Nature-Based Solutions (NBS), such as green roofs and green walls, offer sustainable strategies to mitigate Climate Change and to address additional challenges associated with urbanization, such as air pollution, biodiversity loss, noise, and decreased ability to retain rainwater. Despite their widely recognized benefits, usually defined as ecosystem services, their adoption remains limited due to high costs, complex installation processes, and maintenance requirements. This work presents the EU-funded LIFE CAPT-TILE project, which introduces an innovative and modular tile solution to overcome these constraints by developing a lightweight terracotta-based tile named “CAPT-TILE” to be integrated with soil layers to support vegetation growth on facades and on pitched and flat roofs. The project follows a multi-phase methodology, including laboratory trials, operational testing through the realization of two mock-ups, and practical in situ demonstration of CAPT-TILEs’ performance. This includes their installation on roofs and a green façade on two buildings located in Italy and Poland. The project aims to demonstrate the environmental and socio-economic benefits of CAPT-TILEs, including GHG sequestration (i.e., each m2 of CAPT-TILE surface has the potential to absorb 1.1–1.8 kg of CO2 from the atmosphere and store it as plant biomass), biodiversity promotion, and mitigation of noise and temperature. Furthermore, market validation and sustainability assessments, conducted through Life Cycle Assessment and Life Cycle Costing, will support the CAPT-TILE technologies’ commercialization and facilitate their widespread adoption. The LIFE CAPT-TILE project represents a significant effort toward democratizing urban greening systems by bridging the gap between environmental goals and socioeconomic constraints.

Azolla spp. are floating ferns used for centuries as biofertilizers to enrich the soil with inorganic nitrogen and improve rice yields. However, the molecular interactions between Azolla and co-cultivated rice plants only recently started to be thoroughly investigated. In this study, we exploited an experiment in which rice plants were grown together with Azolla by maintaining a low and constant concentration of inorganic nitrogen. We employed a combination of non-targeted metabolomics, chemometrics, and molecular networking to dissect the impact of Azolla co-cultivation on the metabolome of rice roots- and leaves, as well as to annotate the metabolites released by Azolla into the growing medium. Our analyses showed that Azolla can synthesize and release a broad range of metabolites in the culture medium, mainly comprising small peptides (i.e., di- and tri-peptides) and flavonoids, that may have stimulated the rice plant growth. We also observed a systemic response in the upregulation of rice metabolites, first in the roots and then in the leaves. Metabolomics analysis indicated that during the first stages of co-cultivation, the impact of Azolla on rice mainly resulted in the accumulation of small peptides, lipids and carbohydrates in roots, as well as flavonoid glycosides and carbohydrates in leaves. Consistent with these results, transcriptomics analysis of rice roots indicated significant changes in the expressions of genes coding for small peptide and lipid transporters and genes involved in the pathways of amino acid salvage and biosynthesis. Overall, our study provides new insights into Azolla's beneficial and growth-promoting effects on rice. Understanding the molecular mechanisms by which Azolla functions as a biostimulant in rice co-culture will facilitate the development of more sustainable and environmentally friendly techniques to increase yields.

Bryophytes desiccate rapidly when relative humidity decreases. The capacity to withstand dehydration depends on several ecological and physiological factors. Volatile organic compounds (VOCs) may have a role in enhancing tolerance to desiccating bryophytes. However, the functions of VOCs in bryophytes have received little attention so far. We aimed to investigate the impact of a dehydration-rehydration treatment on primary carbon metabolism and volatile terpenes (VTs) in three bryophytes with contrasting ecological traits: Vessicularia dubyana, Porella platyphylla and Pleurochaete squarrosa. First, we confirmed the desiccation sensitivity gradient of the species. Under fully hydrated conditions, the photosynthetic rate (A) was inversely associated with stress tolerance, with a lower rate in more tolerant species. The partial recovery of A in P. platyphylla and P. squarrosa after rehydration confirmed the desiccation tolerance of these two species. On the other hand, A did not recover after rehydration in V. dubyana. Regarding VT, each species exhibited a distinct VT profile under optimum hydration, with the highest VT pool found in the more desiccation-sensitive species (V. dubyana). However, the observed species-specific VT pattern could be associated with the ecological habitat of each species. P. squarrosa, a moss of dry habitats, may synthesize mainly non-volatile secondary metabolites as stress-defensive compounds. On the other hand, V. dubyana, commonly found submerged, may need to invest photosynthetically assimilated carbon to synthesize a higher amount of VTs to cope with transient water stress occurrence. Further research on the functions of VTs in bryophytes is needed to deepen our understanding of their ecological significance.

