Exploring the genetic basis of resistance to Neopestalotiopsis species in strawberry.

Aggressive strains of Neopestalotiopsis sp. have recently emerged as devastating pathogens of strawberry (Fragaria × ananassa Duchesne ex Rozier), infecting nearly all plant parts and causing severe outbreaks of leaf spot and fruit rot in Florida and globally. The development of host resistance is imperative due to the absence of fungicides that effectively inhibit Neopestalotiopsis sp. growth on an infected strawberry crop. Here, we analyzed 1578 individuals from the University of Florida's (UF) strawberry breeding program to identify and dissect genetic variation for resistance to Neopestalotiopsis sp. and to explore the feasibility of genomic selection. We found that less than 12% of elite UF germplasm exhibited resistance, with narrow-sense heritability estimates ranging from 0.28 to 0.69. Through genome-wide association studies (GWAS), we identified two loci accounting for 7%-16% of phenotypic variance across four trials and 3 years. Several candidate genes encoding pattern recognition receptors, intra-cellular nucleotide-binding leucine-rich repeats, and downstream components of plant defense pathways co-localized with the Neopestalotiopsis sp. resistance loci. Interestingly, favorable alleles at the largest-effect locus were rare in elite UF material and had previously been unintentionally introduced from an exotic cultivar. The array-based markers and candidate genes described herein provide the foundation for targeting this locus through marker-assisted selection. The predictive abilities of genomic selection models, with and without explicitly modeling peak GWAS markers as fixed effects, ranged between 0.25 and 0.59, suggesting that genomic selection holds promise for enhancing resistance to Neopestalotiopsis sp. in strawberry.

Detection of Strawberry necrotic shock virus using conventional and TaqMan(®) quantitative RT-PCR.

Graft-indexing of an advanced selection from the University of Florida strawberry breeding program produced virus-like symptoms on Fragaria vesca. However; RT-PCR testing of the material did not detect the presence of any of 16 strawberry virus species or members of virus groups for which strawberries are routinely indexed. Large scale sequencing of the material revealed the presence of an isolate of Strawberry necrotic shock virus. The nucleotide sequence of this isolate from Florida shows a significant number of base changes in the annealing sites of the primers compared to the primers currently in use for the detection of SNSV thereby explaining the most probable reason for the inability to detect the virus in the original screening. RT-PCR and Taqman(®) qPCR assays were developed based on conserved virus sequences identified in this isolate from Florida and other sequences for SNSV currently present in GenBank. The two assays were applied successfully on multiple samples collected from several areas across the United States as well as isolates from around the world. Comparison between the RT-PCR and the qPCR assays revealed that the qPCR assay is at least 100 times more sensitive than conventional PCR.