Erwinia amylovora: the molecular basis of fireblight disease

Taxonomy: Bacteria; Proteobacteria; γ subdivision; order Enterobacteriales; family Enterobacteriaceae; genus Erwinia .
Microbiological properties: Gram-negative, motile rods.
Related species: E. carotovora (soft-rot diseases) , E. chrysanthemi (soft-rot diseases) , E. (Pantoea) stewartii (Stewart's wilt of corn) , E. (Pantoea) herbicola (epiphyte).
Host range: Affects rosaceous plants, primarily members of the Pomoideae . Economically important hosts are apple and pear. The commercial implications of fireblight outbreaks are aggravated by the limited effectiveness of current control measures.
Disease symptoms: E. amylovora infection is characterized by water soaking of infected tissue, followed by wilting and tissue necrosis. Necrosis gives tissue a scorched, blackened appearance, giving rise to the name fireblight. Symptoms are often localized to blossom bracts or young shoots but, in highly susceptible hosts, can spread systemically resulting in death of the entire tree. Infections can vary in severity depending on climatic conditions and host susceptibility.
Useful web site: http://www.agric.gov.ab.ca     

PA 20, a semi-selective medium for isolation and enumeration of Pantoea ananatis

A semi-selective medium, PA 20, was developed for the isolation and enumeration of Pantoea ananatis from plant material, specifically from onion seed. The medium has a pH of 8.0 and contains NH4H2PO4, K2HPO4, magnesium sulphate, NaCl, d (+) arabitol, crystal violet, bromothymol blue and thallium nitrate. All P. ananatis strains from a variety of hosts produced characteristic yellow colonies in 6-7 days at 25 degrees C. Plating efficiencies on PA 20 in comparison to nutrient agar ranged from 92 to 112%. Recovery from naturally infested and artificially contaminated onion seed was high, with an almost total reduction of saprophytes.     

Production of quorum-sensing-related signal molecules by epiphytic bacteria inhabiting wheat heads

The production of quorum-sensing-related signal molecules (QSRMs) among culturable bacteria comprising the community on wheat heads was investigated. The taxonomic position of 186 bacterial isolates obtained from ten heads was inferred based on 16S rRNA gene sequences, and their QSRM production was determined using two bioreporter strains of N-acylhomoserine lactones. Approximately 33% of isolates produced QSRMs, though the proportion of QSRM-producing isolates on a wheat head was significantly negatively correlated with population size. Most of the producing isolates were Pantoea species, most commonly Pantoea ananatis. Furthermore, the proportion of Pantoea ananatis that produced QSRMs was significantly negatively correlated with the number of bacterial genera (community richness) on each head. Finally, community richness was positively correlated with population size. Qualitative analysis using thin-layer-chromatography revealed that the QSRMs of Pantoea isolates were composed of at least two compounds. This is the first report indicating that Pantoea ananatis isolates inhabiting wheat heads are capable of producing QSRMs. QSRM production by Pantoea spp. may contribute to the predominance of this genus on wheat heads, particularly at relatively low population densities and community diversity.     

Complete genome sequence of clinical isolate Pantoea ananatis LMG 5342

The enterobacterium Pantoea ananatis is an ecologically versatile species. It has been found in the environment, as plant epiphyte and endophyte, as an emerging phytopathogen, and as a presumptive, opportunistic human pathogen. Here, we report the complete genome sequence of P. ananatis LMG 5342, isolated from a human wound.     

Isolation of Enterobacter cowanii from Eucalyptus showing symptoms of bacterial blight and dieback in Uruguay

