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Phytophthora cinnamomi and Phytophthora multivora are considered important forest pathogens in different areas of the world. However, while P. cinnamomi presence and impact are widely reported, little is known about the presence of Phytophthora multivora in the Mediterranean evergreen oak forests and its possible role in their decline syndrome. The present study was conducted to identify the presence of P. cinnamomi, P. multivora, and other Phytophthora species in the Castelporziano Presidential Estate in Italy using classical (baiting) and metabarcoding (HTS). A total of 52 rhizosphere soil samples were collected from declining oak stands, from which 254 isolates of oomycetes taxa were obtained among which Phytophthora cinnamomi, P. multivora, P. plurivora, P. gonapodyides, and P. bilorbang were identified based on morphological traits and barcoding. The results of HTS analysis revealed the presence in the rhizosphere of 20 different Phytophthora spp. including all the taxa obtained through baiting. Interestingly, the Phytophthora soilborne community did not differ among tree hosts. Phytophthora cinnamomi was the most common species in the investigated site. The aggressiveness of P. multivora to holm and cork oak was evaluated on 3 months old seedlings using soil infestation and keeping P. cinnamomi as the positive control. Soil inoculation test demonstrated the aggressiveness of P. multivora on both holm oak and cork oak. Phytophthora multivora resulted by far less aggressive than P. cinnamomic. Holm oak resulted the most sensitive host to both pathogens. While P. multivora seems to play a relevant role in the decline of forest and urban ecosystems in Australia, its role in evergreen oak decline seems to be limited at the moment. However, in consideration of its sensible activity as a fine root feeder and adaptation to dry environments, its role in holm and cork oak decline could become relevant in the medium-term specifically in association with climate changes.

Aerial drones are systems that have been largely employed in a number of civil applications, from surveillance and environmental monitoring to day-to-day service. In view of this, the 2020 Mohamed Bin Zayed International Robotics Challenge (MBZIRC) held in Abu Dhabi (UAE) was a competition that aimed to push forward the technological limits of the actual aerial devices. We present the design and development of drone prototypes to meet one of the goals of Challenge 1, namely autonomously identifying and attacking randomly-positioned colored targets. This paper reports the main hardware/software design choices to accomplish the highest flexibility and usability of our aerial platforms, namely the strategies to control the overall dynamics and vision for the eventual interaction with targets. We achieved the fourth place among 22 international teams, since we successfully dispatched all the targets, which were dispersed throughout an area of 6000 m2 , in 365 s. Although focused on specific goals, the methodologies developed in view of the competition are suitable for further improvements towards other application fields.

Truffle cultivation has quickly grown in European countries and elsewhere, so optimization of the nursery inoculation method to cover the high demand for mycorrhized seedlings is needed. The suitability of two compost-based potting mixes to produce Quercus pubescens Willd. mycorrhized with the black Périgord truffle T. melanosporum Vittad. was tested as an alternative to the employ of traditional potting mix used for mycorrhization and considered as the control treatment. The effects on the mycorrhizal development and the morphometric assessment of the root and shoot system of the downy oak seedlings were investigated 8 months after the spore slurry inoculation in a glasshouse experiment using the root analysis software WinRHIZO®. From the results obtained, the compost mix containing green organic residues from pruning and mowing (Mix 2) achieved better performance with a higher mycorrhization percentage than the control (59.7% and 46.7%, respectively) and significantly higher root growth with enhanced root length (531 cm), number of root tips (3.329), and root forks (3.941) both with respect to the control and the potting mix containing municipal and green organic residues (Mix 1). In conclusion, from this work is evident that potting mix containing recycled organic matter, which is readily available, cheap, and environmentally sustainable can offer excellent mycorrhization performances and may be included in the mycorrhization process of Q. pubescens seedlings with T. melanosporum under controlled conditions.

Bacterial pathogens employ a general strategy to overcome host defenses by coordinating the virulence gene expression using dedicated regulatory systems that could raise intricate networks. During the last twenty years, many studies of

Biochar (BC) soil amendments could partially counteract soil carbon (C) stock decrease in broad-leaved forests in Italy; however, its effects on the growth of representative tree species

In the present study, the sentinel arboreta strategy was applied, and its efficacy was evaluated at the Ataturk Arboretum (Istanbul, Turkey), having as a study case the interaction Tubakia spp.—Quercus spp. Thirty-four oak species native of America and Eurasia were sampled within the Fagaceae collection of the arboretum. Isolation trials were conducted from leaf necroses, and High Throughput Sequencing for fungal taxa was carried out from asymptomatic leaf blades. Four Tubakia species were identified, T. dryina, T. suttoniana, T. hallii, and T. macnabbii. Three out of four are of recent description and the present study contributed to updating their host-range. Thirty-two oak-Tubakia interactions new to science were described. Hypotheses were formulated on the possible movement across geographic areas of these species and on the risk posed in case of introduction in the distribution range of susceptible host species. As a conclusive remark, the present study confirmed the efficacy of the sentinel arboreta strategy to highlight new host–pathogen interactions and the risk of host-shift events.

Over recent years, αvβ6 and αvβ8 Arg-Gly-Asp (RGD) integrins have risen to prominence as interchangeable co-receptors for the cellular entry of herpes simplex virus-1 (HSV-1). In fact, the employment of subtype-specific integrin-neutralizing antibodies or gene-silencing siRNAs has emerged as a valuable strategy for impairing HSV infectivity. Here, we shift the focus to a more affordable pharmaceutical approach based on small RGD-containing cyclic pentapeptides. Starting from our recently developed αvβ6-preferential peptide [RGD-Chg-E]-CONH2 (1), a small library of N-methylated derivatives (2–6) was indeed synthesized in the attempt to increase its affinity toward αvβ8. Among the novel compounds, [RGD-Chg-(NMe)E]-CONH2 (6) turned out to be a potent αvβ6/αvβ8 binder and a promising inhibitor of HSV entry through an integrin-dependent mechanism. Furthermore, the renewed selectivity profile of 6 was fully rationalized by a NMR/molecular modeling combined approach, providing novel valuable hints for the design of RGD integrin ligands with the desired specificity profile.

