Biology subjects

Dar, D.

Publications and source records attributed to Dar, D..

6 recordsLinked to original sources

bioRxiv2026

Single-cell transcriptome imaging reveals conserved and virulence-linked phenotypic states in bacteria

Single-cell phenotypic diversity underlies fundamental bacterial behaviors. Resolving this heterogeneity at scale while maintaining sensitive detection remains a central challenge. Here, we address this gap by combining the strengths of single-molecule fluorescence in situ hybridization (smFISH) and transcriptome-wide sequencing. We present par2FISH, a framework for discovering transcriptionally defined subpopulations across conditions by multiplexing tens of thousands of smFISH reactions. Through comparative single-cell transcriptomics in Escherichia coli and Salmonella, we uncover conserved and species-specific patterns of heterogeneity, including mutually exclusive states, coordinated metabolic specialization, and differential activation of virulence programs. We further obtain complete transcriptomes for selected subpopulations using marker-based cell sorting and RNA-sequencing (FACS-RNA-seq). This integrated strategy paves the way for exposing bacterial phenotypic landscapes and the environmental, physiological, and evolutionary factors that shape them.

microbiology↗

bioRxiv2025

Single-Cell Phenotypic Heterogeneity Shapes Quorum Signaling Dynamics in Pseudomonas aeruginosa

Quorum sensing (QS) orchestrates collective responses in bacteria. Yet, how individuality arises and functions within a system designed to unify behavior remains poorly understood. Here, we use imaging-transcriptomics to comprehensively profile the QS transcriptional response of the opportunistic pathogen Pseudomonas aeruginosa at single-cell resolution. We observe heterogeneity across all stages of QS, from signal production to public goods expression. While most cells cooperate, the extent of public goods contribution varies substantially, primarily due to transcriptional noise. Additionally, cells expressing high levels of a given exoproduct were often enriched for multiple other public goods. Expression of QS signal synthases, particularly in the Las and PQS systems, exhibits extreme cell-cell variability indicative of regulation, and exceeding levels observed in known differentiation into hypervirulent or motile subpopulations. These patterns are dynamic, peaking during early signaling, suggesting that a subset of highly committed cells may influence key decision points in the QS response. We find that cellular memory carried from prior growth cycles shapes QS dynamics via signaling-primed cells but does not influence de novo formation of hyper-signaling cells. Differentiation into hyper-signaling subpopulations is also robust to exogenous autoinducers across a wide concentration range, suggesting it is likely mediated by an internal mechanism. Despite variation in QS network outputs between strains, hyper-signaling subpopulations consistently emerge across diverse isolates, pointing to an evolutionarily conserved strategy. Our findings recast QS as a system that embeds a deliberately heterogeneous entry point within an otherwise synchronizing program, offering new insight into how bacterial populations balance cooperation and conflict.

microbiology↗

bioRxiv2022

An expanding arsenal of immune systems that protect bacteria from phages

Bacterial anti-phage defense systems are frequently clustered in microbial genomes, forming defense islands. This genomic property enabled the recent discovery of multiple defense systems based on their genomic co-localization with known systems, but the full arsenal of anti-phage mechanisms in bacteria is still unknown. In this study we report the discovery of 21 new defense systems that protect bacteria from phages, based on computational genomic analyses and phage infection experiments. We find multiple systems with protein domains known to be involved in eukaryotic anti-viral immunity, including ISG15-like proteins, dynamin-like proteins, and SEFIR domains, and show that these domains participate in bacterial defense against phages. Additional systems include protein domains predicted to manipulate DNA and RNA molecules, as well as multiple toxin-antitoxin systems shown here to function in anti-phage defense. The systems we discovered are widely distributed in bacterial and archaeal genomes, and in some bacteria form a considerable fraction of the immune arsenal. Our data substantially expand the known inventory of defense systems utilized by bacteria to counteract phage infection.

microbiology↗

bioRxiv2021

In situ single-cell activities of microbial populations revealed by spatial transcriptomics

Microbial populations and communities are heterogeneous, yet capturing their diverse activities has proven challenging at the relevant spatiotemporal scales. Here we present par-seqFISH, a targeted transcriptome-imaging approach that records both gene-expression and spatial context within microscale assemblies at a single-cell and molecule resolution. We apply this approach to the opportunistic bacterial pathogen, Pseudomonas aeruginosa, analyzing [~]600,000 individuals across dozens of physiological conditions in planktonic and biofilm cultures. We explore the phenotypic landscape of this bacterium and identify metabolic and virulence related cell-states that emerge dynamically during growth. We chart the spatial context of biofilm-related processes including motility and kin-exclusion mechanisms and identify extensive and highly spatially-resolved metabolic heterogeneity. We find that distinct physiological states can co-exist within the same biofilm, just a few microns away, underscoring the importance of the microenvironment. Together, our results illustrate the complexity of microbial populations and present a new way of studying them at high-resolution.

microbiology↗

bioRxiv2020

Global landscape of phenazine biosynthesis and biodegradation reveals species-specific colonization patterns in agricultural soils and crop microbiomes

Phenazines are natural bacterial antibiotics that can protect crops from disease. However, for most crops it is unknown which producers and specific phenazines are ecologically relevant, and whether phenazine biodegradation can counter their effects. To better understand their ecology, we developed and environmentally-validated a quantitative metagenomic approach to mine for phenazine biosynthesis and biodegradation genes, applying it to >800 soil and plant-associated shotgun-metagenomes. We discover novel producer-crop associations and demonstrate that phenazine biosynthesis is prevalent across habitats and preferentially enriched in rhizospheres, whereas biodegrading bacteria are rare. We validate an association between maize and Dyella japonica, a putative producer abundant in crop microbiomes. D. japonica upregulates phenazine biosynthesis during phosphate limitation and robustly colonizes maize seedling roots. This work provides a global picture of phenazines in natural environments and highlights plant-microbe associations of agricultural potential. Our metagenomic approach may be extended to other metabolites and functional traits in diverse ecosystems.

microbiology↗

bioRxiv2018

Whole blood and hypertonic saline resuscitation improve the outcomes of controlled hemorrhagic shock combined with penetrating brain injury in rat

One of the most frequent injury patterns leading to early death of polytraumatized patients is hemorrhagic shock combined with traumatic brain injury. Currently, there is no agreement on the best treatment protocol of such injury. This study was aimed to compare the outcomes of resuscitation with various volumes of hypertonic saline (HTS), whole blood (WB) and fresh frozen plasma (FFP) in a rat model of penetrating brain injury combined with controlled hemorrhagic shock in order to look for optimal treatment. Anesthetized rats were intubated and ventilated. 30% of circulating blood volume was withdrawn following open brain injury, and intravenous resuscitation with various volumes of HTS, WB and FFP (3, 9, and 18 ml/kg) was administered. Blood samples were collected during the experiment for measurements of lactate and base excess levels, and before sacrifice for neutrophil counts. Twenty four and 48 hours after the injury, rats neurological status was examined. Rats were, then, sacrificed and pathological studies of brain and lung sections were performed. Regardless of fluid type, resuscitation improved the mean arterial pressure and lactate levels in accordance to fluids volume. However, base excess was improved only when rats were treated with WB and FFP. Improvement in behavioral performance of the rats was observed when treated with 3 and 9 ml/kg HTS or with 18 ml/kg WB or FFP. Blood and lung neutrophils were reduced in rats treated with 9 or 18 ml/kg WB. Mortality rates were reduced in rats administered with 9 and 18 ml/kg of HTS or WB. Taken together, resuscitation with WB at 9-18ml/kg or HTS at 9ml/kg is optimal for treatment of combined injury.

physiology↗