Functional and evolutionary significance of unknown genes from uncultivated taxa.

Many of the Earth's microbes remain uncultured and understudied, limiting our understanding of the functional and evolutionary aspects of their genetic material, which remain largely overlooked in most metagenomic studies1. Here we analysed 149,842 environmental genomes from multiple habitats2-6 and compiled a curated catalogue of 404,085 functionally and evolutionarily significant novel (FESNov) gene families exclusive to uncultivated prokaryotic taxa. All FESNov families span multiple species, exhibit strong signals of purifying selection and qualify as new orthologous groups, thus nearly tripling the number of bacterial and archaeal gene families described to date. The FESNov catalogue is enriched in clade-specific traits, including 1,034 novel families that can distinguish entire uncultivated phyla, classes and orders, probably representing synapomorphies that facilitated their evolutionary divergence. Using genomic context analysis and structural alignments we predicted functional associations for 32.4% of FESNov families, including 4,349 high-confidence associations with important biological processes. These predictions provide a valuable hypothesis-driven framework that we used for experimental validatation of a new gene family involved in cell motility and a novel set of antimicrobial peptides. We also demonstrate that the relative abundance profiles of novel families can discriminate between environments and clinical conditions, leading to the discovery of potentially new biomarkers associated with colorectal cancer. We expect this work to enhance future metagenomics studies and expand our knowledge of the genetic repertory of uncultivated organisms.

A Nitrate-Sensing Domain-Containing Chemoreceptor Is Required for Successful Entry and Virulence of Dickeya dadantii 3937 in Potato Plants.

Nitrate metabolism plays an important role in bacterial physiology. During the interaction of plant-pathogenic bacteria with their hosts, bacteria face variable conditions with respect to nitrate availability. Perception mechanisms through the chemosensory pathway drive the entry and control the colonization of the plant host in phytopathogenic bacteria. In this work, the identification and characterization of the nitrate- and nitrite-sensing (NIT) domain-containing chemoreceptor of Dickeya dadantii 3937 (Dd3937) allowed us to unveil the key role of nitrate sensing not only for the entry into the plant apoplast through wounds but also for infection success. We determined the specificity of this chemoreceptor to bind nitrate and nitrite, with a slight ligand preference for nitrate. Gene expression analysis showed that nitrate perception controls not only the expression of nitrate reductase genes involved in respiratory and assimilatory metabolic processes but also the expression of gyrA, hrpN, and bgxA, three well-known virulence determinants in Dd3937.

Pseudomonas syringae pv. tomato infection of tomato plants is mediated by GABA and l-Pro chemoperception.

Foliar bacterial pathogens have to penetrate the plant tissue and access the interior of the apoplast in order to initiate the pathogenic phase. The entry process is driven by chemotaxis towards plant-derived compounds in order to locate plant openings. However, information on plant signals recognized by bacterial chemoreceptors is scarce. Here, we show that the perception of GABA and l-Pro, two abundant components of the tomato apoplast, through the PsPto-PscC chemoreceptor drives the entry of Pseudomonas syringae pv. tomato into the tomato apoplast. The recognition of both compounds by PsPto-PscC caused chemoattraction to both amino acids and participated in the regulation of GABA catabolism. Mutation of the PsPto-PscC chemoreceptor caused a reduced chemotactic response towards these compounds which in turn impaired entry and reduced virulence in tomato plants. Interestingly, GABA and l-Pro levels significantly increase in tomato plants upon pathogen infection and are involved in the regulation of the plant defence response. This is an example illustrating how bacteria respond to plant signals produced during the interaction as cues to access the plant apoplast and to ensure efficient infection.

Prevalence and Specificity of Chemoreceptor Profiles in Plant-Associated Bacteria.

