Biology subjects

Delavat, F.

Publications and source records attributed to Delavat, F..

8 recordsLinked to original sources

bioRxiv2025

Prophage-marine diazotroph interplay shapes both biofilm structure and nitrogen release

Marine environments are frequently oligotrophic, characterized by low amount of bioassimilable Nitrogen sources. At the global scale, the microbial fixation of N2, or diazotrophy, represents the primary source of fixed nitrogen in pelagic marine ecosystems, playing a key role in supporting primary production and driving the export of organic matter to the deep ocean. However, given the high energetic cost of N2 fixation, the active release of fixed nitrogen by diazotrophs appears counterintuitive, suggesting the existence of alternative, passive release pathways, that remain understudied to date. Here, we show that the marine Non Cyanobacterial Diazotroph Vibrio diazotrophicus is endowed with a prophage belonging to the Myoviridae family, whose expression is induced under anoxic and biofilm-forming conditions. We demonstrate that this prophage can spontaneously excise from the genome of its host and that it forms intact and infective phage particles. Moreover, phage-mediated host cell lysis leads to increased biofilm production as compared to a prophage-free derivative mutant, and to increased release of Dissolved Organic Carbon and of ammonium. Altogether, we provide evidences that viruses may play a previously unrecognized role in oceanic ecosystem dynamics by structuring micro-habitats suitable for diazotrophy and by contributing to the recycling of (in-)organic matter. ImportanceDiazotrophs are key players in ocean functioning by providing fixed nitrogen to ecosystems and fueling primary production. However, from a physiological point-of-view, the active release of nitrogenous compounds by diazotrophs is paradoxical, since they would invest in an energy-intensive process and supply nutrient to non-sibling cells, with the risk of being outcompeted. Therefore, alternative ways leading to the release of fixed nitrogen must exist. Here, we show that the marine Non Cyanobacterial Diazotroph Vibrio diazotrophicus possesses one prophage, whose activation leads to cell death, to increased biofilm production and to the release of Dissolved Organic Compounds and of ammonium. Taken together, our results provide evidences that marine phage-diazotroph interplay leads to the creation of micro-habitats suitable for diazotrophy like biofilm and to nutrient cycling, and contribute to better understanding of the role of viruses in marine ecosystems.

microbiology↗

bioRxiv2023

Development and utilization of new O2-independent bioreporters

Fluorescent proteins have revolutionized science since their discovery in 1962. They have enabled imaging experiments to decipher the function of proteins, cells and organisms, as well as gene regulation. GFP and all its derivatives are now standard tools in cell biology, immunology, molecular biology and microbiology laboratories around the world. A common feature of these proteins is their O2-dependent maturation allowing fluorescence, which precludes their use in anoxic contexts. In this work, we report the development and in cellulo characterization of genetic circuits encoding the O2-independent KOFP-7 protein, a flavin-binding fluorescent protein. We have optimized the genetic circuit for high bacterial fluorescence at population and single-cell level, implemented this circuit in various plasmids differing in host range, and quantified their fluorescence under both aerobic and anaerobic conditions. Finally, we showed that KOFP-7 based constructions can be used to produce fluorescing cells of V. diazotrophicus, a facultative anaerobe, demonstrating the usefulness of the genetic circuits for various anaerobic bacteria. These genetic circuits can thus be modified at will, both to solve basic and applied research questions, opening a highway to shed light on the obscure anaerobic world. ImportanceFluorescent proteins are used since decades, and have allowed major discoveries in biology in a wide variety of fields, and are used in environmental as well as clinical contexts. GFP and all its derivatives share a common feature: they rely on the presence of O2 for protein maturation and fluorescence. This dependency precludes their use in anoxic environments. Here, we constructed a series of genetic circuits allowing production of KOFP-7, an O2-independant Flavin-Binding Fluorescent Protein. We demonstrated that Escherichia coli cells producing KOFP-7 are fluorescent, both at the population and single-cell levels. Importantly, we showed that, unlike cells producing GFP, cells producing KOFP-7 are fluorescent in anoxia. Finally, we demonstrated that Vibrio diazotrophicus NS1, a facultative anaerobe, is fluorescent in the absence of O2 when KOFP-7 is produced. Altogether, the development of new genetic circuits allowing O2-independent fluorescence will open new perspective to study anaerobic processes.

