N-Acyl Homoserine Lactones and Lux Solos Regulate Social Behaviour and Virulence of Pseudomonas syringae pv. actinidiae

The phyllosphere is a complex environment where microbes communicate through signalling molecules in a system, generally known as quorum sensing (QS). One of the most common QS systems in Gram-negative proteobacteria is based on the production of N-acyl homoserine lactones (AHLs) by a LuxI synthase and their perception by a LuxR sensor. Pseudomonas syringae pv. actinidiae (Psa), the aetiological agent of the bacterial canker of kiwifruit, colonises plant phyllosphere before penetrating via wounds and natural openings. Since Psa genome encodes three LuxR solos without a cognate LuxI, this bacterium may perceive diffusible signals, but it cannot produce AHLs, displaying a non-canonical QS system. The elucidation of the mechanisms underlying the perception of environmental cues in the phyllosphere by this pathogen and their influence on the onset of pathogenesis are of crucial importance for a long-lasting and sustainable management of the bacterial canker of kiwifruit. Here, we report the ability of Psa to sense its own population density and the presence of surrounding bacteria. Moreover, we show that Psa can perceive AHLs, indicating that AHL-producing neighbouring bacteria may regulate Psa virulence in the host. Our results suggest that the ecological environment is important in determining Psa fitness and pathogenic potential. This opens new perspectives in the use of more advanced biochemical and microbiological tools for the control of bacterial canker of kiwifruit

Comparative transcriptome analysis of the interaction between Actinidia chinensis var. chinensis and Pseudomonas syringae pv. Actinidiae in absence and presence of acibenzolar-S-methyl

Background: Since 2007, bacterial canker caused by Pseudomonas syringae pv. actinidiae (Psa) has become a pandemic disease leading to important economic losses in every country where kiwifruit is widely cultivated. Options for controlling this disease are very limited and rely primarily on the use of bactericidal compounds, such as copper, and resistance inducers. Among the latter, the most widely studied is acibenzolar-S-methyl. To elucidate the early molecular reaction of kiwifruit plants (Actinidia chinensis var. chinensis) to Psa infection and acibenzolar-S-methyl treatment, a RNA seq analysis was performed at different phases of the infection process, from the epiphytic phase to the endophytic invasion on acibenzolar-S-methyl treated and on non-treated plants. The infection process was monitored in vivo by confocal laser scanning microscopy. Results: De novo assembly of kiwifruit transcriptome revealed a total of 39,607 transcripts, of which 3360 were differentially expressed during the infection process, primarily 3 h post inoculation. The study revealed the coordinated changes of important gene functional categories such as signaling, hormonal balance and transcriptional regulation. Among the transcription factor families, AP2/ERF, MYB, Myc, bHLH, GATA, NAC, WRKY and GRAS were found differentially expressed in response to Psa infection and acibenzolar-S-methyl treatment. Finally, in plants treated with acibenzolar-S-methyl, a number of gene functions related to plant resistance, such as PR proteins, were modulated, suggesting the set-up of a more effective defense response against the pathogen. Weighted-gene coexpression network analysis confirmed these results. Conclusions: Our work provides an in-depth description of the plant molecular reactions to Psa, it highlights the metabolic pathway related to acibenzolar-S-methyl-induced resistance and it contributes to the development of effective control strategies in open field

Large expert-curated database for benchmarking document similarity detection in biomedical literature search

Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency-Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research.Peer reviewe

Using fundamental knowledge of induced resistance to develop control strategies for bacterial canker of kiwifruit caused by Pseudomonas syringae pv. actinidiae