Chemosensory pathways are among the most abundant prokaryotic signal transduction systems, allowing bacteria to sense and respond to environmental stimuli. Signaling is typically initiated by the binding of specific molecules to the ligand binding domain (LBD) of chemoreceptor proteins (CRs). Although CRs play a central role in plant-microbiome interactions such as colonization and infection, little is known about their phylogenetic and ecological specificity. Here, we analyzed 82,277 CR sequences from 11,806 representative microbial species covering the whole prokaryotic phylogeny, and we classified them according to their LBD type using a de novo homology clustering method. Through phylogenomic analysis, we identified hundreds of LBDs that are found predominantly in plant-associated bacteria, including several LBDs specific to phytopathogens and plant symbionts. Functional annotation of our catalogue showed that many of the LBD clusters identified might constitute unknown types of LBDs. Moreover, we found that the taxonomic distribution of most LBD types that are specific to plant-associated bacteria is only partially explained by phylogeny, suggesting that lifestyle and niche adaptation are important factors in their selection. Finally, our results show that the profile of LBD types in a given genome is related to the lifestyle specialization, with plant symbionts and phytopathogens showing the highest number of niche-specific LBDs. The LBD catalogue and information on how to profile novel genomes are available at https://github.com/compgenomicslab/CRs. IMPORTANCE Considering the enormous variety of LBDs at sensor proteins, an important question resides in establishing the forces that have driven their evolution and selection. We present here the first clear demonstration that environmental factors play an important role in the selection and evolution of LBDs. We were able to demonstrate the existence of LBD families that are highly enriched in plant-associated bacteria but show a wide phylogenetic spread. These findings offer a number of research opportunities in the field of single transduction, such as the exploration of similar relationships in chemoreceptors of bacteria with a different lifestyle, like those inhabiting or infecting the human intestine. Similarly, our results raise the question whether similar LBD types might be shared by members of different sensor protein families. Lastly, we provide a comprehensive catalogue of CRs classified by their LBD region that includes a large number of putative new LBD types.

Blue-light perception by epiphytic Pseudomonas syringae drives chemoreceptor expression, enabling efficient plant infection.

Adaptation and efficient colonization of the phyllosphere are essential processes for the switch to an epiphytic stage in foliar bacterial pathogens. Here, we explore the interplay among light perception and global transcriptomic alterations in epiphytic populations of the hemibiotrophic pathogen Pseudomonas syringae pv. tomato DC3000 (PsPto) following contact with tomato leaves. We found that blue-light perception by PsPto on leaf surfaces is required for optimal colonization. Blue light triggers the activation of metabolic activity and increases the transcript levels of five chemoreceptors through the function of light oxygen voltage and BphP1 photoreceptors. The inactivation of PSPTO_1008 and PSPTO_2526 chemoreceptors causes a reduction in virulence. Our results indicate that during PsPto interaction with tomato plants, light perception, chemotaxis, and virulence are highly interwoven processes.

Chemoperception of Specific Amino Acids Controls Phytopathogenicity in Pseudomonas syringae pv. tomato.

Chemotaxis has been associated with the pathogenicity of bacteria in plants and was found to facilitate bacterial entry through stomata and wounds. However, knowledge regarding the plant signals involved in this process is scarce. We have addressed this issue using Pseudomonas syringae pv. tomato, which is a foliar pathogen that causes bacterial speck in tomato. We show that the chemoreceptor P. syringae pv. tomato PscA (PsPto-PscA) recognizes specifically and with high affinity l-Asp, l-Glu, and d-Asp. The mutation of the chemoreceptor gene largely reduced chemotaxis to these ligands but also altered cyclic di-GMP (c-di-GMP) levels, biofilm formation, and motility, pointing to cross talk between different chemosensory pathways. Furthermore, the PsPto-PscA mutant strain showed reduced virulence in tomato. Asp and Glu are the most abundant amino acids in plants and in particular in tomato apoplasts, and we hypothesize that this receptor may have evolved to specifically recognize these compounds to facilitate bacterial entry into the plant. Infection assays with the wild-type strain showed that the presence of saturating concentrations of d-Asp also reduced bacterial virulence.IMPORTANCE There is substantive evidence that chemotaxis is a key requisite for efficient pathogenesis in plant pathogens. However, information regarding particular bacterial chemoreceptors and the specific plant signal that they sense is scarce. Our work shows that the phytopathogenic bacterium Pseudomonas syringae pv. tomato mediates not only chemotaxis but also the control of pathogenicity through the perception of the plant abundant amino acids Asp and Glu. We describe the specificity of the perception of l- and d-Asp and l-Glu by the PsPto-PscA chemoreceptor and the involvement of this perception in the regulation of pathogenicity-related traits. Moreover, a saturating concentration of d-Asp reduces bacterial virulence, and we therefore propose that ligand-mediated interference of key chemoreceptors may be an alternative strategy to control virulence.