Forty-six strains of Trichoderma spp. have been isolated from soils in different locations and ecosystems of Algeria. They were identified at species level by analysis of their Internal Transcribed Spacers regions 1 and 2 (ITS1 and ITS2) of rDNA and a partial sequence of the Translation Elongation Factor 1-alpha (TEF 1-α) gene. The 46 Trichoderma spp. were assigned to Trichoderma atroviride (12 strains), T. gamsii (10), T. orientale (1) and 23 to T. harzianum species complex (T. harzianum, T. afroharzianum, T. atrobrunneum and T. guizhouense). In the present study we highlight that T. gamsii, T. orientale, T. atrobrunneum and T. guizhouense are being reported for the first time in Algeria. Trichoderma spp. isolates growth was evaluated at temperatures ranging from 10 to 40 °C on PDA medium. The optimum growth was recorded at 25 °C and 30 °C, and only T. orientale was able to grow at 40 °C. The in vitro test revealed the potential antagonist of Trichoderma spp. isolates against four pathogenic species associated with strategic crops in Algeria, Fusarium culmorum, Botrytis cinerea, Alternaria solani and Rhizoctonia solani. In direct confrontation, the growth rate inhibition was ranked between 37.22% and 80.95% while in indirect confrontation was between 00% and 88.89%. The biocontrol assay carried out on wheat plant showed that T. atroviride (Ta.09), T. orientale (To.15), T. afroharzianum (T af. 17 and T af. 37) and T. gamsii (T g. 39) performed well against F. culmorum the crown rot and head blight pathogen of wheat in Algeria. This finding is based on the significant decrease in disease severity compared to the control (> 82%). Data recorded, have also shown that T. atroviride Ta.09 recorded the highest percentage of disease reduction (97.28%).

Cork oak decline and mortality are increasingly prevalent in the Mediterranean basin. This require particular attention due to the great economic value of this plant species. Biscogniauxia mediterranea is one of the most common pathogens highly associated to cork oak decline. However, documentation on its incidence is still scarce, especially in Tunisia. In this context, our research aimed at detecting its presence in Tunisian natural cork oak forests and assessing factors influencing its occurrence. Investigations were carried out in six forests in northwestern Tunisia. Dendrometric parameters of trees, ecological factors (rainfall, temperature, and altitude), and the presence of B. mediterranea as pathogen were determined. The endophytic presence of this fungus was also established. It was detected in all investigated forests, with significantly different frequencies, and was reported to be related to the dendrometric structure of the cork oak trees. On the other hand, the presence of B. mediterranea as pathogen was only detected in Ain Beya and Ain Sarouia forests. It was very limited with 4% of trees per site showing the disease signs. In conclusion, measures to prevent the dispersal of B. mediterranea throughout other cork oak forests shall be implemented in order to conserve such valuable forest resources.

In the Circeo National Park (Italy), a survey assessed the occurrence of the ambrosia beetle Xylosandrus germanus in addition to the conspecifics X. compactus and X. crassiusculus. This is the first report in Europe of the co‐presence of three Xylosandrus species, in an evergreen Mediterranean maquis dominated by holm oaks (Quercus ilex L.). Although no damage caused by X. germanus has been observed, the richness of plant species in the Circeo National Park may allow the establishment of this highly polyphagous ambrosia beetle, possibly endangering this native ecosystem. The dominance in the associated fungal community of Ambrosiella grosmanniae and Fusarium spp. reflects a stable symbiotic relationship of these taxa with X. germanus.

Authors explored the potential use of Vis/NIR hyperspectral imaging (HSI) and Fourier-transform Near-Infrared (FT-NIR) spectroscopy to be used as in-line tools for the detection of unsound chestnut fruits (i.e. infected and/or infested) in comparison with the traditional sorting technique. For the intended purpose, a total of 720 raw fruits were collected from a local company. Chestnut fruits were preliminarily classified into sound (360 fruits) and unsound (360 fruits) batches using a proprietary floating system at the facility along with manual selection performed by expert workers. The two batches were stored at 4 ± 1 °C until use. Samples were left at ambient temperature for at least 12 h before measurements. Subsequently, fruits were subjected to non-destructive measurements (i.e. spectral analysis) immediately followed by destructive analyses (i.e. microbiological and entomological assays). Classification models were trained using the Partial Least Squares Discriminant Analysis (PLS-DA) by pairing the spectrum of each fruit with the categorical information obtained from its destructive assay (i.e., sound, Y = 0; unsound, Y = 1). Categorical data were also used to evaluate the classification performance of the traditional sorting method. The performance of each PLS-DA model was evaluated in terms of false positive error (FP), false negative error (FN) and total error (TE) rates. The best result (8% FP, 14% FN, 11% TE) was obtained using Savitzky-Golay first derivative with a 5-points window of smoothing on the dataset of raw reflectance spectra scanned from the hilum side of fruit using the Vis/NIR HSI setup. This model showed similarity in terms of False Negative error rate with the best one computed using data from the FT-NIR setup (i.e. 15% FN), which, however, had the lowest global performance (17% TE) due to the highest False Positive error rate (19%). Finally, considering that the total error rate committed by the traditional sorting system was about 14.5% with a tendency of misclassifying unsound fruits, the results indicate the feasibility of a rapid, in-line detection system based on spectroscopic measurements.