molecular biology↗

bioRxiv2023

Cooperation and cheating orchestrate Vibrio assemblages and polymicrobial synergy in oysters infected with OsHV-1 virus

Polymicrobial diseases significantly impact the health of humans and animals but remain understudied in natural systems. We recently described the Pacific Oyster Mortality Syndrome (POMS), a polymicrobial disease that impacts oyster production and is prevalent worldwide. Analysis of POMS-infected oysters on the French North Atlantic coast revealed that the disease involves co-infection with the endemic ostreid herpesvirus 1 (OsHV-1) and virulent bacterial species such as Vibrio crassostreae. However, it is unknown whether consistent Vibrio populations are associated with POMS in different regions, how Vibrio contribute to POMS, and how they interact with the OsHV-1 virus during pathogenesis. We resolved the Vibrio population structure in oysters from a Mediterranean ecosystem and investigated their functions in POMS development. We find that Vibrio harveyi and Vibrio rotiferianus are the predominant species found in OsHV-1-diseased oysters and show that OsHV-1 is necessary to reproduce the partition of the Vibrio community observed in the field. By characterizing the interspecific interactions between OsHV-1, V. harveyi and V. rotiferianus, we find that only V. harveyi synergizes with OsHV-1. When co-infected, OsHV-1 and V. harveyi behave cooperatively by promoting mutual growth and accelerating oyster death. V. harveyi showed high virulence potential in oysters and dampened host cellular defenses, making oysters a more favorable niche for microbe colonization. We next investigated the interactions underlying the co-occurrence of diverse Vibrio species in diseased oysters. We found that V. harveyi harbors genes responsible for the biosynthesis and uptake of a key siderophore called vibrioferrin. This important resource promotes the growth of V. rotiferianus, a cheater that efficiently colonizes oysters during POMS without costly investment in host manipulation nor metabolite sharing. By connecting field-based approaches, laboratory infection assays and functional genomics, we have uncovered a web of interdependencies that shape the structure and function of the POMS pathobiota. We showed that cooperative behaviors contribute to synergy between bacterial and viral co-infecting partners. Additional cheating behaviors further shape the polymicrobial consortium. Controlling such behaviors or countering their effects opens new avenues for mitigating polymicrobial diseases.

microbiology↗

bioRxiv2022

Genetic and physiological insights into the diazotrophic activity of a non-cyanobacterial marine diazotroph

Nitrogen (N2) fixation, or diazotrophy, supports a large part of primary production in oceans. Culture-independent approaches highlighted the presence in abundance of marine non-cyanobacterial diazotrophs (NCD) but their ecophysiology remains elusive, mostly because of the low number of isolated NCD and because of the lack of available genetic tools for these isolates. Here, a dual genetic and functional approach allowed unveiling the ecophysiology of a marine NCD affiliated to the species Vibrio diazotrophicus. Physiological characterization of the first marine NCD mutant obtained so far was performed using a soft-gellan assay, demonstrating that a {Delta}nifH mutant in not able to grow in nitrogen-deprived media. Furthermore, we demonstrated that V. diazotrophicus produces a thick biofilm under diazotrophic conditions, suggesting biofilm production as an adaptive response of this NCD to cope with the inhibition of nitrogen-fixation by molecular oxygen. Finally, the genomic signature of V. diazotrophicus is essentially absent from metagenomic data of Tara Ocean expeditions, despite having been isolated from various marine environments. We think that the genetically tractable V. diazotrophicus strain used in this study may serve as an ideal model to study the ecophysiology of these overlooked procaryotic group.

microbiology↗

bioRxiv2021

A bistable orthogonal prokaryotic differentiation system underlying development of conjugative transfer competence

The mechanisms and impact of horizontal gene transfer processes to distribute gene functions with potential adaptive benefit among prokaryotes have been well documented. In contrast, little is known about the life-style of mobile elements mediating horizontal gene transfer, whereas this is the ultimate determinant for their transfer fitness. Here, we investigate the life-style of an integrative and conjugative element (ICE) within the genus Pseudomonas that stands model for a widespread family transmitting genes for xenobiotic compound metabolism and antibiotic resistances. The ICE only transfers from a small fraction of cells in a population, which we uncover here, results from a dedicated transfer competence program imposed by the ICE. Transfer competence is orthogonally maintained in individual cells in which it is activated, making them the centerpiece of ICE conjugation. The components mediating transfer competence are widely conserved, underscoring their selected fitness for efficient transfer of this class of mobile elements.

