Synthesis and in-vitro antifungal activity of 6-trifluoromethyipyrazolo[3,4-6d]pyrimidin-4(5H)-thiones

Twelve 6-trifluoromethylpyrazolo[3,4-d]pyrimidin-4(5H)-thiones were prepared by the reaction between 4-thiocarbamoyl-5-aminopyrazoles and trifluoroacetic anhydride. They were tested in vitro for antifungal activity against a series of phytopathogenic fungi of different taxonomic classes. The EC50 and MIC values of four compounds were comparable or inferior to those of reference commercial fungicides in controlling Sclerotinia minor, Corticium solani and Phytium ultimum.     

Pseudomonas syringae pv. coryli, the Causal Agent of Bacterial Twig Dieback of Corylus avellana

ABSTRACT Thirty-eight bacterial strains isolated from hazelnut (Corylus avellana) cv. Tonda Gentile delle Langhe showing a twig dieback in Piedmont and Sardinia, Italy, were studied by a polyphasic approach. All strains were assessed by fatty acids analysis and repetitive sequence-based polymerase chain reaction (PCR) fingerprinting using BOX and ERIC primer sets. Representative strains also were assessed by sequencing the 16S rDNA and hrpL genes, determining the presence of the syrB gene, testing their biochemical and nutritional characteristics, and determining their pathogenicity to hazelnut and other plants species or plant organs. Moreover, they were compared with reference strains of other phytopathogenic pseudomonads. The strains from hazelnut belong to Pseudomonas syringae (sensu latu), LOPAT group Ia. Both fatty acids and repetitive-sequence-based PCR clearly discriminate such strains from other Pseudomonas spp., including P. avellanae and other P. syringae pathovars as well as P. syringae pv. syringae strains from hazelnut. Also, the sequencing of 16S rDNA and hrpL genes differentiated them from P. avellanae and from P. syringae pv. syringae. They did not possess the syrB gene. Some nutritional tests also differentiated them from related P. syringae pathovars. Upon artificial inoculation, these strains incited severe twig diebacks only on hazelnut. Our results justify the creation of a new pathovar because the strains from hazelnut constitute a homogeneous group and a discrete phenon. The name of P. syringae pv. coryli is proposed and criteria for routine identification are presented.     

Characterization and degradation of lignin from steam explosion of pine and corn stalk of lignin: The role of superoxide ion and ozone

Steam explosion of corn stalk in the presence of 3% sulphuric acid at 200 °C for 5 min gave the highest recovery of lignin. Lignin has M_w = 2640 and M_z = 93,994. In the UV spectrum absorptions at λ = 231 and 280 nm were recorded. ¹H NMR spectrum of lignin showed signals attributable to cinnamaldehyde units, guaiacyl units, and syringyl units. Syringyl and guaiacyl units are in 1:1 ratio. ¹³C NMR spectrum showed signals for guaiacyl, syringyl, and p-hydroxyphenyl units. The spectrum showed a prevalence of guaiacyl units. The ¹³C NMR spectrum is in agreement with the presence of cinnamic units. The same characterization was performed on lignin from pine. The irradiation of lignin from pine from steam explosion process in the presence of oxygen, in conditions described for the formation of superoxide ion, for different irradiation time was followed isolating the lignin and determining the average molecular weight. The experiments showed that, until 8 h irradiation, M_n decreases, while M_w and M_z increases. After 8 h irradiation an inverse behaviour was observed, with an increase of M_n and a decrease of M_w and M_z. These results are in agreement with an initial polymerization process followed by a photoinduced degradation. Ozonization was carried out in acetonitrile–methanol solution. The reaction showed a zero-order kinetics. After 50 min the average molecular weight of lignin is the half. The reaction mixture was analyzed by using GC–MS. Oxalic acid was determined.     

Mixtures of organic amendments and biochar promote beneficial soil microbiota and affect Fusarium oxysporum f. sp. lactucae,Rhizoctonia solani and Sclerotinia minor disease suppression

The use of soil fumigants and fungicides to control soilborne pathogens is reduced due to awareness of their negative drawbacks. Long-term application of such agrochemicals negatively affects soil microbiota and reduces natural soil suppressiveness. We investigated long-term impacts of biochar and not-pyrogenic organic amendment (OAs: manure, alfalfa straw and glucose), on disease suppression compared with conventional management in three pathosystems: Fusarium oxysporum f. sp. lactucae (FOL)–lettuce, Rhizoctonia solani–tomato and Sclerotinia sclerotiorum–lettuce, by conditioning soil for 2 years. Soil analyses included pH, electrical conductivity, organic carbon, total nitrogen, C:N ratio, N-NO₃⁻, N-NH₄⁺, cation exchange capacity, available phosphorus, Ca²⁺, Mg²⁺, K⁺, Na⁺, fluorescein diacetate (FDA), and total and active limestone. Soil microbiota was characterized by combining BIOLOG EcoPlates with next-generation sequencing of 16S and 18S rRNA genes. Soil amended with OAs generally suppressed disease by S. sclerotiorum and FOL compared to fumigants and synthetic fertilizers. However, the incidence of R. solani infection was lower in soil treated with synthetic fertilizers than soil amended with OAs. EC, pH, C:N, N-NO₃⁻, N-NH₄⁺, FDA and BIOLOG were significantly correlated with disease, depending on pathosystem. Bacterial richness and diversity, presence of some genera like Acidobacteria, Chloracidobacteria, Solibacteres, Anaerolineae, Nitrospira and Deltaproteobacteria were negatively related to disease incidence of FOL and S. sclerotiorum, whereas damping-off caused by R. solani was negatively affected by the presence of Flavobacterium and Chitinofagha. Therefore, we concluded that long-term application of OAs can effectively improve soil suppressiveness and reduce disease incidence against root pathogens, although the effects vary considerably depending upon pathosystem.     

