First report of Phytophthora niederhauserii causing root rot of carob tree (Ceratonia siliqua L.) in Spain.

Carob tree (Ceratonia siliqua L.) is a crop native to the Mediterranean Basin mainly being used as livestock feed, in pharmaceutical industry, and also a valuable source of human food due to the high dietary fibre and sugar contents. In January 2023, one-year-old 'Duraio' carob trees grafted on 'Rojal' rootstock on pots showing dark brown to necrotic lesions on the secondary roots, decline symptoms and leading to the death of some plants, were detected in a nursery located at the Valencian Community region in Spain. Disease severity was 40 to 50% of root area and disease incidence was approximately 50% on approximately 1000 plants. Six representative plants were randomly collected. Roots were washed under running tap water to rinse the soil away, and small symptomatic pieces were cut, disinfected with 70% ethanol, and then dried on absorbent paper. Two-mm-long segments were plated on CMA-PARBPH, a Phytophthora-selective medium, and incubated at 25ºC. After 2-3 days, growing colonies were transferred to potato dextrose agar (PDA) medium. A Phytophthora-like organism was consistently isolated (50% of root segments, n= 120). Colonies were whitish with irregular margins, had coenocityc mycelium, irregularly branched hyphal swellings, and chlamydospores were absent. Sporangia were non-papillate, persistent, ellipsoid and measuring 25 to 40 × 50 to 90 µm (average: 35.5 × 74.7 µm, n = 50). They proliferated internally with both nested and extended proliferation. These morphological features were similar to those of Phytophthora niederhauserii (Abad et al., 2014). Internal transcribed spacer (ITS) and cytochrome c oxidase I (COX1) regions of a representative isolate CF3 were sequenced to confirm the identity. Both sequences were deposited in GenBank under accession numbers OR763816 for ITS and OR783697 for COX1. OR763816 showed 99.87% sequence identity to P. niederhauserii strain Ex-type MG865552 and OR783697 showed 99.85% sequence identity to P. niederhauserii strain Ex-type MH136944 (Abad et al., 2023). Pathogenicity test was performed on one-year-old carob tree seedlings (Duraio/Rojal) grown in 13 cm-diameter pots to fulfill Koch's postulates. The inoculum was prepared in 1 L glass flasks with a mixture of 200 ml vermiculite, 20 ml oat grains, and 175 ml of V8 broth (20% V8 juice and 0.2% of CaCO3 in demineralized water) (Jung et al., 1996). Glass flasks with vermiculite/oat/V8 mixture were autoclaved three times for 20 min at 120 ºC. These mixtures were inoculated with the isolate CF3 which was previously grown on V8 agar medium and incubated for six weeks in the dark at room temperature (Pérez-Sierra et al., 2013). For inoculation, twenty-gram inoculum were mixed with 200 g autoclaved potting mix (peat, vermiculite and sand; 1:1:1, v/v/v), and added to the pots to plant the seedlings. Seven plants were inoculated and non-infested vermiculite/oat/V8 mixture was used to prepare seven control plants. Two months after inoculation one of the inoculated plants died. Six months after inoculation, only the inoculated plants showed decline symptoms with dry leaves and root necrosis. Isolates resembling P. niederhauserii were recovered by plating the roots from all inoculated plants on CMA-PARBPH, and the identity of the isolates was confirmed as P. niederhauserii based on ITS and COX1 sequencing. To our knowledge, this is the first report of P. niederhauserii causing root rot on carob tree. The detection of this pathogen in nurseries is relevant because its dissemination to orchards could have a negative impact in carob crop production.

First Report of Shoot Blight of Grapevine Caused by Sclerotinia sclerotiorum in Illes Balears, Mallorca, Spain.

