Biology subjects

Haney, C. H.

Publications and source records attributed to Haney, C. H..

17 recordsLinked to original sources

bioRxiv2025

Predicting bacterial-mediated entomopathogenicity through comparative genomics and statistical modeling

Bacterial genomes encode for vast functional diversity and have both beneficial and detrimental effects on insect hosts. However, connecting bacterial genotype to a host-associated phenotype can be experimentally time-consuming, particularly if insecticidal mechanisms are highly host-specific. To streamline the identification of virulence genes on a new host, we tested if merging existing mechanistic knowledge with in vivo tests on a small number of bacterial isolates could predict bacterial genes associated with entomopathogenesis. We used a model consisting of Drosophila melanogaster interactions with pathogenic and commensal genome-sequenced strains of Pseudomonas bacteria. We compiled a database of previously described insecticidal and biocontrol genes within the Pseudomonas genus and used comparative genomics to probe the distribution of these genes across Pseudomonas strains. We found natural variation in the presence of known insecticidal genes across the genus. We tested the insect-killing capacity of 13 Pseudomonas spp. strains against D. melanogaster and found natural variation in insecticidal activity. To identify bacterial genes associated with fly mortality, we employed two statistical models to correlate bacterial virulence with the presence of previously described insecticidal activity. To validate our predictions, we used a P. aeruginosa PAO1 transposon mutant library and identified 8 operons that are necessary for killing D. melanogaster. We show that by combining existing literature with phenotyping a small number of strains, we can identify genes necessary for insecticidal activity in a previously untested insect model. More broadly these findings illustrate a discovery pipeline for bacterial virulence mechanisms, accelerating the discovery of insect pest biocontrol mechanisms.

microbiology↗

bioRxiv2024

In vitro biofilm formation only partially predicts beneficial Pseudomonas fluorescens protection against rhizosphere pathogens

Plant roots form associations with both beneficial and pathogenic soil microorganisms. While members of the rhizosphere microbiome can protect against pathogens, the mechanisms are poorly understood. We hypothesized that the ability to form a robust biofilm on the root surface is necessary for the exclusion of pathogens; however, it is not known if the same biofilm formation components required in vitro are necessary in vivo. Pseudomonas fluorescens WCS365 is a beneficial strain that is phylogenetically closely related to an opportunistic pathogen P. fluorescens N2C3 and confers robust protection against P. fluorescens N2C3 in the rhizosphere. We used this plant-mutualist-pathogen model to screen collections of P. fluorescens WCS365 increased attachment mutants (iam) and surface attachment defective (sad) transposon insertion mutants that form increased or decreased levels of biofilm on an abiotic surface, respectively. We found that while the P. fluorescens WCS365 mutants had altered biofilm formation in vitro, only a subset of these mutants, including those involved in large adhesion protein (Lap) biosynthesis, flagellin biosynthesis and O-antigen biosynthesis, lost protection against P. fluorescens N2C3. We found that the inability of P. fluorescens WCS365 mutants to grow in planta, and the inability to suppress pathogen growth, both partially contributed to loss of plant protection. We did not find a correlation between the extent of biofilm formed in vitro and pathogen protection in planta indicating that biofilm formation on abiotic surfaces may not fully predict pathogen exclusion in planta. Collectively, our work provides insights into mechanisms of biofilm formation and host colonization that shape the outcomes of host-microbe-pathogen interactions.

microbiology↗

bioRxiv2024

Order among chaos: high throughput MYCroplanters can distinguish interacting drivers of host infection in a highly stochastic system

While microbiomes provide diverse benefits for their host, they are notoriously variable in structure and function. As a result, substantial experimental replication and scalability are required to identify the contributions of and interactions between microbiota, the host and the environment. Here, we describe a novel high throughput plant growth system (MYCroplanters) to test how multiple host, microbiota, and pathogen variables predict host health. Using an Arabidopsis-Pseudomonas host-microbiome-pathogen model, we found that host genotype and order of arrival predict competition outcome between strains in the rhizosphere, but pathogen and microbiota dose can overwhelm these effects. Regardless of host or inoculation conditions, final microbial community structure emerged as a consistent predictor of host health. We demonstrate that high-throughput tools like MYCroplanters can isolate interacting drivers of host health. Increasing the scale at which we can screen components of microbiome-mediated host benefits will facilitate building microbiome engineering solutions for medicine and agricultural applications.

