Allergenicity at component level of sub-pollen particles from different sources obtained by osmolar shock: A molecular approach to thunderstorm-related asthma outbreaks.

BACKGROUND: The so-called "thunderstorm asthma" (TA) is an uncommon but dramatic outbreak of asthma attacks occurring during a thunderstorm in the pollen and moulds season. Mechanisms which make the pollen able to enter the deeper airways and provoke severe asthma symptoms are still unclear. OBJECTIVE: To test the hypothesis that sub-pollen particles (SPPs) originated from the rupture by an osmotic shock of pollen associated with TA contain allergens. METHODS: After hydration, SPPs released from pollen grains of grass, pellitory, olive, cypress, ragweed and birch were isolated and determined by microscopy. Allergens were determined by in vitro ELISA inhibition tests indirectly using the sera from 10 polyreactive patients. An inhibition <50% was considered as negative, 50%-75% moderate and > 75% complete. RESULTS: The inhibition experiments showed that the SPPs from birch and cypress were unable to inhibit serum IgE reactivity to Bet v 1 and Cup a 1, respectively. Ragweed SPPs inhibited ragweed pollen extract and Amb a 1 by 75.8 ± 0.11% and 81.2 ± 0.15%, respectively. Olive and pellitory SPPs retained almost the whole IgE-binding capability in all cases tested. Grass SPPs inhibited 32 ± 0.06% of Lolium perenne Lol p 1 and 65% of Phleum pratense extracts, but results were highly variable for individual allergens (97.5%-0.03% for Phl p 2, 45.3 ± 0.12% for Phl p 5, 24.7 ± 0.22% for Phl p 6, and 38.3 ± 0.2% for Phl p 1). CONCLUSIONS: Inhibition experiments confirm the hypothesis that SSPs obtained after the osmotic shock of pollen involved in TA, namely grass, pellitory and olive tree pollen, contain allergens and therefore they can induce severe asthma attacks during thunderstorms.

Ambrosia artemisiifolia L. temperature-responsive traits influencing the prevalence and severity of pollinosis: a study in controlled conditions.

BACKGROUND: Ambrosia artemisiifolia L. is one of the most important sources of allergenic pollen in many regions of the world. Its health impact increased over the last decades and is expected to further increase in consequence of climate change. However little information is available on the specific role played by temperature on allergy rising. The aim of this work was to evaluate the effect of temperature on A. artemisiifolia growth, flowering and pollen allergenicity, the major plant functional traits influencing the prevalence and severity of pollinosis. RESULTS: Plants were grown in controlled conditions at three thermal regimes: "Low" (LT: 18-14 °C light-dark), "Intermediate" (IT: 24-20 °C light-dark) and "High" (HT: 30-26 °C light-dark). During plant development, plant vegetative and reproductive morpho-functional traits were measured and, at the end of plant life-cycle, mature pollen was collected and analyzed for its allergenic properties by slot blot, 1D- and 2D-western blot (by using a pool of sera from ragweed-allergic patients) and liquid chromatography-tandem mass spectrometry. A. artemisiifolia showed a great development plasticity leading to a broad temperature tolerance. Shoot architecture, growth rate, number of male inflorescence and pollen allergenicity were temperature-responsive traits. Pollen allergenicity increased in parallel with temperature and differences were related to allergen synthesis and Amb a 1-IgE-binding. Flavonoids whose concentration in pollen decreased with the increase of temperature, were recognized as the cause of the negligible Amb a 1-IgE binding in LT pollen. CONCLUSIONS: Results show that temperature governs plant development and pollen allergenicity influencing the temporal and spatial magnitude of subject exposure to allergens.

Ecological filtering and plant traits variation across quarry geomorphological surfaces: implication for restoration.