Bio-stimulants are showing growing success and are gradually replacing synthetic fertilizers in agriculture. Wood distillate (WD), also known as wood vinegar or pyroligneous acid, is a by-product of biomass pyrolysis and is increasingly used as a bio-stimulant for crop production. This study investigated whether weekly foliar applications of 0.25% and 0.50% WD have a differential effect on the chlorophyll and sugar content as well as biomass production in lettuce (Lactuca sativa L.). Moreover, the additional beneficial effect from the addition of corroborants of plant origin such as 3% soy lecithin and 5% flavonoid-rich wood glycolic extract to WD (BF) was investigated. Moreover, the possible toxicological concern from some potentially toxic elements (PTEs), namely Cd, Cu, Fe, Pb, and Zn, which may be abundant in WD was verified. After four weeks, we found that 0.25% WD not only increases lettuce biomass, which has an economic value, but also has beneficial effects on other qualitative parameters such as sugars and total sweetness. On the other hand, the use of 0.5% WD decreased the content of soluble sugars, suggesting a hormetic-type effect. We did not find evidence of further beneficial effects from the addition to WD of plant-derived corroborants, nor of any enrichment in the content of the investigated PTEs.

Biochar (BC) soil amendments could partially counteract soil carbon (C) stock decrease in broad-leaved forests in Italy; however, its effects on the growth of representative tree species—Fagussylvatica L. and Quercuscerris L.—has not yet been addressed. We examine whether seed germination and growth of these species are affected by addition of BC obtained from deciduous broadleaf trees. Seeds were left to germinate in greenhouse conditions under three different BC amendments: 0% (control), 10% and 20% (v/v). Seedlings were then subjected to controlled conditions under the same BC percentage. Biochar effects on seed germination were assessed measuring germination time and percentage, while effects on photosynthesis were assessed using leaf chlorophyll content (mg/m2) and photosynthetic efficiency (FV/FM). Plant growth was estimated by recording leaf number, longest leaf length and plant height. Biochar treatments had no negative effects on germination and early growth stage of the two species. Positive effects were found on the chlorophyll content of both species (ca. +8%) regardless of the treatment and on the leaf number (+30%), leaf length (+14%) and plant height (+48%) of Q. cerris (only with 10% BC). Biochar applications seem, therefore, a suitable method for increasing broad-leaved forest C stock in Italy.

Bacterial pathogens employ a general strategy to overcome host defenses by coordinating the virulence gene expression using dedicated regulatory systems that could raise intricate networks. During the last twenty years, many studies of Helicobacter pylori, a human pathogen responsible for various stomach diseases, have mainly focused on elucidating the mechanisms and functions of virulence factors. In parallel, numerous studies have focused on the molecular mechanisms that regulate gene transcription to attempt to understand the physiological changes of the bacterium during infection and adaptation to the environmental conditions it encounters. The number of regulatory proteins deduced from the genome sequence analyses responsible for the correct orchestration of gene transcription appears limited to 14 regulators and three sigma factors. Furthermore, evidence is accumulating for new and complex circuits regulating gene transcription and H. pylori virulence. Here, we focus on the molecular mechanisms used by H. pylori to control gene transcription as a function of the principal environmental changes.

BACKGROUND The brown rot fungus, Gnomoniopsis castanea, is the main organism responsible for the outbreak of chestnut postharvest decay that is threatening the sustainability of the chestnut market in Europe. Currently, no specific strategy is available to mitigate the impact and remediate the high losses of fruits in postharvest storage. In the present study, the different phases of chestnut handling in a standard facility plant were analyzed by evaluating the amount of fruit rot and infection by G. castanea at each phase. RESULTS The warm bath (48 °C) was identified as the critical phase, requiring strict parametrization to effectively inactivate G. castanea in fruits. Laboratory tests indicated that maintaining fruits at 50 °C for a maximum of 45 min provided optimal conditions to completely inactivate G. castanea inoculum during postharvest handling. However, the warm bath at 50 °C and over was not effective in inactivating the complex of fungal taxa responsible for contamination and development of molds. Higher temperatures and extended treatment times caused significant losses in fruit quality, as indicated by taste panel evaluation. Upscaling of postharvest facilities is discussed and critically evaluated. CONCLUSION The warm bath (50 °C for 45 min) is effective in completely inactivating G. castanea in fruits but did not reduce the impacts of the complex of molds responsible for external contamination and mycotoxin production. © 2021 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Biscogniauxia mediterranea is a xylariaceous ascomycete responsible for ‘charcoal disease’, the most frequent disease of cork oak in the Mediterranean basin. However, little is known regarding this fungus in North Africa. This is the first report on the morphological and genetic diversity of B. mediterranea populations in Tunisia. A total of forty-eight isolates from cork oak trees, corresponding to three different sites and three tree parts, were identified as B. mediterranea based on morphological and molecular diagnosis. Variability among the isolates was evaluated using morphological traits and microsatellite-primed PCR (MSP-PCR). B. mediterranea isolates showed high variability in culture, specifically in colony color. A high level of genetic diversity within populations and within tree parts was detected (H = 0.417, I = 0.605, and H = 0.415, I = 0.601 respectively). Significant genetic differentiation among populations (GST = 0.116 and ΦST = 0.137; p