ecology↗

bioRxiv2023

Innate immunity can distinguish beneficial from pathogenic rhizosphere microbiota

For optimal growth and development, hosts depend on their ability to promote healthy symbiotic interactions while restricting pathogen growth. To ask whether hosts can distinguish phylogenetically similar pathogens and beneficial bacteria, we used two closely related plant-associated strains of Pseudomonas fluorescens where one is beneficial and the other exhibits toxin-dependent virulence. We show that while the two strains co-exist in vitro, the beneficial outcompetes that pathogen in planta. Using several readouts for plant innate immunity, we found that the beneficial and pathogenic strains elicit mechanistically distinct immune responses that occur in distinct root compartments. We show that while both the pathogenic and beneficial bacterial have plant recognizable MAMPs, the pathogen uniquely induces MAMP-independent immune responses. We found that the pathogen induces both a toxin-independent and a unique toxin-dependent defense response that remains intact in immune signaling mutants including bak1/bkk1 and npr1/4D. We conclude that hosts can distinguish between phylogenetically similar microbes.

plant biology↗

bioRxiv2022

PSKR1 balances the plant growth-defense trade-off in the rhizosphere microbiome

Microbiota provide benefits to their hosts including nutrient uptake and protection against pathogens. How hosts balance an appropriate immune response to avoid microbiota overgrowth while avoiding autoimmunity is not well understood. Here we show that Arabidopsis pskr1 (phytosulfokine receptor 1) loss-of-function mutants display autoimmunity and reduced rhizosphere bacterial growth when inoculated with normally growth-promoting Pseudomonas fluorescens. Transcriptional profiling demonstrated that PSKR1 regulates the plant growth-defense trade-off during plant-microbiome interactions: PSKR1 upregulates plant photosynthesis and root growth but suppresses salicylic acid (SA)-mediated defense responses. Genetic epistasis experiments showed that PSRK1 inhibition of microbiota-induced autoimmunity is fully dependent on SA signaling. Finally, using a transgenic reporter, we showed that P. fluorescens induces PSKR1 expression in roots, suggesting P. fluorescens might manipulate plant signaling to promote its colonization. Our data demonstrate a genetic mechanism to coordinate beneficial functions of the microbiome while preventing autoimmunity.

plant biology↗

bioRxiv2021

Amino acid availability determines plant immune homeostasis in the rhizosphere microbiome

Microbes possess conserved microbe-associated molecular patterns (MAMPs) such as flagellin that are recognized by plant receptors to induce immunity. Despite containing the same MAMPs as pathogens, commensals thrive in the plant rhizosphere microbiome indicating they must suppress or evade host immunity. The beneficial bacteria Pseudomonas capeferrum WCS358 can suppress Arabidopsis root immunity via acidification by secreting gluconic acid. While gluconic acid is sufficient to suppress immunity, we found that it is not necessary in a second beneficial strain Pseudomonas simiae WCS417, which produces more gluconic acid than WCS358. To uncover mechanisms that contribute to the suppression of Arabidopsis immunity, we performed a forward genetic screen in EMS-mutagenized P. simiae WCS417 using a flagellin-inducible CYP71A12pro:GUS reporter as an Arabidopsis immune readout. We identified a mutant that cannot suppress flagellin-elicited CYP71A12pro:GUS expression or acidify the rhizosphere. Next generation sequencing revealed a mutation in the catabolic site of an ornithine carbamoyltransferase argF, which is required for arginine biosynthesis. The mutant could be complemented by expression of argF from a plasmid, and a {Delta}argF mutant could not suppress immunity. Fungal pathogens can use alkalization through production of ammonia and glutamate, the arginine biosynthetic precursors, to promote their own growth and virulence. Therefore, we hypothesized that the biosynthesis of specific amino acids may be necessary to reduce levels of ammonia and glutamate to prevent rhizosphere alkalization and bacterial overgrowth. Genetically blocking arginine, glutamine, or proline biosynthesis, or by adding corresponding exogenous amino acids, resulted in rhizosphere alkalization. Interestingly, exogenous amino acids caused bacterial overgrowth in a gluconic acid-deficient mutants. Our findings show that bacterial amino acid biosynthesis contributes to acidification by preventing accumulation of glutamate precursors and the resulting alkalization. Collectively this work shows that by regulating nutrient availability, plants have the potential to regulate their immune homeostasis in the rhizosphere microbiome.

