Draft Genome Sequence of Erwinia oleae, a Bacterium Associated with Olive Knots Caused by Pseudomonas savastanoi pv. savastanoi.
ABSTRACT: Erwinia oleae is a nonpathogenic bacterial species isolated from olive knots caused by Pseudomonas savastanoi pv. savastanoi. Since the presence of E. oleae in the knots increases disease severity, interspecies interactions with the pathogen are hypothesized. Here, we report the first draft genome sequence of the E. oleae type strain.
Project description:Erwinia toletana was first reported in 2004 as a bacterial species isolated from olive knots caused by the plant bacterium Pseudomonas savastanoi pv. savastanoi. Recent studies have shown that the presence of this bacterium in the olive knot environment increases the virulence of the disease, indicating possible interspecies interactions with P. savastanoi pv. savastanoi. Here, we report the first draft genome sequence of an E. toletana strain.
Project description:The olive knot disease (Olea europea L.) is caused by the bacterium Pseudomonas savastanoi pv. savastanoi. P. savastanoi pv. savastanoi in the olive knot undergoes interspecies interactions with the harmless endophyte Erwinia toletana; P. savastanoi pv. savastanoi and E. toletana colocalize and form a stable community, resulting in a more aggressive disease. P. savastanoi pv. savastanoi and Etoletana produce the same type of the N-acylhomoserine lactone (AHL) quorum sensing (QS) signal, and they share AHLs in planta In this work, we have further studied the AHL QS systems of P. savastanoi pv. savastanoi and Etoletana in order to determine possible molecular mechanism(s) involved in this bacterial interspecies interaction/cooperation. The AHL QS regulons of P. savastanoi pv. savastanoi and Etoletana were determined, allowing the identification of several QS-regulated genes. Surprisingly, the P. savastanoi pv. savastanoi QS regulon consisted of only a few loci whereas in Etoletana many putative metabolic genes were regulated by QS, among which are several involved in carbohydrate metabolism. One of these loci was the aldolase-encoding gene garL, which was found to be essential for both colocalization of P. savastanoi pv. savastanoi and Etoletana cells inside olive knots as well as knot development. This study further highlighted that pathogens can cooperate with commensal members of the plant microbiome.IMPORTANCE This is a report on studies of the quorum sensing (QS) systems of the olive knot pathogen Pseudomonas savastanoi pv. savastanoi and olive knot cooperator Erwinia toletana These two bacterial species form a stable community in the olive knot, share QS signals, and cooperate, resulting in a more aggressive disease. In this work we further studied the QS systems by determining their regulons as well as by studying QS-regulated genes which might play a role in this cooperation. This represents a unique in vivo interspecies bacterial virulence model and highlights the importance of bacterial interspecies interaction in disease.
Project description:Pseudomonas savastanoi pv. savastanoi strain NCPPB 3335 is a model bacterial pathogen for studying the molecular basis of disease production in woody hosts. We report the sequencing of the hrpS-to-hrpZ region of NCPPB 3335, which has allowed us to determine the phylogenetic position of this pathogen with respect to previously sequenced Pseudomonas syringae hrp clusters. In addition, we constructed a mutant of NCPPB 3335, termed T3, which carries a deletion from the 3' end of the hrpS gene to the 5' end of the hrpZ operon. Despite its inability to multiply in olive tissues and to induce tumor formation in woody olive plants, P. savastanoi pv. savastanoi T3 can induce knot formation on young micropropagated olive plants. However, the necrosis and formation of internal open cavities previously reported in knots induced by the wild-type strain were not observed in those induced by P. savastanoi pv. savastanoi T3. Tagging of P. savastanoi pv. savastanoi T3 with green fluorescent protein (GFP) allowed real-time monitoring of its behavior on olive plants. In olive plant tissues, the wild-type strain formed aggregates that colonized the intercellular spaces and internal cavities of the hypertrophic knots, while the mutant T3 strain showed a disorganized distribution within the parenchyma of the knot. Ultrastructural analysis of knot sections revealed the release of extensive outer membrane vesicles from the bacterial cell surface of the P. savastanoi pv. savastanoi T3 mutant, while the wild-type strain exhibited very few vesicles. This phenomenon has not been described before for any other bacterial phytopathogen during host infection.
