Detection, Characterization, and Biological Effect of Quorum-Sensing Signaling Molecules in Peanut-Nodulating Bradyrhizobia

Bacteria of the genus Bradyrhizobium are able to establish a symbiotic relationship with peanut (Arachis hypogaea) root cells and to fix atmospheric nitrogen by converting it to nitrogenous compounds. Quorum sensing (QS) is a cell-cell communication mechanism employed by a variety of bacterial species to coordinate behavior at a community level through regulation of gene expression. The QS process depends on bacterial production of various signaling molecules, among which the N-acylhomoserine lactones (AHLs) are most commonly used by Gram-negative bacteria. Some previous reports have shown the production of QS signaling molecules by various rhizobia, but little is known regarding mechanisms of communication among peanut-nodulating strains. The aims of this study were to identify and characterize QS signals produced by peanut-nodulating bradyrhizobial strains and to evaluate their effects on processes related to cell interaction. Detection of AHLs in 53 rhizobial strains was performed using the biosensor strains Agrobacterium tumefaciens NTL4 (pZLR4) and Chromobacterium violaceum CV026 for AHLs with long and short acyl chains, respectively. None of the strains screened were found to produce AHLs with short acyl chains, but 14 strains produced AHLs with long acyl chains. These 14 AHL-producing strains were further studied by quantification of β-galactosidase activity levels (AHL-like inducer activity) in NTL4 (pZLR4). Strains displaying moderate to high levels of AHL-like inducer activity were subjected to chemical identification of signaling molecules by high-performance liquid chromatography coupled to mass spectrometry (LC-MS/MS). For each AHL-producing strain, we found at least four different AHLs, corresponding to N-hexanoyl-dl-homoserine lactone (C6), N-(3-oxodecanoyl)-l-homoserine lactone (3OC10), N-(3-oxododecanoyl)-l-homoserine lactone (3OC12), and N-(3-oxotetradecanoyl)-l-homoserine lactone (3OC14). Biological roles of 3OC10, 3OC12, and 3OC14 AHLs were evaluated in both AHL-producing and -non-producing peanut-nodulating strains. Bacterial processes related to survival and nodulation, including motility, biofilm formation, and cell aggregation, were affected or modified by the exogenous addition of increasing concentrations of synthetic AHLs. Our results clearly demonstrate the existence of cell communication mechanisms among bradyrhizobial strains symbiotic of peanut. AHLs with long acyl chains appear to be signaling molecules regulating important QS physiological processes in these bacteria

Central Role of the Holliday Junction Helicase RuvAB in vlsE Recombination and Infectivity of Borrelia burgdorferi

Antigenic variation plays a vital role in the pathogenesis of many infectious bacteria and protozoa including Borrelia burgdorferi, the causative agent of Lyme disease. VlsE, a 35 kDa surface-exposed lipoprotein, undergoes antigenic variation during B. burgdorferi infection of mammalian hosts, and is believed to be a critical mechanism by which the spirochetes evade immune clearance. Random, segmental recombination between the expressed vlsE gene and adjacent vls silent cassettes generates a large number of different VlsE variants within the infected host. Although the occurrence and importance of vlsE sequence variation is well established, little is known about the biological mechanism of vlsE recombination. To identify factors important in antigenic variation and vlsE recombination, we screened transposon mutants of genes known to be involved in DNA recombination and repair for their effects on infectivity and vlsE recombination. Several mutants, including those in BB0023 (ruvA), BB0022 (ruvB), BB0797 (mutS), and BB0098 (mutS-II), showed reduced infectivity in immunocompetent C3H/HeN mice. Mutants in ruvA and ruvB exhibited greatly reduced rates of vlsE recombination in C3H/HeN mice, as determined by restriction fragment polymorphism (RFLP) screening and DNA sequence analysis. In severe combined immunodeficiency (C3H/scid) mice, the ruvA mutant retained full infectivity; however, all recovered clones retained the ‘parental’ vlsE sequence, consistent with low rates of vlsE recombination. These results suggest that the reduced infectivity of ruvA and ruvB mutants is the result of ineffective vlsE recombination and underscores the important role that vlsE recombination plays in immune evasion. Based on functional studies in other organisms, the RuvAB complex of B. burgdorferi may promote branch migration of Holliday junctions during vlsE recombination. Our findings are consistent with those in the accompanying article by Dresser et al., and together these studies provide the first examples of trans-acting factors involved in vlsE recombination