microbiology↗

bioRxiv2021

The ColR/S two-component system is a conserved determinant of host association across Pseudomonas species

Members of the bacterial genus Pseudomonas form mutualistic, commensal and pathogenic associations with diverse hosts. The prevalence of host association across the genus suggests that symbiosis may be a conserved ancestral trait and that distinct symbiotic lifestyles may be more recently evolved. Here we show that the ColR/S two-component system, part of the Pseudomonas core genome, is functionally conserved between Pseudomonas aeruginosa and Pseudomonas fluorescens. Using plant rhizosphere colonization and virulence in a murine abscess model, we show that colR is required for commensalism with plants and virulence in animals. Comparative transcriptomics revealed that the ColR regulon has diverged between P. aeruginosa and P. fluorescens and deleting components of the ColR regulon revealed strain-specific, but not hostspecific, requirements for ColR-dependent genes. Collectively, our results suggest that ColR/S allows Pseudomonas to sense and respond to a host, but that the ColR-regulon has diverged between Pseudomonas strains with distinct lifestyles.

microbiology↗

bioRxiv2021

A screen for mutants deficient in coronatine-mediated suppression of root immunity identifies Arabidopsis SDA1 as a novel integrator of immunity and phytohormone signaling

Despite the importance of the root immune system in the interaction with rhizosphere microbes, the majority of genetic screens for immunity regulators have been performed in leaves. A previous screen identified 27 hsm (hormone-mediated suppression of MAMP-triggered immunity) mutants that are impaired in jasmonic acid (JA)-mediated suppression of pattern-triggered immunity (PTI) in roots. Here we characterized 16 of the hsm mutants that retain JA sensitivity and are potential negative regulators of root immunity. We found that the majority of hsm mutants show enhanced resistance to Fusarium, a root fungal pathogen; however, only a subset are more resistant to a foliar pathogen. Surprisingly, 12 of 16 hsm mutants are also impaired in abscisic acid (ABA)-mediated suppression of PTI, suggesting a largely shared pathway between JA- and ABA-mediated immune suppression in roots. Although all hsm mutants are insensitive to JA-mediated suppression of root immunity, hsm4 shows hypersensitivity to JA-mediated root growth inhibition and JA-induced gene expression. Consistently, hsm4 is more resistant to leaf pathogens, suggesting that HSM4 is a negative regulator of both root and leaf immunity. Hsm4 was mapped to a mutation in a conserved ARM-repeat protein homologous to yeast SDA1, which has been reported to regulate 60S ribosome biogenesis. As translational reprogramming is a critical layer of immune regulation, this work suggests that AtSDA1 is a novel negative translational regulator of immunity. Additionally, a comprehensive characterization of all 16 hsm mutants provides a genetic toolkit to identify novel mechanisms that regulate root immunity.

plant biology↗

bioRxiv2021

Natural variation in Arabidopsis ISR1 affects iron localization and induced systemic resistance

