Biology subjects

Chen, S. L.

Publications and source records attributed to Chen, S. L..

14 recordsLinked to original sources

bioRxiv2026

A recipient-based anti-conjugation factor triggers an abortive mechanism by targeting the Type IV secretion system

Many bacterial defense (immune) systems prevent the entry of foreign DNA by directly recognizing and targeting nucleic acids, effectively blocking all mechanisms of horizontal gene transfer1. However, systems defending specifically against conjugation, a major route for gene dissemination, have heretofore not been reported. We have discovered a novel defense factor, which we name AbjA (Abortive conjugation protein A), that specifically limits successful plasmid conjugation into a recipient bacterium. AbjA interacts directly with and targets the ATPase component TrbE of the Type IV secretion system (T4SS)2,3 to induce cell death; this contrasts with most other defense systems that act at the nucleic acid level. AbjA therefore represents the first member of a new class of bacterial defense factors that trigger what we term abortive conjugation. Previously, recipient bacteria were viewed largely as defenseless against the mechanism of conjugation4-6; our discovery and characterization of AbjA demonstrates that recipient bacteria can block conjugation to limit the transfer (and thus spread) of plasmids. Discovery of this class of defense systems thus has implications for bacterial defense, plasmid evolution, and possible strategic alternatives to rationally target plasmid spread, particularly with respect to virulence and antibiotic resistance.

microbiology↗

bioRxiv2026

Home is where the host is: Evolutionary history of geographic spread, host switching, and adaptive genomic signatures in two generalist Group B Streptococcus clonal groups

Group B Streptococcus (GBS) is a pathogen of global relevance in neonatal and maternal disease as well as bovine mastitis. Two closely related clonal groups, denoted 103 and 314 (CG103/314) have been detected in humans and cattle on multiple continents in recent decades but are poorly characterised compared to other host-generalist clades. We examined their potential origins, host-switching events and presence of a suite of genetic markers for antimicrobial resistance, virulence and host association using a newly assembled dataset of 248 CG103/314 genomes from humans, cattle, and food originating from five continents. We detected multiple host switches between humans and cattle, and significant regional differences in AMR gene distribution, possibly reflecting local differences in antimicrobial use across countries and hosts and indicating a capacity for regional adaptation to selective pressures. Across the evolutionary history of CG103/314 from both host species, the prevalence of the Lac.2 operon, a genetic marker associated with bovine host adaptation, was high, whereas the prevalence of the scpB-lmb gene pair, a genetic marker of human host adaptation in other GBS clonal groups, was very low. All isolates with scpB-lmb were associated with human disease rather than carriage. Our dataset displayed biases typical of research into multi-host pathogens, when sampling is often focused on a specific host species or setting. Consistent, balanced, contemporaneous and sympatric sampling efforts across host species and sources are needed for a full understanding of the distribution and emergence of CG103/314 and similar multi-host pathogens impacting food safety and public health. Impact statementThis study provides a comprehensive, global genomic overview of generalist clonal groups 103/314 of the human and animal pathogen Group B Streptococcus (GBS). By analysing host switching, antimicrobial resistance and virulence-associated markers, we show that these clonal groups display adaptation patterns shaped by region- and host-specific selective pressures. Our findings include potential expansion of the host range from humans and cattle into porcupines and pigs, and provides detailed discussion around anthropocentric sampling bias, highlighting the importance of balanced, multi-host sampling of generalist GBS lineages and One Health pathogens in general. This work reinforces the need for coordinated One Health surveillance to monitor emerging sub-lineages with relevance for food safety, human and animal health.

evolutionary biology↗

bioRxiv2024

Design of an intracellular aptamer-based fluorescent biosensor to track burden in E. coli

Cell burden impacts the performance of engineered synthetic systems. For this reason, there is great interest toward the development of tools to track burden and improve biotechnology applications. Fluorogenic RNA aptamers are excellent candidates for live monitoring of burden because their production is expected to impose a negligible load on transcription resources. Here we characterise the performance of a library of aptamers when expressed from different promoters in E. coli. We find that aptamer relative performance is dependent on the promoter and the strain, and that, contrary to expectation, aptamer expression impacts host fitness. By selecting two of the aptamers with brighter output and lower impact, we then design an intracellular biosensor able to report on the activation of the burden response in engineered cells. The sensor developed here adds to the collection of tools available for burden mitigation and may support bioprocessing applications where improved host performance is sought.

synthetic biology↗

bioRxiv2023

The HtrA chaperone monitors sortase-assembled pilus biogenesis in Enterococcus faecalis