Aims:  This study was performed to identify bacterial strains isolated simultaneously with Pantoea species from Eucalyptus trees showing symptoms of bacterial blight and dieback in Uruguay.
Methods and Results:  Several molecular techniques including 16S rRNA and rpoB gene sequencing and DNA–DNA hybridization were used to characterize the Gram-negative, facultatively anaerobic, slime-producing bacterial strains isolated along with Pantoea species from Eucalyptus . Hypersensitivity reactions (HR) and pathogenicity tests were performed on tobacco and Eucalyptus seedlings, respectively. The isolates clustered closely with the type strain of Enterobacter cowanii in both phylogenetic trees constructed. The DNA–DNA similarity between the isolates and the type strain of Ent. cowanii ranged from 88% to 92%. A positive HR was observed on the tobacco seedlings, but no disease symptoms were visible on the inoculated Eucalyptus seedlings.
Conclusions:  Enterobacter cowanii was isolated from trees with symptoms of bacterial blight although strains of this bacterial species do not appear to be the causal agent of the disease.
Significance and Impact of the Study:  This study provides the first report of Ent. cowanii isolated from Eucalyptus . Its presence in Eucalyptus tissue suggests that it is an endophyte in trees showing symptoms of blight.     

Identification of Pantoea ananatis gene encoding membrane pyrroloquinoline quinone (PQQ)-dependent glucose dehydrogenase and pqqABCDEF operon essential for PQQ biosynthesis

Pantoea ananatis accumulates gluconate during aerobic growth in the presence of glucose. Computer analysis of the P. ananatis SC17(0) sequenced genome revealed an ORF encoding a homologue (named gcd) of the mGDH (EC 1.1.99.17) apoenzyme from Escherichia coli and a putative pyrroloquinoline quinone (PQQ) biosynthetic operon homologous to pqqABCDEF from Klebsiella pneumoniae. Construction of Δgcd and Δpqq mutants of P. ananatis confirmed the proposed functions of these genetic elements. The P. ananatis pqqABCDEF was cloned in vivo and integrated into the chromosomes of P. ananatis and E. coli according to the Dual In/Out strategy. Introduction of a second copy of pqqABCDEF to P. ananatis SC17(0) doubled the accumulation of PQQ. Integration of the operon into E. coli MG1655ΔptsGΔmanXY restored the growth of bacteria on glucose. The obtained data show the essential role of pqqABCDEF in PQQ biosynthesis in P. ananatis and E. coli. We propose that the cloned operon could be useful for an efficient phosphoenolpyruvate-independent glucose consumption pathway due to glucose oxidation and construction of E. coli strains with the advantage of phosphoenolpyruvate-derived metabolite production.     

Complete genome sequence of the rice pathogen Pantoea ananatis strain PA13

Pantoea ananatis is the causative agent of sheath and grain rot in rice. Here, we present the complete genome sequence of P. ananatis strain PA13, originally isolated from a diseased rice grain.     

Exopolysaccharide production is influenced by sugars, N-acylhomoserine lactone, and transcriptional regulators RcsA and RcsB, but does not affect pathogenicity in the plant pathogen Pantoea ananatis

Pantoea ananatis SK-1 produced EPS by AHL-mediated quorum sensing on an LB agar plate containing glucose, fructose, and sucrose. rcsA and rcsB mutants did not produce EPS with or without AHLs and with or without sugars, but they induced necrotic symptoms in onion leaves. These results indicate that EPS production does not relate to the pathogenicity of SK-1.     

Draft genome sequence of Pantoea ananatis B1-9, a nonpathogenic plant growth-promoting bacterium

Pantoea ananatis B1-9 is an endophytic Gram-negative rhizobacterium that was isolated for its ability to promote plant growth and improve crop yield in the field. Here we report the draft genome sequence of P. ananatis B1-9. Comparison of this sequence to the sequenced genome of a plant-pathogenic P. ananatis strain, LMG20103, indicated that the pathogenesis-related genes were absent, but a subset of gene functions that may be related to its plant growth promotion were present.     

The complete genome sequence of Pantoea ananatis AJ13355, an organism with great biotechnological potential

