Incidence and effects of Colletotrichum acutatum J.H. Simmonds 1968, in Dryocosmus kuriphilus Yatsumatsu (Hymenoptera Cynipidae) galls on Castanea sativa Mill

Castanea sativa Mill., is a typical nut crop of the Mediterranean basin and one of the most common forest trees in the hilly parts of Italy (Borghetti et al., 1986). In Lombardy, is present in Como, Lecco, Sondrio, Varese, Pavia and Brescia provincies mainly in mixed forests (Boriani et al.,2013). C.sativa is one of the best example of a forest tree species threatened by invasive pathogens and insect pests, with major impacts caused by Cryphonectria parasitica (Murrill M.E.) Barr, Phytophthora spp. and Dryocosmus kuriphilus Yasumatsu (ACGW) (Turchetti et al., 2008). ACGW causes on C.sativa, galls on young twigs, on leaf petioles or on leaves. They measure 5-20 mm in diameter and are green or rose-colour and are readily detected on plants. In 2014, during the insect monitoring, the occurrence of Colletotrichum acutatum in necrotized galls of ACGW in chestnut stands was recorded for the first time in Italy (Gaffuri et al., 2015). Thus, the overall goal of this work, has been to investigate the necrotized galls in order to isolate and identify the causative agent. Consequently, it has been necessary to organize a monitoring and sampling of shoots bark and galls on C.sativa. For the first time, we discoved C.acutatum in Lombardia and Trentino orchards, and from monitoring data, this fungus was found to be spread in all chestnut-growing areas monitored on necrotic galls. Furthermore, during this survey, single or few nuts showed a still undescribed symptom: “Pink rot”, a clear and sometimes intense pink coloration of the endosperm. This symptoms never been reported previously. C.acutatum is commonly found on cultivated and weeds plants and it was associated to anthracnosis, with subsequent and progressive necrosis of green tissues. However, these symptoms were not observed on chestnut. Since this study represents the first reporting of C.acutatum in chestnut and galls of ACGW, other molecular analysis on barcode genes were necessary to confirm the identification. The genus Colletotrichum has undergone frequent taxonomic changes in the past decades with the merging and addition of many species (Baroncelli et al., 2017) and now are recognized as species complex (Damm et al., 2012). A total of 360 samples from shoots and necrotic or healthy galls were taken in order to determine the presence of fungi. From the isolation it was possible to identify the presence of cultures referred to Colletotrichum sp. in nineteen area out of the monitored forty. Positive isolation of other fungi as Trichoderma sp, Fusarium sp., Cryphonectria parasitica and Gnomognopsis sp. were generally obtained in all the areas. Colletotrichum coltures were obtained from the province of Trento, Bergamo, Como and Lecco. Molecular analisys of Colletotrichum isolates have been identified as C. fioiriniae and the analysis of internal transcribed spacer (ITS), tubulin (TUB) and calmodulin (CAL) nucleotide sequence data strongly supported these results. C. fioriniae, as shown in Baroncelli et al., 2014, belongs to C. acutatum species complex and from recorded data, we can state that C.fioriniae is present in Italy on C.sativa. Despite C.fioriniae causes anthracnose on different plants (Pszczółkowska et al.,2016) to date, there is still no correlation between symptoms and the presence of C.acutatum on C.sativa though it was observed a specific symptom called “pink rot“ (Gaffuri et al., 2016). The discovering of this almost worldwide pathogen associated to a new symptomatology on chestnuts confirms a possible risk related to its presence, because the endophytic isolates showed the same pathogenicity of those obtained from infected nuts. Considering the potential pathogenic role of this fungus, many questions still remain unanswered: its presence on healthy chestnut trees opens interesting views on its ecological role and impact both on chestnut ecosystem and on other host.