A multidisciplinary approach to tackling invasive species: barcoding, morphology, and metataxonomy of the leafhopper Arboridia adanae.

The leafhopper genus Arboridia includes several species that feed on Vitis vinifera and cause leaf chlorosis. We report the first alien Arboridia infestation in Italy in 2021 in an Apulian vineyard. To confirm the taxonomic status of the species responsible for crop damage, and reconstruct its demographic history, we barcoded individuals from Apulia together with Arboridia spp. from Crete (Greece), A. adanae from Central Turkey and other specimens of the presumed sister species, A. dalmatina from Dalmatia (Croatia). Molecular phylogenies and barcoding gap analysis identified clades not associated with sampling locations. This result is incongruent with classical specimen assignment and is further supported by morphological analyses, which did not reveal significant differences among the populations. Therefore, we propose A. dalmatina as a junior synonym of A. adanae, which would become the only grapevine-related Arboridia species in the eastern Mediterranean. To further characterise A. adanae evolution, we performed a molecular clock analysis that suggested a radiation during the Pleistocene glaciations. Finally, to assess whether the Apulian individuals carried microorganisms of agricultural relevance, we sequenced their bacterial microbiota using 16S rRNA amplicon sequencing identifying three phytopathogens not generally associated with Arboridia activities as well as Wolbachia in one Apulian haplogroup. We discuss the agricultural implications of this infestation.

LC-Q-TOF-MS/MS data of phyllobilins in apple peels cv. 'Gala' during ripening under shelf-life conditions.

During senescence and ripening, higher plants degrade the green pigment chlorophyll to linear tetrapyrrols, referred to as phyllobilins (PBs). This dataset provides chromatograms and mass spectral data of PBs acquired from methanolic extracts of cv. Gala apple peels at five different shelf life (SL) stages. Data were obtained using a (ultra-high) pressure liquid chromatograph (UHPLC) coupled to high resolution quadrupole-time of flight-mass spectrometer (HRMS-Q-TOF). A data-dependent inclusion list (IL) ex professo containing all known masses of PBs was applied to address PBs, and fragmentation patterns were studied to confirm their identity operating a MS2 method. Mass accuracy was set to 5 ppm for parent ion peaks, this parameter was adopted as inclusion criterium. The detection of PBs' appearance during ripening can be helpful for assessing the quality and maturity of the apples.

Chlorophyll breakdown during fruit ripening: Qualitative analysis of phyllobilins in the peel of apples (Malus domestica Borkh.) cv. 'Gala' during different shelf life stages.

Chlorophyll (Chl) degradation is an important process of fruit ripening, mediated by the pheophorbide a oxygenase/phyllobilin (PaO/PB) pathway. Chl is catabolized to linear tetrapyrrolic phyllobilins that resemble the bile pigments. Determining the optimal harvest date is of pivotal importance for the fruit industry. Among the physiological changes at the onset of ripening and during shelf life, the breakdown of Chl appears as a promising indicator for fruit maturity. In addition, PBs are a frequently overlooked component of the human diet with antioxidant and antiproliferative activities. However, their relevance for a healthy diet and their potential as ripening indicator in fruit cannot be evaluated yet, as details on emergence and composition of PBs in fruit are yet elusive. Using apple (Malus × domestica Borkh.) cv. 'Gala' as a model fruit, the Chl content and the emergence and identity of PBs were analyzed during a controlled shelf life period using UV/Vis spectroscopy and ultra-high pressure liquid chromatography coupled to high resolution quadrupole-time of flight-mass spectrometry (UHPLC-Q-TOF-MS). An in-house database with chromatographic and MS data from 51 PBs, revealed ten chlorophyll catabolites, including five NCCs, one YCC, and four DNCCs (including a previously unknown one). PBs were identified with increasing abundance and diversity from the onset of Chl degradation, suggesting a potential role as ripening indicators.

Non-destructive fluorescence sensing for assessing microclimate, site and defoliation effects on flavonol dynamics and sugar prediction in Pinot blanc grapes.