The Streptococcus pneumoniae Pilus-1 Displays a Biphasic Expression Pattern

The Streptococcus pneumoniae pilus-1 is encoded by pilus islet 1 (PI-1), which has three clonal variants (clade I, II and III) and is present in about 30% of clinical pneumococcal isolates. In vitro and in vivo assays have demonstrated that pilus-1 is involved in attachment to epithelial cells and virulence, as well as protection in mouse models of infection. Several reports suggest that pilus-1 expression is tightly regulated and involves the interplay of numerous genetic regulators, including the PI-1 positive regulator RlrA. In this report we provide evidence that pilus expression, when analyzed at the single-cell level in PI-1 positive strains, is biphasic. In fact, the strains present two phenotypically different sub-populations of bacteria, one that expresses the pilus, while the other does not. The proportions of these two phenotypes are variable among the strains tested and are not influenced by genotype, serotype, growth conditions, colony morphology or by the presence of antibodies directed toward the pilus components. Two sub-populations, enriched in pilus expressing or not expressing bacteria were obtained by means of colony selection and immuno-detection methods for five strains. PI-1 sequencing in the two sub-populations revealed the absence of mutations, thus indicating that the biphasic expression observed is not due to a genetic modification within PI-1. Microarray expression profile and western blot analyses on whole bacterial lysates performed comparing the two enriched sub-populations, revealed that pilus expression is regulated at the transcriptional level (on/off regulation), and that there are no other genes, in addition to those encoded by PI-1, concurrently regulated across the strains tested. Finally, we provide evidence that the over-expression of the RrlA positive regulator is sufficient to induce pilus expression in pilus-1 negative bacteria. Overall, the data presented here suggest that the observed biphasic pilus expression phenotype could be an example of bistability in pneumococcus

Rôle du quorum-sensing et prévalence des bactériophages chez la bactérie phytostimulatrice Azospirillum

Le but de ce travail était d identifier les fonctions régulées par quorum-sensing (QS) chez la bactérie phytostimulatrice Azospirillum. Les effets phytobénéfiques in vitro des souches B518 et TVV3 (isolées du riz) ne sont pas altérées par l inactivation des molécules signal impliquées dans le QS. La combinaison d une approche ciblée et d une approche globale par protéomique montre que le QS régule des fonctions liées à l adaptation à la plante, notamment à la colonisation racinaire chez B518. Chez TVV3, aucune fonction régulée par QS n a pu être identifiée mais les gènes impliqués dans le QS sont localisés dans un environnement atypique, constitué de gènes prophagiques. La mise en évidence d un prophage chez TVV3 a conduit à la caractérisation de phages tempérés chez dix autres souches et au séquençage du premier génome d un bactériophage isolé d Azospirillum. Ce travail montre que la régulation de type QS est souche spécifique et révèle la prévalence des phages chez AzospirillumThe aim of this work was to identify quorum-sensing (QS) regulated functions in the phytobeneficial bacterium Azospirillum. Inactivation of signal molecules involved in QS has no deleterious effect on the in vitro phytostimulatory properties of strains B518 and TVV3 (isolated from rice). By combining a targeted approach and a global proteomic approach, QS was shown to regulate functions linked to adaptation to the plant, notably to root colonization, in strain B518. In strain TVV3, no QS-regulated function was identified but QS genes were localized in an atypical environment containing genes of prophage origin. The isolation of phage particles in strain TVV3 led to the characterization of temperate phages in ten other strains and to the sequencing of the first bacteriophage genome isolated from an Azospirillum strain. This work indicates that QS regulation is strain-specific and reveals bacteriophage prevalence in AzospirillumLYON1-BU.Sciences (692662101) / SudocSudocFranceF

Bacterial hybrid histidine kinases in plant-bacteria interactions

Two-component signal transduction systems are essential for many bacteria to maintain homeostasis and adapt to environmental changes. Two-component signal transduction systems typically involve a membrane-bound histidine kinase that senses stimuli, autophosphorylates in the transmitter region and then transfers the phosphoryl group to the receiver domain of a cytoplasmic response regulator that mediates appropriate changes in bacterial physiology. Although usually found on distinct proteins, the transmitter and receiver modules are sometimes fused into a so-called hybrid histidine kinase (HyHK). Such structure results in multiple phosphate transfers that are believed to provide extra-fine-tuning mechanisms and more regulatory checkpoints than classical phosphotransfers. HyHK-based regulation may be crucial for finely tuning gene expression in a heterogeneous environment such as the rhizosphere, where intricate plant-bacteria interactions occur. In this review, we focus on roles fulfilled by bacterial HyHKs in plant-associated bacteria, providing recent findings on the mechanistic of their signalling properties. Recent insights into understanding additive regulatory properties fulfilled by the tethered receiver domain of HyHKs are also addressed

Ecological conditions and molecular determinants involved in agrobacterium lifestyle in tumors

The study of pathogenic agents in their natural niches allows for a better understanding of disease persistence and dissemination. Bacteria belonging to the Agrobacterium genus are soil-borne and can colonize the rhizosphere. These bacteria are also well known as phytopathogens as they can cause tumors (crown gall disease) by transferring a DNA region (T-DNA) into a wide range of plants. Most reviews on Agrobacterium are focused on virulence determinants, T-DNA integration, bacterial and plant factors influencing the efficiency of genetic transformation. Recent research papers have focused on the plant tumor environment on the one hand, and genetic traits potentially involved in bacterium-plant interactions on the other hand. The present review gathers current knowledge about the special conditions encountered in the tumor environment along with the Agrobacterium genetic determinants putatively involved in bacterial persistence inside a tumor. By integrating recent metabolomic and transcriptomic studies, we describe how tumors develop and how Agrobacterium can maintain itself in this nutrient-rich but stressful and competitive environment