First report of Colletotrichum grossum causing apple bitter rot worldwide (Italy).

Apple bitter rot is a globally widespread disease that is observed on fruits both pre-harvest and post-harvest, contributing to considerable economic losses. While the Colletotrichum acutatum species complex are predominant in Europe (Baroncelli et al. 2014; Amaral Carneiro and Baric 2021), in recent years, the Colletotrichum gloeosporioides species complex are emerging, raising many concerns (Amaral Carneiro et al. 2023). Circular, slightly sunken, brown lesions with acervuli produced in concentric spots were observed on 'Story® Inored' cultivar harvested in September 2022 from an organic orchard in Masi (Padova province, Italy), with a disease incidence close to 30%. From ten diseased apples, tissue samples were excised under aseptic conditions from surface-cleaned fruit at the margin between healthy and diseased pulp tissue, transferred to potato dextrose agar medium and incubated in the dark at 25 °C for 7 days, whereafter five single-spore cultures were obtained. Pure colonies grown at 25 °C for 7 days appeared light gray-white on the upper side with floccose aerial mycelium, while the reverse side was dark gray with a distinct margin. Conidia were hyaline, cylindrical in shape with both ends rounded or one end acute and measured 16.6 ± 1.4 × 6.1 ± 0.5 µm [mean ± SD] (n=50) as described by Diao et al. 2017. To identify the species, genomic DNA of a representative isolate (C38) was extracted, beta-tubulin (TUB2), calmodulin (CAL), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), glutamine synthetase (GS), Apn2-Mat1-2 intergenic spacer (ApnMat) genes and the internal transcribed spacer (ITS) region, were amplified by PCR and Sanger sequenced (Rojas et al. 2010; Weir et al. 2012). The obtained DNA sequences of, TUB2, CAL, GAPDH, GS, ApnMat and ITS were submitted to GenBank under the accession numbers OR025589, OR025586, OR025587, OR025588, OR025585 and OR004800, respectively. A MegaBLAST analysis resulted 100 % identity to the epitype CAUG7 of Colletotrichum grossum (Diao et al. 2017) for GAPDH (KP890159), for TUB2 (KP890171), 99.85% for CAL (KP890147) and 99,5 % for ITS (KP890165). The phylogenetic tree constructed by concatenation with the obtained sequences, as well as references, revealed that the C38 isolate clustered within C. grossum, confirming the BLAST approach. Pathogenicity tests were performed on 40 'Story® Inored' apples cleaned and wounded with a sterilized needle and exposed to two different conditions: 20 apples (10 inoculated with 20 µl of spore suspension (105 ml-1) and 10 with sterile water as control) were incubated at 20°C with a 12-hour photoperiod for 14 days, while the remaining 20 apples, prepared with same approach, were placed at 1°C for 3 months, then at room temperature for 14 days. Symptoms appeared after 6 days on apples incubated at 20°C, whereas those stored at 1°C displayed symptoms at 11 days after being placed at room temperature. In both conditions, lesions were similar to those observed on the original fruits; while the controls remained asymptomatic. Identity of reisolated fungal colonies was confirmed by CAL, GAPDH and GS region sequence analysis. C. grossum has been reported rarely: in 2017 on Capsicum annuum var. grossum in China, in 2018 on Mangifera indica leaves in Cuba, and in 2021 on Rhyncospermum jasminoides in Italy (Diao et al. 2017; Manzano León et al. 2018; Guarnaccia et al. 2021). To the best of our knowledge, this is the first report of apple bitter rot caused by Colletotrichum grossum worldwide.

Irrigation Targeted to Provoke Ejection of Ascospores of Venturia inaequalis Shortens the Season for Ascospore Release and Results in Less Apple Scab.