Beneficial root-associated bacteria can induce systemic resistance (ISR) to foliar pathogens and there is known transcriptional and genetic overlap in the root response to iron deficiency and ISR. A previous study found that there is natural variation in ISR among Arabidopsis accessions. The Ws accession is deficient in ISR, and the responsible recessive genetic locus, named ISR1, was mapped to chromosome 3. To find candidate genes that may underlie ISR deficiency in Ws, we identified genes that are induced in response to the ISR-triggering bacterium Pseudomonas simiae WCS417 and to iron stress and that have non-synonymous mutations in the Ws genome with respect to the ISR-responsive Col-0. We identified a kelch-domain containing protein encoded by At3g07720 that has a genomic rearrangement in Ws. We found that overexpression of Col-0 At3g07720 restores ISR to Ws, indicating that At3g07720 encodes ISR1. Isr1 loss of function mutants do not affect plant growth under iron limiting conditions but have increased levels of apoplastic iron. We found that iron supplementation, P. simiae WCS417, or a loss of isr1 enhance ROS production in a non-additive manner, suggesting they work through the same mechanism to enhance resistance. Our findings show that ISR1 is required for iron localization, immunity, and ISR, and suggest that increased iron uptake induced by ISR-eliciting bacteria may directly contribute to immunity through increased reactive oxygen production.

plant biology↗

bioRxiv2021

Commensal Pseudomonas fluorescens protect Arabidopsis from closely-related Pseudomonas pathogens in a colonization-dependent manner

Plants form commensal associations with soil microorganisms, creating a root microbiome that provides benefits to the host including protection against pathogens. While bacteria can inhibit pathogens through production of antimicrobial compounds in vitro, it is largely unknown how microbiota contribute to pathogen protection in planta. We developed a gnotobiotic model system consisting of Arabidopsis thaliana, and an opportunistic pathogen Pseudomonas sp. N2C3, to identify mechanisms that determine the outcome of plant-pathogen-microbiome interactions in the rhizosphere. We screened 25 phylogenetically diverse Pseudomonas strains for their ability to protect against N2C3 and found that commensal strains closely related to N2C3 were more likely to protect against pathogenesis. We used a comparative genomics approach to identify unique genes in the protective strains that revealed no genes that correlate with protection, suggesting that variable regulation of components of the core Pseudomonas genome may contribute to pathogen protection. We found that commensal colonization level was highly predictive of protection and so tested deletions in genes previously shown to be required for Arabidopsis rhizosphere colonization. We identified a response regulator colR that is required for Pseudomonas protection from N2C3 and fitness in competition with N2C3 indicating that competitive exclusion may contribute to pathogen protection. We found that Pseudomonas WCS365 also protects against the agricultural pathogen Pseudomonas fuscovaginae SE-1, the causal agent of bacterial sheath brown rot of rice. This work establishes a gnotobiotic model to uncover mechanisms by which members of the microbiome can protect hosts from pathogens and informs our understanding of the use of beneficial strains for microbiome engineering in dysbiotic soil systems.

microbiology↗

bioRxiv2021

Sticky Pi, an AI-powered smart insect trap for community chronoecology

In the face of severe environmental crises that threaten insect biodiversity, new technologies are imperative to monitor both the identity and ecology of insect species. Traditionally, insect surveys rely on manual collection of traps, which provide abundance data but mask the large intra- and inter-day variations in insect activity, an important facet of their ecology. Although laboratory studies have shown that circadian processes are central to insects biological functions, from feeding to reproduction, we lack the high-frequency monitoring tools to study insect circadian biology in the field. To address these issues, we developed the Sticky Pi, a novel, autonomous, open-source, insect trap that acquires images of sticky cards every twenty minutes. Using custom deep-learning algorithms, we automatically and accurately scored where, when and which insects were captured. First, we validated our device in controlled laboratory conditions with a classic chronobiological model organism, Drosophila melanogaster. Then, we deployed an array of Sticky Pis to the field to characterise the daily activity of an agricultural pest, Drosophila suzukii, and its parasitoid wasps. Finally, we demonstrate the wide scope of our smart trap by describing the sympatric arrangement of insect temporal niches in a community, without targeting particular taxa a priori. Together, the automatic identification and high sampling rate of our tool provide biologists with unique data that impacts research far beyond chronobiology; with applications to biodiversity monitoring and pest control as well as fundamental implications for phenology, behavioural ecology, and ecophysiology. We released the Sticky Pi project as an open community resource on https://doc.sticky-pi.com.