Volatile organic compounds (VOCs) are important phytochemicals emitted by plants to cope with a multitude of (a)biotic stresses and to communicate with the surrounding ecosystem. The biosynthesis and emission of VOCs are closely intertwined with photosynthetic and other cellular metabolic processes. While only some plant species constitutively release large amounts of VOCs, their emissions can be induced in any species following exposure to stressful conditions. The complex bouquet of VOCs is shaped during the plant's defense response and represents a chemical fingerprint that identifies species and specific physiological states of plants. Analytical approaches coupling mass spectrometry with either (static and dynamic) enclosure systems or micrometeorological methods provide accurate measurements of VOC emissions at plant or ecosystem levels. Off-line thermal desorption gas chromatography mass spectrometry (TD–GC–MS) relies on adsorbents and pre-concentration during sample collection and is ideal for the chemical identification of VOCs or large-scale sampling. Proton transfer reaction mass spectrometry (PTR–MS) is a real-time technique that allows direct measurements of VOCs without sample collection, which is useful for monitoring changes in VOC emissions and investigating their biosynthesis. Traditionally, analyses of VOC emissions from tree leaves, branches, and above the canopy have been employed to investigate the plant response - to (a)biotic stresses and to evaluate the impact of reactive VOCs (e.g. terpenes) in the atmosphere. However, recent findings highlight how measuring stress-induced VOC emissions can be adopted as a non-invasive method to assess the occurrence of stress conditions in plants, which could be used to monitor the health of forests.

The bacterium Stenotrophomonas rhizophila is known to be beneficial for plants and has been frequently isolated from the rhizosphere of crops. In the present work, we isolated from the phyllosphere of an ornamental plant an epiphytic strain of S. rhizophila that we named Ep2.2 and investigated its possible application in crop protection. Compared to S. maltophilia LMG 958, a well-known plant beneficial species which behaves as opportunistic human pathogen, S. rhizophila Ep2.2 showed distinctive features, such as different motility, a generally reduced capacity to use carbon sources, a greater sensitivity to fusidic acid and potassium tellurite, and the inability to grow at the human body temperature. S. rhizophila Ep2.2 was able to inhibit in vitro growth of the plant pathogenic fungi Alternaria alternata and Botrytis cinerea through the emission of volatile compounds. Simultaneous PTR-MS and GC-MS analyses revealed the emission, by S. rhizophila Ep2.2, of volatile organic compounds (VOCs) with well-documented antifungal activity, such as furans, sulphur-containing compounds and terpenes. When sprayed on tomato leaves and plants, S. rhizophila Ep2.2 was able to restrict B. cinerea infection and to prime the expression of Pti5, GluA and PR1 plant defense genes.