In 2021, grapevines (Vitis vinifera L.) cv. Callet growing in a commercial vineyard located at Pollença (northeast of the island of Majorca, Spain) showed severe symptoms of shoot blight during spring and early summer, with an incidence of 70%. Symptoms consisted of elongated cankered-like lesions, surrounded by water-soaked darker tissues, that developed at the base or around the middle nodes of the shoot. For fungal isolation, shoot samples with lesions were collected, surface disinfected with 2% NaCl for 90s, rinsed twice with deionized water and placed in Petri plates containing potato dextrose agar (PDA). The plates were incubated at 25°C under 12 h light-darkness for 6 days. Isolations consistently yielded on kind of fungal colonies that produced white mycelium and black spherical to elongated sclerotia (2 to 10 mm in diameter). Morphological characterization was consistent with the description of Sclerotinia sclerotiorum (Lib.) de Bary (Bolton et al. 2006). Three isolates (UIB 118-1, UIB 118-26, and UIB 129-41) were preserved and deposited in the Culture Collection of Microbiology-Faculty of Sciences, University of Balearic Islands, Spain. Genomic DNA was extracted from isolates UIB 118-26 and UIB 129-41 using the EZNA Miniprep Kit (Omega Bio-Tek, Norcross, GA). The internal transcribed spacer (ITS) region of ribosomal DNA, ß-tubulin (BTUB) and calmodulin (CAL) gene regions were amplified using ITS1F-ITS4 (Gardes and Bruns, 1996; White et al. 1990), Bt-2a/Bt-2b (Glass and Donaldson 1995) and CAL228F/CAL737R (Carbone and Kohn 1999) primer sets, respectively. Amplicons were sequenced and deposited in GenBank with accession numbers MZ604647 and MZ604648 for ITS, OK634402 and OK634403 for BTUB and OK634404 and OK634405 for CAL. BLASTn search showed that isolates were >99 % (ITS, BTUB and CAL) identical to S. sclerotiorum GenBank accession no. KF859933, CP017815 and KF871381, respectively. Pathogenicity tests were conducted using eight one-year old grapevines cv. Cabernet Sauvignon. Old and new green shoots were inoculated by inserting a 6-mm plug of mycelium taken from actively growing cultures on PDA into cuts made at the base and at the distal part of each shoot with a sterile scalpel with a total of eight inoculation points per plant. Inoculated wounds were sealed with Parafilm tape to avoid rapid dehydration. Inoculated plants and an equal number of wounded but non-inoculated plants (negative controls) were maintained at 25 ± 1°C for 48 h in plastic containers to ensure a high relative humidity (>90%). After 5 days, the infection girdled and rotted the green new shoots, whereas the older partially lignified shoots developed a localized long brown lesion that reached 16 cm in length. Due to the rotting of the basal part of the petiole, leaves turned gray, wilted, and died, easily detaching from the stem. In advanced stages of the disease, 7 days after infection, branches died and fell with the leaves remained attached (Fig 1 A, B). Reisolations from diseased shoots were successfully performed on PDA to fulfill Koch's postulates. S. slerotiorum was previously reported on grapevine causing shoot blight in Chile (Latorre and Guerrero, 2001), Korea (Jong-Han et al. 2009), California-USA (Boland and Hall, 1994) and Australia (Hall et al. 2002). AlsoS. sclerotiorum was reported among the endophytic mycobiota associated with Vitis vinifera in the Iberian Peninsula (Gonzalez and Tello, 2011) but not as a pathogen causing visible symptoms on that crop. So, this is the first report of the occurrence of S. slerotiorum as a pathogen of grapevines in Spain causing symptoms of canker and shoot blight. This finding highlights a potential risk of this fungal disease for the wine industry in the Mediterranean region and specially for Spain, the country with the largest acreage devoted to grapevines. Although chemical and biological are suitable control strategies, disease management is difficult as sclerotia of Sclerotinia can remain in the soil for up to eight years (Adams and Ayears, 1979), and preventive surveys are greatly recommended as an important epidemiological tool to monitor the epidemiology of disease and identify potential outbreaks of this new pathogen on grapevine in Spain.

Exploring Factors Conditioning the Expression of Botryosphaeria Dieback in Grapevine for Integrated Management of the Disease.