Pseudomonas syringae pv. actinidiae (Psa) which causes bacterial canker of kiwifruit (Actinidia deliciosa and A. chinensis) was first isolated in Japan in 1984 (Takikawa et al., 1989), and soon after in Korea (Koh et al., 1994) and Italy (Scortichini, 1994). The economic impact on the global production of kiwifruit of those early occurrences was relatively limited (Vanneste et al., 2011). However, the latest outbreak of Psa which started in Italy in 2008 and rapidly spread throughout most of the kiwifruit growing regions of the world, represents a major threat to the global kiwifruit industry (Vanneste, 2012). The pathovar actinidiae is not a genetically homogeneous pathovar; strains can be grouped in four biovars based on their molecular, microbiological and pathogenic characteristics (Vanneste et al., 2013) which is consistent with MLST and whole genome sequence analysis (Ferrante and Scortichini, 2010; Mazzaglia et al., 2011; Chapman et al., 2012). The recent outbreak of bacterial canker on kiwifruit in Europe and New Zealand is caused by the same biovar of Psa (biovar 3) (Chapman et al., 2012; Vanneste et al., 2013). During the 2 years that the pathogen has been present in New Zealand, over 60% of the area planted in kiwifruit has been affected (Kiwifruit Vine Health, 2012). This rapid spread may be attributable to the virulence of biovar 3 and to the scarcity of products available for control of plant pathogenic bacteria in general, and Psa in particular. Many products used for control of plant pathogenic bacteria contain antibiotics (mostly streptomycin) or heavy metals (mostly copper). Both types of products do have limitations because of phytotoxicity or because they are not authorized in some countries (e.g., antibiotics in Europe). This has led to a large screening programme in New Zealand for the identification of potentially effective products to control Psa. The products tested included a number of commercially available potential elicitors of host resistance. One of the most effective elicitors in glasshouse trials on A. chinensis and A. deliciosa was acibenzolar-S-methyl [ASM], sold under the names of Bion\uae or Actigard\uae (Syngenta)

Integrated Use of Aureobasidium pullulans Strain CG163 and Acibenzolar-S-Methyl for Management of Bacterial Canker in Kiwifruit

An isolate of Aureobasidium pullulans (strain = CG163) and the plant defence elicitor acibenzolar-S-methyl (ASM) were investigated for their ability to control leaf spot in kiwifruit caused by Pseudomonas syringae pv. actinidiae biovar 3 (Psa). Clonal Actinidia chinensis var. deliciosa plantlets (‘Hayward’) were treated with ASM, CG163 or ASM + CG163 at seven and one day before inoculation with Psa. ASM (0.2 g/L) was applied either as a root or foliar treatments and CG163 was applied as a foliar spray containing 2 × 107 CFU/mL. Leaf spot incidence was significantly reduced by all treatments compared with the control. The combination of ASM + CG163 had greater efficacy (75%) than either ASM (55%) or CG163 (40%) alone. Moreover, treatment efficacy correlated positively with the expression of defence-related genes: pathogenesis-related protein 1 (PR1), β-1,3-glucosidase, Glucan endo 1,3-β-glucosidase (Gluc_PrimerH) and Class IV chitinase (ClassIV_Chit), with greater gene upregulation in plants treated with ASM + CG163 than by the individual treatments. Pathogen population studies indicated that CG163 had significant suppressive activity against epiphytic populations of Psa. Endophytic populations were reduced by ASM + CG163 but not by the individual treatments, and by 96–144 h after inoculation were significantly lower than the control. Together these data suggest that ASM + CG163 have complementary modes of action that contribute to greater control of leaf spotting than either treatment alone

Host-specific signal perception by PsaR2 LuxR solo induces Pseudomonas syringae pv. actinidiae virulence traits

Plant-associated bacteria, including pathogens, recognise host-derived signals to activate specific responses. The genome of Pseudomonas syringae pv. actinidiae (Psa), the aetiological agent of bacterial canker of kiwifruit, encodes for three putative LuxR-like receptors. Proteins of this family are usually involved in the quorum sensing system, through the perception of autoinducers (AHLs) produced by a cognate LuxI. However, Psa does not produce AHLs according to the lack of LuxI-encoding gene. It has been proposed that the so-called LuxR solos may be involved in the perception of environmental stimuli. We thus hypothesised that Psa LuxR-like receptors could be involved in host-derived signal sensing.Psa virulence traits, i.e., biofilm formation, motility and endophytic colonisation, were stimulated by growing the pathogen in host plant extracts, but not in non-host plant extracts or rich medium. Moreover, the phenotypic analyses of Psa mutant strains lacking the LuxR solo-encoding genes, demonstrated that PsaR2 plays a major role in host recognition and induction of virulence responses. The heterologous expression of PsaR2, followed by affinity chromatography and fraction activity assessment, confirmed the specific recognition of plant-derived components by this sensor. Overall, these data provide a deeper understanding of the regulation of Psa virulence through interkingdom communication, which represents a interesting target for the development of tolerant/resistant genotypes or innovative control strategies

