Mary E. Mason

Assessment of beech scale resistance in full- and half-sibling American beech families

A beech bark disease infested American beech tree (Fagus grandifolia Ehrh.) and two uninfested trees were selected in a mature natural stand in Michigan, USA, and mated to form two full-sib families for evaluating the inheritance of resistance to beech scale (Cryptococcus fagisuga Lind.), the insect element of beech bark disease. Four half-sib families from both infested and uninfested trees were also evaluated for resistance. Using an artificial infestation technique, adult and egg count data were collected over 2 years and analyzed with generalized linear mixed methods to account for nonnormal distributions of the response variables. A significant effect for family was found for each variable. Family least squares means were computed as a measure of resistance and repeatabilities were calculated to provide an upper limit estimate of broad-sense heritability. The two families that ranked highest for resistance were the full-sib family from two uninfested parents and the half-sib family from a stand where all diseased trees had been removed. Together, the results suggest that selection and breeding may be an effective means to improve populations for artificial regeneration, and silvicultural treatments may provide an effective management option for mitigating beech bark disease through managing the genetic composition of natural regeneration.

Arabidopsis protochlorophyllide oxidoreductase A (PORA) restores bulk chlorophyll synthesis and normal development to a porB porC double mutant

In angiosperms the strictly light-dependent reduction of protochlorophyllide to chlorophyllide is catalyzed by NADPH:protochlorophyllide oxidoreductase (POR). The Arabidopsis thaliana genome encodes three structurally related but differentially regulated POR genes, PORA, PORB and PORC. PORA is expressed primarily early in development—during etiolation, germination and greening. In contrast, PORB and PORC are not only expressed during seedling development but also throughout the later life of the plant, during which they are responsible for bulk chlorophyll synthesis. The Arabidopsis porB-1 porC-1 mutant displays a severe xantha (highly chlorophyll-deficient) phenotype characterized by smaller prolamellar bodies in etioplasts and decreased thylakoid stacking in chloroplasts. Here we have demonstrated the ability of an ectopic PORA overexpression construct to restore prolamellar body formation in the porB-1 porC-1 double mutant background. In response to illumination, light-dependent chlorophyll production, thylakoid stacking and photomorphogenesis are also restored in PORA-overexpressing porB-1 porC-1 seedlings and adult plants. An Arabidopsis porB-1porC-1 double mutant can therefore be functionally rescued by the addition of ectopically expressed PORA, which suffices in the absence of either PORB or PORC to direct bulk chlorophyll synthesis and normal plant development.

Evaluation of Reference Genes for Expression Studies in Ash (Fraxinus spp.)

Ash (Fraxinus spp.) is a dominant tree species in North America, in both managed and natural landscapes. However, due to the rapid invasion by the emerald ash borer (Agrilus planipennis), an exotic invasive insect pest, millions of North American ash trees have been killed. Real-time quantitative polymerase chain reaction (RTq-PCR) is widely used for validating transcript levels in gene expression studies for which a good reference gene is mandatory. In the current study, we evaluated the stability of ten reference genes in at least five different tissues (phloem, roots, shoots, immature leaves, and mature leaves), and two developmental stages (young and old) among three ash species including the resistant Asian Manchurian ash (F. mandshurica) and two susceptible North American ash species (green—F. pennsylvanica and white—F. americana). Of the examined genes, the translation elongation factor alpha (eEF1α) was observed to be most stable and thus is recommended for RTq-PCR based gene expression studies in Fraxinus species. To our knowledge, this is the first report on the stability of reference genes across ash species (in different tissues and during development).

Intraspecific variation in Fraxinus pennsylvanica responses to emerald ash borer (Agrilus planipennis)

The emerald ash borer (EAB; Agrilus planipennis Fairmaire) is a bark and wood boring beetle native to east Asia that was first discovered in North America in 2002. Since then, entire stands of highly susceptible green ash (Fraxinus pennsylvanica Marshall) have been killed within a few years of infestation. We have identified a small number of mature green ash trees which have been attacked by EAB, yet survived the peak EAB infestation that resulted in mortality of the rest of the ash cohort. Adult landing and feeding preference bioassays, leaf volatile quantification and EAB egg bioassay experiments were used to characterize potential differences in responses of these select “lingering” green ash trees relative to known EAB susceptible controls. Three selections were identified as being significantly less preferred for adult feeding, but no specific leaf volatile profile was associated with this reduced preference. Egg bioassays identified two ash selections that had significant differences in larval survival and development; one having a higher number of larvae killed by apparent host tree defenses and the other having lower larval weight. Correlation and validation of the bioassay results in replicated plantings to assess EAB resistance in the field is still necessary. However, the differences between lingering ash selections and susceptible controls measured by these bioassays indicate that more than one mechanism is responsible for the increased resistance to EAB that resulted in these selections surviving longer than their counterparts. Efforts to further increase ash resistance to EAB through use of these selections in a breeding program are underway.

Methods for collecting ash (Fraxinus spp.) seeds

The threat of emerald ash borer (Agrilus planipennis) has necessitated ash seed collection to preserve the ash genetic resource. For ash species found only in natural areas, we have developed seed collection methods that require no special training or certification. Collection techniques for natural areas include pole pruners for branches lower than 30 ft (9 m) and ropes or rope saws for higher branches. Seed collection must be timed correctly by examining samara color, seed coat color, and embryo size to be sure that seeds are fully mature. Documentation of seed identity and post-harvest handling of seeds are discussed.