Tumors utilize a number of effective strategies, including the programmed death 1/PD ligand 1 (PD-1/PD-L1) axis, to evade immune-mediated control of their growth. PD-L1 expression is mainly induced by IFN receptor signaling or constitutively induced. Integrins are an abundantly expressed class of proteins which play multiple deleterious roles in cancer and exert proangiogenic and prosurvival activities. We asked whether αvβ3-integrin positively regulates PD-L1 expression and the anticancer immune response. We report that αvβ3-integrin regulated constitutive and IFN-induced PD-L1 expression in human and murine cancerous and noncancerous cells. αvβ3-integrin targeted STAT1 through its signaling C tail. The implantation of β3-integrin-depleted tumor cells led to a dramatic decrease in the growth of primary tumors, which exhibited reduced PD-L1 expression and became immunologically hot, with increased IFNγ content and CD8+ cell infiltration. In addition, the implantation of β3-integrin-depleted tumors elicited an abscopal immunotherapeutic effect measured as protection from the challenge tumor and durable splenocyte and serum reactivity to B16 cell antigens. These modifications to the immunosuppressive microenvironment primed cells for checkpoint (CP) blockade. When combined with anti-PD-1, β3-integrin depletion led to durable therapy and elicited an abscopal immunotherapeutic effect. We conclude that in addition to its previously known roles, αvβ3-integrin serves as a critical component of the cancer immune evasion strategy and can be an effective immunotherapy target.

The aim of this study was to estimate the contribution of submicronic (PM1) and coarse (PM > 1) particulate matter deposition to the heavy metal load of lichens exposed along a busy road, based on the assumption that the accumulation of heavy metals occurs mostly as particles. We tested the hypothesis that lichens exposed inside a nylon cover with mesh size of 1 μm accumulate less heavy metals than lichens exposed without any cover. To this purpose, thalli of the lichen Evernia prunastri were transplanted for three months along a busy road of the urban area of Siena (Central Italy), with half of the samples exposed inside a nylon cover with a mesh size of 1 μm. The content of Al, As, Cd, Cr, Cu, Fe, Ni, Pb, Sb, and Zn was measured by ICP-MS before and after the exposure. The results showed that samples exposed inside the nylon cover accumulated a lower amount of most chemical elements and that Sb, along with Cd and Cu were the main traffic-related elements in the study area. Differences in element uptake were not caused by differences in sample vitality caused by the experimental procedure. Using a conversion factor it was possible to estimate element deposition rates for PM1 and PM > 1.

In this work, two hypotheses have been tested: 1) that lichen transplants “travelling” in smokers' cars accumulate relevant amounts of nicotine and heavy metals from cigarette smoke, and 2) that such exposure affects their vitality. Lichen samples (Evernia prunastri) were exposed for two months inside the cabin of 10 volunteer's cars, equally distributed between smokers and non-smokers. Travelling in a smoker's car for two months increased the content of nicotine and heavy metals (Al, As, Cd, Cr, Cu, Ni, Pb and Sb) in the lichen. Exposed to Control (EC) ratios revealed an indoor uptake also for Cu and Sb in non-smoker's cars, caused by traffic pollution. A smoke factor, calculated as the ratio between values of smokers’ and non-smokers’ cars, indicated a 85.6-fold contribution for nicotine and contributions in the range 1.2 (Pb) to 2 (Ni) for heavy metals; in addition, after travelling in smokers' cars, lichens showed a remarkable (60%) reduction of their vitality, as indicated by the potential quantum yield of primary photochemistry. The study demonstrated that the effects of indoor pollution by cigarette smoke can be detected using lichen transplants.

This article is based upon work carried out during COST Action FP1406 PINESTRENGTH (Pine pitch canker‐strategies for management of Gibberella circinata in greenhouses and forests), supported by COST (European Cooperation in Science and Technology). This work is a contribution of URGENTpine (PTDC/AGR‐FOR/2768/2014) funded by FCT – Fundacao para a Ciencia e a Tecnologia, I.P., through national funds, and the co‐funding by the FEDER (POCI‐01‐0145‐FEDER‐016785), within the PT2020 Partnership Agreement and Compete 2020. Thanks are due for the financial support to CESAM (UID/AMB/50017/2019), to FCT/MEC through national funds, and the co‐funding by the FEDER, within the PT2020 Partnership Agreement and Compete 2020. FCT supported J. M.‐G. (SFRH/BPD/122928/2016), J. A. (SFRH/BD/120967/2016), G. P. (SFRH/BPD/101669/2014) and A. A. (IF/00835/2013). Funds from the Spanish Government via the MINECO/FEDER grant NGSFORFUSARIUM (AGL2015‐69370‐R) and FUTURPIN (AGL2015‐68274‐C03‐02‐R) are also acknowledged. E. J. V. received funding from the Academy of Finland (grant decision number 309896). The contribution of M. M. was funded by the Forestry Commission, UK. The authors declare no conflict of interest. S. W. was partly financed by the project PROTREE, which was funded jointly by a grant from BBSRC, Defra, ESRC, the Forestry Commission, NERC and the Scottish Government, under the Tree Health and Plant Biosecurity Initiative.

In the Mediterranean basin, coastal dune systems are a priority habitat for nature conservation. Seirophora villosa is strictly associated with undisturbed dune juniper formations and can be used as an indicator of the status of conservation. Light regime and water availability are generally the main ecological factors modified by habitat fragmentation that can be detrimental to the colonisation and survival of S. villosa populations in coastal dunes. This study aims to investigate how light regime and water availability affect individual specimens of S. villosa, by studying the relationship between photosynthetic activity and water content per thallus area in different sized S. villosa thalli and comparing susceptibility of hydrated and desiccated thalli to light stress. During dehydration, photosynthetic activity decreased, reaching low constant values in smaller thalli more quickly than in larger ones. During the exposure of dry thalli to high light, photoinhibition consistently occurred earlier in smaller specimens than in larger ones. Moreover, larger thalli that were kept dried recovered to pre-treatment values within 3 days, while smaller ones took 5 days to recover initial values. On the other hand, both large and small wet thalli were photoinhibited within 1 day and recovered to pre-treatment values within 4 and 6 days respectively. Our results showed that S. villosa thalli are susceptible to sudden increases in light exposure, especially in the case of small specimens, which after photoinhibition exhibited a reduced ability to recover. Habitat fragmentation therefore represents a significant threat to the species, particularly in dispersal and establishment phases.