Project description:Olive knot (OK) is a widespread bacterial disease, caused by Pseudomonas savastanoi pv. savastanoi (Pss), which currently has not effective control methods. The use of naturally occurring microbial antagonists, such as bacteria, as biocontrol agents could be a strategy to manage this disease. The objective of this work was to select bacteria from olive tree phyllosphere able to antagonize Pss using in vitro and in planta experiments. The elucidation of their modes of action and the potential relationship between antagonism and bacteria origin has been investigated, as well. To this end, 60 bacterial isolates obtained from the surface and inner tissues of different organs (leaves, twigs, and knots), from two olive cultivars of varying susceptibilities to OK, were screened for their in vitro antagonistic effect against Pss. A total of 27 bacterial strains were able to significantly inhibit Pss growth, being this effect linked to bacteria origin. Strains from OK-susceptible cultivar and colonizing the surface of plant tissues showed the strongest antagonistic potential. The antagonistic activity was potentially due to the production of volatile compounds, siderophores and lytic enzymes. Bacillus amyloliquefaciens P41 was the most effective antagonistic strain and their capacity to control OK disease was subsequently assayed using in planta experiments. This strain significantly reduces OK disease severity (43.7%), knots weight (55.4%) and population size of Pss (26.8%), while increasing the shoot dry weight (55.0%) and root water content (39.6%) of Pss-infected olive plantlets. Bacterial isolates characterized in this study, in particular B. amyloliquefaciens P41, may be considered as promising biocontrol candidates for controlling OK disease.
Project description:The olive fruit fly, Bactrocera oleae, is the most destructive pest of olive orchards worldwide. The monophagous larva has the unique capability of feeding on olive mesocarp, coping with high levels of phenolic compounds and utilizing non-hydrolyzed proteins present, particularly in the unripe, green olives. On the molecular level, the interaction between B. oleae and olives has not been investigated as yet. Nevertheless, it has been associated with the gut obligate symbiotic bacterium Candidatus Erwinia dacicola. Here, we used a B.oleae microarray to analyze the gene expression of larvae during their development in artificial diet, unripe (green) and ripe (black) olives. The expression profiles of Ca. E. dacicola were analyzed in parallel, using the Illumina platform. Several genes were found overexpressed in the olive fly larvae when feeding in green olives. Among these, a number of genes encoding detoxification and digestive enzymes, indicating a potential association with the ability of B. oleae to cope with green olives. In addition, a number of biological processes seem to be activated in Ca. E. dacicola during the development of larvae in olives, with the most notable being the activation of amino-acid metabolism.
Project description:In a number of compatible plant-bacterium interactions, a rise in apoplastic Ca2+ levels is observed, suggesting that Ca2+ represents an important environmental clue, as reported for bacteria infecting mammalians. We demonstrate that Ca2+ entry in Pseudomonas savastanoi pv. savastanoi (Psav) strain DAPP-PG 722 is mediated by a Na+ /Ca2+ exchanger critical for virulence. Using the fluorescent Ca2+ probe Fura 2-AM, we demonstrate that Ca2+ enters Psav cells foremost when they experience low levels of energy, a situation mimicking the apoplastic fluid. In fact, Ca2+ entry was suppressed in the presence of high concentrations of glucose, fructose, sucrose or adenosine triphosphate (ATP). Since Ca2+ entry was inhibited by nifedipine and LiCl, we conclude that the channel for Ca2+ entry is a Na+ /Ca2+ exchanger. In silico analysis of the Psav DAPP-PG 722 genome revealed the presence of a single gene coding for a Na+ /Ca2+ exchanger (cneA), which is a widely conserved and ancestral gene within the P. syringae complex based on gene phylogeny. Mutation of cneA compromised not only Ca2+ entry, but also compromised the Hypersensitive response (HR) in tobacco leaves and blocked the ability to induce knots in olive stems. The expression of both pathogenicity (hrpL, hrpA and iaaM) and virulence (ptz) genes was reduced in this Psav-cneA mutant. Complementation of the Psav-cneA mutation restored both Ca2+ entry and pathogenicity in olive plants, but failed to restore the HR in tobacco leaves. In conclusion, Ca2+ entry acts as a 'host signal' that allows and promotes Psav pathogenicity on olive plants.
Project description:Olive knot disease, caused by the bacterium Pseudomonas savastanoi pv. savastanoi, seriously affects olive trees in the Mediterranean basin. Here, we report the draft genome sequence of P. savastanoi pv. savastanoi DAPP-PG 722, a strain isolated in Italy from an olive plant affected by knot disease.