In an era of growing international competition in modern viticulture, the study and implementation of innovative technologies to increase the production of high-quality grapes and wines are of critical importance. In this study, the non-destructive portable sensor Multiplex, based on fluorescence sensing technique, was applied to evaluate grape maturity parameters and flavonol content of the understudied Pinot blanc variety. The effects of environmental and agronomical factors on flavonol content of Pinot blanc grapes were investigated in eight vineyards characterised by different microclimatic and agronomic conditions. Furthermore, the direct impact of canopy management treatment on the flavonol dynamics of the grapes oriented in the four cardinal directions was assessed. Results highlight the positive role of moderate temperatures and direct sunlight exposure on Pinot blanc flavonol content; however, no direct vineyard-elevation effect was observed. The ability to modulate and evaluate the flavonol content in field represent crucial factors because of their potential effect on flavonoids-dependent wine characteristics, such as stability and ageing. In the present study, for the first time, two calibration curves were reported for pre- and post-veraison periods between flavonol indices and the berry skin flavonol content and a good correlation was observed between Multiplex measurement and the total polyphenolic content of grape juice. Moreover, the strong correlation between the chlorophyll index with grape juice sugar content and titratable acidity revealed the practical application of non-destructive sensors to predict the optimal harvest time for Pinot blanc grapes. In conclusion, the non-destructive fluorescence sensor Multiplex is a high-potential tool for innovative viticulture, for evaluating grape skin composition variables in white grape varieties.

Biogenic volatile organic compounds in the grapevine response to pathogens, beneficial microorganisms, resistance inducers, and abiotic factors.

The synthesis of volatile organic compounds (VOCs) in plants is triggered in response to external stimuli, and these compounds can migrate to distal tissues and neighbouring receivers. Although grapevine VOCs responsible for wine aroma and plant-insect communications are well characterized, functional properties of VOCs produced in response to phytopathogens, beneficial microorganisms, resistance inducers, and abiotic factors have been less studied. In this review, we focused on the emission patterns and potential biological functions of VOCs produced by grapevines in response to stimuli. Specific grapevine VOCs are emitted in response to the exogenous stimulus, suggesting their precise involvement in plant defence response. VOCs with inhibitory activities against pathogens and responsible for plant resistance induction are reported, and some of them can also be used as biomarkers of grapevine resistance. Likewise, VOCs produced in response to beneficial microorganisms and environmental factors are possible mediators of grapevine-microbe communications and abiotic stress tolerance. Although further functional studies may improve our knowledge, the existing literature suggests that VOCs have an underestimated potential application as pathogen inhibitors, resistance inducers against biotic or abiotic stresses, signalling molecules, membrane stabilizers, and modulators of reactive oxygen species. VOC patterns could also be used to screen for resistant traits or to monitor the plant physiological status.

The Differential Growth Inhibition of Phytophthora spp. Caused by the Rare Sugar Tagatose Is Associated With Species-Specific Metabolic and Transcriptional Changes.

Tagatose is a rare sugar with no negative impacts on human health and selective inhibitory effects on plant-associated microorganisms. Tagatose inhibited mycelial growth and negatively affected mitochondrial processes in Phytophthora infestans, but not in Phytophthora cinnamomi. The aim of this study was to elucidate metabolic changes and transcriptional reprogramming activated by P. infestans and P. cinnamomi in response to tagatose, in order to clarify the differential inhibitory mechanisms of tagatose and the species-specific reactions to this rare sugar. Phytophthora infestans and P. cinnamomi activated distinct metabolic and transcriptional changes in response to the rare sugar. Tagatose negatively affected mycelial growth, sugar content and amino acid content in P. infestans with a severe transcriptional reprogramming that included the downregulation of genes involved in transport, sugar metabolism, signal transduction, and growth-related process. Conversely, tagatose incubation upregulated genes related to transport, energy metabolism, sugar metabolism and oxidative stress in P. cinnamomi with no negative effects on mycelial growth, sugar content and amino acid content. Differential inhibitory effects of tagatose on Phytophthora spp. were associated with an attempted reaction of P. infestans, which was not sufficient to attenuate the negative impacts of the rare sugar and with an efficient response of P. cinnamomi with the reprogramming of multiple metabolic processes, such as genes related to glucose transport, pentose metabolism, tricarboxylic acid cycle, reactive oxygen species detoxification, mitochondrial and alternative respiration processes. Knowledge on the differential response of Phytophthora spp. to tagatose represent a step forward in the understanding functional roles of rare sugars.