Sortase-assembled pili contribute to virulence in many Gram-positive bacteria. In Enterococcus faecalis, the endocarditis and biofilm-associated pilus (Ebp) is polymerized on the membrane by sortase C (SrtC) and attached to the cell wall by sortase A (SrtA). In the absence of SrtA, polymerized pili remain anchored to the membrane (i.e. off-pathway). Here we show that the high temperature requirement A (HtrA) bifunctional chaperone/protease of E. faecalis is a quality control system that clears aberrant off-pathway pili from the cell membrane. In the absence of HtrA and SrtA, accumulation of membrane-bound pili leads to cell envelope stress and partially induces the regulon of the ceftriaxone resistance-associated CroRS two-component system, which in turn causes hyper-piliation and cell morphology alterations. Inactivation of croR in the {Delta}srtA{Delta}htrA background partially restores the observed defects of the {Delta}srtA{Delta}htrA strain, supporting a role for CroRS in the response to membrane perturbations. Moreover, absence of SrtA and HtrA decreases basal tolerance of E. faecalis against cephalosporins and daptomycin. The link between HtrA, pilus biogenesis and the CroRS two-component system provides new insights into the E. faecalis response to endogenous membrane perturbations. Author summaryTo explore the role of the HtrA chaperone/protease in E. faecalis off-pathway pilus clearance, we deleted htrA in an E. faecalis OG1RF {Delta}srtA strain known to retain polymerized pili on the cell membrane. Cells in the {Delta}srtA{Delta}htrA background are hyper-piliated, possess altered morphology, and are more susceptible to cell envelope-targeting antibiotics as compared to the parent OG1RF strain. RNA sequencing of the {Delta}srtA{Delta}htrA strain revealed transcriptional changes reminiscent of a membrane stress response. This response was pilus-dependent and contained several members of the CroR regulon. Inactivation of the response regulator CroR in the {Delta}srtA{Delta}htrA background restored (at least partially) piliation and cell morphology but not antibiotic susceptibility, linking CroR for the first time to pilus biogenesis and endogenous cell envelope stress.

microbiology↗

bioRxiv2023

Three Accessory Gene Clusters Drive Host-Adaptation in Group B Streptococcus

Streptococcus agalactiae (Group B Streptococcus, GBS) is a major pathogen of humans and animals, posing a threat to human health as well as food security. Here, we investigate the role of genomic mechanisms, including homologous recombination and horizontal gene transfer, in shaping the population structure of GBS and its adaptation to three major host groups (humans, cattle, fishes). We demonstrate that the GBS population comprises host-specialist, host-adapted lineages as well as host generalists, and that these categories differ in their level or recombination. Although the accessory genome at large varies by lineage rather than host, genome wide association studies show that host association is driven by three accessory genome clusters, regardless of lineage or breadth of the host spectrum. These genomic clusters (scpB in human GBS, lactose operon in bovine GBS, Locus 3 in fish GBS) are known (scpB, Lac.2) or shown here (Locus 3) to be functionally relevant and are shared with other streptococcal species occupying the same host niche. These findings demonstrate the importance of considering the role of non-human host species in the evolution of GBS, including high risk clones that may lead to interspecies transmission and affect efficacy of future GBS vaccines.

genomics↗

bioRxiv2022

Depleting cationic lipids involved in antimicrobial resistance drives adaptive lipid remodeling in Enterococcus faecalis