Species from the Botryosphaeriaceae family are the causal agents of Botryosphaeria dieback (BD), a worldwide grapevine trunk disease. Because of their lifestyle and their adaptation to a wide range of temperatures, these fungi constitute a serious threat to vineyards and viticulture, especially in the actual context of climate change. Grapevine plants from both nurseries and vineyards are very susceptible to infections by botryosphaeriaceous fungi due to several cuts and wounds made during their propagation process and their entire life cycle, respectively. When decline becomes chronic or apoplectic, it reduces the longevity of the vineyard and affects the quality of the wine, leading to huge economic losses. Given the environmental impact of fungicides, and their short period of effectiveness in protecting pruning wounds, alternative strategies are being developed to fight BD fungal pathogens and limit their propagation. Among them, biological control has been recognized as a promising and sustainable alternative. However, there is still no effective strategy for combating this complex disease, conditioned by both fungal life traits and host tolerance traits, in relationships with the whole microbiome/microbiota. To provide sound guidance for an effective and sustainable integrated management of BD, by combining the limitation of infection risk, tolerant grapevine cultivars, and biological control, this review explores some of the factors conditioning the expression of BD in grapevine. Among them, the lifestyle of BD-associated pathogens, their pathogenicity factors, the cultivar traits of tolerance or susceptibility, and the biocontrol potential of Bacillus spp. and Trichoderma spp. are discussed.

Harnessing the power of comparative genomics to support the distinction of sister species within Phyllosticta and development of highly specific detection of Phyllosticta citricarpa causing citrus black spot by real-time PCR.

Citrus crops are affected by many fungal diseases. Among them, Citrus Black Spot caused by the ascomycete Phyllosticta citricarpa is particularly economically damaging wherever it occurs. Many other species of Phyllosticta are described on Citrus, but only P. citricarpa is considered a quarantine pest on the European continent. In order to prevent the introduction of this species into Europe, it is essential to have a detection test which can reliably identify it, and not confuse it with other species present on citrus, notably P. paracitricarpa. The latter taxon has recently been described as very close to P. citricarpa, and most detection tests do not allow to distinguish the two species. In this work, we exploited the genomic data of 37 isolates of Phyllosticta spp. from citrus, firstly to assess their phylogenetic relationships, and secondly to search for genomic regions that allowed the definition of species-specific markers of P. citricarpa. Analysis of 51 concatenated genes separated P. citricarpa and P. paracitricarpa in two phylogenetic clades. A locus was selected to define a hydrolysis probe and primers combination that could be used in real-time PCR for the specific detection of the quarantine species, to the exclusion of all others present on Citrus. This test was then thoroughly validated on a set of strains covering a wide geographical diversity, and on numerous biological samples to demonstrate its reliability for regulatory control. The validation data highlighted the need to check the reliability of the test in advance, when a change of reagents was being considered.

Social cognition in first-episode schizophrenia/schizoaffective disorder patients.

INTRODUCTION: People with schizophrenia have neurocognitive as well as social cognition deficits. Numerous studies have shown impairment in these domains in patients with chronic schizophrenia. However, these disturbances during the early phase of the disease have been less studied. OBJECTIVE: The aim of the study is to explore the theory of mind (ToM) and emotional processing in first-episode patients, compared to healthy subjects. METHOD: Forty patients with a first psychotic episode of less than 5 years' duration, and 40 healthy control subjects matched by age and years of schooling were assessed. The measures of social cognition included four stories of false belief, the Reading the Mind in the Eyes Test (RMET) and the Pictures Of Facial Affect (POFA) series. RESULTS: The patients with a first psychotic episode performed significantly worse in all tasks of social cognition, compared to the healthy controls. The second-order ToM was impaired whereas the first-order ToM was preserved in the patients. Happiness was the emotion most easily identified by both patients and controls. Fear was most difficult for the patients, while for the controls it was disgust. CONCLUSIONS: Deficits in ToM and emotional processing are present in patients with a first psychotic episode.

Evaluation of Bacillus subtilis PTA-271 and Trichoderma atroviride SC1 to control Botryosphaeria dieback and black-foot pathogens in grapevine propagation material.

BACKGROUND: Grapevine trunk diseases (GTDs) are a complex group of diseases that lead to major economic losses in all wine-producing countries. The investigation of biocontrol agents (BCAs) capable of forestalling or at least minimizing the development of GTDs has, recently, become a priority. Nursery experiments were set up to (i) assess the biocontrol effect of Trichoderma atroviride (Ta) SC1 and Bacillus subtilis (Bs) PTA-271, alone and in simultaneous application, against Botryosphaeria dieback (BOT)- and black-foot (BF)- associated pathogens during the grapevine propagation process and (ii) evaluate the success of the BCA inoculation during the grapevine propagation process, using quantitative reverse-transcription polymerase chain reaction techniques. RESULTS: The results demonstrated a significant reduction in the percentage of potentially infected plants and the percentage of fungal isolation from wood fragments of BOT and BF pathogens in nursery material treated with Ta SC1 and Bs PTA-271, respectively. In one of the experiments, simultaneous treatments with Bs PTA-271 and Ta SC1 caused a reduction in percentages of potentially infected plants and fungal isolation, from wood fragments containing BOT and BF pathogens. CONCLUSION: These biological treatments may be relevant components of an integrated approach, using complementary management strategies to limit infection by GTD pathogens, but further research is still needed to elucidate the effectiveness of Bs PTA-271 and the benefits of simultaneous application with Ta SC1 for the control of GTD pathogens in nurseries. © 2022 Society of Chemical Industry.