The Pseudomonas syringae pv. tomato DC3000 PSPTO_0820 multidrug transporter is involved in resistance to plant antimicrobials and bacterial survival during tomato plant infection.

Multidrug resistance efflux pumps protect bacterial cells against a wide spectrum of antimicrobial compounds. PSPTO_0820 is a predicted multidrug transporter from the phytopathogenic bacterium Pseudomonas syringae pv. tomato DC3000. Orthologs of this protein are conserved within many Pseudomonas species that interact with plants. To study the potential role of PSPTO_0820 in plant-bacteria interaction, a mutant in this gene was isolated and characterized. In addition, with the aim to find the outer membrane channel for this efflux system, a mutant in PSPTO_4977, a TolC-like gene, was also analyzed. Both mutants were more susceptible to trans-cinnamic and chlorogenic acids and to the flavonoid (+)-catechin, when added to the culture medium. The expression level of both genes increased in the presence of (+)-catechin and, in the case of PSPTO_0820, also in response to trans-cinnamic acid. PSPTO_0820 and PSPTO_4977 mutants were unable to colonize tomato at high population levels. This work evidences the involvement of these two proteins in the resistance to plant antimicrobials, supporting also the importance of chlorogenic acid, trans-cinnamic acid, and (+)-catechin in the tomato plant defense response against P. syringae pv. tomato DC3000 infection.

Pseudomonas syringae pv. tomato exploits light signals to optimize virulence and colonization of leaves.

Light is pervasive in the leaf environment, creating opportunities for both plants and pathogens to cue into light as a signal to regulate plant-microbe interactions. Light enhances plant defences and regulates opening of stomata, an entry point for foliar bacterial pathogens such as Pseudomonas syringae pv. tomato DC3000 (PsPto). The effect of light perception on gene expression and virulence was investigated in PsPto. Light induced genetic reprogramming in PsPto that entailed significant changes in stress tolerance and virulence. Blue light-mediated up-regulation of type three secretion system genes and red light-mediated down-regulation of coronatine biosynthesis genes. Cells exposed to white light, blue light or darkness before inoculation were more virulent when inoculated at dawn than dusk probably due to an enhanced entry through open stomata. Exposure to red light repressed coronatine biosynthesis genes which could lead to a reduced stomatal re-opening and PsPto entry. Photoreceptor were required for the greater virulence of light-treated and dark-treated PsPto inoculated at dawn as compared to dusk, indicating that these proteins sense the absence of light and contribute to priming of virulence in the dark. These results support a model in which PsPto exploits light changes to maximize survival, entry and virulence on plants.

Suppression of Plant Immune Responses by the Pseudomonas savastanoi pv. savastanoi NCPPB 3335 Type III Effector Tyrosine Phosphatases HopAO1 and HopAO2.