Exposure to high light intensity (HL) and cold treatment (CT) induces reddish pigmentation in Azolla filiculoides, an aquatic fern. Nevertheless, how these conditions, alone or in combination, influence Azolla growth and pigment synthesis remains to be fully elucidated. Likewise, the regulatory network underpinning the accumulation of flavonoids in ferns is still unclear. Here, we grew A. filiculoides under HL and/or CT conditions for 20 days and evaluated the biomass doubling time, relative growth rate, photosynthetic and non-photosynthetic pigment contents, and photosynthetic efficiency by chlorophyll fluorescence measurements. Furthermore, from the A. filiculoides genome, we mined the homologs of MYB, bHLH, and WDR genes, which form the MBW flavonoid regulatory complex in higher plants, to investigate their expression by qRT-PCR. We report that A. filiculoides optimizes photosynthesis at lower light intensities, regardless of the temperature. In addition, we show that CT does not severely hamper Azolla growth, although it causes the onset of photoinhibition. Coupling CT with HL stimulates the accumulation of flavonoids, which likely prevents irreversible photoinhibition-induced damage. Although our data do not support the formation of MBW complexes, we identified candidate MYB and bHLH regulators of flavonoids. Overall, the present findings are of fundamental and pragmatic relevance to Azolla's biology.

Biogenic Volatile Organic Compounds (BVOCs) emitted from vegetation are precursors of ozone (O3), photochemical oxidants and secondary organic aerosols (SOA) in the lower troposphere. The interaction between urban polluted air plumes originated along the coasts of the Mediterranean basin with biogenic emission occurring inland contributes to the occurrence of exceedances of the air quality standards for O3 and fine suspended particles (PM10, PM2.5). This interaction is favoured by prevailing sea-land breeze circulation during persistent high-pressure conditions. The actual contribution of BVOC to photochemical pollution is still uncertain because the approaches used to assess the emissions from terrestrial vegetation are quite different. There is some evidence in literature that BVOC emissions from Mediterranean vegetation is not accurately estimated by models based on the plant functional types (PFT) approach. To investigate these issues, a Plant Specific Emission Model (PSEM) was developed and applied to the Campania region in Southern Italy, for which a detailed vegetation inventory has been built. BVOC emission maps estimated by PSEM were compared with those generated using a PFT methodology, evidencing significant differences. BVOC emissions from the two models were then used to predict the concentrations of precursors and products of photochemical smog pollution over the Gulf of Naples (Italy) using a chemical-transport model. Simulations were performed during a period characterised by high pressure conditions that favour an enhanced O3 production under a sea-breeze circulation regime. VOCs concentration profiles predicted by the two models were compared with field data collected over Mount Vesuvius using a tethered balloon coupled with a PTR-MS. The results proved the better capability of PSEM to predict the concentrations of many BVOCs, including isoprene and some of its primary oxidation products over the measuring site, and suggested the significant potential of BVOC emission to produce SOA over the gulf of Naples.

Durum wheat is a rain-fed crop mainly cultivated in the Mediterranean basin and threatened by climate change. In particular, drought stress is one of the major constraints that can negatively affect crops production worldwide. In the present study we characterized drought stress responses in a set of durum wheat genotypes by combining plant growth parameters analysed using a plant phenotyping platform, with physiological parameters derived from gas-exchange measurements, namely exchanges of CO 2 , water vapor, and profile of emitted Volatile Organic Compounds (VOCs), in order to develop new programs for precision agriculture.

Many agronomic trials demonstrated the nitrogen-fixing ability of the ferns Azolla spp. arid its obligate cyanobiont Trichormus azollae. In this study, we have screened the emission of volatile organic compounds (VOCs) and analyzed pigments (chlorophylls, carotenoids) as well as phenolic compounds in Azolla azollae symbionts exposed to different light intensities. Our results revealed VOC emission mainly comprising isoprene and methanol (similar to 82% and similar to 13% of the overall blend, respectively). In particular, by dissecting VOC emission from A. filiculoides and T. azollae, we found that the cyanobacterium does not emit isoprene, whereas it relevantly contributes to the methanol flux. Enhanced isoprene emission capacity (15.95 +/- 2.95 nmol m(-2) s(-1)), along with increased content of both phenolic compounds and carotenoids, was measured in A. filiculoides grown for long-term under high (700 mu mol m(-2) s(-1)) rather than medium (400 mu mol m(-2) s(-1)) and low (100 mu mol m(-2) s(-1)) light intensity. Moreover, lightresponses of chlorophyll fluorescence demonstrated that A. filiculoides was able to acclimate to high growth light. However, exposure of A. filiculoides from low (100 mu mol m(-2) s(-1)) to very high light (1000 mu mol m(-2) s(-1)) did not affect, in the short term, photosynthesis, but slightly decreased isoprene emission and leaf pigment content whereas, at the same time, dramatically raised the accumulation of phenolic compounds (i.e. deoxyanthocyanidins and phlobaphenes). Our results highlight a coordinated photoprotection mechanism consisting of isoprene emission and phenolic compounds accumulation employed by A. filiculoides to cope with increasing light intensities.