Pathogenicity of Oomycete Species to Different Prunus Hybrid Rootstocks.

Diseases caused by soilborne oomycetes are a limiting factor for the cultivation of Prunus spp., which makes the choice of a suitable rootstock a key factor. The objective of this study was to evaluate the pathogenicity of 12 oomycete species belonging to the genera Globisporangium, Phytophthora (Ph.), and Phytopythium (Pp.) to three Prunus hybrid rootstocks: 'Garnem', 'GF-677', and 'Rootpac-40'. These three rootstocks are widely used to grow stone fruit and almond in the Mediterranean Basin. Pathogenicity tests were conducted using 15 oomycete isolates and 1-year-old rootstock seedlings. Ninety days after inoculation, disease symptoms were evaluated on a severity scale, and the area under the disease progression curve and the survival probability of the inoculated seedlings were calculated. Moreover, root dry weight was recorded. All the isolates included in the pathogenicity tests were pathogenic on the rootstock seedlings and were reisolated from root lesions. Large differences in virulence were detected among the different oomycete species and isolates of Ph. niederhauserii for each rootstock. Phytophthora multivora and Pp. helicoides were generally the most virulent species. The results of the present research offer substantial contribution to increase our knowledge about the pathogenicity of several oomycete species that are frequently isolated in Prunus orchards and the potential risks that they pose for Prunus spp. crops.

Pain and glory: Narrative (De)constructions of older gay men in contemporary Spanish culture and cinema.

If (older) gay men have recurrently been stereotyped as hypersexual and as sexually voracious, they have also been represented as weak and effeminate, miserable and lonely, and as less manly than their heterosexual counterparts (Goltz, 2014; Freeman, 2010). Quite often, as Goltz reminds us, the two stereotypes intersect, as in classic films such as Death in Venice, Gods and Monsters, or Love and Death on Long Island, to name but a few, where ageism and homophobia combine to judge intergenerational relations as inappropriate and gay characters as "dirty old men" eager to recover their lost youth. Given these negative images, it is no wonder, then, that both youthism and ableism have become part and parcel of contemporary gay culture, which may also be linked to the few positive cultural images available of aging or disabled gay male bodies (Goltz, 2014). Starting off from the assumption that bodies are shaped and reshaped in complex interactions between physical and symbolic dimensions, the paper will demonstrate, however, how (auto)biographical narratives of older gay men, what we call "egodocuments," may be useful to rethink such traditional (mis)conceptions. Crossing the traditional divide between the Social Sciences and the Humanities, the study will draw on both life stories and film representations of older gay men in Spain, focusing on Spanish director Pedro Almodóvar's (2019) latest film, Pain and Glory, as a (semi-)autobiographical re-vision of traditional representations of gay men's aging. While Almodovar's protagonist and alter-ego, Salvador Mallo, appears as "prematurely aged" (Curtis and Thompson, 2015) due to bodily pain and disability, the film also travels back and forth from Mallos's childhood and youth to his maturity. This shows aging not only as a life-course experience, but also helps redefine it as a "queer" rather than linear or "straight" experience (Halberstam), allowing for both "pain and glory" to coexist in old age. Almodóvar's portraiture of gay aging will be compared to the life stories collected in a number of focus groups with Spanish older gay men, which will provide equally complex, varied, and often contradictory narratives.

Susceptibility of Almond (Prunus dulcis) Cultivars to Twig Canker and Shoot Blight Caused by Diaporthe amygdali.