‘Microbigation’: delivery of biological control agents through drip irrigation systems

The uniform and precise application of microbial particles close to the target organism and to the plant to be protected can increase the success of a biological control treatment. The use of systems or technologies which are usually available in agriculture could influence the acceptability of biocontrol agents by farmers, and enlarge the market. A pilot system was realized using dripper lines, drippers, filters and other tools commonly used in irrigation and precision agriculture in the greenhouse to evaluate their suitability for applying and distributing microbial biocontrol agents. Conidial suspensions of marketed or marketable agents were used, i.e. Fusarium oxysporum, F. solani, Trichoderma harzianum and Paecilomyces lilacinus. The experiments carried out demonstrated that conidial suspensions (10⁶ conidia ml⁻¹) can pass through the drippers without causing clogging, regardless of their size, and remained viable. The term ‘microbigation’ is here proposed for this kind of microbial application technique.     

Kiwellin, a modular protein from green and gold kiwi fruits: evidence of in vivo and in vitro processing and IgE binding

Kiwellin, an allergenic protein formerly isolated from green kiwi fruit, has been identified as the most abundant component of the gold kiwi species. A protein named KiTH, showing a 20 kDa band on reducing SDS-PAGE and 100% identity with the C-terminal region of kiwellin, has been identified in the extract of the ripe green species. In vitro treatment of purified kiwellin with the protease actinidin from green kiwi fruit originated KiTH and kissper, a recently described pore-forming peptide. Primary structure analysis and experimental evidence suggest that kiwellin is a modular protein with two domains. It may undergo in vivo proteolytic processing by actinidin, thus producing KiTH and kissper. When probed with sera recognizing kiwellin from green kiwi fruit, KiTH showed IgE binding, with reactivity levels sometimes different from those of kiwellin. The IgE-binding capacity of kiwellin from gold kiwi fruit appears to be similar to that of the green species.     

Evaluation of phenolic compounds in virgin olive oil by direct injection in high-performance liquid chromatography with fluorometric detection

Hydrophilic phenols are the most abundant natural antioxidants of virgin olive oil (VOO), in which tocopherols and carotenes are also present. The prevalent classes of hydrophilic phenols found in VOO are phenyl alcohols, phenolic acids, secoiridoids such as the dialdehydic form of decarboxymethyl elenolic acid linked to (3,4-dihydroxyphenyl)ethanol or (p-hydroxypheny1)ethanol (3,4-DHPEA-EDA or p-HPEA-EDA) and an isomer of the oleuropein aglycon (3,4-DHPEA-EA), lignans such as (+)-1-acetoxypinoresinol and (+)-pinoresinol, and flavonoids. A new method for the analysis of VOO hydrophilic phenols by direct injection in high-performance liquid chromatography (HPLC) with the use of a fluorescence detector (FLD) has been proposed and compared with the traditional liquid-liquid extraction technique followed by the HPLC analysis utilizing a diode array detector (DAD) and a FLD. Results show that the most important classes of phenolic compounds occurring in VOO can be evaluated using HPLC direct injection. The efficiency of the new method, as compared to the liquid-liquid extraction, was higher to quantify phenyl alcohols, lignans, and 3,4-DHPEA-EA and lower for the evaluation of 3,4-DHPEA-EDA and p-HPEA-EDA.     

The impact of clonality on an endangered tree (Elaeocarpus williamsianus) in a fragmented rainforest

The occurrence and distribution of clonality in the endangered rainforest tree Elaeocarpus williamsianus (Elaeocarpaceae) was investigated using SSR and RAPD analyses for 170 apparent individual trees found across seven sites. The results obtained with the two molecular techniques were in complete agreement in showing that single clones are present in most of the E. williamsianus sites with two genets occurring at the largest and most adequately protected site. In addition, seed production, viability and germinability were determined for four of these populations. Fruit were produced in all four populations tested although sterile fruit were very common. Only two E. williamsianus trees representing different genets within the same site produced viable seed. The overall genetic diversity within E. williamsianus is much lower than expected and thus the potential for sexual reproduction has been significantly diminished. It is concluded that habitat fragmentation removed an existing balance between vegetative and sexual reproduction in this species. Such findings have added urgency to the management of this species which could include a reintroduction program incorporating the collation of all clones within selected sites.     

On the chemical composition and nutritional value of pleurotus taxa growing on umbelliferous plants (apiaceae)

Unpublished data on the chemical composition and nutritional value of Pleurotus mushrooms, growing on Umbelliferous plants (Apiaceae), are here reported. Cultivated basidiomata of four different Pleurotus taxa were analyzed in order to evaluate the composition in lipids, sugars, nitrogen, water, vitamins, ashes, and energetic values. The results showed that Pleurotus mushrooms are suitable in every type of diet thanks to their low caloric content, gastronomic value, vitamins, and mineral salt contents. The presence of a high content of vitamin B(12) and riboflavin in Pleurotus nebrodensis is noteworthy.