The effector repertoire of the olive pathogen P. savastanoi pv. savastanoi NCPPB 3335 includes two members of the HopAO effector family, one of the most diverse T3E families of the P. syringae complex. The study described here explores the phylogeny of these dissimilar members, HopAO1 and HopAO2, among the complex and reveals their activities as immune defense suppressors. Although HopAO1 is predominantly encoded by phylogroup 3 strains isolated from woody organs of woody hosts, both HopAO1 and HopAO2 are phylogenetically clustered according to the woody/herbaceous nature of their host of isolation, suggesting host specialization of the HopAO family across the P. syringae complex. HopAO1 and HopAO2 translocate into plant cells and show hrpL-dependent expression, which allows their classification as actively deployed type III effectors. Our data also show that HopAO1 and HopAO2 possess phosphatase activity, a hallmark of the members of this family. Both of them exert an inhibitory effect on early plant defense responses, such as ROS production and callose deposition, and are able to suppress ETI responses induced by the effectorless polymutant of P. syringae pv. tomato DC3000 (DC3000D28E) in Nicotiana. Moreover, we demonstrate that a ΔhopAO1 mutant of P. savastanoi NCPBB 3335 exhibits a reduced fitness and virulence in olive plants, which supports the relevance of this effector during the interaction of this strain with its host plants. This work contributes to the field with the first report regarding functional analysis of HopAO homologs encoded by P. syringae or P. savastanoi strains isolated from woody hosts.

Differential modulation of plant immune responses by diverse members of the Pseudomonas savastanoi pv. savastanoi HopAF type III effector family.

The Pseudomonas savastanoi pv. savastanoi NCPPB 3335 type III secretion system (T3SS) effector repertoire includes 33 candidates, seven of which translocate into host cells and interfere with plant defences. The present study was performed to investigate the co-existence of both plasmid- and chromosomal-encoded members of the HopAF effector family, HopAF1-1 and HopAF1-2, respectively, in the genome of NCPPB 3335. Here, we show that the HopAF1 paralogues are widely distributed in the Pseudomonas syringae complex, where HopAF1-1 is most similar to the homologues encoded by other P. syringae pathovars infecting woody hosts that belong to phylogroups 1 and 3. We show that the expression of both HopAF1-1 and HopAF-2 is transcriptionally dependent on HrpL and demonstrate their delivery into Nicotiana tabacum leaves. Although the heterologous delivery of either HopAF1-1 or HopAF1-2 significantly suppressed the production of defence-associated reactive oxygen species levels, only HopAF1-2 reduced the levels of callose deposition. Moreover, the expression of HopAF1-2 by functionally effectorless P. syringae pv. tomato DC3000D28E completely inhibited the hypersensitive response in tobacco and significantly increased the competitiveness of the strain in Nicotiana benthamiana. Despite their functional differences, subcellular localization studies reveal that green fluorescent protein (GFP) fusions to either HopAF1-1 or HopAF1-2 are targeted to the plasma membrane when they are expressed in plant cells, a process that is completely dependent on the integrity of their N-myristoylation motif. Our results further support the notion that highly similar T3SS effectors might differentially interact with diverse plant targets, even when they co-localize in the same cell compartment.

Prediction of bacterial associations with plants using a supervised machine-learning approach.

Recent scenarios of fresh produce contamination by human enteric pathogens have resulted in severe food-borne outbreaks, and a new paradigm has emerged stating that some human-associated bacteria can use plants as secondary hosts. As a consequence, there has been growing concern in the scientific community about these interactions that have not yet been elucidated. Since this is a relatively new area, there is a lack of strategies to address the problem of food-borne illnesses due to the ingestion of fruits and vegetables. In the present study, we performed specific genome annotations to train a supervised machine-learning model that allows for the identification of plant-associated bacteria with a precision of ∼93%. The application of our method to approximately 9500 genomes predicted several unknown interactions between well-known human pathogens and plants, and it also confirmed several cases for which evidence has been reported. We observed that factors involved in adhesion, the deconstruction of the plant cell wall and detoxifying activities were highlighted as the most predictive features. The application of our strategy to sequenced strains that are involved in food poisoning can be used as a primary screening tool to determine the possible causes of contaminations.

