Real-time PCR assay for Colletotrichum acutatum sensu stricto quantification in olive fruit samples

Olive anthracnose is caused by fungal species within the Colletotrichum acutatum, C. gloeosporioides and C. boninense complexes. Anthracnose causes severe pre- and post-harvest olive drupe fall. This study aimed to design a species-specific qPCR assay, based on klap1 gene, suitable for C. acutatum s.s. quantification in cv. Galega Vulgar fruit samples. The developed qPCR assay presented a detection limit of 10.14 fg/reaction, and a linear cycle threshold of R-2 = 0.996. C. acutatum inoculum was detected in pulverized olive fruits, and in early infection stages, before symptom appearance, 16 h after inoculation (Ct values = 28.29 +/- 1.1). In olive samples, the derived melting curve was specific presenting a single dissociation peak (T-metting = 88.7 degrees C). The designed assay was effectively applied in C. acutatum detection and quantification using infected olive samples, with a LOD of 0.59 ng and a LOQ of 1.8 ng, allowing its application to orchard management.

Automated summary

This study successfully designed a species-specific qPCR assay for quantification of C. acutatum s.s. in cv. Galega Vulgar olive fruit samples, with high sensitivity and linear relationship. The assay detected C. acutatum inoculum in infected olive samples, providing a valuable tool for orchard management.