Revegetation patterns after quarry abandonment have been widely studied from several ecological points of view, but a trait-based approach is still lacking. The aim of this study was to characterise the plant species assemblages and the associated functional traits filtered on different geomorphological surfaces in abandoned limestone quarry areas: artificial cliffs, embankments, and platforms. We then verified if species with certain traits were better able to overcome the dispersal and environmental filters necessary for establishment. To this aim, we analyzed 113 vegetation plots and collected data on 25 morphological, ecological, and dispersal traits to detect species adaptaions across these man-made environments. As a case study, we investigated the extraction basin of Botticino (Lombardy, Italy), the second largest in Italy. The results obtained by SIMPER and CCA analyses showed that rockiness, stoniness, slope, elevation, and time of surfaces are the main filters that varied across quarries and affected plant assemblages at the macro-scale level. Across the three geomorphological surfaces (meso-scale) of quarries, more specific abiotic filters selecting species were found. In turn, traits differentiation according to the three main geomorphological surfaces of quarry emphasized that further filters acting at the micro-scale imply differences in dispersal mechanisms and resource availability. This work highlighted the utility to study species assemblages and environmental filters to address quarry restoration according to the type of geomorphological surface. The investigation of some traits (chorological form, life forms, seed dispersal,s and plant height) can furnish some interesting indications for practice individuating further abiotic filters acting at the micro-scale.

Concomitant sensitization to ragweed and mugwort pollen: who is who in clinical allergy?

BACKGROUND: In many areas of Europe, double sensitization to ragweed and mugwort is common, and because of the overlapping flowering periods of the 2 plants, it is not possible to diagnose the primary sensitizing allergen source and hence to determine the proper immunotherapy. OBJECTIVES: To elucidate whether double-sensitized patients are cosensitized or cross-sensitized and, in the latter case, to define the primary sensitizer. METHODS: Serum samples from 34 patients with late summer respiratory allergy underwent skin prick testing with whole ragweed, and mugwort extracts were analyzed for their reactivity to recombinant Art v 1 and Amb a 1 by ImmunoCAP and then to Amb a 1, Art v 6, and Art v 1 isoforms by a proteomic approach. In double reactors, the primary sensitizing sources were detected by inhibition experiments. RESULTS: Serum samples from patients monosensitized to ragweed contained IgE to epitopes specific of all Amb a 1 isoforms. In contrast, serum samples from double reactors found to be primarily sensitized to mugwort reacted to Art v 1 and Art v 6 and cross-reacted to a few Amb a 1 isoforms. Finally, serum samples from double reactors found to be primarily sensitized to ragweed contained IgE reacting to all Amb a 1 isoforms, part of which cross-reacted to Art v 6. We did not find cosensitized patients. CONCLUSION: This study found that Art v 6 plays an important role in mugwort allergy and that the cross-reactivity between Art v 6 and Amb a 1 is frequent, bidirectional, and clinically relevant in the area of Milan.

The rotation of white lupin (Lupinus albus L.) with metal-accumulating plant crops: a strategy to increase the benefits of soil phytoremediation.

Most of the plants employed to remove metals from contaminated soils are annuals and have a seed-to-seed life cycle of a few months, usually over spring and summer. Consequently, for most of the year, fields are not actively cleaned but are completely bare and subject to erosion by water and wind. The objective of this study was to evaluate the benefits of using Lupinus albus as a winter crop in a rotation sequence with a summer crop ideally selected for phytoextraction, such as industrial hemp. Lupin plants were grown in two alkaline soil plots (heavy metal-contaminated and uncontaminated) of approximately 400 m(2) each after the cultivation and harvest of industrial hemp. A smaller-scale parallel pot experiment was also performed to better understand the lupin behavior in increasing concentrations of Cd, Cu, Ni and Zn. White lupin grew well in alkaline conditions, covering the soil during the winter season. In few months plants were approximately 40-50 cm high in both control and contaminated plots. In fields where the bioavailable fraction of metals was low (less than 12%), plants showed a high tolerance to these contaminants. However, their growth was affected in some pot treatments in which the concentrations of assimilable Cu, Zn and Ni were higher, ranging from approximately 40-70% of the total concentrations. The lupin's ability to absorb heavy metals and translocate them to shoots was negligible with respect to the magnitude of contamination, suggesting that this plant is not suitable for extending the period of phytoextraction. However, it is entirely exploitable as green manure, avoiding the application of chemical amendments during phytoremediation. In addition, in polluted fields, white lupin cultivation increased the soil concentration of live bacteria and the bioavailable percentage of metals. On average live bacteria counts per gram of soil were 65×10(6)±18×10(6) and 99×10(6)±22*10(6) before and after cultivation, respectively. The percentages of bioavailable Cu, Pb, Ni, Zn and Cr, which were 5.7±0.7, 5.3±1.7, 1.2±0.1, 12±1.5 and 0.1±0.02%, respectively, before lupin growth, increased to 9.6±1.6, 7±2, 2±0.3, 14±1.5 and 0.1±0.02% after lupin harvest. On the whole, our results indicate that the winter cultivation of white lupin in sequence with a metal-accumulator summer crop can improve the recovery of soil quality during the phytoextraction period. It improves the safety of the area, limiting additional ecological and human health problems, and enhances soil health by avoiding the use of chemical amendments and by increasing the levels of viable microorganisms.