Role of Dickeya dadantii 3937 chemoreceptors in the entry to Arabidopsis leaves through wounds.

Chemotaxis enables bacteria to move towards an optimal environment in response to chemical signals. In the case of plant-pathogenic bacteria, chemotaxis allows pathogens to explore the plant surface for potential entry sites with the ultimate aim to prosper inside plant tissues and to cause disease. Chemoreceptors, which constitute the sensory core of the chemotaxis system, are usually transmembrane proteins which change their conformation when sensing chemicals in the periplasm and transduce the signal through a kinase pathway to the flagellar motor. In the particular case of the soft-rot pathogen Dickeya dadantii 3937, jasmonic acid released in a plant wound has been found to be a strong chemoattractant which drives pathogen entry into the plant apoplast. In order to identify candidate chemoreceptors sensing wound-derived plant compounds, we carried out a bioinformatics search of candidate chemoreceptors in the genome of Dickeya dadantii 3937. The study of the chemotactic response to several compounds and the analysis of the entry process to Arabidopsis leaves of 10 selected mutants in chemoreceptors allowed us to determine the implications of at least two of them (ABF-0020167 and ABF-0046680) in the chemotaxis-driven entry process through plant wounds. Our data suggest that ABF-0020167 and ABF-0046680 may be candidate receptors of jasmonic acid and xylose, respectively.

N2O and CH4 emissions from a fallow-wheat rotation with low N input in conservation and conventional tillage under a Mediterranean agroecosystem.

Conservation agriculture that includes no tillage (NT) or minimum tillage (MT) and crop rotation is an effective practice to increase soil organic matter in Mediterranean semiarid agrosystems. But the impact of these agricultural practices on greenhouse gases (GHGs), such as nitrous oxide (N2O) and methane (CH4), is variable depending mainly on soil structure and short/long-term tillage. The main objective of this study was to assess the long-term effect of three tillage systems (NT, MT and conventional tillage (CT)) and land-covers (fallow/wheat) on the emissions of N2O and CH4 in a low N input agricultural system during one year. This was achieved by measuring crop yields, soil mineral N and dissolved organic C contents, and fluxes of N2O and CH4. Total cumulative N2O emissions were not significantly different (P>0.05) among the tillage systems or between fallow and wheat. The only difference was produced in spring, when N2O emissions were significantly higher (P<0.05) in fallow than in wheat subplots, and NT reduced N2O emissions (P<0.05) compared with MT and CT. Taking into account the water filled pore space (WFPS), both nitrification and denitrification could have occurred during the experimental period. Denitrification capacity in March was similar in all tillage systems, in spite of the higher DOC content maintained in the topsoil of NT. This could be due to the similar denitrifier densities, targeted by nirK copy numbers at that time. Cumulative CH4 fluxes resulted in small net uptake for all treatments, and no significant differences were found among tillage systems or between fallow and wheat land-covers. These results suggest that under a coarse-textured soil in low N agricultural systems, the impact of tillage on GHG is very low and that the fallow cycle within a crop rotation is not a useful strategy to reduce GHG emissions.

Temporal and Spatial Resolution of Activated Plant Defense Responses in Leaves of Nicotiana benthamiana Infected with Dickeya dadantii.

The necrotrophic bacteria Dickeya dadantii is the causal agent of soft-rot disease in a broad range of hosts. The model plant Nicotiana benthamiana, commonly used as experimental host for a very broad range of plant pathogens, is susceptible to infection by D. dadantii. The inoculation with D. dadantii at high dose seems to overcome the plant defense capacity, inducing maceration and death of the tissue, although restricted to the infiltrated area. By contrast, the output of the defense response to low dose inoculation is inhibition of maceration and limitation in the growth, or even eradication, of bacteria. Responses of tissue invaded by bacteria (neighboring the infiltrated areas after 2-3 days post-inoculation) included: (i) inhibition of photosynthesis in terms of photosystem II efficiency; (ii) activation of energy dissipation as non-photochemical quenching in photosystem II, which is related to the activation of plant defense mechanisms; and (iii) accumulation of secondary metabolites in cell walls of the epidermis (lignins) and the apoplast of the mesophyll (phytoalexins). Infiltrated tissues showed an increase in the content of the main hormones regulating stress responses, including abscisic acid, jasmonic acid, and salicylic acid. We propose a mechanism involving the three hormones by which N. benthamiana could activate an efficient defense response against D. dadantii.

