S. Burruano

Fungal Assemblages Associated with Roots of Halophytic and Non-halophytic Plant Species Vary Differentially Along a Salinity Gradient

Structure of fungal communities is known to be influenced by host plants and environmental conditions. However, in most cases, the dynamics of these variation patterns are poorly understood. In this work, we compared richness, diversity, and composition between assemblages of endophytic and rhizospheric fungi associated to roots of two plants with different lifestyles: the halophyte Inula crithmoides and the non-halophyte I. viscosa (syn. Dittrichia viscosa L.), along a spatially short salinity gradient. Roots and rhizospheric soil from these plants were collected at three points between a salt marsh and a sand dune, and fungi were isolated and characterized by ITS rDNA sequencing. Isolates were classified in a total of 90 operational taxonomic units (OTUs), belonging to 17 fungal orders within Ascomycota and Basidiomycota. Species composition of endophytic and soil communities significantly differed across samples. Endophyte communities of I. crithmoides and I. viscosa were only similar in the intermediate zone between the salt marsh and the dune, and while the latter displayed a single, generalist association of endophytes, I. crithmoides harbored different assemblages along the gradient, adapted to the specific soil conditions. In the lower salt marsh, root assemblages were strongly dominated by a single dark septate sterile fungus, also prevalent in other neighboring salt marshes. Interestingly, although its occurrence was positively correlated to soil salinity, in vitro assays revealed a strong inhibition of its growth by salts. Our results suggest that host lifestyle and soil characteristics have a strong effect on endophytic fungi and that environmental stress may entail tight plant–fungus relationships for adaptation to unfavorable conditions.

A cytological and ultrastructural study on the maturation and germination of oospores of Plasmopara viticola from overwintering vine leaves

Observations on cytological and ultrastructural changes in Plasmopara viticola oospores were carried out during the overwintering period. Three types of oospores were observed. Type I, characterized by a thin inner oospore wall (IOW), large lipid globules and two nuclei, was recovered only in samples collected in October. These oospores were considered to be immature. Maturation occurred during November and involved a noticeable increase in thickness of the IOW, fusion of nuclei, formation of an ooplast and break up of large lipid globules into smaller ones (type II oospores). A few oospores (type III) showed abnormal organization with very large lipid globules and less frequently discernible nuclei. IOW solubilization, dissolution of the ooplast and lipid globules and nuclear division were the first detectable events during oospore germination. Germinating oospores produce a germ tube which was terminated by a sporangium. In its young stage, the sporangium had a thick wall and an unusual multi-layered membrane. During this phase, nuclear divisions took place in the sporangium. While sporangium development progressed, the ribosome density in the cytoplasm decreased and mitochondria, initially roundish with evident cristae, became their usual tubular profile. The plasma membrane had a typical structure and storage organelles, such as finger print vacuoles and lipid globules, became more numerous in the cytoplasm. Larger vacuoles contained the flagella of differentiating zoospores.

Presence of endophytic bacteria in Vitis vinifera leaves as detected by fluorescence in situ hybridization

Fluorescence in situ hybridization (FISH) in combination with confocal laser scanning microscopy (CLSM) was applied to detect and localize bacterial colonies in leaf tissues of Vitis vinifera. Leaves were cleared to minimize the autofluorescence of plant fragments. The use of fluorescently labeled bacterial probe EUB338 on discolored grapevine leaf disks allowed the estimation of the spatial distribution of different bacterial colonies. In particular, bacterial colonies were found in veins, cells, hairs, intercellular spaces, and in cut edges of leaf disks of both non-Acremonium byssoides-colonized and A. byssoides-colonized leaves of five different cultivars. Furthermore, CLSM confirmed that bacteria were located in different layers of the leaf tissue. At the same time, one cleared disk of each foliar sample was crushed and plated on Plate Count Agar to isolate cultivable endophytic bacteria. Some of these microorganisms were presumptively identified, by partial 16S rRNA gene sequencing, as Burkholderia phytofirmans, thus suggesting their potential role as plant growth promoters and sources of resistance against pathogenic agents, such as fungi.

Pathogenicity bioassays of isolates of Beauveria bassiana on Rhynchophorus ferrugineus

BACKGROUND The control of Rhynchophorus ferrugineus (Olivier), the main palm pest in the Mediterranean Basin, is problematic because of its biology and the current restrictions in many European countries on the use of chemical insecticides in urban areas. Entomopathogenic fungi have been studied as potential biological control agents, but information on their natural incidence is limited. Strains of Beauveria bassiana (Balsamo) Vuillemin were isolated from symptomatic insects collected on dead palms, and their pathogenicity against different instars of R. ferrugineus was evaluated in the laboratory. RESULTS The overall percentage of infected insects found in Canary palms was 7%. In laboratory bioassays, hatching of eggs treated with three different isolates of B. bassiana was 41.2, 26.8 and 29.9%, significantly lower than the control (62.4%). Larvae and adults were treated with a single isolate in two ways: spraying each insect with a conidial suspension or feeding them with fruit portions previously immersed in the same conidial suspension. At the end of the two trials, the mortality of treated larvae was 88 and 92%, and the mean survival time was 10.4 and 11.8 days, significantly different from the control, where no insect died during the trials. Mortality and survival time recorded in either trial on adults did not significantly differ between treatment and control. CONCLUSION This study shows that the pathogenicity of wild isolates of B. bassiana differs among the tested R. ferrugineus instars. The low mortality of treated adults supports their use as vectors of B. bassiana as a potential tool for reducing R. ferrugineus populations.

Lasiolactols A and B Produced by the Grapevine Fungal Pathogen Lasiodiplodia mediterranea

A strain of Lasiodiplodia mediterranea, a fungus associated with grapevine decline in Sicily, produced several metabolites in liquid medium. Two new dimeric γ‐lactols, lasiolactols A and B (1 and 2), were characterized as (2S*,3S*,4R*,5R*,2′S*,3′S*,4′R*,5′R*)‐ and (2R*,3S*,4R*,5R*,2′R*,3′S*,4′R*,5′R*)‐(5‐(4‐hydroxymethyl‐3,5‐dimethyl‐tetrahydro‐furan‐2‐yloxy)‐2,4‐dimethyl‐tetrahydro‐furan‐3‐yl]‐methanols by IR, 1D‐ and 2D‐NMR, and HR‐ESI‐MS. Other four metabolites were identified as botryosphaeriodiplodin, (5R)‐5‐hydroxylasiodiplodin, (–)‐(1R,2R)‐jasmonic acid, and (–)‐(3S,4R,5R)‐4‐hydroxymethyl‐3,5‐dimethyldihydro‐2‐furanone (3 – 6, resp.). The absolute configuration (R) at hydroxylated secondary C‐atom C(7) was also established for compound 3. The compounds 1 – 3, 5, and 6, tested for their phytotoxic activities to grapevine cv. Inzolia leaves at different concentrations (0.125, 0.25, 0.5, and 1 mg/ml) were phytotoxic and compound 5 showed the highest toxicity. All metabolites did not show in vitro antifungal activity against four plant pathogens.