Project description:Pseudomonas savastanoi pv. savastanoi strains harbor native plasmids belonging to the pPT23A plasmid family (PFPs) which are detected in all pathovars of the related species Pseudomonas syringae examined and contribute to the ecological and pathogenic fitness of their host. However, there is a general lack of information about the gene content of P. savastanoi pv. savastanoi plasmids and their role in the interaction of this pathogen with olive plants. We designed a DNA macroarray containing 135 plasmid-borne P. syringae genes to conduct a global genetic analysis of 32 plasmids obtained from 10 P. savastanoi pv. savastanoi strains. Hybridization results revealed that the number of PFPs per strain varied from one to four. Additionally, most strains contained at least one plasmid (designated non-PFP) that did not hybridize to the repA gene of pPT23A. Only three PFPs contained genes involved in the biosynthesis of the virulence factor indole-3-acetic acid (iaaM, iaaH, and iaaL). In contrast, ptz, a gene involved in the biosynthesis of cytokinins, was found in five PFPs and one non-PFP. Genes encoding a type IV secretion system (T4SS), type IVA, were found in both PFPs and non-PFPs; however, type IVB genes were found only on PFPs. Nine plasmids encoded both T4SSs, whereas seven other plasmids carried none of these genes. Most PFPs and non-PFPs hybridized to at least one putative type III secretion system effector gene and to a variety of additional genes encoding known P. syringae virulence factors and one or more insertion sequence transposase genes. These results indicate that non-PFPs may contribute to the virulence and fitness of the P. savastanoi pv. savastanoi host. The overall gene content of P. savastanoi pv. savastanoi plasmids, with their repeated information, mosaic arrangement, and insertion sequences, suggests a possible role in adaptation to a changing environment.
Project description:A bacterial isolate (SCU-B244T) was obtained in China from crickets (Teleogryllus occipitalis) living in cropland deserted for approximately 10 years. The isolated bacteria were Gram-negative, facultatively anaerobic, oxidase-negative rods. A preliminary analysis of the 16S rRNA gene sequence indicated that the strain belongs to either the genus Erwinia or Pantoea. Analysis of multilocus sequence typing based on concatenated partial atpD, gyrB and infB gene sequences and physiological and biochemical characteristics indicated that the strain belonged to the genus Erwinia, as member of a new species as it was distinct from other known Erwinia species. Further analysis of the 16S rRNA gene showed SCU-B244T to have 94.71% identity to the closest species of that genus, Erwinia oleae (DSM 23398T), which is below the threshold of 97% used to discriminate bacterial species. DNA-DNA hybridization results (5.78±2.52%) between SCU-B244T and Erwinia oleae (DSM 23398T) confirmed that SCU-B244T and Erwinia oleae (DSM 23398T) represent different species combined with average nucleotide identity values which range from 72.42% to 74.41. The DNA G+C content of SCU-B244T was 55.32 mol%, which also differs from that of Erwinia oleae (54.7 to 54.9 mol%). The polyphasic taxonomic approach used here confirmed that the strain belongs to the Erwinia group and represents a novel species. The name Erwinia teleogrylli sp. nov. is proposed for this novel taxon, for which the type strain is SCU-B244T (= CGMCC 1.12772T = DSM 28222T = KCTC 42022T).
Project description:Pseudomonas savastanoi pv. savastanoi NCPPB 3335 is a model for the study of the molecular basis of disease production and tumor formation in woody hosts, and its draft genome sequence has been recently obtained. Here we closed the sequence of the plasmid complement of this strain, composed of three circular molecules of 78,357 nt (pPsv48A), 45,220 nt (pPsv48B), and 42,103 nt (pPsv48C), all belonging to the pPT23A-like family of plasmids widely distributed in the P. syringae complex. A total of 152 coding sequences were predicted in the plasmid complement, of which 38 are hypothetical proteins and seven correspond to putative virulence genes. Plasmid pPsv48A contains an incomplete Type IVB secretion system, the type III secretion system (T3SS) effector gene hopAF1, gene ptz, involved in cytokinin biosynthesis, and three copies of a gene highly conserved in plant-associated proteobacteria, which is preceded by a hrp box motif. A complete Type IVA secretion system, a well conserved origin of transfer (oriT), and a homolog of the T3SS effector gene hopAO1 are present in pPsv48B, while pPsv48C contains a gene with significant homology to isopentenyl-diphosphate delta-isomerase, type 1. Several potential mobile elements were found on the three plasmids, including three types of MITE, a derivative of IS801, and a new transposon effector, ISPsy30. Although the replication regions of these three plasmids are phylogenetically closely related, their structure is diverse, suggesting that the plasmid architecture results from an active exchange of sequences. Artificial inoculations of olive plants with mutants cured of plasmids pPsv48A and pPsv48B showed that pPsv48A is necessary for full virulence and for the development of mature xylem vessels within the knots; we were unable to obtain mutants cured of pPsv48C, which contains five putative toxin-antitoxin genes.