Exploring new roles for the rpoS gene in the survival and virulence of the fire blight pathogen Erwinia amylovora.

Erwinia amylovora causes fire blight in economically important plants of the family Rosaceae. This bacterial pathogen spends part of its life cycle coping with starvation and other fluctuating environmental conditions. In many Gram-negative bacteria, starvation and other stress responses are regulated by the sigma factor RpoS. We obtained an E. amylovora rpoS mutant to explore the role of this gene in starvation responses and its potential implication in other processes not yet studied in this pathogen. Results showed that E. amylovora needs rpoS to develop normal starvation survival and viable but nonculturable (VBNC) responses. Furthermore, this gene contributed to stationary phase cross-protection against oxidative, osmotic, and acid stresses and was essential for cross-protection against heat shock, but nonessential against acid shock. RpoS also mediated regulation of motility, exopolysaccharide synthesis, and virulence in immature loquats, but not in pear plantlets, and contributed to E. amylovora survival in nonhost tissues during incompatible interactions. Our results reveal some unique roles for the rpoS gene in E. amylovora and provide new knowledge on the regulation of different processes related to its ecology, including survival in different environments and virulence in immature fruits.

Jasmonate-dependent modifications of the pectin matrix during potato development function as a defense mechanism targeted by Dickeya dadantii virulence factors.

The plant cell wall constitutes an essential protection barrier against pathogen attack. In addition, cell-wall disruption leads to accumulation of jasmonates (JAs), which are key signaling molecules for activation of plant inducible defense responses. However, whether JAs in return modulate the cell-wall composition to reinforce this defensive barrier remains unknown. The enzyme 13-allene oxide synthase (13-AOS) catalyzes the first committed step towards biosynthesis of JAs. In potato (Solanum tuberosum), there are two putative St13-AOS genes, which we show here to be differentially induced upon wounding. We also determine that both genes complement an Arabidopsis aos null mutant, indicating that they encode functional 13-AOS enzymes. Indeed, transgenic potato plants lacking both St13-AOS genes (CoAOS1/2 lines) exhibited a significant reduction of JAs, a concomitant decrease in wound-responsive gene activation, and an increased severity of soft rot disease symptoms caused by Dickeya dadantii. Intriguingly, a hypovirulent D. dadantii pel strain lacking the five major pectate lyases, which causes limited tissue maceration on wild-type plants, regained infectivity in CoAOS1/2 plants. In line with this, we found differences in pectin methyl esterase activity and cell-wall pectin composition between wild-type and CoAOS1/2 plants. Importantly, wild-type plants had pectins with a lower degree of methyl esterification, which are the substrates of the pectate lyases mutated in the pel strain. These results suggest that, during development of potato plants, JAs mediate modification of the pectin matrix to form a defensive barrier that is counteracted by pectinolytic virulence factors from D. dadantii.

Translocation and functional analysis of Pseudomonas savastanoi pv. savastanoi NCPPB 3335 type III secretion system effectors reveals two novel effector families of the Pseudomonas syringae complex.