Arundo donax has been recognized as a promising crop for biomass production on marginal lands due to its superior productivity and stress tolerance. However, salt stress negatively impacts A. donax growth and photosynthesis. In this study, we tested whether the tolerance of A. donax to salinity stress can be enhanced by the addition of 5-aminolevulinic acid (ALA), a known promoter of plant growth and abiotic stress tolerance. Our results indicated that root exposure to ALA increased the ALA levels in leaves along the A. donax plant profile. ALA enhanced Na+ accumulation in the roots of salt-stressed plants and, at the same time, lowered Na+ concentration in leaves, while a reduced callose amount was found in the root tissue. ALA also improved the photosynthetic performance of salt-stressed apical leaves by stimulating stomatal opening and preventing an increase in the ratio between abscisic acid (ABA) and indol-3-acetic acid (IAA), without affecting leaf methanol emission and plant growth. Supply of ALA to the roots reduced isoprene fluxes from leaves of non-stressed plants, while it sustained isoprene fluxes along the profile of salt-stressed A. donax. Thus, ALA likely interacted with the methylerythritol 4-phosphate (MEP) pathway and modulate the synthesis of either ABA or isoprene under stressful conditions. Overall, our study highlights the effectiveness of ALA supply through soil fertirrigation in preserving the young apical developing leaves from the detrimental effects of salt stress, thus helping of A. donax to cope with salinity and favoring the recovery of the whole plant once the stress is removed.

Arundo donax L. is an invasive species that has been recently employed for biomass production due to its wellknown ability to colonize harsh environment. Based on previous observations, the present study investigated the potential role of phenylpropanoids and class III peroxidases to confer adaptation through biochemical and transcriptomic analysis in A. donax after Na+ and P excess supply, both in single stress and in combination, and after growth at low P level. The levels of hydrogen peroxide, flavonoids (i.e., quercetin, apigenin and kaempferol derivatives) and the activity of class III peroxidases, as well as the expression of several genes encoding for their enzymes involved in their biosynthesis, increased when Na+ was supplied in combination with P. These results suggest that those biomolecules are involved in the response of A. donax, to the presence of +Na and P in the soil. Moreover, even though at the sampling time no significant accumulation of lignin has been determined, the trend of accumulation of such metabolite and most of all the increase of several transcripts involved in its synthesis was found. This work for the first time indicates the need for further investigation devoted to elucidating whether the strengthening of cell walls via lignin synthesis is one of the mechanisms used by A. donax to adapt to harsh environments.

Bryophytes life cycle and physiology strongly depend on water availability and most species are desiccation-tolerant. Volatile organic compounds (VOCs) may have a role in helping plants overcome stresses, including desiccation. We aimed to investigate the impact of a dehydration-rehydration treatment on primary metabolism and VOCs in three different bryophytes species: the desiccation sensitive (DS) moss Vessicularia dubyana , and the desiccation-tolerant (DT) liverwort Porella platyphylla and the DT moss Pleurochaete squarrosa . We found a different sensitivity of CO 2 photosynthetic assimilation (A) both under photorespiratory (normoxic) and non-photorespiratory conditions (hypoxic), to dehydration/rehydration in DT and DS bryophytes, as A did not recover in DS. In all the investigated species, monoterpenes were the main components of the VOCs that correlated with A in the three species. However, the DS species constitutively contained a higher content of VOCs than the two DT species. However, a markedly species-specific pattern was found, independent of the desiccation tolerance strategy and probably linked to the ecological strategies and attributes. When bryophytes were desiccated, the total VOCs content increased, with a fingerprint monoterpenes characteristic of desiccation in the DT- species (terpinolene, β-pinene, and p-cymene), which possibly reveal VOCs functions similar to those characteristics of higher plants.