Analysis of Petrogenic Hydrocarbons in Plant Tissues: A Simple GC-MS-Based Protocol to Distinguish Biogenic Hydrocarbons from Diesel-Derived Compounds.

Diesel contamination of farming soils is of great concern because hydrocarbons are toxic to all forms of life and can potentially enter the food web through crops or plants used for remediation. Data on plant ability to uptake, translocate and accumulate diesel-derived compounds are controversial not only due to the probable diverse attitude of plant species but also because of the lack of a reliable method with which to distinguish petrogenic from biogenic compounds in plant tissues. The purpose of this study was to set up a GC-MS-based protocol enabling the determination of diesel-derived hydrocarbons in plants grown in contaminated soil for assessing human and ecological risks, predicting phytoremediation effectiveness and biomass disposal. To this end, two plant species, Vicia sativa L. and Secale cereale L., belonging to two diverse vascular plant families, were used as plant models. They were grown in soil spiked with increasing concentrations of diesel fuel, and the produced biomass was used to set up the hydrocarbon extraction and GC-MSD analysis. The developed protocol was also applied to the analysis of Typha latifolia L. plants, belonging to a different botanical family and grown in a long-time and highly contaminated natural soil. Results showed the possibility of distinguishing diesel-derived compounds from biogenic hydrocarbons in most terrestrial vascular plants, just considering the total diesel compounds in the n-alkanes carbon range C10-C26, where the interference of biogenic compounds is negligible. Diesel hydrocarbons quantification in plant tissues was strongly correlated (0.92 < r2 < 0.99) to the concentration of diesel in spiked soils, suggesting a general ability of the considered plant species to adsorb and translocate relatively low amounts of diesel hydrocarbons and the reliability of the developed protocol.

Ragweed is in the Air: Ambrosia L. (Asteraceae) and Pollen Allergens in a Changing World.

Globally Ambrosia species (Asteraceae), commonly called ragweed, are recognized to be one of the most problematic groups of invasive weeds and one of the main allergenic genus. Climate and land-use change and air pollution are expected to promote ragweed spread, increase airborne ragweed pollen concentrations (the source of allergens), extend the pollen season, and promote longdistance transport of pollen or sub-pollen particles containing allergens. The allergenicity of pollen itself is going to increase. Likely, all these factors will have meaningful effects in the exacerbation of the sensitization to ragweed pollen and the severity of allergy symptoms. Globally the major health concern regards A. artemisiifolia, because of its very wide global distribution and highly invasive behavior. Together with A. artemisiifolia, also A. trifida and A. psilostachya are species of health concern distributed across different continents, widespread and invasive in several regions. The present review summarizes the characteristics of these species and gives an overview of factors contributing to their allergenicity.

Does Caulerpa prolifera with Its Bacterial Coating Represent a Promising Association for Seawater Phytoremediation of Diesel Hydrocarbons?