Pseudomonas savastanoi pv. savastanoi NCPPB 3335 causes olive knot disease and is a model pathogen for exploring bacterial infection of woody hosts. The type III secretion system (T3SS) effector repertoire of this strain includes 31 effector candidates plus two novel candidates identified in this study which have not been reported to translocate into plant cells. In this work, we demonstrate the delivery of seven NCPPB 3335 effectors into Nicotiana tabacum leaves, including three proteins from two novel families of the P. syringae complex effector super-repertoire (HopBK and HopBL), one of which comprises two proteins (HopBL1 and HopBL2) that harbor a SUMO protease domain. When delivered by P. fluorescens heterologously expressing a P. syringae T3SS, all seven effectors were found to suppress the production of defense-associated reactive oxygen species. Moreover, six of these effectors, including the truncated versions of HopAA1 and HopAZ1 encoded by NCPPB 3335, suppressed callose deposition. The expression of HopAZ1 and HopBL1 by functionally effectorless P. syringae pv. tomato DC3000D28E inhibited the hypersensitive response in tobacco and, additionally, expression of HopBL2 by this strain significantly increased its competitiveness in N. benthamiana. DNA sequences encoding HopBL1 and HopBL2 were uniquely detected in a collection of 31 P. savastanoi pv. savastanoi strains and other P. syringae strains isolated from woody hosts, suggesting a relevant role of these two effectors in bacterial interactions with olive and other woody plants.

Light regulates motility, attachment and virulence in the plant pathogen Pseudomonas syringae pv tomato DC3000.

Pseudomonas syringae pv tomato DC3000 (Pto) is the causal agent of the bacterial speck of tomato, which leads to significant economic losses in this crop. Pto inhabits the tomato phyllosphere, where the pathogen is highly exposed to light, among other environmental factors. Light represents a stressful condition and acts as a source of information associated with different plant defence levels. Here, we analysed the presence of both blue and red light photoreceptors in a group of Pseudomonas. In addition, we studied the effect of white, blue and red light on Pto features related to epiphytic fitness. While white and blue light inhibit motility, bacterial attachment to plant leaves is promoted. Moreover, these phenotypes are altered in a blue-light receptor mutant. These light-controlled changes during the epiphytic stage cause a reduction in virulence, highlighting the relevance of motility during the entry process to the plant apoplast. This study demonstrated the key role of light perception in the Pto phenotype switching and its effect on virulence.

The role of secretion systems and small molecules in soft-rot Enterobacteriaceae pathogenicity.

Soft-rot Enterobacteriaceae (SRE), which belong to the genera Pectobacterium and Dickeya, consist mainly of broad host-range pathogens that cause wilt, rot, and blackleg diseases on a wide range of plants. They are found in plants, insects, soil, and water in agricultural regions worldwide. SRE encode all six known protein secretion systems present in gram-negative bacteria, and these systems are involved in attacking host plants and competing bacteria. They also produce and detect multiple types of small molecules to coordinate pathogenesis, modify the plant environment, attack competing microbes, and perhaps to attract insect vectors. This review integrates new information about the role protein secretion and detection and production of ions and small molecules play in soft-rot pathogenicity.

A bacterial cysteine protease effector protein interferes with photosynthesis to suppress plant innate immune responses.

The bacterial pathogen Pseudomonas syringae pv tomato DC3000 suppresses plant innate immunity with effector proteins injected by a type III secretion system (T3SS). The cysteine protease effector HopN1, which reduces the ability of DC3000 to elicit programmed cell death in non-host tobacco, was found to also suppress the production of defence-associated reactive oxygen species (ROS) and callose when delivered by Pseudomonas fluorescens heterologously expressing a P. syringae T3SS. Purified His(6) -tagged HopN1 was used to identify tomato PsbQ, a member of the oxygen evolving complex of photosystem II (PSII), as an interacting protein. HopN1 localized to chloroplasts and both degraded PsbQ and inhibited PSII activity in chloroplast preparations, whereas a HopN1(D299A) non-catalytic mutant lost these abilities. Gene silencing of NtPsbQ in tobacco compromised ROS production and programmed cell death by DC3000. Our data reveal PsbQ as a contributor to plant immunity responses and a target for pathogen suppression.