For the first time, emission/deposition fluxes of volatile organic compounds (VOCs) and H2S from a historic closed landfill site in Southern Italy were determined by Eddy Covariance (EC) using Proton Transfer Reaction Time-of-Flight Mass Spectrometry (PTR-TOF-MS). This was done in two field campaigns of one week performed in July and October 2016, where fluxes of CO2 and CH4 were also measured. Many compounds not previously identified in the biogas were detected by PTR-TOF-MS, but only in July some of them produced positive fluxes exceeding the flux limit of detection. Methanol was the most emitted compound with an average flux of 44.20 +/- 4.28 mu g m-2 h-1, followed by toluene with a mean flux of 18.97 +/- 2.47 mu g m-2 h-1. Toluene fluxes were 10 times higher than those of benzene, fitting rather well with values previously measured in the biogas. VOCs emission fluxes of monoterpenes and highly reactive arenes did not reflect, however, the biogas composition. This, combined with tiny emissions of VOC oxidation products, suggests that landfill emissions underwent some photochemical degradation before being dispersed in the atmospheric boundary layer (ABL). Deposition fluxes of some VOCs emitted from the sea was also observed in July. No relevant VOC fluxes were instead measured in October, suggesting that temperature was the variable controlling most landfill emission. Albeit small, summer landfill emissions from the investigated site can have an impact on the population living nearby, because they contain or still generate compounds that causing nuisance.

Climate change will affect the growing season and increase the occurrence of extreme stressful events, thus altering crop phenological phases and the associated emission of biogenic volatile organic compounds (BVOC). BVOC exchange has been poorly investigated in field crops, especially in the Mediterranean area. In this study we report continuous measurements of BVOC fluxes and CO2 net ecosystem exchange (NEE), together with environmental variables, green area index (GAI) and aboveground biomass (AGB) during a whole growing season in a grain sorghum (Sorghum bicolor x Sorghum sudangrass., cv. Nicol, Pioneer) plantation located in Southern Europe. Results of this intensive field campaign showed that, while the bare soil of our site was a sink of BVOC, the sorghum plantation became a source of oxygenated BVOC, mainly methanol and acetaldehyde, which were emitted over the season at an average rate of 0.137 +/- 0.013 and 0.070 +/- 0.004 nmol m(-2) s(-1), respectively. In addition, the application of the advanced data mining method of Self-Organizing Maps (SOM) revealed distinctive patterns of BVOC fluxes correlating with sorghum growth stages (GS): in the first stage (GS1), developing plantlets emitted a mixture of BVOC uniquely characterized by monoterpenes; in GS2, adult plants forming an homogeneous dense canopy emitted the most abundant fluxes of a mixture of oxygenated BVOC comprising methanol, acetaldehyde, formic acid, acetone, acetic acid and n-pentenol; once plants entered the flowering stage (in GS3), only a few BVOC continued to be emitted at the highest rates (i.e. formic acid, acetone, acetic acid, n-pentenol). Moreover, the application of SOM to a sub-set of BVOC fluxes highlighted the possibility to qualitatively differentiate stressful events of plant lodging and harvest cutting. In fact, enhanced emission of acetaldehyde distinguished the BVOC mixture emitted from lodged rather than from cut and harvested sorghum plants in the field.