Pantoea ananatis AJ13355 is a newly identified member of the Enterobacteriaceae family with promising biotechnological applications. This bacterium is able to grow at an acidic pH and is resistant to saturating concentrations of L-glutamic acid, making this organism a suitable host for the production of L-glutamate. In the current study, the complete genomic sequence of P. ananatis AJ13355 was determined. The genome was found to consist of a single circular chromosome consisting of 4,555,536 bp [DDBJ: AP012032] and a circular plasmid, pEA320, of 321,744 bp [DDBJ: AP012033]. After automated annotation, 4,071 protein-coding sequences were identified in the P. ananatis AJ13355 genome. For 4,025 of these genes, functions were assigned based on homologies to known proteins. A high level of nucleotide sequence identity (99%) was revealed between the genome of P. ananatis AJ13355 and the previously published genome of P. ananatis LMG 20103. Short colinear regions, which are identical to DNA sequences in the Escherichia coli MG1655 chromosome, were found to be widely dispersed along the P. ananatis AJ13355 genome. Conjugal gene transfer from E. coli to P. ananatis, mediated by homologous recombination between short identical sequences, was also experimentally demonstrated. The determination of the genome sequence has paved the way for the directed metabolic engineering of P. ananatis to produce biotechnologically relevant compounds.     

Isolation and characterization of plant growth-promoting strain Pantoea NII-186. From Western Ghat Forest soil, India

Aims:  To isolate plant growth-promoting bacterium from Western Ghat forests in India.
Methods and Results:  A Gram-negative, rod shaped, cream white coloured strain Pantoea NII-186 isolated from Western Ghat soil sample. The taxonomic position of the bacterium was confirmed by sequencing of 16S rRNA and phylogenetic analysis. A strain grew at a wide range of temperature ranging from 5–40°C, but optimum growth was observed at 28–30°C. It showed multiple plant growth-promoting attributes such as phosphate solubilization activity, indole acetic acid (IAA) production, siderophore production and HCN production. It was able to solubilize (28 μg of Ca₃PO₄ ml⁻¹ day⁻¹), and produce IAA (59 μg) at 28°C. The solubilization of insoluble phosphate was associates with a drop in the pH of the culture medium. Pantoea sp. NII-186 tolerate to different environmental stresses like 5–40°C, 0–7% salt concentration and 4–12 pH range.
Conclusions:  The 16S rRNA gene sequence confirmed that the isolate NII-186 was belongs to Pantoea genus and showed considerable differences in physiological properties with previously reported species of this genus. Isolate NII-186 possessed multiple attributes of plant growth-promoting activity.
Significance and Impact of the Study:  Hence in the context it is proposed that Pantoea sp. NII-186, could be deployed as an inoculant to attain the desired plant growth-promoting activity in agricultural environment.     

Carotenoids' influence on radiotolerance of Pantoea agglomerans, a plant pathogen

Aim:  The aim of this study was to evaluate the effect of γ radiation on the carotenoid content of two strains of the Enterobacteriaceae : Pantoea agglomerans .
Methods and Results:  Pantoea agglomerans strains ATCC 49174 and RL1 were used for this study. Successive radiation treatments were performed to study the radiotolerance. Total carotenoids were obtained by multiple extraction using chloroform/methanol (2 : 1), quantified by measuring the optical density at 453 nm and their antioxidant activity measured by a colorimetric method. The D ₁₀ studies were conducted using a UC-15A irradiator loaded with ⁶⁰Co. Bacterial counts from various dilutions were carried out after irradiation. Strain ATCC 49174 irradiated at 1 kGy produced 4·3 times more carotenoids than the control, whereas carotenoid synthesis increased by 2·9-fold in the strain RL1. However, there was no significant difference in the D ₁₀ values.
Conclusion:  Carotenoid increased production is influenced by γ radiation but does not modify the tolerance to radiations.
Significance and Impact of the Study:  To our knowledge, this is the first study to demonstrate the effects of γ radiation on carotenoid production levels.     

Exotic wildland weeds serve as reservoirs for a newly introduced cole crop pest,Bagrada hilaris (Hemiptera: Pentatomidae)

The role of wildland weeds that serve as alternate hosts for insect pests has not been directly examined for the potential to sustain pest populations or facilitate pest outbreaks. The development of weed biological control programmes is also complicated by weed species that are closely related to economically important or native plants, especially rare or special status species. In recent surveys in southern California, USA, we found a newly introduced insect pest of cole crops, Bagrada hilaris Burmeister (Bagrada bug), feeding on Brassicaceae weeds in riparian areas adjacent to agricultural fields where cole crops are routinely grown. Insect populations grew to levels well over action thresholds and caused severe damage to populations of the invasive weed, Lepidium latifolium (perennial pepperweed). The numerical response of B. hilaris on L. latifolium and other Brassicaceae weeds in natural areas may pose a significant challenge to effectively managing pest populations in crops. However, the accidental introduction of this insect provides the opportunity to examine plant–insect interactions with important implications for development of biologically based control methods for weeds.     

