The length and complexity of tuberculosis (TB) therapy, as well as the propensity of Mycobacterium tuberculosis to develop drug resistance, are major barriers to global TB control efforts. M. tuberculosis is known to have the ability to enter into a drug-tolerant state, which may explain many of these impediments to TB treatment. We have identified a novel mechanism of genetically encoded but rapidly reversible drug-tolerance in M. tuberculosis caused by transient frameshift mutations in a homopolymeric tract (HT) of seven cytosines (7C) in the glpK gene. Inactivating frameshift mutations associated with the 7C HT in glpK produce small colonies that exhibit heritable multi-drug increases in minimal inhibitory concentrations and decreases in drug-dependent killing; however, reversion back to a fully drug-susceptible large-colony phenotype occurs rapidly through the introduction of additional insertions or deletions in the same glpK HT region. These reversible frameshift mutations in the 7C HT of M. tuberculosis glpK occur in clinical isolates, accumulate in M. tuberculosis infected mice with further accumulation during drug treatment, and exhibit a reversible transcriptional profile including induction of dosR and sigH and repression of kstR regulons, similar to that observed in other in vitro models of M. tuberculosis tolerance. These results suggest that GlpK phase variation may contribute to drug-tolerance, treatment failure and relapse in human TB. Drugs effective against phase-variant M. tuberculosis may hasten TB treatment and improve cure rates.\n\nSIGNIFICANCEThe ability of M. tuberculosis to survive during prolonged treatment has been attributed to either transient stress responses or fixed heritable drug-resistance producing mutations. We show that phase-variation in the M. tuberculosis glpK gene represents a third type of resistance mechanism. The ability of these glpK mutants to grow slowly and then rapidly revert suggests that these transiently-heritable changes may also explain how a hidden population of drug-tolerant bacteria develops during TB treatment. As a genetically trackable cause of drug-tolerance, M. tuberculosis glpK mutants provides a unique opportunity to study these phenomena at a cellular and mechanistic level. These mutants could also be used for developing drugs that target tolerant populations, leading to more rapid and effective TB treatments.

microbiology↗