ecology↗

bioRxiv2020

Loss of a plant receptor kinase recruits beneficial rhizosphere-associated Pseudomonas

Maintaining microbiome structure is critical for the health of both plants1 and animals2. In plants, enrichment of beneficial bacteria is associated with advantageous outcomes including protection from biotic and abiotic stress3,4. However, the genetic and molecular mechanisms by which plants enrich for specific beneficial microbes without general dysbiosis have remained elusive. Here we show that through regulation of NADPH oxidase, FERONIA kinase negatively regulates beneficial Pseudomonas fluorescens in the Arabidopsis rhizosphere microbiome. By rescreening a collection of Arabidopsis mutants that affect root immunity under gnotobiotic conditions, followed by microbiome sequencing in natural soil, we identified a FERONIA mutant (fer-8) with a rhizosphere microbiome enriched in P. fluorescens without phylum-level dysbiosis. Using microbiome transplant experiments, we found that the fer-8 microbiome was beneficial and promoted plant growth. The effect of FER on rhizosphere Pseudomonads was independent of its immune coreceptor function, role in development, and jasmonic acid autoimmunity. We found that the fer-8 mutant has reduced basal levels of reactive oxygen species (ROS) in roots and that mutants deficient in NADPH oxidase showed elevated rhizosphere Pseudomonad levels. Overexpression of the ROP2 gene (encoding a client of FER and positive regulator of NADPH oxidase5) in fer-8 plants suppressed Pseudomonad overgrowth. This work shows that FER-mediated ROS production regulates levels of beneficial Pseudomonads in the rhizosphere microbiome.

plant biology↗

bioRxiv2020

Comparative genomics identified a genetic locus in plant-associated Pseudomonas spp. that is necessary for induced systemic susceptibility

Plant root-associated microbes promote plant growth and induce systemic resistance (ISR) to foliar pathogens. In an attempt to find novel growth-promoting and ISR-inducing strains, we previously identified strains of root-associated Pseudomonas spp. that promote plant growth but unexpectedly induced systemic susceptibility (ISS) rather than ISR to foliar pathogens. Here we demonstrate that the ISS-inducing phenotype is common among root-associated Pseudomonas spp. Using comparative genomics, we identified a single P. fluorescens locus that is unique to ISS strains. We generated a clean deletion of the 11-gene ISS locus and found that it is necessary for the ISS phenotype. Although the functions of the predicted genes in the locus are not apparent based on similarity to genes of known function, the ISS locus is present in diverse bacteria and a subset of the genes have previously been implicated in pathogenesis in animals. Collectively these data show that a single bacterial locus contributes to modulation of systemic plant immunity. ImportanceMicrobiome-associated bacteria can have diverse effects on health of their hosts, yet the genetic and molecular basis of these effects have largely remained elusive. This work demonstrates that a novel bacterial locus can modulate systemic plant immunity. Additionally, this work demonstrates that growth promoting strains may have unanticipated consequences on plant immunity and this is critical to consider when engineering the plant microbiome for agronomic improvement.

microbiology↗

bioRxiv2019

Ectomycorrhizal fungi induce systemic resistance against insects on a non-mycorrhizal plant in a CERK1-dependent manner

Below-ground microbes can induce systemic resistance (ISR) against foliar pests and pathogens on diverse plant hosts. The prevalence of ISR among plant-microbe-pest systems raises the question of host specificity in microbial induction of ISR. To test whether ISR is limited by plant host range, we tested the ISR-inducing ectomycorrhizal (ECM) fungus Laccaria bicolor on the non-mycorrhizal plant Arabidopsis. We found that root inoculation with L. bicolor triggered ISR against the insect herbivore Trichoplusia ni and induced systemic susceptibility (ISS) against the bacterial pathogen Pseudomonas syringae pv. tomato DC3000 (Pto). We found that L. bicolor-triggered ISR against T. ni was dependent on jasmonic acid (JA) signaling and salicylic acid (SA) biosynthesis and signaling. We found that heat killed L. bicolor and chitin are sufficient to trigger ISR against T. ni and ISS against Pto and that the chitin receptor CERK1 is necessary for L. bicolor-mediated effects on systemic immunity. Collectively our findings suggest that some ISR responses might not require intimate co-evolution of host and microbe, but rather might be the result of root perception of conserved microbial signals.