Twenty-five almond cultivars were assessed for susceptibility to Diaporthe amygdali, causal agent of twig canker and shoot blight disease. In laboratory experiments, growing twigs were inoculated with four D. amygdali isolates. Moreover, growing shoots of almond cultivars grafted onto INRA 'GF-677' rootstock were used in 4-year field inoculations with one D. amygdali isolate. In both types of experiments, inoculum consisted of agar plugs with mycelium, which were inserted underneath the bark, and the lesion lengths caused by the fungus were measured. Necrotic lesions were observed in the inoculated almond cultivars in both laboratory and field tests, confirming the susceptibility of all evaluated cultivars to all inoculated isolates of D. amygdali. Cultivars were grouped as susceptible or very susceptible according to a cluster analysis. The relationship between some agronomic traits and cultivar susceptibility was also investigated. Blooming and ripening times were found to be relevant variables explaining cultivar performance related to D. amygdali susceptibility. Late and very late blooming and early and medium ripening cultivars were highly susceptible to D. amygdali. Our results may provide valuable information that could assist in ongoing breeding programs of this crop and in the selection of cultivars for new almond plantations.

Cultivar Contributes to the Beneficial Effects of Bacillus subtilis PTA-271 and Trichoderma atroviride SC1 to Protect Grapevine Against Neofusicoccum parvum.

Grapevine trunk diseases (GTDs) are a big threat for global viticulture. Without effective chemicals, biocontrol strategies are developed as alternatives to better cope with environmental concerns. A combination of biological control agents (BCAs) could even improve sustainable disease management through complementary ways of protection. In this study, we evaluated the combination of Bacillus subtilis (Bs) PTA-271 and Trichoderma atroviride (Ta) SC1 for the protection of Chardonnay and Tempranillo rootlings against Neofusicoccum parvum Bt67, an aggressive pathogen associated to Botryosphaeria dieback (BD). Indirect benefits offered by each BCA and their combination were then characterized in planta, as well as their direct benefits in vitro. Results provide evidence that (1) the cultivar contributes to the beneficial effects of Bs PTA-271 and Ta SC1 against N. parvum, and that (2) the in vitro BCA mutual antagonism switches to the strongest fungistatic effect toward Np-Bt67 in a three-way confrontation test. We also report for the first time the beneficial potential of a combination of BCA against Np-Bt67 especially in Tempranillo. Our findings highlight a common feature for both cultivars: salicylic acid (SA)-dependent defenses were strongly decreased in plants protected by the BCA, in contrast with symptomatic ones. We thus suggest that (1) the high basal expression of SA-dependent defenses in Tempranillo explains its highest susceptibility to N. parvum, and that (2) the cultivar-specific responses to the beneficial Bs PTA-271 and Ta SC1 remain to be further investigated.

Relationship Between the Xylem Anatomy of Grapevine Rootstocks and Their Susceptibility to Phaeoacremonium minimum and Phaeomoniella chlamydospora.

Fungal grapevine trunk diseases (GTDs) are some of the most pressing threats to grape production worldwide. While these diseases are associated with several fungal pathogens, Phaeomoniella chlamydospora and Phaeoacremonium minimum are important contributors to esca and Petri diseases. Recent research has linked grapevine xylem diameter with tolerance to Pa. chlamydospora in commercial rootstocks. In this study, we screen over 25 rootstocks for xylem characteristics and tolerance to both Pa. chlamydospora and Pm. minimum. Tolerance was measured by fungal incidence and DNA concentration (quantified via qPCR), while histological analyses were used to measure xylem characteristics, including xylem vessels diameter, density, and the proportion of the stem surface area covered by xylem vessels. Rootstocks were grouped into different classes based on xylem characteristics to assess the potential association between vasculature traits and pathogen tolerance. Our results revealed significant differences in all the analyzed xylem traits, and also in DNA concentration for both pathogens among the tested rootstocks. They corroborate the link between xylem vessels diameter and tolerance to Pa. chlamydospora. In Pm. minimum, the rootstocks with the widest xylem diameter proved the most susceptible. This relationship between vasculature development and pathogen tolerance has the potential to inform both cultivar choice and future rootstock breeding to reduce the detrimental impact of GTDs worldwide.

Cultivar Susceptibility to Natural Infections Caused by Fungal Grapevine Trunk Pathogens in La Mancha Designation of Origin (Spain).