Wood distillate (WD) is an environmentally safe bio-based product stimulating plant growth and yield and allowed in Italy in organic farming. To the best of our knowledge, there are no studies on the effects of WD on spontaneous plants growing among crops, including their functional traits such as biomass. To test such effects, we carried out a lab experiment on artificially reconstructed arable plant communities composed of five species of conservation interest, which are specialist winter cereal crops: Bromus secalinus L., Centaurea cyanus L., Lathyrus aphaca L., Legousia speculum-veneris (L.) Chaix, and Scandix pecten-veneris L. After sowing 45 pots under controlled conditions, we applied WD at three concentrations (0%, 0.2%, and 0.5%) six times over 7 weeks. The number of emerged plants in each pot was counted every two weeks. Finally, we harvested all plants and measured the fresh and dry above-ground weight of each species in each pot. The resulting data were analyzed by Permutational Analysis of Variance. The application of 0.2% and 0.5% WD modified the community composition after two weeks, but such differences later disappeared. Both 0.2% and 0.5% WD had a positive effect on the dry weight of S. pecten-veneris and a negative effect on that of L. speculum-veneris. Moreover, 0.2% and 0.5% WD increased seedling emergence in L. aphaca, and 0.5% WD increased seedling emergence in S. pecten-veneris. Both 0.2% and 0.5% WD enhanced seedling emergence in the entire community. We suggest that the use of WD at low concentrations in winter cereals may be a sustainable agricultural practice that benefits crops without harming the associated plant diversity.

The great hopes raised by the discovery of the immunoregulatory cytokine interleukin 12 (IL-12) as an anticancer agent were marred during early clinical experimentation because of severe adverse effects, which prompted a search for alternative formulations and routes of administration. Onco-immunotherapeutic viruses (OIVs) are wild-type or genetically engineered viruses that exert antitumor activity by causing death of the tumor cells they infect and by overcoming a variety of immunosuppressive mechanisms put in place by the tumors. OIVs have renewed the interest in IL-12, as they offer the opportunity to encode the cytokine transgenically from the viral genome and to produce it at high concentrations in the tumor bed. A large body of evidence indicates that IL-12 serves as a potent adjuvant for the immunotherapeutic response elicited by OIVs in murine tumor models. The list of OIVs includes onco-immunotherapeutic herpes simplex, adeno, measles, Newcastle disease, and Maraba viruses, among others. The large increase in IL-12-mediated adjuvanticity was invariably observed for all the OIVs analyzed. Indirect evidence suggests that locally delivered IL-12 may also increase tumor antigenicity. Importantly, the OIV/IL-12 treatment was not accompanied by adverse effects and elicited a long-lasting immune response capable of halting the growth of distant tumors. Thus, OIVs provide an avenue for reducing the clinical toxicity associated with systemic IL-12 therapy, by concentrating the cytokine at the site of disease. The changes to the tumor microenvironment induced by the IL-12-armed OIVs primed the tumors to an improved response to the checkpoint blockade therapy, suggesting that the triple combination is worth pursuing in the future. The highly encouraging results in preclinical models have prompted translation to the clinic. How well the IL-12-OIV-checkpoint inhibitors' combination will perform in humans remains to be fully investigated.

This study examined whether foliar applications of wood distillate (WD) have a protective effect on photosynthesis and the antioxidant power of lettuce when exposed to an ecologically relevant O3 concentration. Seedlings of lettuce (Lactuca sativa L.) were fumigated daily with 60 ppb of O3 for 30 days, five hours per day. Once per week, 50% of the fumigated plants were treated with foliar applications of 0.2% WD, while control plants were treated with water. The results clearly showed the ability of WD to protect lettuce plants from ozone-induced damage. Specifically, WD-treated plants exhibited lower damage to the photosynthetic machinery, assessed through a series of chlorophyll fluorescence parameters, a higher chlorophyll content, higher antioxidant power, as well as antioxidant molecules, i.e., caffeic acid and quercetin, and higher biomass. Counteracting the overproduction of ozone-generated reactive oxygen species (ROS) is speculated to be the main mechanism by which WD protects the plant from ozone-induced damage.

In the previous chapter, we describe the engineering of a HSV-BAC genome by galK recombineering. Here we describe the procedures to reconstitute, or regenerate, the replicating recombinant virus, and the methods to purify it and characterize it for the correct expression of the transgene. We present the example of R-115, a recombinant expressing murine interleukin 12 (mIL12) from the US1-US2 intergenic region. A specific method for the production of highly purified virions by iodixanol gradient, suitable for in vivo applications, is also detailed.

Since the cloning of the herpes simplex virus (HSV) genome as BAC (bacterial artificial chromosome), the genetic engineering of the viral genome has become readily feasible. The advantage is that the modification of the animal virus genome is carried out in bacteria, with no replication or production of viral progeny, and is separated from the reconstitution or regeneration of the recombinant virus in mammalian cells. This allows an easy engineering of essential genes, as well. Many technologies have been developed for herpesvirus BAC engineering. In our hands the most powerful is galK recombineering that exploits a single marker (galK) for positive and negative selection and PCR amplicons for seamless modification in the desired genome locus. Here we describe the engineering of the HSV recombinant BAC 115 by the insertion of a heterologous cassette for the expression of murine interleukin 12 (mIL12) in the intergenic sequence between US1 and US2 ORFs.

