Quorum sensing, virulence and secondary metabolite production in plant soft-rotting bacteria

Quorum sensing describes the ability of bacteria to sense their population density and respond by modulating gene expression. In the plant soft-rotting bacteria, such as Erwinia, an arsenal of plant cell wall-degrading enzymes is produced in a cell density-dependent manner, which causes maceration of plant tissue. However, quorum sensing is central not only to controlling the production of such destructive enzymes, but also to the control of a number of other virulence determinants and secondary metabolites. Erwinia synthesizes both N-acylhomoserine lactone (AHL) and autoinducer-2 types of quorum sensing signal, which both play a role in regulating gene expression in the phytopathogen. We review the models for AHL-based regulation of carbapenem antibiotic production in Erwinia. We also discuss the importance of quorum sensing in the production and secretion of virulence determinants by Erwinia, and its interplay with other regulatory systems.     

Acyl-homoserine lactone-dependent eavesdropping promotes competition in a laboratory co-culture model

Many Proteobacteria use acyl-homoserine lactone (AHL)-mediated quorum sensing to activate the production of antibiotics at high cell density. Extracellular factors like antibiotics can be considered public goods shared by individuals within a group. Quorum-sensing control of antibiotic production may be important for protecting a niche or competing for limited resources in mixed bacterial communities. To begin to investigate the role of quorum sensing in interspecies competition, we developed a dual-species co-culture model using the soil saprophytes Burkholderia thailandensis (Bt) and Chromobacterium violaceum (Cv). These bacteria require quorum sensing to activate the production of antimicrobial factors that inhibit growth of the other species. We demonstrate that quorum-sensing-dependent antimicrobials can provide a competitive advantage to either Bt or Cv by inhibiting growth of the other species in co-culture. Although the quorum-sensing signals differ for each species, we show that the promiscuous signal receptor encoded by Cv can sense signals produced by Bt, and that this ability to eavesdrop on Bt can provide Cv an advantage in certain situations. We use an in silico approach to investigate the effect of eavesdropping in competition, and show conditions where early activation of antibiotic production resulting from eavesdropping can promote competitiveness. Our work supports the idea that quorum sensing is important for interspecies competition and that promiscuous signal receptors allow eavesdropping on competitors in mixed microbial habitats.     

Molecular phylogeny based on the 16S rRNA gene of elite rhizobial strains used in Brazilian commercial inoculants

Nitrogen is often a limiting nutrient, therefore the sustainability of food crops, forages and green manure legumes is mainly associated with their ability to establish symbiotic associations with stem and root-nodulating N2-fixing rhizobia. The selection, identification and maintenance of elite strains for each host are critical. Decades of research in Brazil resulted in a list of strains officially recommended for several legumes, but their genetic diversity is poorly known. This study aimed at gaining a better understanding of phylogenetic relationships of 68 rhizobial strains recommended for 64 legumes, based on the sequencing of the 16S rRNA genes. The strains were isolated from a wide range of legumes, including all three subfamilies and 17 tribes. Nine main phylogenetic branches were defined, seven of them related to the rhizobial species: Bradyrhizobium japonicum, B. elkanii, Rhizobium tropici, R. leguminosarum, Sinorhizobium meliloti/S. fredii, Mesorhizobium ciceri/M. loti, and Azorhizobium caulinodans. However, some strains differed by up to 35 nucleotides from the type strains, which suggests that they may represent new species. Two other clusters included bacteria showing similarity with the genera Methylobacterium and Burkholderia, and amplification with primers for nifH and/or nodC regions was achieved with these strains. Host specificity of several strains was very low, as they were capable of nodulating legumes of different tribes and subfamilies. Furthermore, host specificity was not related to 16S rRNA, therefore evolution of ribosomal and symbiotic genes may have been diverse. Finally, the great diversity observed in this study emphasizes that tropics are an important reservoir of N2-fixation genes.     