microbiology↗

bioRxiv2020

An analog to digital converter controls bistable transfer competence of a widespread integrative and conjugative element

Conjugative transfer of the integrative and conjugative element ICEclc in Pseudomonas requires development of a transfer competence state in stationary phase, which arises only in 3-5% of individual cells. The mechanisms controlling this bistable switch between non-active and transfer competent cells have long remained enigmatic. Using a variety of genetic tools and epistasis experiments in P. putida, we uncovered an upstream cascade of three consecutive transcription factor-nodes, which controls transfer competence initiation. Initiation activates a feedback loop, which stochastic modeling and experimental data demonstrated acts as a scalable converter of unimodal input to bistable output. The feedback loop further enables prolonged production of a transcription factor that ensures downstream transfer competence formation in activated cells. Phylogenetic analyses showed that the ICEclc regulatory factors are widespread among Gamma- and Beta-proteobacteria, highlighting its evolutionary conservation and prime importance to control the behaviour of this wide family of conjugative elements.

genomics↗

bioRxiv2019

A four-step regulatory cascade controls bistable transfer competence development of the integrative and conjugative element ICEclc in Pseudomonas

Genetic bistability controls different phenotypic programs in defined subpopulations of genetically identical bacteria. Conjugative transfer of the integrative and conjugative element ICEclc in Pseudomonas requires development of a transfer competence state in stationary phase, but this state arises only in 3-5% of individual cells. The mechanisms controlling and underlying the bistable switch between non-active and transfer competence cells have long remained enigmatic. Using a variety of genetic tools combined with stochastic modeling, we characterize here the factors and overall network architecture controlling bistable ICEclc activation of transfer competence. Two new key regulators (BisR and BisDC) were uncovered, that link the hierarchical cascade of ICEclc transfer competence activation to in total four regulatory nodes. The final activator complex named BisDC drives a positive feedback on its own transcription, and directly controls the "late" ICE promoters for excision and transfer. Stochastic mathematical modeling conceptually explained the arisal and maintenance of bistability by the feedback loop, and demonstrated its importance to guarantee consistent prolonged downstream output in activated cells. A minimized gene set allowing controllable bistable output in a Pseudomonas putida in absence of the ICEclc largely confirmed model predictions. Phylogenetic analyses further showed that the two new ICEclc regulatory factors are widespread among putative ICEs found in Gamma- and Beta- proteobacteria, highlighting the conceptual importance of our findings for the behaviour of this wide family of conjugative elements. Author summaryIntegrative and conjugative elements (ICEs) are mobile genetic elements present in virtually every bacterial species, which can confer adaptive functions to their host, such as antibiotic resistance or xenometabolic pathways. Integrated ICEs maintain by replication along with the genome of their bacterial host, but in order to transfer, the ICE excises and conjugates into a new recipient cell. Single-cell studies on a unique but widely representative ICE model from Pseudomonas (ICEclc) showed that transfer only occurs from a small dedicated subpopulation of cells that arises during stationary phase conditions. This bistable subpopulation differentiation is highly significant for ICE behaviour and fitness, but how it is regulated has remained largely unknown. The present work unveiled the architecture of the ICEclc transfer competence regulation, and showed its widespread occurrence among ICEs of the same family. Stochastic mathematical modeling explained how bistability is generated and maintained, prolonging the capacity of stationary phase cells to complete all stages of ICE activation.

microbiology↗

bioRxiv2018

Transient Replication in Specialized Cells Favors Conjugative Transfer of a Selfish DNA Element

Bacterial evolution is driven to a large extent by horizontal gene transfer (HGT) - the processes that distribute genetic material between species rather than by vertical descent. HGT is mostly mediated by an assortment of different selfish DNA elements, several of which have been characterized in great molecular detail. In contrast, very little is known on adaptive features optimizing horizontal fitness. By using single DNA molecule detection and time-lapse microscopy, we analyze here the fate of an integrative and conjugative element (ICE) in individual cells of the bacterium Pseudomonas putida. We uncover how the ICE excises and irregularly replicates, exclusively in a sub-set of specialized host cells. As postulated, ICE replication is dependent on its origin of transfer and its DNA relaxase. Rather than being required for ICE maintenance, however, we find that ICE replication serves more effective conjugation to recipient cells, providing selectable benefit to its horizontal transfer.

microbiology↗