Treatments of crops with additives to increase their productivity may pose environmental risks and induce negative effects also on non-target organisms. In this study, we investigated the potential effect of chestnut wood distillate (pyroligneous acid) used in agriculture, on the accumulation of trace elements in aquatic plants. As a model species, the common water fern Azolla filiculoides Lam. was selected, being often used also in phytoremediation processes. The content of selected elements of toxicological concern (As, Ba, Cd, Cu, Fe, Mn, Ni, Pb, Zn) was assessed in the fern after short-term treatments (1–3 days) over a range of wood distillate concentrations 1:300 (3.33 mL/L), 1:500 (2.00 mL/L), 1:700 (1.43 mL/L). A statistically significant accumulation of Cd, Cu, Mn, Pb, Zn (1:700) and Pb (1:300) was recorded after three days of incubation, despite the concentrations remained overall low. Using treatment vs. control ratios, a trend of increasing temporal uptake was detected for As, Ba, Fe, Mn, Pb (1:700); Mn, Pb (1:500), and only Pb at 1:300. The results suggested that, under the experimental conditions, element uptake is positively influenced by time and negatively by increasing concentrations of wood distillate, likely due to the acidification of the medium. On the whole, the element concentrations measured in A. filiculoides were low and did not pose any toxicological concern.

In summer 2016 a severe infestation of the alien ambrosia beetle Xylosandrus compactus was recorded from the Mediterranean maquis in the Circeo National Park in Central Italy. Trees and shrubs were infested and displayed wilting and necrosis of terminal branches caused by the combined impact of the insect and associated pathogenic fungi. A preliminary screening carried out on captured adults resulted in the isolation of a discrete number of fungal taxa with different life strategies, ranging from true mutualist (e.g. Ambrosiella xylebori) to plant pathogens (Fusarium spp.). In the present study, high-throughput sequencing was applied to determine the total diversity and functionality of the fungal community associated with X. compactus adults collected in the galleries of three Mediterranean woody hosts, Quercus ilex, Laurus nobilis, and Ceratonia siliqua. The effect of season and host in determining the composition of the associated fungal community was investigated. A total of 206 OTUs composed the fungal community associated with X. compactus. Eighteen OTUs were shared among the three hosts, including A. xylebori and members of the Fusarium solani complex. All but two were previously associated with beetles. Sixty-nine out of 206 OTUs were resolved to species level, identifying 60 different fungal species, 22 of which already reported in the literature as associated with beetles or other insects. Functional guild assigned most of the fungal species to saprotrophs and plant pathogens. Effects of seasonality and host on fungal community assemblage were highlighted suggesting the acquisition by the insect of new fungal taxa during the invasion process. The consequences of enriched fungal community on the risk of the insurgence of novel threatful insect–fungus association are discussed considering direct and indirect effects on the invaded habitat.

Forests play a key role in the climate system thanks to their large carbon uptake and storage. On the other hand, forests are vulnerable to climate extremes and pest attacks, causing early tree mortality which in turn could reduce their carbon uptake capacity.Early tree mortality is often associated to a complex interaction of predisposing stress factors (poor site quality, unfavourable stand conditions), inciting factors (frost, drought, mechanical damage) and contributing factors (fungi, insect borers).In this context, the aim of the present work was to investigate the processes underlying the tree mortality observed in an evergreen mixed forest stand dominated by Quercus ilex, located in the Circeo National Park (central Italy).The forest has the typical structure of an old-coppice not more managed (actual rotation time about 2 times that the normal), and was recently (2016) affected by an outbreak of Asian ambrosia beetle (Xylosandrus compactus) and Granulate ambrosia beetle (Xylosandrus crassiusculus) that caused an extensive trees crown browning. In 2019, plots were set in the area to monitor the beetle population dynamic and their impact on tree mortality. In each plot, species, dimension (DBH), stage of dieback, stem origin (resprouts after coppicing or from seed), presence of epicormic shoots and subcortical fungi stroma, were recorded for each woody plant.The plot survey revealed a high frequency of stems classified in a declining stage or dead, on average 42% of the standing stems, with significant differences among the species: 97%, 85%, 74% and 47% for Arbutus unedo, Quercus ilex, Phyllirea latifolia and Fraxinus ornus respectively.The higher stem mortality of Q. ilex was recorded in the smaller diameter classes, suggesting that the self-thinning process played an important role on the observed mortality as typical in the old not more managed coppices.To disentangle the role of the interruption of the management from the climatic and biological drivers, time trends on NDVI index were constrained with the duration of the summer dry seasons and comparing our forest with similar Q. ilex forest coppices in the region and regularly managed.Furthermore, the contribution of recent ambrosia beetles attack was assessed identifying the presence of twigs with signs of previous beetle attack on healthy, declining and dead plants.Our findings point towards complex tree mortality dynamics, in which the competition generated by the stand abandonment predisposed the forest to the insect attack, leading to the general decline of the forest stand.