Anthropic diesel-derived contamination of Mediterranean coastal waters is of great concern. Nature-based solutions such as phytoremediation are considered promising technologies to remove contaminants from marine environments. The aim of this work was to investigate the tolerance of the Mediterranean autochthonous seaweed Caulerpa prolifera (Forsskal) Lamouroux to diesel fuel and its hydrocarbon degradation potential. Changes in C. prolifera traits, including its associated bacterial community abundance and structure, were determined by fluorescence microscopy and next-generation sequencing techniques. Thalli of C. prolifera artificially exposed to increasing concentration of diesel fuel for 30 days and thalli collected from three natural sites with different levels of seawater diesel-derived hydrocarbons were analysed. Gas chromatography was applied to determine the seaweed hydrocarbon degradation potential. Overall, in controlled conditions the lower concentration of diesel (0.01%) did not affect C. prolifera survival and growth, whereas the higher concentration (1%) resulted in high mortality and blade damages. Similarly, only natural thalli, collected at the most polluted marine site (750 mg L-1), were damaged. A higher abundance of epiphytic bacteria, with a higher relative abundance of Vibrio bacteria, was positively correlated to the health status of the seaweed as well as to its diesel-degradation ability. In conclusion, C. prolifera tolerated and degraded moderate concentrations of seawater diesel-derived compounds, especially changing the abundance and community structure of its bacterial coating. The protection and exploitation of this autochthonous natural seaweed-bacteria symbiosis represents a useful strategy to mitigate the hydrocarbon contamination in moderate polluted Mediterranean costal environments.

China-US grain trade shapes the spatial genetic pattern of common ragweed in East China cities.

Common ragweed is an invasive alien species causing severe allergies in urban residents. Understanding its urban invasion pathways is crucial for effective control. However, knowledge is limited, with most studies focusing on agricultural and natural areas, and occurrence record-based studies exhibiting uncertainties. We address this gap through a study in East China cities, combining population genetics and occurrence records. Leaf samples from 37 urban common ragweed populations across 15 cities are collected. Genomic and chloroplast DNA extraction facilitate analysis of spatial genetic patterns and gene flows. Additionally, international grain trade data is examined to trace invasion sources. Results indicate spatial genetic patterns impacted by multiple introductions over time. We infer the modern grain trade between the United States and China as the primary invasion pathway. Also, cities act as transportation hubs and ports of grain importation might disperse common ragweed to urban areas. Invasive species control should account for cities as potential landing and spread hubs of common ragweed.

Overexpression of PhEXPA1 increases cell size, modifies cell wall polymer composition and affects the timing of axillary meristem development in Petunia hybrida.

• Expansins are cell wall proteins required for cell enlargement and cell wall loosening during many developmental processes. The involvement of the Petunia hybrida expansin A1 (PhEXPA1) gene in cell expansion, the control of organ size and cell wall polysaccharide composition was investigated by overexpressing PhEXPA1 in petunia plants. • PhEXPA1 promoter activity was evaluated using a promoter-GUS assay and the protein's subcellular localization was established by expressing a PhEXPA1-GFP fusion protein. PhEXPA1 was overexpressed in transgenic plants using the cauliflower mosaic virus (CaMV) 35S promoter. Fourier transform infrared (FTIR) and chemical analysis were used for the quantitative analysis of cell wall polymers. • The GUS and GFP assays demonstrated that PhEXPA1 is present in the cell walls of expanding tissues. The constitutive overexpression of PhEXPA1 significantly affected expansin activity and organ size, leading to changes in the architecture of petunia plants by initiating premature axillary meristem outgrowth. Moreover, a significant change in cell wall polymer composition in the petal limbs of transgenic plants was observed. • These results support a role for expansins in the determination of organ shape, in lateral branching, and in the variation of cell wall polymer composition, probably reflecting a complex role in cell wall metabolism.

The Arabidopsis BET bromodomain factor GTE4 is involved in maintenance of the mitotic cell cycle during plant development.

Bromodomain and Extra Terminal domain (BET) proteins are characterized by the presence of two types of domains, the bromodomain and the extra terminal domain. They bind to acetylated lysines present on histone tails and control gene transcription. They are also well known to play an important role in cell cycle regulation. In Arabidopsis (Arabidopsis thaliana), there are 12 BET genes; however, only two of them, IMBIBITION INDUCIBLE1 and GENERAL TRANSCRIPTION FACTOR GROUP E6 (GTE6), were functionally analyzed. We characterized GTE4 and show that gte4 mutant plants have some characteristic features of cell cycle mutants. Their size is reduced, and they have jagged leaves and a reduced number of cells in most organs. Moreover, cell size is considerably increased in the root, and, interestingly, the root quiescent center identity seems to be partially lost. Cell cycle analyses revealed that there is a delay in activation of the cell cycle during germination and a premature arrest of cell proliferation, with a switch from mitosis to endocycling, leading to a statistically significant increase in ploidy levels in the differentiated organs of gte4 plants. Our results point to a role of GTE4 in cell cycle regulation and specifically in the maintenance of the mitotic cell cycle.