Criteria for release of genetically-improved phytoseiids: an examination of the risks associated with release of biological control agents

Biological control of agricultural pests by phytoseiid predators has been achieved through classical introductions, conservation of indigenous and established foreign species, and augmentation of both introduced and indigenous species. Laboratory selection of phytoseiids has produced several strains that have been mass reared and released for pest management programs in glasshouses and agricultural cropping systems. Concerns over risks of classical biological control have developed recently. The development of recombinant DNA (rDNA) techniques for the genetic manipulation of crops and microorganisms also has inaugurated a debate on the safety of releasing transgenic organisms into the environment. This debate will extend to the release of phytoseiids that have been manipulated with rDNA techniques. Risks associated with releasing phytoseiids for augmentation or classical biological control programs are minimal and the benefits are great. Research initiated to answer questions about the risks of releasing transgenic phytoseiids into the environment provides opportunities to expand our understanding of the ecological impact of phytoseiids in agricultural and natural environments and could lead to improved pest management tactics.     

A framework to gauge the epidemic potential of plant pathogens in environmental reservoirs: the example of kiwifruit canker

New economically important diseases on crops and forest trees emerge recurrently. An understanding of where new pathogenic lines come from and how they evolve is fundamental for the deployment of accurate surveillance methods. We used kiwifruit bacterial canker as a model to assess the importance of potential reservoirs of new pathogenic lineages. The current kiwifruit canker epidemic is at least the fourth outbreak of the disease on kiwifruit caused by Pseudomonas syringae in the mere 50 years in which this crop has been cultivated worldwide, with each outbreak being caused by different genetic lines of the bacterium. Here, we ask whether strains in natural (non‐agricultural) environments could cause future epidemics of canker on kiwifruit. To answer this question, we evaluated the pathogenicity, endophytic colonization capacity and competitiveness on kiwifruit of P. syringae strains genetically similar to epidemic strains and originally isolated from aquatic and subalpine habitats. All environmental strains possessing an operon involved in the degradation of aromatic compounds via the catechol pathway grew endophytically and caused symptoms in kiwifruit vascular tissue. Environmental and epidemic strains showed a wide host range, revealing their potential as future pathogens of a variety of hosts. Environmental strains co‐existed endophytically with CFBP 7286, an epidemic strain, and shared about 20 virulence genes, but were missing six virulence genes found in all epidemic strains. By identifying the specific gene content in genetic backgrounds similar to known epidemic strains, we developed criteria to assess the epidemic potential and to survey for such strains as a means of forecasting and managing disease emergence.     

Distribution Patterns of Wolbachia Endosymbionts in the Closely Related Flower Bugs of the Genus Orius: Implications for Coevolution and Horizontal Transfer

Vertical transmission is the primary route of the endosymbiont Wolbachia for its own spread among invertebrate hosts, but horizontal transmission between different hosts is believed to have occurred multiple times. However, it is not well known how Wolbachia commonly spread among closely related hosts. We focused on the closely related species of the minute pirate bugs belonging to the genus Orius, which are important biological control agents in agricultural crops because they are the most useful natural enemy of various tiny pests, such as thrips. Here, we examined five Orius species (Orius sauteri, Orius nagaii, Orius minutus, Orius strigicollis, and Orius tantillus) from eight geographic localities in Japan for Wolbachia infection. Two distinct strains, wOus1 and wOus2, were detected based on Wolbachia surface protein (wsp) gene sequencing. Furthermore, multilocus sequence typing revealed that each of the strains comprised two variants that differed in a single nucleotide. The overall distribution patterns of the two Wolbachia strains were found to differ among host species: prevalent double infection with wOus1 and wOus2 in O. strigicollis; fixation of single infection with wOus2 in O. nagaii; occurrence of single infection with wOus1 in O. sauteri; prevalence of single infection with wOus1 in O. minutus with an exception in a single population; and lack of Wolbachia infection in O. tantillus. Such differences in the distribution patterns of Wolbachia may reflect the evolutionary history of Wolbachia infection among Orius species and/or ecological and physiological differences among the Orius species that determine the invasiveness and maintenance of the two Wolbachia strains.     