The bacterial cell membrane is an interface for cell envelope synthesis, protein secretion, virulence factor assembly and a target for host cationic antimicrobial peptides (CAMPs). To resist CAMP killing, several Gram-positive pathogens encode the multiple peptide resistance factor (MprF) enzyme that covalently attaches cationic amino acids to anionic phospholipids in the cell membrane. While E. faecalis encodes two mprF paralogs, MprF2 plays a dominant role in conferring resistance to killing by the CAMP human {beta}-defensin 2 (hBD-2) in E. faecalis strain OG1RF. The goal of the current study is to understand the broader lipidomic and functional roles of E. faecalis mprF. We analyzed the lipid profiles of parental wild type and mprF mutant strains and show that while {triangleup}mprF2 and {triangleup}mprF1 {triangleup}mprF2 mutants completely lacked cationic lysyl-phosphatidylglycerol (L-PG), the {triangleup}mprF1 mutant synthesized [~]70% of L-PG compared to the parent. Unexpectedly, we also observed a significant reduction of PG in {triangleup}mprF2 and {triangleup}mprF1 {triangleup}mprF2. In the mprF mutants, particularly {triangleup}mprF1 {triangleup}mprF2, the decrease in L-PG and PG is compensated by an increase in the phosphorus-containing lipid, GPDGDAG, and D-ala-GPDGDAG. These changes were accompanied by a downregulation of de novo fatty acid biosynthesis and an accumulation of long-chain acyl-acyl carrier proteins (long-chain acyl-ACPs), suggesting that the suppression of fatty acid biosynthesis was mediated by the transcriptional repressor FabT. Growth in chemically defined media lacking fatty acids revealed severe growth defects in the {triangleup}mprF1 {triangleup}mprF2 mutant strain, but not the single mutants, which was partially rescued through supplementation with palmitic and stearic acids. Changes in lipid homeostasis correlated with lower membrane fluidity, impaired protein secretion, and increased biofilm formation in both {triangleup}mprF2 and {triangleup}mprF1 {triangleup}mprF2, compared to wild type and {triangleup}mprF1. Collectively, our findings reveal a previously unappreciated role for mprF in global lipid regulation and cellular physiology, which could facilitate the development of novel therapeutics targeting MprF. Significance StatementThe cell membrane plays a pivotal role in protecting bacteria against external threats, such as antibiotics. Cationic phospholipids such as lysyl-phosphatidyglycerol (L-PG) resist the action of cationic antimicrobial peptides through electrostatic repulsion. Here we demonstrate that L-PG depletion has several unexpected consequences in Enterococcus faecalis, including a reduction of phosphatidylglycerol (PG), enrichment of a phosphorus-containing lipid, reduced fatty acid synthesis accompanied by an accumulation of long-chain acyl-acyl carrier proteins (long chain acyl-ACPs), lower membrane fluidity, and impaired secretion. These changes are not deleterious to the organism as long as exogenous fatty acids are available for uptake from the culture medium. Our findings suggest an adaptive mechanism involving compensatory changes across the entire lipidome upon removal of a single phospholipid modification. Such adaptations must be considered when devising antimicrobial strategies that target membrane lipids.

microbiology↗

bioRxiv2021

GBS-SBG - GBS Serotyping by Genome Sequencing

Group B Streptococcus agalactiae (GBS; Streptococcus agalactiae) is the most common cause of neonatal meningitis and a rising cause of sepsis in adults. Recently, it has also been shown to cause foodborne disease. As with many other bacteria, the polysaccharide capsule of GBS is antigenic, enabling its use for strain serotyping. Recent advances in DNA sequencing have made sequence-based typing attractive (as has been implemented for several other bacteria, including Escherichia coli, Klebsiella pneumoniae species complex, Streptococcus pyogenes, and others). For GBS, existing WGS-based serotyping systems do not provide complete coverage of all known GBS serotypes (specifically including subtypes of serotype III), and none are simultaneously compatible with the two most common data types, raw short reads and assembled sequences. Here, we create a serotyping database (GBS-SBG, GBS Serotyping by Genome Sequencing), with associated scripts and running instructions, that can be used to call all currently described GBS serotypes, including subtypes of serotype III, using both direct short-read- and assembly-based typing. We achieved higher concordance using GBS-SBG on a previously reported data set of 790 strains. We further validated GBS-SBG on a new set of 572 strains, achieving 99.8% concordance with PCR-based molecular serotyping using either short-read- or assembly-based typing. The GBS-SBG package is publicly available and will accelerate and simplify serotyping by sequencing for GBS. DATA SUMMARYO_LIThe GBS-SBG package is open source and available for at Github under the MIT license (URL - https://github.com/swainechen/GBS-SBG) C_LIO_LIAccession numbers of the sequencing reads and reference sequences used in the study from earlier reports have been provided within the article and the supplementary data C_LIO_LIThe WGS data for the 572 isolates used in the study is available at https://www.ncbi.nlm.nih.gov/bioproject/PRJNA293392 C_LI

bioinformatics↗

bioRxiv2020

Mass allelic exchange: enabling sexual genetics in Escherichia coli

Despite dramatic advances in genomics, connecting genotypes to phenotypes is still challenging. Sexual genetics combined with linkage analysis is a powerful solution to this problem but generally unavailable in bacteria. We build upon a strong negative selection system to invent Mass Allelic Exchange (MAE), which enables hybridization of arbitrary (including pathogenic) strains of E. coli. MAE reimplements the natural phenomenon of random crossovers, enabling classical linkage analysis. We demonstrate the utility of MAE with virulence-related gain-of-function screens, discovering that transfer of a single operon from a uropathogenic strain is sufficient for enabling a commensal E. coli to form large intracellular bacterial collections within bladder epithelial cells. MAE thus enables assaying natural allelic variation in E. coli (and potentially other bacteria), complementing existing loss-of-function genomic techniques. One Sentence SummaryWe create F1 hybrids of E. coli using MAE, bringing the power of linkage analysis to bear on phenotypic diversity (including virulence)

microbiology↗

bioRxiv2020

A faster and more accurate algorithm for calculating population genetics statistics requiring sums of Stirling numbers of the first kind