Comparative genomics-informed design of two LAMP detection assays for detection of the kiwifruit pathogen Pseudomonas syringae pv. actinidiae and discrimination of isolates belonging to the pandemic biovar 3

The aim of this study was to develop a rapid, sensitive and reliable field-based assay for detection of the quarantine pathogen Pseudomonas syringae pv. actinidiae, the causal agent of the most destructive and economically important bacterial disease of kiwifruit. A comparative genomic approach was used on the publicly available P. syringae pv. actinidiae genomic data to select unique target regions for the development of two loop-mediated isothermal amplification (LAMP) assays able to detect P. syringae pv. actinidiae and to discriminate strains belonging to the highly virulent and globally spreading P. syringae pv. actinidiae biovar 3. Both LAMP assays showed specificity in accordance to their target and were able to detect reliably 125 CFU per reaction in less than 30 min. The developed assays were able to detect the presence of P. syringae pv. actinidiae in symptomatic as well as in asymptomatic naturally infected kiwifruit material, thus increasing the potential of the LAMP assays for phytosanitary purposes

Comparative transcriptome analysis of the interaction between Actinidia chinensis var. chinensis and Pseudomonas syringae pv. actinidiae in absence and presence of acibenzolar-S-methyl

Abstract Background Since 2007, bacterial canker caused by Pseudomonas syringae pv. actinidiae (Psa) has become a pandemic disease leading to important economic losses in every country where kiwifruit is widely cultivated. Options for controlling this disease are very limited and rely primarily on the use of bactericidal compounds, such as copper, and resistance inducers. Among the latter, the most widely studied is acibenzolar-S-methyl. To elucidate the early molecular reaction of kiwifruit plants (Actinidia chinensis var. chinensis) to Psa infection and acibenzolar-S-methyl treatment, a RNA seq analysis was performed at different phases of the infection process, from the epiphytic phase to the endophytic invasion on acibenzolar-S-methyl treated and on non-treated plants. The infection process was monitored in vivo by confocal laser scanning microscopy. Results De novo assembly of kiwifruit transcriptome revealed a total of 39,607 transcripts, of which 3360 were differentially expressed during the infection process, primarily 3 h post inoculation. The study revealed the coordinated changes of important gene functional categories such as signaling, hormonal balance and transcriptional regulation. Among the transcription factor families, AP2/ERF, MYB, Myc, bHLH, GATA, NAC, WRKY and GRAS were found differentially expressed in response to Psa infection and acibenzolar-S-methyl treatment. Finally, in plants treated with acibenzolar-S-methyl, a number of gene functions related to plant resistance, such as PR proteins, were modulated, suggesting the set-up of a more effective defense response against the pathogen. Weighted-gene coexpression network analysis confirmed these results. Conclusions Our work provides an in-depth description of the plant molecular reactions to Psa, it highlights the metabolic pathway related to acibenzolar-S-methyl-induced resistance and it contributes to the development of effective control strategies in open field

Postharvest Application of Acibenzolar-S-Methyl Activates Salicylic Acid Pathway Genes in Kiwifruit Vines

The plant defence inducer Actigard® (acibenzolar-S-methyl [ASM]) is applied before flowering and after fruit harvest to control bacterial canker in kiwifruit caused by Pseudomonas syringae pv. actinidiae. Pre-flowering application of ASM is known to upregulate defence gene expression; however, the effect of postharvest ASM on defence gene expression in the vine is unknown. In this study, the expression of eight ”defence marker” genes was measured in the leaves of Actinidia chinensis var. chinensis, ”Zesy002,” and Actinidia chinensis var. deliciosa, “Hayward,” vines after postharvest treatment with ASM and/or copper. There were two orchards per cultivar with harvest dates approximately three weeks apart for investigating potential changes in responsiveness to ASM during the harvest period. In all trials, postharvest ASM induced the expression of salicylic-acid-pathway defence genes PR1, PR2, PR5, BAD, DMR6, NIMIN2, and WRKY70. Gene upregulation was the greatest at 1 day and 7 days after treatment and declined to the control level after 3 weeks. In “Zesy002”, the ASM-induced response was greater at the early harvest site than at the late harvest site. This decline was concomitant with leaf yellowing and a reduction in RNA yield. Effects of postharvest ASM on gene expression did not persist into the following spring, nor were vines conditioned to respond more strongly to pre-flowering ASM application