High Phenotypic Plasticity in a Prominent Plant Invader along Altitudinal and Temperature Gradients.

Studies on plant growth and trait variation along environmental gradients can provide important information for identifying drivers of plant invasions and for deriving management strategies. We used seeds of the annual plant invader Ambrosia artemisiifolia L. (common ragweed) collected from an agricultural site in Northern Italy (226 m. a.s.l; Mean Annual Air Temperature: 12.9 °C; precipitations: 930 mm) to determine variation in growth trajectories and plant traits when grown along a 1000-m altitudinal gradient in Northern Italy, and under different temperature conditions in the growth chamber (from 14/18 °C to 26/30 °C, night/day), using a non-liner modeling approach. Under field conditions, traits related to plant height (maximum height, stem height, number of internodes) followed a three-parameter logistic curve. In contrast, leaf traits (lateral spread, number of leaves, leaf length and width) followed non-monotonic double-Richards curves that captured the decline patterns evident in the data. Plants grew faster, reaching a higher maximum plant height, and produced more biomass when grown at intermediate elevations. Under laboratory conditions, plants exhibited the same general growth trajectory of field conditions. However, leaf width did not show the recession after the maximum value shown by plants grown in the field, although the growth trajectories of some individuals, particularly those grown at 18 °C, showed a decline at late times. In addition, the plants grown at lower temperatures exhibited the highest value of biomass and preserved reproductive performances (e.g., amount of male inflorescence, pollen weight). From our findings, common ragweed exhibits a high phenotypic plasticity of vegetative and reproductive traits in response to different altitudes and temperature conditions. Under climate warming, this plasticity may facilitate the shift of the species towards higher elevation, but also the in situ resistance and (pre)adaptation of populations currently abundant at low elevations in the invasive European range. Such results may be also relevant for projecting the species management such as the impact by possible biocontrol agents.

Integrated biological and chemical characterisation of a pair of leonardesque canal lock gates.

The Museo Nazionale della Scienza e della Tecnologia "Leonardo da Vinci" in Milan is exposing two pairs of canal lock gates, used to control the water flow in Milan canal system, whose design appears in the Leonardo's Codex Atlanticus. The wood present in the gates has been deeply characterised by mean of a multidisciplinary investigation involving i) DNA barcoding of wood fragments; ii) microbial community characterisation, and iii) chemical analyses. DNA barcoding revealed that two fragments of the gates belonged to wood species widely used in the middle age: Fagus sylvatica and Picea abies. The chemical characterisations were based on the use of ionic liquid as dissolving medium in order to analyse the entire cell wall material by means of Gel Permeation Chromatography (GPC) and 2D-NMR-HSQC techniques. This multidisciplinary analytical approach was able to highlight the complex nature of the degradation occurred during the gate operation (XVI-XVIII centuries): an intricate interplay between microbial populations (i.e. Shewanella), inorganic factors (i.e. iron from nails), physical factors and the lignocellulosic material.

Seeding and Overseeding Native Hayseed Support Plant and Soil Arthropod Communities in Agriculture Areas.

Using native seed mixtures to create or recover grassland habitats in rotation to crops or in strips surrounding fields is considered a cost-effective practice to enhance ecosystem resilience and agro-biodiversity. The aim of this research was to assess the effects of native hayseed mixtures on plant and microarthropod communities in an agricultural area of Northern Italy. Three different experimental treatments were set up. The first was a control (C) (i.e., non-seeded plots left to spontaneous vegetation succession after ploughing no deeper than 15 cm). The second, hayseed seeded (Hs) after ploughing no deeper than 15 cm. The third experimental treatment was hayseed overseeded (Ov) on the resident plant community after only a superficial harrowing. Ov plots exhibited the preeminent positive effects on the total productivity and quality of the grassland in terms of total vegetation cover, cover and richness of typical grassland species (i.e., Molinio-Arrhenatheretea species), and cover of legumes, grasses and perennial species. Moreover, Ov sites exhibited the highest abundance of microarthropod taxa and soil biological quality (QBS-ar) but only in spring, when the disturbance of ploughing negatively affected Hs and C plots. On the other hand, Hs sites showed a great reduction of invasive alien (i.e., Ambrosia artemisiifolia and Artemisia verlotiorum) and segetal weed species (i.e., Capsella bursa-pastoris and Spergula arvensis) in terms of cover. This study provides valuable indication on using hayseed mixtures to create grassland habitats as reservoir of native flora and soil biodiversity in agriculture areas.

