Effect of Cultivar Resistance and Soil Management on Spatial-Temporal Development of Verticillium Wilt of Olive: A Long-Term Study.

Verticillium wilt, caused by Verticillium dahliae, challenges olive cultivation and an Integrated Disease Management (IDM) approach is the best-suited tool to combat it. Since 1998, an IDM strategy in an orchard (called Granon, Spain) of the susceptible cv. Picual was conducted by increasing planting density with moderately resistant cv. Frantoio, chemical weed control, and replanting of dead olives with cv. Frantoio following soil solarization. The Verticillium wilt epidemic in Granon orchard was compared to the epidemic in a non-IDM orchard (called Ancla, Spain) with plowed soil and dead Picual olives replanted with the same cultivar. Field evaluations (2012-2013) showed an incidence and severity of the disease as Picual-Ancla > Picual-Granon > Frantoio-Granon. The spatiotemporal dynamics of the Verticillium epidemics from 1998 to 2010 were monitored with digital images using SIG. The annual tree mortalities were 5.6% for Picual olives in Ancla orchard, and 3.1 and 0.7% for Picual and Frantoio olives in Granon orchard, respectively. There was a negative relationship between the mortality of olive trees (%) by the pathogen and the height (m) above sea level. The annual mortality of cv. Picual olives was positively correlated with spring rainfalls. The Index of Dispersion and beta-binomial distribution showed aggregation of Verticillium-dead olives. In conclusion, this IDM strategy considerably reduced the disease in comparison with traditional agronomic practices.

First Report of Colletotrichum karstii Causing Fruit Anthracnose of Carissa grandiflora in Spain.

The species Carissa grandiflora A. DC., commonly called Natal plum, is a shrub native to the coastal region of Natal, South Africa. In southern Spain, Natal plum is used as an ornamental plant due to its beautiful flowers and red ripen fruits. In March 2019 and 2020, we surveyed nine public gardens in the cities of Cadiz and Sanlucar de Barrameda (Andalusia, Spain); and Natal plum fruit showing anthracnose symptoms were observed in six (55% prevalence) of them. Affected fruits showed necrotic and circular lesions with acervuli in the center (Fig. 1a) causing the complete mummification of the fruit (Fig. 1b). Affected fruits were collected from four gardens and disinfested according to Moral et al. (2010). Six fungal isolates were recovered from small (3-4 × 1-2 mm) pieces of the affected fruits in Potato Dextrose Agar (PDA), and hyphal tips from them were transferred to fresh PDA to obtain pure cultures. The six isolates were initially identified as Colletotrichum karstii according to their morphology and the sequences of the ITS1-5.8S-ITS2 (ITS) region (Damm et al. 2012). The six Colletotrichum isolates showed similar colony morphology and their ITS sequences were identical. Overall, C. karstii isolates showed cylindrical and straight conidia that were 12.1 to 14.2 µm long and 4.9 to 5.6 µm wide (n = 50). The aerial mycelia of the fungus varied from grayish-white to dark gray. A multilocus approach was conducted for more precise identification of the Colletotrichum species. For that, ITS, beta-tubulin (TUB2), actin (ACT), partial sequences of the chitin synthase 1 (CHS-1), histone 3 (HIS3), and a 200-bp intron fragment of the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) of a representative isolate (FITP19001) were amplified and sequenced according to Damm et al. (2012). GenBank Accession Nos. for ITS, TUB2, ACT, CHS-1, HIS3 and GADPH: MT757643, MT759805, MT759806, MT759807, MT759808 and MT759809, respectively. Sequences showed 100% identity with homologous sequences belonging to C. karstii (GenBank taxid:1095194). To test Koch's postulates, 10 unripen and 10 ripen C. grandiflora fruits, harvested from asymptomatic plants, were inoculated. For each group, five fruits were inoculated using a drop of 10 µl of 5 × 104 conidia per ml suspension of C. karstii (FITP19001) and another five fruits were inoculated using a mycelial plug of the same isolate. Inoculated fruits were incubated in a humid chamber at room temperature (19-24ºC) under light for two weeks. Non-inoculated control fruits were treated with sterile water or a PDA plug and incubated under the same conditions. The pathogenicity test was conducted twice. After 10 days, typical anthracnose symptoms developed on both unripen and ripen inoculated fruits, but not on non-inoculated controls. Overall, the severity of anthracnose lesions was higher on ripen fruits than in the unripen fruits. Likewise, the severity of symptoms was higher on the fruits inoculated using a mycelial plug than on those fruits inoculated with a spore suspension. The species C. karstii was reisolated from lesions of all inoculated fruits as described above but not from non-inoculated fruits. The species C. karstii has been described affecting numerous species worldwide (Damm et al., 2012). Previously, C. gloeosporioides was reported causing fruit anthracnose of Natal plum in Florida (Alfieri et al., 1984). To our knowledge, this is the first report of C. karstii causing anthracnose on the fruit of Natal plum in Spain and worldwide.