Analysis of the Pantoea ananatis pan-genome reveals factors underlying its ability to colonize and interact with plant,insect and vertebrate hosts

Background

Pantoea ananatis is found in a wide range of natural environments, including water, soil, as part of the epi- and endophytic flora of various plant hosts, and in the insect gut. Some strains have proven effective as biological control agents and plant-growth promoters, while other strains have been implicated in diseases of a broad range of plant hosts and humans. By analysing the pan-genome of eight sequenced P. ananatis strains isolated from different sources we identified factors potentially underlying its ability to colonize and interact with hosts in both the plant and animal Kingdoms.

Results

The pan-genome of the eight compared P. ananatis strains consisted of a core genome comprised of 3,876 protein coding sequences (CDSs) and a sizeable accessory genome consisting of 1,690 CDSs. We estimate that ~106 unique CDSs would be added to the pan-genome with each additional P. ananatis genome sequenced in the future. The accessory fraction is derived mainly from integrated prophages and codes mostly for proteins of unknown function. Comparison of the translated CDSs on the P. ananatis pan-genome with the proteins encoded on all sequenced bacterial genomes currently available revealed that P. ananatis carries a number of CDSs with orthologs restricted to bacteria associated with distinct hosts, namely plant-, animal- and insect-associated bacteria. These CDSs encode proteins with putative roles in transport and metabolism of carbohydrate and amino acid substrates, adherence to host tissues, protection against plant and animal defense mechanisms and the biosynthesis of potential pathogenicity determinants including insecticidal peptides, phytotoxins and type VI secretion system effectors.

Conclusions

P. ananatis has an ‘open’ pan-genome typical of bacterial species that colonize several different environments. The pan-genome incorporates a large number of genes encoding proteins that may enable P. ananatis to colonize, persist in and potentially cause disease symptoms in a wide range of plant and animal hosts.

Electronic supplementary material

The online version of this article (doi: 10.1186/1471-2164-15-404) contains supplementary material, which is available to authorized users.     

Effect of γ-irradiation on membrane fatty acids and peptidoglycan's muropeptides of Pantoea agglomerans, a plant pathogen

Aims:  The aim of this study was to evaluate the effect of γ-irradiation on the fatty acids (FA) and muropeptides content of two strains of an Enterobacteriacea : Pantoea agglomerans .
Methods and Results:  Pantoea agglomerans strains ATCC 49174 and RL1 isolated from irradiated carrots were used for this study. Radiation treatments (1 and 3·5 kGy) were performed to study the radiotolerance. Total lipids were obtained by multiple extractions using methanol/chloroform (2 : 1) and were quantified by GC. Muropeptides were purified by successive enzymatic digestions and analysed using a reverse phase C₁₈ column in high performance liquid chromatography. A significant ( P  ≤ 0·05) modification of the bacterial wall was noticed for the membrane FA composition and the muropeptides.
Conclusion:  Effects of irradiation on the bacterial membrane are noticeable and could play an important role on the cellular response and ability to survive this harsh environment.
Significance and Impact of the Study:  To our knowledge, it is the first study to demonstrate the effects of ionizing irradiation on the modification of the FA and one of the few to confirm its effects on the muropeptides of the peptidoglycan.     

A FAFLP system for the improved identification of plant-pathogenic and plant-associated species of the genus Pantoea

The majority of Pantoea species are either plant-pathogenic or plant-associated and cause a wide variety of symptoms on a range of hosts. Identification of Pantoea species is difficult due to minor differences in phenotypic characteristics between them and related Enterobacteriaceae. Fluorescent amplified fragment length polymorphism (FAFLP) analysis was investigated for use as a rapid, molecular-based identification technique to the species level of the genus Pantoea. Following analysis of the band patterns generated by FAFLP, seven distinct clusters were observed, one for each validly published species of the genus. FAFLP has proven to be a rapid, reproducible identification technique for all species of the genus Pantoea.