The interactive effect of herbivory, nutrient enrichment and mucilage on shallow rocky macroalgal communities.

This paper focuses on the interactive short and long-term effect of three different stressors on a macroalgal assemblage. Three stressors are considered: herbivory, nutrients and mucilage. The experiment was conducted in Tavolara Punta Coda Cavallo Marine Protected Area (Mediterranean Sea) during a bloom of the benthic mucilage-producing microalga Chrysophaeum taylorii (Pelagophyceae); this microalga is recently spreading in the Mediterranean Sea. On a rocky substratum, 36 plots 20 × 20 cm in size were prepared. Factorial combinations of three experimental treatments were applied in triplicate, including three grazing levels crossed with two nutrient enrichment and two mucilage removal treatments. Significant differences were observed among treatments 8 weeks later, at the end of summer. In particular, dark filamentous algae were more abundant in all enriched plots, especially where mucilage and macroalgae had been removed; a higher percent cover of crustose coralline algae was instead observed where nutrients had been increased and no grazing pressure acted. Furthermore, the abundance of Dictyota spp. and Laurencia spp. was significantly higher in enriched mucilage-free plots where the grazing pressure was null or low. However, the effects of the treatments on the overall assemblage of the macroalgal community were not long persistent (36 weeks later). These results illustrate the capacity of a shallow-water macroalgal community to quickly recover from the simultaneous impacts of herbivory, nutrient enrichment, and mucilage.

Resolution of Viable and Membrane-Compromised Free Bacteria in Aquatic Environments by Flow Cytometry.

In aquatic environments, free heterotrophic bacteria play an extremely important role due to their high biomass, wide panel of metabolisms, and ubiquity, as well as the toxicity of certain species. This unit presents a nucleic-acid double-staining protocol (NADS) for flow cytometry that can distinguish fractions of viable, damaged, or membrane-compromised cells within the free-bacterial community. The NADS protocol is based on the simultaneous utilization of two nucleic acid stains-membrane-permeant SYBR Green and membrane-impermeant propidium iodide (PI). The efficiency of the double staining on fresh samples is magnified by the FRET from SYBR Green to PI when both are bound to the nucleic acids. Full quenching of SYBR Green fluorescence by PI identifies cells with a compromised membrane, partial quenching indicates cells with a slightly damaged membrane, and lack of quenching characterizes cells with an intact membrane. Samples do not require any pretreatment and this protocol can be performed almost anywhere. © 2018 by John Wiley & Sons, Inc.

Effect of Soil pH on the Growth, Reproductive Investment and Pollen Allergenicity of Ambrosia artemisiifolia L.

Despite the importance of soil reaction for alien plant establishment, few and incomplete studies have included this key factor so far. In this study, we investigated the effects of soil pH on the germination, growth (plant height, width, dry weight, etc.) and reproductive investment (inflorescence size and n° of flowers) of Ambrosia artemisiifolia (common ragweed), an allergenic species that is highly invasive and alien in Europe, through a replicated experiment in controlled conditions. In addition, we determined if soil pH has an effect on the total pollen allergenicity of the species. After preliminary germination tests on agar at different pH (from pH4 to pH8), plants were grown in natural soils with pH values of 5 (acid), 6 (sub-acid) and 7 (neutral) obtained by modifying a natural soil by liming methods (calcium hydroxide solution). Results showed that plants grown at pH7 were shorter and developed leaves at a slower rate than those grown at pH5 and pH6; plants grown at pH7 did not produce flowers and pollen. We also observed that, at pH5 and pH6, larger plants (as assessed by the dry weight of the aerial biomass) had both larger and more numerous inflorescences and emitted pollen earlier. Finally, the IgE-binding signal was higher in pollen samples collected from plants grown at pH5 (Integrated Optical Density, IOD, range: 1.12-1.25) than in those grown at pH6 (IOD range: 0.86 -1.03). Although we acknowledge the limitations of only testing the effects of pH in controlled conditions, this study suggests that soil pH greatly affects the growth and development of A. artemisiifolia and indicates that it may have a role in limiting the distribution and hazardousness of this plant. Future field tests should therefore assess the effectiveness of liming in the management and control of ragweed and other alien species.