Cross-kingdom signalling: exploitation of bacterial quorum sensing molecules by the green seaweed Ulva

The green seaweed Ulva has been shown to detect signal molecules produced by bacteria. Biofilms that release N-acylhomoserine lactones (AHLs) attract zoospores--the motile reproductive stages of Ulva. The evidence for AHL involvement is based on several independent lines of evidence, including the observation that zoospores are attracted to wild-type bacteria that produce AHLs but are not attracted to mutants that do not produce signal molecules. Synthetic AHL also attracts zoospores and the attraction is lost in the presence of autoinducer inactivation (AiiA) protein. The mechanism of attraction is not chemotactic but involves chemokinesis. When zoospores detect AHLs, the swimming rate is reduced and this results in accumulation of cells at the source of the AHL. It has been demonstrated that the detection of AHLs results in calcium influx into the zoospore. This is the first example of a calcium signalling event in a eukaryote in response to bacterial quorum sensing molecules. The role of AHLs in the ecology of Ulva is discussed. It is probable that AHLs act as cues for the settlement of zoospores, rather than being directly involved as a signalling mechanism.     

Quorum sensing in nitrogen-fixing rhizobia.

Members of the rhizobia are distinguished for their ability to establish a nitrogen-fixing symbiosis with leguminous plants. While many details of this relationship remain a mystery, much effort has gone into elucidating the mechanisms governing bacterium-host recognition and the events leading to symbiosis. Several signal molecules, including plant-produced flavonoids and bacterially produced nodulation factors and exopolysaccharides, are known to function in the molecular conversation between the host and the symbiont. Work by several laboratories has shown that an additional mode of regulation, quorum sensing, intercedes in the signal exchange process and perhaps plays a major role in preparing and coordinating the nitrogen-fixing rhizobia during the establishment of the symbiosis. Rhizobium leguminosarum, for example, carries a multitiered quorum-sensing system that represents one of the most complex regulatory networks identified for this form of gene regulation. This review focuses on the recent stream of information regarding quorum sensing in the nitrogen-fixing rhizobia. Seminal work on the quorum-sensing systems of R. leguminosarum bv. viciae, R. etli, Rhizobium sp. strain NGR234, Sinorhizobium meliloti, and Bradyrhizobium japonicum is presented and discussed. The latest work shows that quorum sensing can be linked to various symbiotic phenomena including nodulation efficiency, symbiosome development, exopolysaccharide production, and nitrogen fixation, all of which are important for the establishment of a successful symbiosis. Many questions remain to be answered, but the knowledge obtained so far provides a firm foundation for future studies on the role of quorum-sensing mediated gene regulation in host-bacterium interactions.     

Look who's talking: communication and quorum sensing in the bacterial world

For many years bacteria were considered primarily as autonomous unicellular organisms with little capacity for collective behaviour. However, we now appreciate that bacterial cells are in fact, highly communicative. The generic term 'quorum sensing' has been adopted to describe the bacterial cell-to-cell communication mechanisms which co-ordinate gene expression usually, but not always, when the population has reached a high cell density. Quorum sensing depends on the synthesis of small molecules (often referred to as pheromones or autoinducers) that diffuse in and out of bacterial cells. As the bacterial population density increases, so does the synthesis of quorum sensing signal molecules, and consequently, their concentration in the external environment rises. Once a critical threshold concentration has been reached, a target sensor kinase or response regulator is activated (or repressed) so facilitating the expression of quorum sensing-dependent genes. Quorum sensing enables a bacterial population to mount a co-operative response that improves access to nutrients or specific environmental niches, promotes collective defence against other competitor prokaryotes or eukaryotic defence mechanisms and facilitates survival through differentiation into morphological forms better able to combat environmental threats. Quorum sensing also crosses the prokaryotic-eukaryotic boundary since quorum sensing-dependent signalling can be exploited or inactivated by both plants and mammals.     

