Team members:<\/p>\n
The core set of accessions representing Cucurbita pepo<\/em> diversity in the NPGS has been self-pollinated and is being combined with heirloom cultivars to anchor market classes and enrich for cultivar genetics. Sources of resistance and other representatives of diversity in other species to extend the utility of the panel and being selfed pollinated. The goal of representatives from other species is both because squash improvement often involves crosses between species and to extend the benefits of CucCAP investments to those that work with other species such as C. moschata<\/em> (Puerto Rico). The second round of self pollination will likely take place with a subcontractor. The process is now including strain purification within the stocks for accessions that do not match their descriptors for hull-less seeded accessions that aren\u2019t, we are creating new selfed stocks from hull-less segregants.<\/p>\n Three projects are already taking advantage of the GWAS population. For CucCAP, given the lack of phenotypic data, we have phenotyped the collection for qualitativetraits of bush growth habit and hull-less seeds. Markers were created with this material validated in breeding populations to include as part of the MS. A separate study is mapping cotyledon cucurbitacin content to support results from biparental populations.<\/p>\n Roughly 4,000 -40,000 quality SNPs were called in each of the species collections (C. pepo., C. moschata <\/em>and C. maxima<\/em>), which ranged in size from 314 to 829 accessions. Filtered SNPs were used for population structure analysis. Available geographical, phenotypic, and other metadata were retrieved from GRIN and were used to help interpret structure results. These data support five ancestral groups in each of the species. Population structure was driven mostly by geography, except in where the presence of different subspecies was responsible for some of the structure. Filtering all available historical data from GRIN traits with at least 100 entries resulted in 21 traits for C. pepo<\/em>, 5 for C. moschata<\/em> and 16 for C. maxima<\/em> for further analysis. Traits spanned fruit and agronomic-related characteristics, as well as pest resistances. Fruit traits included fruit width, length, surface color and texture, and flesh color and thickness. Agronomic data included plant vigor and vining habit, andseveral phenotypes related to maturity. Pest-related traits included susceptibility to cucumber beetle and squash bug in C. pepo<\/em> and Watermelon mosaic virus (WMV) and powdery mildew (PM) in C. maxima<\/em>. Marker-based narrow-sense heritability was calculated for each trait and ranged from 0.12 to close to 1. Most traits had moderate to high heritabilities (\u22650.4). Regression of trait data on the Q matrix obtained from structure analysis was used to determine the amount of phenotypic variation explained by population structure. In C. pepo<\/em>, traits related to fruit morphology tended to have high correlations with population structure. Genome wide association was conducted for all traits using standard mixed-model analysis. No significant signals were detected in C. moschata<\/em>. A weak signal was detected in C. maxima<\/em> for fruit set on chromosome 12 and fruit ribbing on chromosome 17. Three phenotypes were significantly associated with SNPs in C. pepo<\/em>: bush\/vine plant architecture on chromosome 10, fruit flesh color on chromosome 5, and fruit width on chromosome 3.<\/p>\n Unlike powdery mildew resistance, virus resistance has not been as widely deployed in commercial breeding lines. Therefore, while we were able to implicate a genomic region in PRSV resistance using a cultivar based association mapping approach, this was not definitive. To validate this region, we created a biparental F2:3 mapping population from a cross between Whitaker (C. pepo<\/em> subsp pepo<\/em>) and Success PM (C. pepo<\/em> subsp ovifera<\/em>). One hundred three of these families were inoculated with PRSV and phenotyped for the recessive single generesistance derived from C. equadorensis<\/em>. We designed SNP markers that we can test in this population once campus reopens to research.<\/p>\n Lauren Brzozowski, a recent PhD in the Mazourek group, used the genotype data from the C. pepo<\/em> collection to do targeted phenotyping of cucurbitacin content. She currently has a paper in review describing the mapping and candidate genes for the Bi-4<\/em> locus in squash which drives striped cucumber beetle herbivory in cotyledons mediated by the presense of cucurbitacins (Brzozowski et al, in review). Striped cucumber beetles are a vector of Squash mosaic virus<\/em>. There is no known genetic resistance to this virus in Cucurbita and the neonicotinoid chemistry used to control these insects have increased regulatory scrutiny in the US and usage restrictions in Europe.