Trials were carried out in apple orchards of Emilia-Romagna and Trentino-Alto Adige in northern Italy to investigate the effects of sprinkler irrigation on possible reduction in inoculum and subsequent disease pressure of Venturia inaequalis, the ascomycete causing apple scab. In spring, volumetric spore traps were placed above apple leaf litter containing pseudothecia with ascospores of the fungus. Pseudothecia matured more rapidly in irrigated plots, and 95% of the total number of spores trapped in a season was reached on average 164 degree days (base temperature 0°C) earlier in irrigated compared to non-irrigated plots. On average for seven location/year combinations, more than 50% of the ascospores were trapped following irrigations carried out for two hours on sunny days before a forecasted rainfall. Subsequently, a much lower number of spores were trapped on rainy days following irrigation. Field trials with scab susceptible apple cultivars were carried out in the two regions to evaluate the efficacy of sprinkler irrigation on disease. Irrigated and non-irrigated plots were either treated with different fungicide control strategies or not treated. Irrigation significantly reduced the incidence of apple scab at both sites, and the overall number of infected leaves and fruit was reduced by more than 50%. Mid-day sprinkler irrigation can significantly reduce the inoculum pressure of V. inaequalis in apple orchards. This may be a sustainable management strategy, especially in areas with extended dry periods.

A public decision support system for the assessment of plant disease infection risk shared by Italian regions.

Integrated pest management (IPM) practices proved to be efficient in reducing pesticide use and ensuring economic farming sustainability. Digital decision support systems (DSS) to support the adoption of IPM practices from plant protection services are required by European legislation. Available DSSs used by Italian plant protection services are heterogeneous with regards to disease forecasting models, datasets for their calibration, and level of integration in operational decision-making. This study presents the MISFITS-DSS, which has been jointly developed by a public research institution and nine regional plant protection services with the objective of harmonizing data collection and decision support for Italian farmers. Participatory approach allowed designing a predictive workflow relying on specific domain expertise, in order to explicitly match actual user needs. The DSS calibration entailed the risk of grapevine downy mildew infection (5-point scale from very low to very high), and phenological observations in 2012-2017 as reference data. Process-based models of primary and secondary infections have been implemented and tested via sensitivity analysis (Morris method) under contrasting weather conditions. Hindcast simulations of grapevine phenology, host susceptibility and disease pressure were post-processed by machine-learning classifiers to predict the reference infection risk. Results indicate that IPM principles are implemented by plant protection services since years. The accurate reproduction of grapevine phenology (RMSE = 4-14 days), which drove the dynamic of host susceptibility, and the use of weather forecasts as model inputs contributed to reliably predict the reference infection risk (88% balanced accuracy). We did a pioneering effort to homogenize the methodology to deliver decision support to Italian farmers, by involving plant protection services in the DSS definition, to foster a further adoption of IPM practices.

Evaluation of a Warning System for Early-Season Control of Grapevine Powdery Mildew.

In several grape-growing areas of the world, including northern Italy, powdery mildew epidemics, caused by Erysiphe necator, are mainly triggered by the ascospores produced in overwintered chasmothecia. Growers in northern Italy usually control the disease with fixed-interval fungicide applications. A warning system was developed for early-season powdery mildew control based on (i) short-term weather forecasts, (ii) a model that simulates the severity of each E. necator ascosporic infection, and (iii) a mobile phone short-message system. This warning system was evaluated in six vineyards in northern Italy from 2006 to 2008, between bud break of vines and early berry development; an unsprayed control was compared with "low-risk" and "high-risk" model-driven sprays and a calendar-based "grower" spray program. Use of the warning system reduced disease severity on leaves and bunches compared with the unsprayed control and resulted in the same level of control of powdery mildew as the grower's spray program, with reduced fungicide applications and costs. On average, 5.7 sprays were applied following the grower's spray program (with an average cost of 221 €/ha/year); use of the warning system reduced fungicide applications by 36% (low-risk program, saving of 56 €/ha/year) or 75% (high-risk program, saving of 161 €/ha/year).