Quorum-sensing in Rhizobium

Quorum-sensing signals are found in many species of legume-nodulating rhizobia. In a well-characterized strain of R. leguminosarum biovar viciae, a variety of autoinducers are synthesised, and all have been identified as N-acyl-homoserine lactones. One of these N-acyl-homoserine lactones, is N-(3-hydroxy-7-cis-tetradecenoyl)-L-homoserine lactone, previously known as small bacteriocin, which inhibits the growth of several R. leguminosarum strains. The cinRI locus is responsible for the production of small bacteriocin. CinR induces cinI in response to the AHL made by CinI, thus forming a positive autoregulatory induction loop. A complex cascade of quorum-sensing loops was characterized, in which the cinIR locus appears to be the master control for three other AHL-dependent quorum-sensing control systems. These systems include the raiI/raiR, traI/triR and rhiI/rhiR. Other rhizobial strains appear to share some of these quorum sensing loci, but not all loci are found in all strains. Small bacteriocin along with the other N-acyl-homoserine lactones produced by these three AHL-based control systems regulate (i) growth inhibition of sensitive strains, (ii) transfer of the symbiotic plasmid pRL1JI, and (iii) expression of the rhizosphere-expressed (rhi) genes that influence nodulation. Some of the genes regulated by these systems have been identified. While the functions of some, such as the trb operon regulated by triR are clear, several of the regulated genes have no homologues of known function. It is anticipated that several other genes regulated by these systems have yet to be identified. Therefore, despite the regulation of one of the most complex quorum-sensing cascade being understood, several of the functions regulated by the quorum-sensing genes remain to be elucidated. This revised version was published online in August 2006 with corrections to the Cover Date.     

Mutualism versus pathogenesis: the give-and-take in plant–bacteria interactions

Pathogenic bacteria and mutualistic rhizobia are able to invade and establish chronic infections within their host plants. The success of these plant–bacteria interactions requires evasion of the plant innate immunity by either avoiding recognition or by suppressing host defences. The primary plant innate immunity is triggered upon recognition of common microbe-associated molecular patterns. Different studies reveal striking similarities between the molecular bases underlying the perception of rhizobial nodulation factors and microbe-associated molecular patterns from plant pathogens. However, in contrast to general elicitors, nodulation factors can control plant defences when recognized by their cognate legumes. Nevertheless, in response to rhizobial infection, legumes show transient or local defence-like responses suggesting that Rhizobium is perceived as an intruder although the plant immunity is controlled. Whether these responses are involved in limiting the number of infections or whether they are required for the progression of the interaction is not yet clear. Further similarities in both plant–pathogen and Rhizobium –legume associations are factors such as surface polysaccharides, quorum sensing signals and secreted proteins, which play important roles in modulating plant defence responses and determining the outcome of the interactions.     

The quorum-sensing system in a plant bacterium Mesorhizobium huakuii affects growth rate and symbiotic nodulation

Mesorhizobium huakuii is a free-living bacterium which is capable of establishing a specific symbiotic relationship with Astragalus sinicus, an important winter green manure widely used in Eastern Asia, allowing for nitrogen fixing during this process. Previous studies demonstrate that M.␣huakuii produces quorum-sensing molecules at high cell density and quorum sensing plays a role in biofilm formations. In this study, we isolated and characterized two quorum-sensing deficient mutants in M. huakuii. Analysis of the flanking region of transposon insertions indicated that autoinducer synthase related genes are not homologous to acyl homoserine lactone (AHL) synthase genes that are shared among many Gram-negative bacteria, but related to peptide synthesis, indicating that M. huakuii quorum-sensing signals are distinct from AHLs. Compared with the wild-type strains, these quorum-sensing deficient mutants promoted their growth rate and were defective in nodule formation on host plants, indicative of a critical role of quorum sensing in M.␣huakuii during the host–bacterium symbiotic interaction.Yijing Gao and Zengtao Zhong contributed equally to this work.     