Helical peptides from VEGF and Vammin hotspots for modulating the VEGF-VEGFR interaction.

The design, synthesis, conformational studies and binding affinity for VEGF receptors of a collection of linear and cyclic peptide analogues of the N-terminal α-helix fragments 13-25 of VEGF and 1-13 of Vammin are described. Linear 13(14)-mer peptides were designed with the help of an AGADIR algorithm and prepared following peptide solid-phase synthetic protocols. Cyclic peptide derivatives were prepared on-resin from linear precursors with conveniently located Glu and Lys residues, by the formation of amide linkages. Conformational analysis, CD and NMR, showed that most synthesized peptides have a clear tendency to be structured as α-helices in solution. Some of the peptides were able to bind a VEGFR-1 receptor with moderate affinity. In addition to the described key residues (Phe17, Tyr21 and Tyr25), Val14 and Val20 seem to be relevant for affinity.

Disulfide and amide-bridged cyclic peptide analogues of the VEGF81₋91 fragment: synthesis, conformational analysis and biological evaluation.

The design, synthesis, conformational studies and binding affinity for VEGFR-1 receptors of a collection of linear and cyclic peptide analogues of the ß-hairpin fragment VEGF(81-91) are described. Cyclic 11-mer peptide derivatives were prepared from linear precursors with conveniently located Cys, Asp or Dap residues, by the formation of disulfide and amide bridges, using solid-phase synthesis. Molecular modelling studies indicated a tendency to be structured around the central ß-turn of the VEGF(81-91) ß-hairpin in most synthesized cyclic compounds. This structural behavior was confirmed by NMR conformational analysis. The NHCO cyclic derivative 7 showed significant affinity for VEGFR-1, slightly higher than the native linear fragment, thus supporting the design of mimics of this fragment as a valid approach to disrupt the VEGF/VEGFR-1 interaction.

Parallel solid-phase synthesis of a small library of linear and hydrocarbon-bridged analogues of VEGF(81-91): potential biological tools for studying the VEGF/VEGFR-1 interaction.

The design, synthesis and binding affinity for VEGFR-1 receptors of a small library of linear and cyclic analogues of the VEGF(81-91) fragment are described. Cyclic 11- and 10-mer peptide derivatives were prepared using parallel solid-phase protocols. The formation of hydrocarbon alkene-bridged cyclic peptides was achieved through optimized ring-closing metathesis reactions from linear derivatives with conveniently located allylGly residues. Alkane-bridged analogues were successfully obtained by ulterior on-resin hydrogenation. Binding assays showed that some of these compounds were able to compete with labeled VEGF for interaction with the VEGFR-1 receptor. Several peptide derivatives, 2, 7 and 8, showed modest but significant binding affinity, indicating that the designed peptide could mimic the VEGF(81-91) fragment and therefore disrupt the VEGF/VEGFR-1 interaction. This fact opens the way for using these peptides as the starting point for biological/pharmacological tools to deeply investigate this protein-protein system.

Gly-Pro-Glu protects beta-amyloid-induced somatostatin depletion in the rat cortex.

The effect of Gly-Pro-Glu (GPE) on the somatostatinergic system of the temporal cortex in amyloid beta-peptide (Abeta) treated rats was investigated. Intracerebroventricular Abeta25-35 administration for 14 days (300 pmol/day) to ovariectomized rats produced a marked reduction in somatostatin (SRIF) content, SRIF receptor density and reduced the inhibitory effect of SRIF on adenylyl cyclase activity. I.p. injection of three doses (300 microg) of GPE on days 0, 6 and 12 resulted in a partial recovery of the parameters affected by Abeta25-35 administration. These results indicate that GPE may have an in vivo effect protecting the temporal cortical somatostatinergic system from Abeta insult.