Stirling numbers of the first kind are used in the derivation of several population genetics statistics, which in turn are useful for testing evolutionary hypotheses directly from DNA sequences. Here, we explore the cumulative distribution function of these Stirling numbers, which enables a single direct estimate of the sum, using representations in terms of the incomplete beta function. This estimator enables an improved method for calculating an asymptotic estimate for one useful statistic, Fus Fs. By reducing the calculation from a sum of terms involving Stirling numbers to a single estimate, we simultaneously improve accuracy and dramatically increase speed.

bioinformatics↗

bioRxiv2019

Cartography of opportunistic pathogens and antibiotic resistance genes in a tertiary hospital environment

There is growing attention surrounding hospital acquired infections (HAIs) due to high associated healthcare costs, compounded by the scourge of widespread multi-antibiotic resistance. Although hospital environment disinfection is well acknowledged to be key for infection control, an understanding of colonization patterns and resistome profiles of environment-dwelling microbes is currently lacking. We report the first extensive genomic characterization of microbiomes (428), common HAI-associated microbes (891) and transmissible drug resistance cassettes (1435) in a tertiary hospital environment based on a 3-timepoint sampling (1 week and >1 year apart) of 179 sites from 45 beds. Deep shotgun metagenomic sequencing unveiled two distinct ecological niches of microbes and antibiotic resistance genes characterized by biofilm-forming and human microbiome influenced environments that display corresponding patterns of divergence over space and time. To study common nosocomial pathogens that were typically present at low abundances, a combination of culture enrichment and long-read nanopore sequencing was used to obtain thousands of high contiguity genomes (2347), phage sequences (1693) and closed plasmids (5910), a significant fraction of which (>60%) are not represented in current sequence databases. These high-quality assemblies and metadata enabled a rich characterization of resistance gene combinations, phage diversity, plasmid architectures, and the dynamic nature of hospital environment resistomes and their reservoirs. Phylogenetic analysis identified multidrug resistant strains as being more widely distributed and stably colonizing across hospital sites. Further genomic comparisons with clinical isolates across multiple species supports the hypothesis that multidrug resistant strains can persist in the hospital environment for extended periods (>8 years) to opportunistically infect patients. These findings highlight the importance of characterizing antibiotic resistance reservoirs in the hospital environment and establishes the feasibility of systematic genomic surveys to help target resources more efficiently for preventing HAIs.

microbiology↗

bioRxiv2018

Comprehensive Identification of Fim-Mediated Inversions in Uropathogenic Escherichia coli with Structural Variation Detection Using Relative Entropy

Most urinary tract infections (UTIs) are caused by uropathogenic Escherichia coli (UPEC), which depend on an extracellular organelle (Type 1 pili) for adherence to bladder cells during infection. Type 1 pilus expression is partially regulated by inversion of a piece of DNA referred to as fimS, which contains the promoter for the fim operon encoding Type 1 pili. fimS inversion is regulated by up to five recombinases collectively known as Fim recombinases. These Fim recombinases are currently known to regulate two other switches: the ipuS and hyxS switches. A long-standing question has been whether the Fim recombinases regulate the inversion of other switches, perhaps to coordinate expression for adhesion or virulence. We answered this question using whole genome sequencing with a newly developed algorithm (Structural Variation detection using Relative Entropy, SVRE) for calling structural variations using paired-end short read sequencing. SVRE identified all of the previously known switches, refining the specificity of which recombinases act at which switches. Strikingly, we found no new inversions that were mediated by the Fim recombinases. We conclude that the Fim recombinases are each highly specific for a small number of switches. We hypothesize that the unlinked Fim recombinases have been recruited to regulate fimS, and fimS only, as a secondary locus; this further implies that regulation of Type 1 pilus expression (and its role in gastrointestinal and/or genitourinary colonization) is important enough, on its own, to influence the evolution and maintenance of multiple additional genes within the accessory genome of E. coli.\n\nImportanceUTIs are a common ailment that affects more than half of all women during their lifetime. The leading cause of UTIs is UPEC, which rely on Type 1 pili to colonize and persist within the bladder during infection. The regulation of Type 1 pili is remarkable for an epigenetic mechanism in which a section of DNA containing a promoter is inverted. The inversion mechanism relies on what are thought to be dedicated recombinase genes; however, the full repertoire for these recombinases is not known. We show here that there are no additional targets beyond those already identified for the recombinases in the entire genome of two UPEC strains, arguing that Type 1 pilus expression itself is the driving evolutionary force for the presence of these recombinase genes. This further suggests that targeting the Type 1 pilus is a rational alternative non-antibiotic strategy for the treatment of UTI.