Phylogenetic diversity of Rhizobium strains nodulating diverse legume species growing in Ethiopia

The taxonomic diversity of thirty-seven Rhizobium strains, isolated from nodules of leguminous trees and herbs growing in Ethiopia, was studied using multilocus sequence analyses (MLSA) of six core and two symbiosis-related genes. Phylogenetic analysis based on the 16S rRNA gene grouped them into five clusters related to nine Rhizobium reference species (99–100% sequence similarity). In addition, two test strains occupied their own independent branches on the phylogenetic tree (AC86a2 along with R. tibeticum; 99.1% similarity and AC100b along with R. multihospitium; 99.5% similarity). One strain from Milletia ferruginea was closely related (>99%) to the genus Shinella, further corroborating earlier findings that nitrogen-fixing bacteria are distributed among phylogenetically unrelated taxa. Sequence analyses of five housekeeping genes also separated the strains into five well-supported clusters, three of which grouped with previously studied Ethiopian common bean rhizobia. Three of the five clusters could potentially be described into new species. Based on the nifH genes, most of the test strains from crop legumes were closely related to several strains of Ethiopian common bean rhizobia and other symbionts of bean plants (R. etli and R. gallicum sv. phaseoli). The grouping of the test strains based on the symbiosis-related genes was not in agreement with the housekeeping genes, signifying differences in their evolutionary history. Our earlier studies revealing a large diversity of Mesorhizobium and Ensifer microsymbionts isolated from Ethiopian legumes, together with the results from the present analysis of Rhizobium strains, suggest that this region might be a potential hotspot for rhizobial biodiversity.     

Messing with bacterial quorum sensing.

Quorum sensing is widely recognized as an efficient mechanism to regulate expression of specific genes responsible for communal behavior in bacteria. Several bacterial phenotypes essential for the successful establishment of symbiotic, pathogenic, or commensal relationships with eukaryotic hosts, including motility, exopolysaccharide production, biofilm formation, and toxin production, are often regulated by quorum sensing. Interestingly, eukaryotes produce quorum-sensing-interfering (QSI) compounds that have a positive or negative influence on the bacterial signaling network. This eukaryotic interference could result in further fine-tuning of bacterial quorum sensing. Furthermore, recent work involving the synthesis of structural homologs to the various quorum-sensing signal molecules has resulted in the development of additional QSI compounds that could be used to control pathogenic bacteria. The creation of transgenic plants that express bacterial quorum-sensing genes is yet another strategy to interfere with bacterial behavior. Further investigation on the manipulation of quorum-sensing systems could provide us with powerful tools against harmful bacteria.     

厌氧氨氧化菌群体感应系统研究

厌氧氨氧化(Anammox)是以铵为电子供体将亚硝酸盐转化为氮气的生物过程。厌氧氨氧化菌(AAOB)生理代谢和细胞结构均十分特殊,且在氮素循环中起着十分重要的作用。厌氧氨氧化已成为环境学、微生物学、海洋学等领域的研究热点。但是,至今人们未能对厌氧氨氧化菌进行纯培养,这严重限制了对厌氧氨氧化菌的深入研究。群体感应是一种普遍存在于微生物细胞之间的通讯机制,它具有根据菌群密度和周围环境变化调节基因表达,以控制细菌群体行为的功能。厌氧氨氧化菌活性的细胞密度效应和生物团聚行为与细菌中普遍存在的群体感应现象相符。探讨了厌氧氨氧化菌群体感应系统存在的可能性、工作机制及其生态学意义,以期为厌氧氨氧化菌的分离培养、团聚体培育等提供理论指导。     

Diversity of rhizobia and interactions among the host legumes and rhizobial genotypes in an agricultural-forestry ecosystem

To investigate the diversity of rhizobia and interactions among the host legumes and rhizobial genotypes in the same habitat, a total of 97 rhizobial strains isolated from nine legume species grown in an agricultural-forestry ecosystem were identified into seven genomic species and 12 symbiotic genotypes within the genera Bradyrhizobium, Mesorhizobium, Rhizobium and Sinorhizobium based upon analyses of genomic DNA regions and symbiotic genes. The results evidenced that the symbiotic genotypes of rhizobia were consistent with their hosts of origin; revealed that vertical transfer was the main mechanism in rhizobia to maintain the symbiotic genes but lateral transfer of symbiotic genes might have happened between the closely related rhizobial species; suggested the existence of co-distribution and co-evolution among the legume hosts and compatible rhizobia. All of these data demonstrated that the biogeography of rhizobia was a result of interactions among the host legumes, bacterial genomic backgrounds and environments.     