One of the main impacts of climate change on agriculture production is the dramatic increase of saline (Na + ) content in substrate, that will impair crop performance and productivity. Here we demonstrate how the application of smart technologies such as an in vivo sensor, termed bioristor, allows to continuously monitor in real-time the dynamic changes of ion concentration in the sap of Arundo donax L. (common name giant reed or giant cane), when exposed to a progressive salinity stress. Data collected in vivo by bioristor sensors inserted at two different heights into A. donax stems enabled us to detect the early phases of stress response upon increasing salinity. Indeed, the continuous time-series of data recorded by the bioristor returned a specific signal which correlated with Na + content in leaves of Na-stressed plants, opening a new perspective for its application as a tool for in vivo plant phenotyping and selection of genotypes more suitable for the exploitation of saline soils.

Arundo donax L. is an invasive grass species with high tolerance to a wide range of environmental stresses. The response of potted A. donax plants to soil stress characterized by prolonged exposure (43 days) to salinity (+Na), to high concentration of phosphorus (+P), and to the combination of high Na and P (+NaP) followed by 14 days of recovery under optimal nutrient solution, was investigated along the entire time-course of the experiment. After an exposure of 43 days, salinity induced a progressive decline in stomatal conductance that hampered A. donax growth through diffusional limitations to photosynthesis and, when combined with high P, reduced the electron transport rate. Isoprene emission from A. donax leaves was stimulated as Na+ concentration raised in leaves. Prolonged growth in P-enriched substrate did not significantly affect A. donax performance, but decreased isoprene emission from leaves. Prolonged exposure of A. donax to + NaP increased the leaf level of H2O2, stimulated the production of carbohydrates, phenylpropanoids, zeaxanthin and increased the de-epoxidation state of the xanthophylls. This might have resulted in a higher stress tolerance that allowed a fast and full recovery following stress relief. Moreover, the high amount of ABA-glucose ester accumulated in leaves of A. donax exposed to + NaP might have favored stomata re-opening further sustaining the observed prompt recovery of photosynthesis. Therefore, prolonged exposure to high P exacerbated the negative effects of salt stress in A. donax plants photosynthetic performances, but enhanced activation of physiological mechanisms that allowed a prompt and full recovery after stress.

Tropospheric ozone (O3) impairs physiological processes of plants while nitrogen (N) deposition may cause imbalances in soil N and other nutrients such as phosphorus (P) suggesting an increase of P demand for plants. However, the combined effect of O3, soil N and P on isoprene emission from leaves has never been tested. We therefore examined isoprene emission in leaves of Oxford poplar clone exposed to O3 (ambient, AA [35.0 nmol mol-1 as daily mean]; 1.5 × AA; 2.0 × AA), soil N (0 and 80 kg N ha-1) and soil P (0, 40 and 80 kg P ha-1) in July and September in a Free-Air Controlled Exposure (FACE) facility. We also investigated the response of isoprene emission to foliar N, P and abscisic acid (ABA) contents in September because the 2-C-methylerythritol-5-phosphate (MEP) pathway of isoprenoid biosynthesis produces ABA. We found that O3 increased isoprene emission in July, which was associated to increased dark respiration, suggesting an activation of metabolism against O3 stress as an initial response. However, O3 decreased isoprene emission in September which was associated to reduced net photosynthesis. In September, isoprene emission was positively correlated with leaf N content and negatively correlated with leaf P content in AA. However, no response of isoprene emission to foliar N and P was found in elevated O3, suggesting that the isoprene responses to foliar N and P depended on the O3 exposure levels. Isoprene emission rate in 1.5 × AA and 2.0 × AA increased with increasing leaf ABA content, indicating accelerated senescence of injured leaves to favor new leaf growth when high O3 and nutritional availability in the soil were combined. Even though foliar N and P usually act as a proxy for isoprene emission rate, the impact of recent abiotic factors such as O3 should be always considered for modeling isoprene emission under climate change.