Grapevine trunk diseases (GTDs) are one of the main biotic stress factors affecting this crop. The use of tolerant grapevine cultivars would be an interesting and sustainable alternative strategy to control GTDs. To date, most studies about cultivar susceptibility have been conducted under controlled conditions, and little information is available about tolerance to natural infections caused by GTD fungi. The objectives of this study were: (i) to identify tolerant cultivars to GTD fungi within a Spanish germplasm collection, based on external symptoms observed in the vineyard; and (ii) to characterize the pathogenic mycoflora associated with symptomatic vines. For this purpose, a grapevine germplasm collection including 22 white and 25 red cultivars was monitored along three growing seasons, and their susceptibility for esca foliar symptoms was assessed. Fungi were identified by using morphological and molecular methods. Cultivars such as, 'Monastrell', 'Graciano', 'Cabernet Franc', 'Cabernet Sauvignon', 'Syrah', 'Moscatel de Alejandría', 'Sauvignon Blanc', and 'Airén' displayed high susceptibility to GTDs, whereas others such as 'Petit Verdot', 'Pinot Noir', 'Chardonnay', and 'Riesling' were considered as tolerant. The prevalent fungal species isolated from symptomatic vines were Phaeomoniella chlamydospora (27.9% of the fungal isolates), Cryptovalsa ampelina (24.6%), and Dothiorella sarmentorum (21.3%).

Evaluation of Sown Cover Crops and Spontaneous Weed Flora as a Potential Reservoir of Black-Foot Pathogens in Organic Viticulture.

(1) Background. An extensive survey of grapevine-sown cover crops and spontaneous weed flora was conducted from 2019 to 2020 in organic vineyards in six European countries (France, Italy, Romania, Slovenia, Spain, Switzerland). Our main objective was to detect and identify the presence of Cylindrocarpon-like asexual morphs species associated with black-foot disease on their roots. (2) Methods. Fungal isolations from root fragments were performed on culture media. Cylindrocarpon-like asexual morph species were identified by analyzing the DNA sequence data of the histone H3 (his3) gene region. In all, 685 plants belonging to different botanical families and genera were analyzed. Cylindrocarpon-like asexual morphs were recovered from 68 plants (9.9% of the total) and approximately 0.97% of the plated root fragments. (3) Results. Three fungal species (Dactylonectria alcacerensis, Dactylonectria torresensis, Ilyonectria robusta) were identified. Dactylonectria torresensis was the most frequent, and was isolated from many cover crop species in all six countries. A principal component analysis with the vineyard variables showed that seasonal temperatures and organic matter soil content correlated positively with Cylindrocarpon-like asexual morphs incidence. (4) Conclusions. The presence of Cylindrocarpon-like asexual morphs on roots of cover crops suggests that they can potentially act as alternative hosts for long-term survival or to increase inoculum levels in vineyard soils.

Cadophora sabaouae sp. nov. and Phaeoacremonium Species Associated with Petri Disease on Grapevine Propagation Material and Young Grapevines in Algeria.

A field survey conducted on asymptomatic grapevine propagation material from nurseries and symptomatic young grapevines throughout different regions of Algeria yielded a collection of 70 Phaeoacremonium-like isolates and three Cadophora-like isolates. Based on morphology and DNA sequence data of ß-tubulin (tub2) and actin, five Phaeoacremonium species were identified including Phaeoacremonium minimum (22 isolates), Phaeoacremonium venezuelense (19 isolates), Phaeoacremonium parasiticum (17 isolates), Phaeoacremonium australiense (8 isolates), and Phaeoacremonium iranianum (4 isolates). The latter two species (P. australiense and P. iranianum) were reported for the first time in Algeria. Multilocus phylogenetic analyses (internal transcribed spacer, tub2, and translation elongation factor 1-α) and morphological features, allowed the description of the three isolates belonging to the genus Cadophora (WAMC34, WAMC117, and WAMC118) as a novel species, named Cadophora sabaouae sp. nov. Pathogenicity tests were conducted on grapevine cuttings cultivar Cardinal. All the identified species were pathogenic on grapevine cuttings.

First Report of Diaporthe amygdali Associated with Twig Canker and Shoot Blight of Nectarine in Spain.