microbiology↗

bioRxiv2018

Long-read whole genome sequencing and comparative analysis of six strains of the human pathogen Orientia tsutsugamushi

BackgroundOrientia tsutsugamushi is a clinically important but neglected obligate intracellular bacterial pathogen of the Rickettsiaceae family that causes the potentially life-threatening human disease scrub typhus. In contrast to the genome reduction seen in many obligate intracellular bacteria, early genetic studies of Orientia have revealed one of the most repetitive bacterial genomes sequenced to date. The dramatic expansion of mobile elements has hampered efforts to generate complete genome sequences using short read sequencing methodologies, and consequently there have been few studies of the comparative genomics of this neglected species.\n\nResultsWe report new high-quality genomes of Orientia tsutsugamushi, generated using PacBio single molecule long read sequencing, for six strains: Karp, Kato, Gilliam, TA686, UT76 and UT176. In comparative genomics analyses of these strains together with existing reference genomes from Ikeda and Boryong strains, we identify a relatively small core genome of 657 genes, grouped into core gene islands and separated by repeat regions, and use the core genes to infer the first whole-genome phylogeny of Orientia.\n\nConclusionsComplete assemblies of multiple Orientia genomes verify initial suggestions that these are remarkable organisms. They have large genomes with widespread amplification of repeat elements and massive chromosomal rearrangements between strains. At the gene level, Orientia has a relatively small set of universally conserved genes, similar to other obligate intracellular bacteria, and the relative expansion in genome size can be accounted for by gene duplication and repeat amplification. Our study demonstrates the utility of long read sequencing to investigate complex bacterial genomes and characterise genomic variation.

genomics↗

bioRxiv2017

Lacer: Accurate Base Quality Score Recalibration For Improving Variant Calling From Next-Generation Sequencing Data In Any Organism

Next-generation sequencing data is accompanied by quality scores that quantify sequencing error. Inaccuracies in these quality scores propagate through all subsequent analyses; thus base quality score recalibration is a standard step in many next-generation sequencing workflows, resulting in improved variant calls. Current base quality score recalibration algorithms rely on the assumption that sequencing errors are already known; for human resequencing data, relatively complete variant databases facilitate this. However, because existing databases are still incomplete, recalibration is still inaccurate; and most organisms do not have variant databases, exacerbating inaccuracy for non-human data. To overcome these logical and practical problems, we introduce Lacer, which recalibrates base quality scores without assuming knowledge of correct and incorrect bases and without requiring knowledge of common variants. Lacer is the first logically sound, fully general, and truly accurate base recalibrator. Lacer enhances variant identification accuracy for resequencing data of human as well as other organisms (which are not accessible to current recalibrators), simultaneously improving and extending the benefits of base quality score recalibration to nearly all ongoing sequencing projects. Lacer is available at: https://github.com/swainechen/lacer.

genomics↗

bioRxiv2016

Systematic integration of biomedical knowledge prioritizes drugs for repurposing

The ability to computationally predict whether a compound treats a disease would improve the economy and success rate of drug approval. This study describes Project Rephetio to systematically model drug efficacy based on 755 existing treatments. First, we constructed Hetionet (neo4j.het.io), an integrative network encoding knowledge from millions of biomedical studies. Hetionet v1.0 consists of 47,031 nodes of 11 types and 2,250,197 relationships of 24 types. Data was integrated from 29 public resources to connect compounds, diseases, genes, anatomies, pathways, biological processes, molecular functions, cellular components, pharmacologic classes, side effects, and symptoms. Next, we identified network patterns that distinguish treatments from non-treatments. Then we predicted the probability of treatment for 209,168 compound-disease pairs (het.io/repurpose). Our predictions validated on two external sets of treatment and provided pharmacological insights on epilepsy, suggesting they will help prioritize drug repurposing candidates. This study was entirely open and received realtime feedback from 40 community members.

bioinformatics↗