Ceratocystis platanicauses canker stain of plane trees, and it represents a serious disease ofPlatanusspp. both in the United States and Europe. Current chemical or biological controls do not effectively manageC. platani, so new preventive methods need to be developed in order to limit this pathogen spreading. In this work, we have characterized the main volatile organic compounds (VOC) emitted in vitro from pure cultures ofC. plataniand other common pathogenic fungal species of hosts plants growing in the same ecosystems as plane trees. We found thatC. plataniemitted a similar blend of VOC compared with phylogenetically similar speciesC. populicola. In particular,C. plataniwas characterized by emission of isoamyl acetate and isobutyl acetate whileC. populicolaby ethyl acetate and isobutyl acetate, which were not released by any of the other out-group fungal species grown on the same medium. Moreover, following a targeted approach based on the main VOC found in vitro, we have successfully validated in vivo that VOC uniquely emitted byC. platani(i.e. isobutyl acetate along with isoamyl alcohol) were released from the bark of plane trees followingC. plataniinoculation. Our results highlight the possibility to exploit VOC emitted specifically byC. platanias biomarkers to recognizePlatanus x acerifoliaplants infected by this pathogen.

Tomato plants can establish symbiotic interactions with arbuscular mycorrhizal fungi (AMF) able to promote plant nutrition and prime systemic plant defenses against pathogens attack; the mechanism involved is known as mycorrhiza-induced resistance (MIR). However, studies on the effect of AMF on viral infection, still limited and not conclusive, indicate that AMF colonization may have a detrimental effect on plant defenses against viruses, so that the term "mycorrhiza-induced susceptibility" (MIS) has been proposed for these cases. To expand the case studies to a not yet tested viral family, that is, Bromoviridae, we investigated the effect of the colonization by the AMFFunneliformis mosseaeon cucumber mosaic virus (CMV) infection in tomato by phenotypic, physiological, biochemical, and transcriptional analyses. Our results showed that the establishment of a functional AM symbiosis is able to limit symptoms development. Physiological and transcriptomic data highlighted that AMF mitigates the drastic downregulation of photosynthesis-related genes and the reduction of photosynthetic CO(2)assimilation rate caused by CMV infection. In parallel, an increase of salicylic acid level and a modulation of reactive oxygen species (ROS)-related genes, toward a limitation of ROS accumulation, was specifically observed in CMV-infected mycorrhizal plants. Overall, our data indicate that the AM symbiosis influences the development of CMV infection in tomato plants and exerts a priming effect able to enhance tolerance to viral infection.

Three morphological mutants (M71a, M71b, M71c) of the antagonist Pseudomonas chlororaphis M71, naturally arose during a biocontrol trial against the phytopathogenic fungus Fusarium oxysporum f.sp. radicis-lycopersisci. In this study, the three mutants were investigated to elucidate their role in the biocontrol of plant pathogens. M71a and M71b phenotypes were generated by a mutation in the two-component system GacS/GacA. The mutation determined an increase in siderophore production and an impaired ability to release proteases, to swarm, to produce phenazine and AHLs and to colonize tomato roots. In vitro antagonistic activity against different plant pathogens was partially reduced in M71a, while M71b resulted effective only against Pythium ultimum. Biocontrol efficacy against Fusarium oxysporum f.sp. radicis-lycopersisci, was partially reduced in M71a and completely lost in M71b. M71c phenotype was impaired in swarming motility, did not produce biofilms and its antagonistic activity was similar to the parental M71 strain. M71c showed an enhanced ability to colonize tomato roots, on which its progeny in part reverted to the M71 parental phenotype. Volatile organic compounds (VOCs) emitted by all four strains, inhibited the growth of Clavibacter michiganensis subsp. michiganensis and Seiridium cardinale in vitro. Real-time screening of VOCs by PTR-MS combined with GC-MS analysis, showed that methanethiol was the main component of the blend produced by all four M71 strains. However, the emissions of hydrogen cyanide, dimethyl disulfide, 1,3-butadiene and acetone were significantly affected by the three different mutations. These findings highlight that the simultaneous presence of different M71 phenotypes may improve, through the integration of different mechanisms, the ecological fitness and biocontrol efficacy of P. chlororaphis M71.