The number of alien species arriving within new regions has increased at unprecedented rates. Managing the pathways through which alien species arrive and spread is important to reduce the threat of biological invasions. Harmonising information on pathways across individual sectors and user groups is therefore critical to underpin policy and action. The European Alien Species Information Network (EASIN) has been developed to easily facilitate open access to data of alien species in Europe. The Convention on Biological Diversity (CBD) Pathway Classification framework has become a global standard for the classification of pathways. We followed a structured approach to assign pathway information within EASIN for a subset of alien species in Europe, which covered 4169 species, spanning taxonomic groups and environments. We document constraints and challenges associated with implementing the CBD Pathway Classification framework and propose potential amendments to increase clarity. This study is unique in the scope of taxonomic coverage and also in the inclusion of primary (independent introductions to Europe) and secondary (means of dispersal for species expansion within Europe, after their initial introduction) modes of introduction. In addition, we summarise the patterns of introduction pathways within this subset of alien species within the context of Europe. Based on the analyses, we confirm that the CBD Pathway Classification framework offers a robust, hierarchical system suitable for the classification of alien species introduction and spread across a wide range of taxonomic groups and environments. However, simple modifications could improve interpretation of the pathway categories ensuring consistent application across databases and information systems at local, national, regional, continental and global scales. Improving consistency would also help in the development of pathway action plans, as required by EU legislation. European Commission ENV.B.2/SER/2015/0037rl COST Action CA17122 Czech Ministry of Environment 20-10349J, RVO 67985939 Natural Environment Research Council NE/R016429/1 Ministerio de Ciencia e Innovación PCI2018-092939, MCIU/AEI/FEDER

Since the discovery of penicillin in the first half of the last century, antibiotics have become the pillars of modern medicine for fighting bacterial infections. However, pathogens resistant to antibiotic treatment have increased in recent decades, and efforts to discover new antibiotics have decreased. As a result, it is becoming increasingly difficult to treat bacterial infections successfully, and we look forward to more significant efforts from both governments and the scientific community to research new antibacterial drugs. This perspective article highlights the high potential of bacterial transcriptional and posttranscriptional regulators as targets for developing new drugs. We highlight some recent advances in the search for new compounds that inhibit their biological activity and, as such, appear very promising for treating bacterial infections.

International audience; The number of alien plant pests and pathogens is rapidly increasing in many countries as a result of increasing trade, particularly the trade in living plants. Sentinel plantings in exporting countries to detect arthropod pests and agents of diseases prior to introduction provide information about the likelihood of introduction and the potential impact on plants native to the importing country. Such plantings can consist of species that are native to exporting or importing countries (in-patria and ex-patria plantings). In-patria plantings consist of young woody plants of species that are commonly exported and can be used to identify pests that may be introduced to new countries via the trade in live plants. Ex-patria plantings consist of exotic young or mature woody plants and surveys may provide information about potential impacts of pests if these were to become established in a new country. We discuss the methods and benefits of this powerful tool and list examples of studies that highlight the large number of unknown organisms and pest-host relationships that can be detected. The usefulness of sentinel plantings is illustrated using examples of arthropod pests and fungal pathogens of European and Asian tree species that were identified in sentinel studies in China and the Asian Russia.

In the 2015–2016 growing seasons, two novel symptoms were assessed on the crown of trees in orchards and coppices of chestnut groves in Central Italy. The first symptom was flagging of annual shoots with green leaves undergoing sudden wilt and turning brown later in the season. The second symptom consisted of leaves on annual shoots turning yellow before wilting in absence of flagging represented the second symptom. Samples were collected along transects in early summer, late summer and winter, and processed in the laboratory. The flagging symptom was associated in early summer with the presence of C. parasitica in cryptic dried buds on stems from the previous year's growth. The pathogen was also found in dormant buds in winter, suggesting that the infection could take place in summer during the Chinese gall wasp oviposition period. Cryphonectria parasitica was also isolated from abandoned galls in winter supporting the hypothesis that galls are a potential source of inoculum for crown infections. Aetiology of yellowing was not clarified and no fungal taxa were specifically associated with this symptom. Gnomoniopsis castanea, C. parasitica and, in early summer, Colletotrichum acutatum were the most abundant fungal taxa isolated from chestnut shoots and buds.

Prostate specific membrane antigen (PSMA) is a specific high frequency cell surface marker of prostate cancers. Theranostic approaches targeting PSMA show no major adverse effects and rule out off-tumor toxicity. A PSMA-retargeted oHSV (R-405) was generated which both infected and was cytotoxic exclusively for PSMA-positive cells, including human prostate cancer LNCaP and 22Rv1 cells, and spared PSMA-negative cells. R-405 in vivo efficacy against LLC1-PSMA and Renca-PSMA tumors consisted of inhibiting primary tumor growth, establishing long-term T immune response, immune heating of the microenvironment, de-repression of the anti-tumor immune phenotype, and sensitization to checkpoint blockade. The in situ vaccination protected from distant challenge tumors, both PSMA-positive and PSMA-negative, implying that it was addressed also to LLC1 tumor antigens. PSMA-retargeted oHSVs are a precision medicine tool worth being additionally investigated in the immunotherapeutic and in situ vaccination landscape against prostate cancers.

In this study we investigated the seasonal variations in the intracellular content of 14 trace elements (Al, As, Ba, Cd, Ce, Cr, Cu, Fe, Mn, Ni, Pb, Pd, Sb, Zn) and physiological parameters (namely chlorophylla, chlorophyllb, ergosterol, photosynthetic efficiency, cell membrane integrity) in the thalli of the lichenEvernia prunastri(L.) Ach. exposed to an urban environment (Siena, central Italy). Lichen thalli were collected before each exposure period from an unpolluted area and transplanted to 16 sites; every 3 months the thalli were retrieved and replaced with new ones. Exposed-to-control ratios of trace elements revealed a marked intracellular accumulation of Cd in summer and autumn, and of Sb in autumn and spring, possibly as a result of vehicular traffic pollution. However, considering the low absolute concentrations of these elements, the intracellular fraction of depositions may hardly have caused an impairment of physiological parameters. As a matter of fact, indicators of photobiont vitality (content of chlorophyllsaandband photosynthetic efficiency) did not show any fluctuation across seasons, while changes in the indicators of mycobiont vitality (cell membrane damage and ergosterol content) overall did reflect some seasonal changes and/or lichen growth.