Direct effects of insecticides on common ragweed-implications for natural enemy exclusion trials.

Experimentally applying pesticides is an important method to assess the efficacy of weed biocontrol agents, but potential direct effects of the chemicals on plant performance are controversial or unknown. We assessed how three broad-spectrum insecticides applied in combination affect the performance of the widely invasive, crop-yield reducing, allergenic common ragweed (Ambrosia artemisiifolia L.) in an insect-free environment. Spraying insecticides had no significant effects on aboveground dry weight, seed and pollen output or pollen allergenicity, and only explained 1-8% of variation in these parameters. Our insecticide treatment can hence be applied to assess biocontrol impact on biomass and reproductive output of common ragweed. As our insecticide treatment delayed senescence, however, other methods of insect exclusion should be preferred when studying common ragweed phenology.

Genetic differentiation, local adaptation and phenotypic plasticity in fragmented populations of a rare forest herb.

BACKGROUND: Due to habitat loss and fragmentation, numerous forest species are subject to severe population decline. Investigating variation in genetic diversity, phenotypic plasticity and local adaptation should be a prerequisite for implementing conservation actions. This study aimed to explore these aspects in ten fragmented populations of Physospermum cornubiense in view of translocation measures across its Italian range. METHODS: For each population we collected environmental data on landscape (habitat size, quality and fragmentation) and local conditions (slope, presence of alien species, incidence of the herbivorous insect Metcalfa pruinosa and soil parameters). We measured vegetative and reproductive traits in the field and analysed the genetic population structure using ISSR markers (STRUCTURE and AMOVA). We then estimated the neutral (FST) and quantitative (PST) genetic differentiation of populations. RESULTS: The populations exhibited moderate phenotypic variation. Population size (range: 16-655 individuals), number of flowering adults (range: 3-420 individuals) and inflorescence size (range: 5.0-8.4 cm) were positively related to Mg soil content. Populations' gene diversity was moderate (Nei-H = 0.071-0.1316); STRUCTURE analysis identified five different clusters and three main geographic groups: upper, lower, and Apennine/Western Po plain. Fragmentation did not have an influence on the local adaptation of populations, which for all measured traits showed PST < FST, indicating convergent selection. DISCUSSION: The variation of phenotypic traits across sites was attributed to plastic response rather than local adaptation. Plant translocation from suitable source populations to endangered ones should particularly take into account provenance according to identified genetic clusters and specific soil factors.

Molecular evidence for benzo[a]pyrene and naphthalene genotoxicity in Trifolium repens L.

Polycyclic aromatic hydrocarbons (PAHs) are among the most dangerous environmental contaminants due to their toxic, carcinogenic and mutagenic effects. Although there are many data in literature that detail the effects of PAHs on animals, little is known about their action on higher plants which are often used as bioindicators. The aim of the present study was to evaluate the genotoxicity of two different PAHs, benzo[a]pyrene (BaP) and naphthalene (Naph), on Trifolium repens L. Clover plants were exposed to soil which had been artificially contaminated with three concentrations of BaP (5, 10 and 20 microg g-1) or Naph (25, 50 and 100 microg g-1). After 15 days, changes in the DNA content and sequence of roots and shoots were evaluated by flow cytometry (FCM) and amplified fragment length polymorphism (AFLP). Root and shoot dry weight were also determined to assess plant growth. Results showed that BaP and Naph were both genotoxic for white clover, inducing significant changes in root and shoot DNA sequence. Damage was more severe in the root than in the shoot suggesting that the translocation of these compounds and their genotoxic metabolites was limited. Ploidy alterations were not detected and the extent of damage caused by all the tested PAH concentrations was not sufficient to affect plant development.