Quorum Sensing in Nitrogen-Fixing Rhizobia

Members of the rhizobia are distinguished for their ability to establish a nitrogen-fixing symbiosis with leguminous plants. While many details of this relationship remain a mystery, much effort has gone into elucidating the mechanisms governing bacterium-host recognition and the events leading to symbiosis. Several signal molecules, including plant-produced flavonoids and bacterially produced nodulation factors and exopolysaccharides, are known to function in the molecular conversation between the host and the symbiont. Work by several laboratories has shown that an additional mode of regulation, quorum sensing, intercedes in the signal exchange process and perhaps plays a major role in preparing and coordinating the nitrogen-fixing rhizobia during the establishment of the symbiosis. Rhizobium leguminosarum, for example, carries a multitiered quorum-sensing system that represents one of the most complex regulatory networks identified for this form of gene regulation. This review focuses on the recent stream of information regarding quorum sensing in the nitrogen-fixing rhizobia. Seminal work on the quorum-sensing systems of R. leguminosarum bv. viciae, R. etli, Rhizobium sp. strain NGR234, Sinorhizobium meliloti, and Bradyrhizobium japonicum is presented and discussed. The latest work shows that quorum sensing can be linked to various symbiotic phenomena including nodulation efficiency, symbiosome development, exopolysaccharide production, and nitrogen fixation, all of which are important for the establishment of a successful symbiosis. Many questions remain to be answered, but the knowledge obtained so far provides a firm foundation for future studies on the role of quorum-sensing mediated gene regulation in host-bacterium interactions.     

Evolutionary genetics of rhizobia: molecular and population aspects

The molecular analysis of the genetic systems controlling the main stages of nodule bacteria (rhizobia) interaction with a legume host (signaling at early stages and symbiotic nitrogen fixation) has shown that the widespread recombination of genetic material in free-living ancestors of rhizobia was an important factor in the evolution of these systems. These recombinations could be conditioned by a high content of repeated DNA sequences and the IS elements in the rhizobial genome. A high recombination activity of rhizobia is manifested in the panmictic structure of their populations, which is associated with frequency-dependent selection favoring rare recombinants. This selection is realized through the competition of virulent strains for the nodule formation and can be controlled by the genes whose expression depends on population density (via the quorum sensing mechanism). A high degree of panmixia in rhizobial populations is associated with their ecotypic polymorphism, manifested as the coexistence of symbiotic and nonsymbiotic strains. This type of polymorphism is caused by individual selection during the periodic changes of ecological niches (soil-plant host) in the rhizobia life cycle. The rhizobia-plant interaction stimulates selection in bacterial populations, which results in the increased levels of their heterogeneity and panmixia. The combination of individual and frequency-dependent selection types resulted in the high rates of symbiosis evolution and polyphyletic origin of diverse rhizobial species.     

Messing with Bacterial Quorum Sensing

Quorum sensing is widely recognized as an efficient mechanism to regulate expression of specific genes responsible for communal behavior in bacteria. Several bacterial phenotypes essential for the successful establishment of symbiotic, pathogenic, or commensal relationships with eukaryotic hosts, including motility, exopolysaccharide production, biofilm formation, and toxin production, are often regulated by quorum sensing. Interestingly, eukaryotes produce quorum-sensing-interfering (QSI) compounds that have a positive or negative influence on the bacterial signaling network. This eukaryotic interference could result in further fine-tuning of bacterial quorum sensing. Furthermore, recent work involving the synthesis of structural homologs to the various quorum-sensing signal molecules has resulted in the development of additional QSI compounds that could be used to control pathogenic bacteria. The creation of transgenic plants that express bacterial quorum-sensing genes is yet another strategy to interfere with bacterial behavior. Further investigation on the manipulation of quorum-sensing systems could provide us with powerful tools against harmful bacteria.