plant biology↗

bioRxiv2019

Bacterially produced spermidine induces plant systemic susceptibility to pathogens

plant biology↗

bioRxiv2018

A genome-wide screen identifies genes in rhizosphere-associated Pseudomonas required to evade plant defenses

Pseudomonas fluorescens and related plant root- (\"rhizosphere\") associated species contribute to plant health by modulating defenses and facilitating nutrient uptake. To identify bacterial fitness determinants in the rhizosphere of the model plant Arabidopsis thaliana, we performed a Tn-Seq screen using the biocontrol and growth-promoting strain Pseudomonas sp. WCS365. The screen, which was performed in parallel on wild-type and an immunocompromised Arabidopsis, identified 231 genes that positively affect fitness in the rhizosphere of wild-type plants. A subset of these genes negatively affect fitness in the rhizosphere of immunocompromised plants. We postulated that these genes might be involved in avoiding plant defenses and verified 7 Pseudomonas sp. WCS365 candidate genes by generating clean deletions. We found that two of these deletion strains, {Delta}morA (encodes a putative diguanylate cyclase/phosphodiesterase) and {Delta}spuC (encodes a putrescine aminotransferase) formed enhanced biofilms and inhibited plant growth. Inhibition of plant growth by {Delta}spuC and {Delta}morA was the result of pattern triggered immunity (PTI) as measured by induction of an Arabidopsis PTI reporter and FLS2/BAK1-dependent inhibition of plant growth. We found that MorA acts as a phosphodiesterase to inhibit biofilm formation suggesting a possible role in biofilm dispersal. We found that both putrescine and its precursor arginine promote biofilm formation that is enhanced in the {Delta}spuC mutant, which cannot break down putrescine suggesting that putrescine might serve as a signaling molecule in the rhizosphere. Collectively, this work identified novel bacterial factors required to evade plant defenses in the rhizosphere.\n\nImportanceWhile rhizosphere bacteria hold the potential to improve plant health and fitness, little is known about the bacterial genes required to evade host immunity. Using a model system consisting of Arabidopsis and a beneficial Pseudomonas sp. isolate, we identified bacterial genes required for both rhizosphere fitness and for evading host immune responses. This work advances our understanding of how evasion of host defenses contributes to survival in the rhizosphere.

microbiology↗

bioRxiv2018

Convergent gain and loss of genomic islands drives lifestyle changes in plant-associated bacteria

Host-associated bacteria can have both beneficial and detrimental effects on host health. While some of the molecular mechanisms that determine these outcomes are known, little is known about the evolutionary histories of pathogenic or mutualistic lifestyles. Using the model plant Arabidopsis, we found that closely related strains within the Pseudomonas fluorescens species complex promote plant growth and occasionally cause disease. To elucidate the genetic basis of the transition between commensalism and pathogenesis, we developed a computational pipeline and identified genomic islands that correlate with outcomes for plant health. One island containing genes for lipopeptide biosynthesis and quorum sensing is required for pathogenesis. Conservation of the quorum sensing machinery in this island allows pathogenic strains to eavesdrop on quorum signals in the environment and coordinate pathogenic behavior. We found that genomic loci associated with both pathogenic and commensal lifestyles were convergently gained and lost in multiple lineages through homologous recombination, possibly constituting an early step in the differentiation of pathogenic and commensal lifestyles. Collectively this work provides novel insights into the evolution of commensal and pathogenic lifestyles within a single clade of host-associated bacteria.

microbiology↗