Nectarine (Prunus persica (L.) Batsch var. nucipersica (Suckow) C. K. Schneid.) is a fruit crop widely cultivated throughout the Mediterranean basin. In Spain, it is mainly grown in eastern regions of the country. In March 2018, 5-year-old nectarine trees showing twig canker symptoms were observed after a rainy spring period in a 0.5 ha orchard located at Alaior, Menorca island (Spain). Cankers were frequent on affected trees (approximately, 80% of the total trees), thus leading to shoot blight. Ten twig segments of one-year old wood with cankers were cut, washed under running tap water, surface disinfected for 1 min in a 1.5% sodium hypochlorite solution and rinsed twice in sterile distilled water. Small pieces (2 mm) of affected tissues were taken from the margin of the cankers and plated on potato dextrose agar (PDA) supplemented with 0.5 g/L of streptomycin sulphate (PDAS). The plates were then incubated at 25 ºC in the dark for 7 to 10 d. Actively growing colonies were first hyphal-tipped and then transferred to PDA and 2% water agar supplemented with sterile pine needles and incubated at 21-22ºC under a 12h/12h near UV / darkness cycle during 21 d (León et al. 2020). Colonies were white at first, becoming light cream, with visible solitary and aggregate pycnidia at maturity. Alpha conidia were aseptate, fusiform, hyaline, multi-guttulated (mean ± SD = 7.4 ± 0.7 × 2.8 ± 0.4 µm, n = 100). Beta and gamma conidia were not observed. The morphological and cultural characteristics of the isolates were congruent with those of Diaporthe spp. (Gomes et al. 2013). The ITS1-5.8S-ITS2 (ITS) region and fragments of ß-tubulin (tub2), the translation elongation factor 1-alpha (tef1-α) gene regions, histone H3 (his3) and calmodulin (cal) genes of representative isolate DAL-59 were amplified and sequenced (Santos et al. 2017). The BLASTn analysis revealed 100% similarity with sequences of D. mediterranea (Synonym D. amygdali) (Hilário et al. 2021) isolate DAL-34 from almond (ITS: MT007489, tub2: MT006686, tef1-α: MT006989, his3: MT007095 and cal: MT006761). Sequences of isolate DAL-59 were deposited in GenBank Database (ITS: MT007491, tub2: MT006688, tef1-α: MT006991, his3: MT007097 and cal: MT006763). Pathogenicity tests were conducted using one-year-old potted plants of nectarine cv. Boreal, which were inoculated with isolate DAL-59. In each plant, a 3 mm wound was made in the center of the main branch (about 30 cm length) with a scalpel. Colonized agar plugs with 3 mm diameter, which were obtained from active 10-day-old colonies growing on PDA, were inserted underneath the epidermis and the wounds sealed with Parafilm. Inoculated plants were incubated in a growth chamber at 23 ºC with 12 h of light per day. Controls were inoculated with uncolonized PDA plugs. There were twelve plants per treatment, which were arranged in a completely randomized design. Five days after inoculation necrosis development was observed in the area of inoculation. Wilting and twig blight symptoms over the lesion occurred 3-wk after inoculation and pycnidia were detected, while the controls remained asymptomatic. Diaporthe amygdali was re-isolated from symptomatic tissues and identified as described above to satisfy Koch's postulates. To our knowledge, this is the first report of D. amygdali causing twig canker and shoot blight disease on nectarine in Spain.

Characterization of Five New Monosporascus Species: Adaptation to Environmental Factors, Pathogenicity to Cucurbits and Sensitivity to Fungicides.

In this study, five new recently described Monosporascus species, M. brasiliensis, M. caatinguensis, M. mossoroensis, M. nordestinus, and M. semiaridus, which were found on weeds collected from cucurbit cultivation fields in northeastern Brazil, are characterized regarding mycelial growth at different pH levels and salinity (NaCl) concentrations, their pathogenicity to selected cucurbit species, and their sensitivity to fungicides with different modes of action. Our results reveal great variability among the representative isolates of each Monosporascus spp. All of them showed a wide range of tolerance to different pH levels, and NaCl significantly reduced their in vitro mycelial growth, although no concentration was able to inhibit them completely. In pathogenicity tests, all seedlings of cucurbits evaluated, melon, watermelon, cucumber, and pumpkin, were susceptible to the five Monosporascus spp. in greenhouse experiments using artificial inoculation of roots. Moreover, all Monosporascus spp. were highly susceptible to the fungicides fludioxonil and fluazinam. Our findings provide relevant information about the response of these new Monosporascus spp. to environmental factors, plant genotypes and fungicides.