<\/p>\n In this report we aggregate data from 2019-2020 with data collected in previous years concerning inheritance of resistance to Papaya ringspot virus (PRSV) in tropical pumpkin (Cucurbita moschata). Some of the conclusions made in previous annual reports have been modified after taking into consideration the additional data. For most of the populations, the new data doubles the number of individuals tested in F2<\/sub> families resulting in much more robust inheritance tests.<\/p>\n Two sources of resistance are well known in C. moschata<\/em>: \u2018Nigerian Local\u2019 and \u2018Menina\u2019. The inheritance of resistance from \u2018Nigerian Local\u2019 has been previously studied, but prior to the CucCAP project inheritance studies have not been reported for \u2018Menina\u2019, nor was it known if resistance to PRSV in \u2018Nigerian Local\u2019 is allelic to that in \u2018Menina\u2019. In our inheritance study susceptible genotypes were \u2018Verde Luz\u2019, \u2018Taina Dorada\u2019 and \u2018TP411\u2019. The third to fifth leaf of inoculated seedlings were rated on a 0 to 4 scale for disease severity and scores were combined to convert to a 0 to 12 scale. Resistant x susceptible F2<\/sub> populations using \u2018Nigerian Local\u2019 as the source of resistance (distributions on the left-hand side of Figure 1) had somewhat normal distributions with an average disease severity of 5.385 (n=231) in Ta\u00edna Dorada x Nigerian Local and 6.02 (n=208) in Verde Luz x Nigerian Local. In contrast, F2populations with \u2018Menina\u2019 (distributions on the right-hand side of Figure 1) were strongly skewed towards resistance with an average severity of 3.52 (n=220) in Ta\u00edna Dorada x Menina, 3.34 (n=214) in Verde Luz x Menina and 2.80 (n=111) in TP411 x Menina. The Nigerian Local x Menina (resistant x resistant) F2<\/sub> population was very strongly skewed toward resistance, with an average combined severity of 0.840 (Figure 2).<\/p>\n To carry out chi-square goodness of fit tests, we grouped plants with an overall severity rating of <4 as resistant and plants with an overall severity rating of >5 as susceptible. The best fit for segregation in F2<\/sub> crosses made with \u2018Nigerian Local\u2019 was to a 7:9 (R:S) genetic model, although the fit was very poor for Verde Luz x Nigerian Local (Table 1). Goodness-of-fit to other two-class models were much worse than for 7:9. All three crosses using \u2019Menina\u2019 fit a 3:1 model. The resistant x resistant cross (Nigerian Local x Menina) fit a 15:1 model. These segregations suggest that at least two genes are involved in the inheritance of resistance to PRSV for \u2018Nigerian Local\u2019 while a single dominant gene might be responsible for the resistance of \u2018Menina\u2019. The data clearly indicate that at least some of the genes for resistance in \u2018Nigerian Local\u2019 and \u2018Menina\u2019 are different. Our data indicates that the resistance to PRSV conferred by \u2018Menina\u2019 is superior to that of \u2018Nigerian Local\u2019. If the resistance of \u2018Menina\u2019 is a single dominant gene as this data suggests, then it will likely be easier to identify resistance markers in \u2018Menina\u2019 than in \u2018Nigerian Local\u2019.<\/p>\nPI collection genomics<\/h4>\n
PRSV resistance – Construction of RIL population for mapping<\/h4>\n
Insect resistance<\/h4>\n
University of Puerto Rico, Mayag\u00fcez, Annual Report \u2013May 2020<\/h3>\n
Figure 1. (below) Distributions of symptom severity in F2<\/sub> populations of tropical pumpkin (Cucurbita moschata) inoculated with Papaya ringspot virus (PRSV). Populations developed with resistant parent \u2018Nigerian Local\u2019 are shown on the left; populations developed with resistant parent \u2018Menina\u2019 are shown on the right. For each plant, disease severity in leaf position 3, 4 and 5 was evaluated on a 0 to 4 scale (0 = no symptoms). Values were summed to produce an overall severity index of 0 to 12.<\/h3>\n
Figure 2. Distribution of combined severity ratings of plants (n=238) from the Nigerian Local x Menina F2<\/sub> population inoculated with Papaya ringspot virus (PRSV). For each plant, disease severity in leaf position 3, 4 and 5 was evaluated on a 0 to 4 scale (0 = no symptoms). Values were summed to produce an overall severity index of 0 to 12<\/h3>\n