Abstract
Apple bitter rot caused by Colletotrichum species is a growing problem worldwide. Colletotrichum spp. are economically important but taxonomically un-resolved. Identification of Colletotrichum spp. is critical due to potential species-level differences in pathogenicity-related characteristics. A 400-isolate collection from New York apple orchards were morphologically assorted to two groups, C. acutatum species complex (CASC) and C. gloeosporioides species complex (CGSC). A sub-sample of 44 representative isolates, spanning the geographical distribution and apple varieties, were assigned to species based on multi-locus phylogenetic analyses of nrITS, GAPDH and TUB2 for CASC, and ITS, GAPDH, CAL, ACT, TUB2, APN2, ApMat and GS genes for CGSC. The dominant species was C. fioriniae, followed by C. chrysophilum and a novel species, C. noveboracense, described in this study. This study represents the first report of C. chrysophilum and C. noveboracense as pathogens of apple. We assessed the enzyme activity and fungicide sensitivity for isolates identified in New York. All isolates showed amylolytic, cellulolytic and lipolytic, but not proteolytic activity. C. chrysophilum showed the highest cellulase and the lowest lipase activity, while C. noveboracense had the highest amylase activity. Fungicide assays showed that C. fioriniae was sensitive to benzovindiflupyr and thiabendazole, while C. chrysophilum and C. noveboracense were sensitive to fludioxonil, pyraclostrobin and difenoconazole. All species were pathogenic on apple fruit with varying lesion sizes. Our findings of differing pathogenicity-related characteristics among the three species demonstrate the importance of accurate species identification for any downstream investigations of Colletotrichum spp. in major apple growing regions.
Highlights
Apple bitter rot caused by Colletotrichum species is a growing problem worldwide
Facing challenges in species delimitation within C. gloeosporioides species complex (CGSC), we increased the number of partially sequenced genes to eight for isolates belonging to CGSC, adding ACT, ApMat, CAL, glutamine synthetase (GS) and APN2, and reduced the number of isolates for identification and downstream analyses to 44 (19 from morphotype 1 and 25 from morphotype 2)
The assorting of Colletotrichum isolates recovered from apple fruit in this study to C. acutatum species complex (CASC) and CGSC using a multiplex-PCR49, confirmed the reliability and affordability of this method to differentiate between these two species complexes
Summary
Apple bitter rot caused by Colletotrichum species is a growing problem worldwide. Colletotrichum spp. are economically important but taxonomically un-resolved. Among the nine major clades of Colletotrichum, the C. gloeosporioides species complex (CGSC) and the C. acutatum species complex (CASC) are the two most common clades that cause bitter rot of apple[2,16,17]. Species such as C. limetticola, C. paranaense, C. melonis in CASC and C. fructicola in CGSC cause Glomerella leaf spot (GLS) of apple[27]. These two diseases are associated with the same fungal genus, differences in pathogenicity, morphology and cultural characteristics of species have been reported[28]. The ITS regions known as the barcode locus for fungi[36,37], is considered insufficient to delimit species in the CGSC37–39
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