Temporal Dispersal Patterns of Phaeomoniella chlamydospora, Causal Agent of Petri Disease and Esca, in Vineyards.

Although the fungus Phaeomoniella chlamydospora is the most commonly detected causal agent of Petri disease and esca, two important fungal grapevine trunk diseases, little is known about the dispersal patterns of P. chlamydospora inoculum. In this work, we studied the dispersal of P. chlamydospora airborne inoculum from 2016 to 2018 in two viticultural areas of eastern (Ontinyent) and northern (Logroño) Spain. The vineyards were monitored weekly from November to April using microscope slide traps, and P. chlamydospora was detected and quantified by a specific real-time quantitative (qPCR) method set up in this work. The method was found to be sensitive, and a good correlation was observed between numbers of P. chlamydospora conidia (counted by microscope) and DNA copy numbers (quantified by qPCR). We consistently detected DNA of P. chlamydospora at both locations and in all seasons but in different quantities. In most cases, DNA was first detected in the last half of November, and most of the DNA was detected from December to early April. When rain was used as a predictor of P. chlamydospora DNA detection in traps, false-negative detections were observed, but these involved only 4% of the total. The dispersal pattern of P. chlamydospora DNA over time was best described (R2 = 0.765 and concordance correlation coefficient = 0.870) by a Gompertz equation, with time expressed as hydrothermal time (a physiological time accounting for the effects of temperature and rain). This equation could be used to predict periods with a high risk of dispersal of P. chlamydospora.

Endoscopic variceal ligation as primary prophylaxis for upper GI bleeding in children.

BACKGROUND AND AIMS: Variceal hemorrhage can be a life-threatening adverse event of chronic liver disease. In contrast to the well-described guidelines for the management of portal hypertension (PH) in adults, there is limited evidence about the optimal prophylactic management of variceal bleeding in children. This study was carried out to assess the efficacy of endoscopic variceal ligation (EVL) as primary prophylaxis to prevent upper GI bleeding in children with PH. METHODS: From January 2014 to April 2018, all pediatric patients with PH disease and medium to large esophageal varices or reddish spots, regardless of the grade of the varix, were prospectively included in the protocol of primary prophylaxis with EVL. A second retrospective group of patients was made after reviewing medical records of 32 pediatric patients with PH that presented esophageal varices in the upper endoscopy and had received propranolol as primary prophylaxis. RESULTS: Twenty-four patients (75%) reached varices eradication in the EVL group, with a median of 2 procedures (range, 1-4) before eradication and a median time to eradication of 3.40 months (range, 1.10-13.33). No EVL-related adverse events were observed. Statistically significant differences were observed in the bleeding rate at 3 years between propranolol and EVL groups (6/32 [21.9%] vs 1/32 [3.2%], P < .02). The hazard ratio for bleeding for patients treated with propranolol compared with those treated with EVL was 2.6 (95% confidence interval, 1.53-3.67). CONCLUSIONS: EVL is a safe and effective treatment to prevent upper GI bleeding in pediatric patients with PH. (Clinical trial registration number: NCT03943784.).

Evaluation of long-term protection from nursery to vineyard provided by Trichoderma atroviride SC1 against fungal grapevine trunk pathogens.

BACKGROUND: Fungal grapevine trunk diseases (GTDs) represent a threat to viticulture, being responsible for important economic losses worldwide. Nursery and vineyard experiments were set up to evaluate the ability of Trichoderma atroviride SC1 to reduce infections of GTD pathogens in grapevine planting material during the propagation process and to assess the long-term protection provided by this biocontrol agent on grapevine plants in young vineyards during two growing seasons. RESULTS: Reductions of some GTD pathogen incidence and severity were found on grapevine propagation material after nursery application of T. atroviride SC1 during the grafting process, and also after additional T. atroviride SC1 treatments performed during two growing seasons in young vineyards, when compared with untreated plants. CONCLUSION: Trichoderma atroviride SC1 showed promise to reduce infections caused by some GTD pathogens in nurseries, and also when establishing new vineyards. This biological control agent could possibly be a valuable component in an integrated management approach where various strategies are combined to reduce GTD infections. © 2019 Society of Chemical Industry.