Abstract
Anthracnose of strawberry, caused primarily by the fungal pathogens belonging to Colletotrichum acutatum species complex (CASC) and C. gloeosporioides species complex (CGSC) is an economically important disease in the Southeast United States. Quiescently infected (QI) planting stock is one of the most important sources of inoculum in the fruiting field that can only be reliably detected by highly sensitive real time quantitative PCR (q-PCR) assay. In this study, a q-PCR assay was developed and optimized that can discriminate anthracnose fruit rot (AFR) and anthracnose crown rot (ACR) causing species based on the difference in post PCR melting temperatures of amplicons. Controlled environment grown plants artificially inoculated with different levels of CASC and CGSC showed a significant (P < 0.001) correlation with levels of quantification expressed by Ct values in q-PCR from petioles and leaf blades. The leaf blade was a significantly larger reservoir of QI than that of the petiole. Both TaqMan and SYBR Green assay showed similar sensitivity and specificity. Detection of QI on leaves at young middle and older stages from inoculation with same number of conidia indicated that middle aged leaves were the best for assessing QI. Quantification of QI from middle aged leaf samples from a strawberry fruiting field that has been planted with pre-inoculated plants at both ends of rows and let inoculum spread showed higher sensitivity and precision by q-PCR compared to that of a traditional paraquat assay. The assay developed and validated in this study offers a new tool for evaluating planting stocks for QI to make decision on preventative control for strawberry anthracnose.
Highlights
The devastating losses caused by anthracnose fruit rot (AFR) and crown rot (ACR), in combination with the regional conducive weather and susceptibility of the current favored cultivars, make it one of the most economically important diseases of strawberry in the Southeast United States (SE) [1] [2]
anthracnose crown rot (ACR) can kill a whole strawberry plant by aggressively invading and producing a reddish-brown to marbled orange necrosis of the crown tissue that causes the plant to wilt and collapse [3]. Both Colletotrichum acutatum species complex (CASC) and C. gloeosporioides species complex (CGSC) can infect all parts of a strawberry plant, fruit rot is the major loss manifested by CASC infection, while crown rot and plant wilting are from CGSC
Optimization of sampling protocol and Quiescently infected (QI) detection methods is essential for advancing diagnostic procedure for anthracnose diseases in strawberry
Summary
The devastating losses caused by anthracnose fruit rot (AFR) and crown rot (ACR), in combination with the regional conducive weather and susceptibility of the current favored cultivars, make it one of the most economically important diseases of strawberry in the Southeast United States (SE) [1] [2]. ACR can kill a whole strawberry plant by aggressively invading and producing a reddish-brown to marbled orange necrosis of the crown tissue that causes the plant to wilt and collapse [3] Both Colletotrichum acutatum species complex (CASC) and C. gloeosporioides species complex (CGSC) can infect all parts of a strawberry plant, fruit rot is the major loss manifested by CASC infection, while crown rot and plant wilting are from CGSC. Contaminated planting stock is not symptomatic at lower level of infection [8] [9], and this can lead to the inadvertent introduction of the pathogen(s) to the fruiting field, a major challenge for fruit growers [6] [10] [11] This contamination can be prevented with a highly sensitive DNA based diagnostic tool.
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