Biology subjects

Vill, A. C.

Publications and source records attributed to Vill, A. C..

5 recordsLinked to original sources

bioRxiv2024

scMicrobe PTA: Near Complete Genomes From Single Bacterial Cells

Microbial genomes produced by single-cell amplification are largely incomplete. Here, we show that primary template amplification (PTA), a novel single-cell amplification technique, generated nearly complete genomes from three bacterial isolate species. Furthermore, taxonomically diverse genomes recovered from aquatic and soil microbiomes using PTA had a median completeness of 81%, whereas genomes from standard amplification approaches were usually <30% complete. PTA-derived genomes also included more associated viruses and biosynthetic gene clusters.

genomics↗

bioRxiv2023

Spatial Mapping of Mobile Genetic Elements and their Cognate Hosts in Complex Microbiomes

The frequent exchange of mobile genetic elements (MGEs) between bacteria accelerates the spread of functional traits, including antimicrobial resistance, within the human microbiome. Yet, progress in understanding these intricate processes has been hindered by the lack of tools to map the spatial spread of MGEs in complex microbial communities, and to associate MGEs to their bacterial hosts. To overcome this challenge, we present an imaging approach that pairs single molecule DNA Fluorescence In Situ Hybridization (FISH) with multiplexed ribosomal RNA FISH, thereby enabling the simultaneous visualization of both MGEs and host bacterial taxa. We used this methodology to spatially map bacteriophage and antimicrobial resistance (AMR) plasmids in human oral biofilms, and we studied the heterogeneity in their spatial distributions and demonstrated the ability to identify their host taxa. Our data revealed distinct clusters of both AMR plasmids and prophage, coinciding with densely packed regions of host bacteria in the biofilm. These results suggest the existence of specialized niches that maintain MGEs within the community, possibly acting as local hotspots for horizontal gene transfer. The methods introduced here can help advance the study of MGE ecology and address pressing questions regarding antimicrobial resistance and phage therapy.

microbiology↗

bioRxiv2023

Evolution of promoter-proximal pausing enabled a new layer of transcription control

Promoter-proximal pausing of RNA polymerase II (Pol II) is a key regulatory step during transcription. Despite the central role of pausing in gene regulation, we do not understand the evolutionary processes that led to the emergence of Pol II pausing or its transition to a rate-limiting step actively controlled by transcription factors. Here we analyzed transcription in species across the tree of life. Unicellular eukaryotes display a slow acceleration of Pol II near transcription start sites that transitioned to a longer-lived, focused pause in metazoans. This event coincided with the evolution of new subunits in the NELF and 7SK complexes. Depletion of NELF in mammals shifted the promoter-proximal buildup of Pol II from the pause site into the early gene body and compromised transcriptional activation for a set of heat shock genes. Our work details the evolutionary history of Pol II pausing and sheds light on how new transcriptional regulatory mechanisms evolve.

evolutionary biology↗

bioRxiv2022

Run-on sequencing reveals nascent transcriptomics of the human microbiome

Precise regulation of transcription initiation and elongation enables bacteria to control cellular responses to environmental stimuli. RNAseq is the most common tool for measuring the transcriptional output of bacteria, comprising predominantly mature transcripts. To gain further insight into transcriptional dynamics, it is necessary to discriminate actively transcribed loci from those represented in the total RNA pool. One solution is to capture RNA polymerase (RNAP) in the act of transcription, but current methods are restricted to culturable and genetically tractable organisms. Here, we apply precision run-on sequencing (PRO-seq) to profile nascent transcription, a method amenable to diverse species. We find that PRO-seq is well-suited to profile small, structured, or post-transcriptionally modified RNAs, which are often excluded from RNAseq libraries. When PRO-seq is applied to the human microbiome, we identify taxon-specific RNAP pause motifs. We also uncover concurrent transcription and cleavage of guide RNAs and tRNA fragments at active CRISPR and tRNA loci. We demonstrate the specific utility of PRO-seq as a tool for exploring transcriptional dynamics in diverse microbial communities.

microbiology↗

bioRxiv2020

Widespread transfer of mobile antibiotic resistance genes within individual gut microbiomes revealed through bacterial Hi-C

The gut microbiome harbors a silent reservoir of antibiotic resistance (AR) genes that is thought to contribute to the emergence of multidrug-resistant pathogens through the process of horizontal gene transfer (HGT). To counteract the spread of AR genes, it is paramount to know which organisms harbor mobile AR genes and with which organisms they engage in HGT. Despite methods to characterize the bulk presence1, abundance2 and function3 of AR genes in the gut, technological limitations of short-read sequencing have precluded linking bacterial taxa to specific mobile genetic elements (MGEs) and their concomitant AR genes. Here, we apply and evaluate a high-throughput, culture-independent method for surveilling the bacterial carriage of MGEs, based on bacterial Hi-C protocols. We compare two healthy individuals with a cohort of seven neutropenic patients undergoing hematopoietic stem cell transplantation, who receive multiple courses of antibiotics throughout their prolonged hospitalizations, and are thus acutely vulnerable to the threat of multidrug-resistant infections4. We find that the networks of HGT are surprisingly distinct between individuals, yet AR and mobile genes are more dispersed across taxa within the neutropenic patients than the healthy subjects. Our data further suggest that HGT is occurring throughout the course of treatment in the microbiomes of neutropenic patients and within the guts of healthy individuals over a similar timeframe. Whereas most efforts to understand the spread of AR genes have focused on pathogenic species, our findings shed light on the role of the human gut microbiome in this process.

systems biology↗