Trichoderma spp. volatile organic compounds protect grapevine plants by activating defense-related processes against downy mildew.

Volatile organic compounds (VOCs) are produced by soil-borne microorganisms and play crucial roles in fungal interactions with plants and phytopathogens. Although VOCs have been characterized in Trichoderma spp., the mechanisms against phytopathogens strongly differ according to the strain and pathosystem. This study aimed at characterizing VOCs produced by three Trichoderma strains used as biofungicides and to investigate their effects against grapevine downy mildew (caused by Plasmopara viticola). A VOC-mediated reduction of downy mildew severity was found in leaf disks treated with Trichoderma asperellum T34 (T34), T. harzianum T39 (T39), and T. atroviride SC1 (SC1) and 31 compounds were detected by head space-solid phase microextraction gas chromatography-mass spectrometry. Among the Trichoderma VOCs annotated, α-farnesene, cadinene, 1,3-octadiene, 2-pentylfuran, and 6-pentyl-2H-pyran-2-one reduced downy mildew severity on grapevine leaf disks. In particular, 6-pentyl-2H-pyran-2-one and 2-pentylfuran increased the accumulation of callose and enhanced the modulation of defense-related genes after P. viticola inoculation, indicating an induction of grapevine defense mechanisms. Moreover, 6-pentyl-2H-pyran-2-one activated the hypersensitive response after P. viticola inoculation, possibly to reinforce the grapevine defense reaction. These results indicate that Trichoderma VOCs can induce grapevine resistance, and these molecules could be further applied to control grapevine downy mildew.

Volatile Organic Compounds From Lysobacter capsici AZ78 as Potential Candidates for Biological Control of Soilborne Plant Pathogens.

The genus Lysobacter includes several bacterial species which show potential for being used in biological control of plant diseases. It was shown recently that several Lysobacter type strains produce volatile organic compounds (VOCs) which controlled the growth of Phytophthora infestans in vitro when the bacteria were grown on a protein rich medium. In the present study, Lysobacter capsici AZ78 (AZ78) has been tested for its potential to produce VOCs that may contribute to the bioactivity against soilborne plant pathogens. To this end, split Petri dish assays of bacterial cultures have been combined with GC-MS measurements with the aim to reveal the identity of the VOCs which inhibit the growth of Pythium ultimum Rhizoctonia solani, and Sclerotinia minor. While AZ78 completely suppressed the growth of P. ultimum and S. minor, the growth of R. solani was still reduced significantly. The GC-MS analysis revealed 22 VOCs to be produced by AZ78, the majority of which were (putatively) identified as mono- and dialkylated methoxypyrazines. Based on additional cultivation and GC-MS experiments, 2,5-dimethylpyrazine, 2-ethyl-3-methoxypyrazine and 2-isopropyl-3-methoxypyrazine were selected as presumable bioactive compounds. Further bioassays employing indirect exposure to standard solutions (1-10 mg per Petri dish) of the synthetic compounds via the gas phase, revealed that each of these pyrazines was able to suppress the growth of the pathogens under investigation. The results of this study highlight the possible future implementation of pyrazine derivatives in the control of soilborne plant diseases and further support the biocontrol potential of L. capsici AZ78.

Downy mildew symptoms on grapevines can be reduced by volatile organic compounds of resistant genotypes.