Control of lichens on stone cultural heritage is mostly achieved by a combination of mechanical removal with biocide applications. However, there is a lack of scientific evidence on the efficacy of different biocides on different species, and on the consistency of biocide effects on heritage sites in different environmental conditions. This results in some uncertainty when conservation interventions to control lichens are routinely defined on the basis of restoration tradition or empirical evaluation, without experimental measures of how lichens respond. In this work, we quantitatively evaluated (a) the efficacy of five commercially-available biocides, applied using a brush or with a cellulose poultice, against two species (Protoparmeliopsis muralis, Verrucaria nigrescens), and (b) whether the effects on the two species were consistent, per treatment, across three Italian heritage sites. Lichen vitality was quantified through analyses of chlorophyll a fluorescence (ChlaF) and ergosterol content. The results indicated that all the tested biocides, and their organic solvents, affected the vitality of both the species. However, most of treatments displayed different efficacy on each species, across the different sites and between brush and poultice applications. Accordingly, when a conservation intervention to control lichen growth is planned, biocide treatments need both species- and site-specific calibrations and lichen vitality should be properly ascertained in situ by monitoring ChlaF parameters (FV/FM and F0) twenty days after trial biocide applications.

The liverwort Lunularia cruciata, known for being a species tolerant to pollution able to colonize urban areas, was collected in the town of Acerra (South Italy) to investigate the biological effects of air pollution in one of the three vertices of the so-called Italian Triangle of Death. The ultrastructural damages observed by transmission electron microscopy in specimens collected in Acerra were compared with samples collected in the city center of Naples and in a small rural site far from sources of air pollution (Riccia, Molise, Southern Italy). The biological response chain to air pollution was investigated considering vitality, photosynthetic efficiency, heat shock protein 70 (Hsp70) induction and gene expression levels, and chlorophyll degradation and related ultrastructural alterations. Particularly, a significant increment in Hsp70 expression and occurrence, and modifications in the chloroplasts' ultrastructure can be strictly related to the environmental pollution conditions in the three sites. The results could be interpreted in relation to the use of these parameters as biomarkers for environmental pollution.

Suppression of root rot diseases of ornamental plants is a potential benefit of formulating soilless container media with compost. A green compost (green nursery compost, GNC), obtained by a circular-economy approach from residues of pruning of woody plants and grass clippings during the nursery activities was analysed for its suppressiveness of root rot diseases using lavender. To this end, a bioassay was develop by formulating potting mixes containing GNC with two rates of peat substitution (25% and 50%) and infested with the root rot pathogens Sclerotinia sclerotiorum, Rhizoctonia solani and Phytophthora nicotianae. Contrasting results were obtained by using both substrates with a significant reduction of root rot by S. sclerotiorum, no effect on the containment of that by P. nicotianae, and an increase of symptoms caused by R. solani. The specific suppressiveness observed may be attributed to the colonisation of compost by specific groups of antagonistic microorganisms. This hypothesis was inv...

Starting from polluting waste, such as those produced by the olive oil industry, composting is able to produce stable organic materials that can be suggested as peat surrogate in the preparation of growth media for potted plants. In this work stable and mature olive mill waste compost (OMWC) was produced by using a simplified novel static composting procedure conducted in gas permeable bags. In order to predict its nutritional and suppressive potential, physico-chemical and biological characteristics of OMWC were investigated as well as its performance when used instead of peat in the preparation of growth media for Photinia fraseri cv. "Red Robin" potted plants. The innovative composting technique showed to be fully comparable to conventional methods of aerobic composting and able to convert OMWs into well-humified, stable, mature and rich in nutrients amendment. OMWC was colonized by different fungal species belonging to the genus Aspergillus and Penicillium that showed an appreciable suppressive potential against Sclerotinia sclerotiorum and Phytophthora cactorum. OMWC also resulted suitable for nursery cultivation of P. fraseri, showing the best performance when used at 33% and 66% peat substitution rate. The recycling method proposed represents a virtuous and low cost model for proper and successful management of olive mill waste.

In September 2016, a survey conducted in the Circeo National Park revealed an outbreak and serious damage caused by the black twig borer (Xylosandrus compactus) and its associated fungi in the Mediterranean maquis. Among the affected hosts, Quercus ilex, Viburnum tinus, Ruscus aculeatus, Pistacia lentiscus, Laurus nobilis and Ceratonia siliqua, showed flagging and wilting of branches and, in younger individuals, the death of the whole plant occurred. In total, 18 different fungal taxa were found associated with the insect. These included Ambrosiella xylebori, Geosmithia pallida, Fusarium spp., Epicoccum nigrum and Bionectria sp. This is the first report in Europe of X. compactus and associated ambrosia fungi in a natural environment.

Gnomoniopsis castanea has been reported as the causal agent of necrosis of chestnut wasp (Dryocosmus kuriphilus) galls. The fungus is frequently observed on galls in chestnut stands infested by the insect in Italy. In the present study the impact of gall necrosis and the dynamic of its development have been studied in mature and young Castanea sativa stands in Central Italy during spring and early summer, before the D. kuriphilus adult flies. Results suggest that gall necrosis develops from resident endophytic inoculum of G. castanea. During the 2 y of monitoring, no differences were found in incidence and severity of the disease. Gall necrosis increased exponentially during the season, reaching 75,4% of galls totally necrotized in the investigated site in mid July. Gall necrosis was shown to have a severe impact on D. kuriphilus vitality, mostly impacting the adults inside the galls. Gall necrosis by G. castanea appears to efficiently control gall wasp in chestnut stands, although the high virulence of the fungus to chestnut fruits precludes its use as biocontrol agent in biological control strategies.