Volatile organic compounds (VOCs) play a crucial role in the communication of plants with other organisms and are possible mediators of plant defence against phytopathogens. Although the role of non-volatile secondary metabolites has been largely characterised in resistant genotypes, the contribution of VOCs to grapevine defence mechanisms against downy mildew (caused by Plasmopara viticola) has not yet been investigated. In this study, more than 50 VOCs from grapevine leaves were annotated/identified by headspace-solid-phase microextraction gas chromatography-mass spectrometry analysis. Following P. viticola inoculation, the abundance of most of these VOCs was higher in resistant (BC4, Kober 5BB, SO4 and Solaris) than in susceptible (Pinot noir) genotypes. The post-inoculation mechanism included the accumulation of 2-ethylfuran, 2-phenylethanol, ß-caryophyllene, ß-cyclocitral, ß-selinene and trans-2-pentenal, which all demonstrated inhibitory activities against downy mildew infections in water suspensions. Moreover, the development of downy mildew symptoms was reduced on leaf disks of susceptible grapevines exposed to air treated with 2-ethylfuran, 2-phenylethanol, ß-cyclocitral or trans-2-pentenal, indicating the efficacy of these VOCs against P. viticola in receiver plant tissues. Our data suggest that VOCs contribute to the defence mechanisms of resistant grapevines and that they may inhibit the development of downy mildew symptoms on both emitting and receiving tissues.

Growth media affect the volatilome and antimicrobial activity against Phytophthora infestans in four Lysobacter type strains.

Bacterial volatile organic compounds (VOCs) play important ecological roles in soil microbial interactions. Lysobacter spp. are key determinants of soil suppressiveness against phytopathogens and the production of non-volatile antimicrobial metabolites has been extensively characterised. However, the chemical composition and antagonistic properties of the Lysobacter volatilome have been poorly investigated. In this work, VOC emission profiles of four Lysobacter type strains grown on a sugar-rich and a protein-rich medium were analysed using solid-phase microextraction gas chromatography-mass spectrometry and proton transfer reaction-time of flight-mass spectrometry. Lysobacter antibioticus, L. capsici, L. enzymogenes and L. gummosus type strains were recognised according to their volatilome assessed using both headspace mass spectrometry methods Moreover, the chemical profiles and functional properties of the Lysobacter volatilome differed according to the growth medium, and a protein-rich substrate maximised the toxic effect of the four Lysobacter type strains against Phytophthora infestans. Antagonistic (pyrazines, pyrrole and decanal) and non-antagonistic (delta-hexalactone and ethanol) VOCs against Ph. infestans or putative plant growth stimulator compounds (acetoin and indole) were mainly emitted by Lysobacter type strains grown on protein- and sugar-rich media respectively. Thus nutrient availability under soil conditions could affect the aggressiveness of Lysobacter spp. and possibly optimise interactions of these bacterial species with the other soil inhabitants.

Emission of volatile sesquiterpenes and monoterpenes in grapevine genotypes following Plasmopara viticola inoculation in vitro.

The grapevine (Vitis vinifera) is one of the most widely cultivated fruit crops globally, and one of its most important diseases in terms of economic losses is downy mildew, caused by Plasmopara viticola. Several wild Vitis species have been found to be resistant to this pathogen and have been used in breeding programs to introduce resistance traits to susceptible cultivars. Plant defense is based on different mechanisms, and volatile organic compounds (VOCs) play a major role in the response to insects and pathogens. Although grapevine resistance mechanisms and the production of secondary metabolites have been widely characterized in resistant genotypes, the emission of VOCs has not yet been investigated following P. viticola inoculation. A Proton Transfer Reaction-Time of Flight-Mass Spectrometer (PTR-ToF-MS) was used to analyze the VOCs emitted by in vitro-grown plants of grapevine genotypes with different levels of resistance. Downy mildew inoculation significantly increased the emission of monoterpenes and sesquiterpenes by the resistant SO4 and Kober 5BB genotypes, but not by the susceptible V. vinifera Pinot noir. Volatile terpenes were implicated in plant defense responses against pathogens, suggesting that they could play a major role in the resistance against downy mildew by direct toxicity or by inducing grapevine resistance. The grapevine genotypes differed in terms of the VOC emission pattern of both inoculated and uninoculated plants, indicating that PTR-ToF-MS could be used to screen hybrids with different levels of downy mildew resistance. Copyright © 2015 John Wiley & Sons, Ltd.