Andrew J. David

Proteomic Comparison of Needles from Blister Rust-Resistant and Susceptible Pinus strobus Seedlings Reveals UpRegulation of Putative Disease Resistance Proteins

In order to characterize a hypersensitive-like reaction in selected Pinus strobus seedlings to Cronartium ribicola, a proteomic comparison of needles from resistant and susceptible seedlings was undertaken using two-dimensional gel electrophoresis (2-DE). The results revealed 19 polypeptides specific to resistant seedlings and seven of these specific to infected resistant seedlings. There were 13 polypeptides up-regulated (> or = 3-fold increase) in resistant family P327 in comparison to needle tissue from susceptible and mock-inoculated seedlings. Electrospray ionization liquid chromatography and tandem mass spectrometry was used to sequence 11 proteins from the 2-DE gels. Sequences obtained from electrospray ionization liquid chromatography and tandem mass spectrometry were used for MS-BLAST and Pro-ID database searches allowing identification with a 95 to 99% confidence level. Six proteins were determined to be homologs of proteins with known roles in disease resistance, five were determined to be homologs of members of the leucine-rich repeat (LRR) superfamily, and one was a homolog of heat shock protein 90, a protein that serves as a cofactor for certain LRR proteins. This is the first report of members of the LRR family with functional homologs in Pinus strobus and of a molecular basis for white pine blister rust resistance in Pinus strobus.

Epicuticular wax and white pine blister rust resistance in resistant and susceptible selections of eastern white pine (Pinus strobus)

ABSTRACT Epicuticular wax on needles was evaluated for its influence on Cronartium ribicola infection of resistant and susceptible selections of Pinus strobus. Environmental scanning electron microscopy comparisons revealed that needles from a resistant selection of eastern white pine, P327, had a significantly higher percentage of stomata that were occluded with wax, fewer basidiospores germinating at 48 h after inoculation, and fewer germ tubes penetrating stomata than needles from a susceptible selection H111. In addition, needles from seedlings that failed to develop symptoms 6 weeks after inoculation, from a cross between P327 and susceptible parent H109, had a significantly higher percentage of stomata occluded with wax compared with needles from seedlings that developed symptoms. In experiments where epicuticular waxes were removed from needles before seedlings were infected, resistant seedlings without wax developed approximately the same number of infection spots (as measured by spot index) as susceptible seedlings with wax intact. Gas chromatography/mass spectrometry comparisons of extracted epicuticular waxes revealed several peaks that were specific to P327 and not found in susceptible H111 suggesting biochemical differences in wax composition. These results implicate the role of epicuticular waxes as a resistance mechanism in P. strobus selection P327 and suggest a role for waxes in reducing spore germination and subsequent infection through stomatal openings.

Histology of White Pine Blister Rust in Needles of Resistant and Susceptible Eastern White Pine

White pine blister rust, Cronartium ribicola, has plagued the forests of North America for almost a century. Over past decades, eastern white pine (Pinus strobus) that appear to tolerate the disease have been selected and incorporated into breeding programs. Seeds from P. strobus with putative resistance were collected from Oconto River Seed Orchard, Nicolet National Forest, WI. Seedlings were grown for 5 months and artificially inoculated with basidiospores of C. ribicola in two replicated greenhouse experiments. Needles from infected seedlings were fixed, sectioned, and stained with a variety of histological reagents, and rate of mortality for the remaining seedlings was monitored. The most susceptible families suffered 50% mortality in approximately half the time of the more resistant families. Extensive inter- and intracellular hyphae were observed in needles from seedlings of susceptible families, whereas hyphal proliferation was restricted in needles of resistant seedlings. Needles from resistant families had pronounced responses to infection. Phenolics, observed with phloroglucinol-HCl staining, were deposited around infection sites where dense mycelial masses were present. Abnormal host cell growth and rapid cell death in the immediate area of infection were also observed in some eastern white pine families.

Comparison of selection methods for optimizing genetic gain and gene diversity in a red pine (Pinus resinosa Ait.) seedling seed orchard

Abstract Four selection methods, individual selection (IS), family selection (FS), family plus within-family selection (FWFS) and combined selection (CS), were used to estimate genetic gain [E(g)] for stem volume and gene diversity (GD) for ten theoretical selection intensities in a 108-family, 12-year-old red pine seedling seed orchard. Estimated genetic gain for stem volume ranged from 4.6% to 11.8% across all selection methods and intensities with CS consistently having the highest gains and FS the lowest for any given selection intensity. Genetic diversity ranged from 0.9797 to 0.9954 across all selection methods and intensities. Individual selection was the best selection method for retaining GD, especially at the higher selection intensities, while FWFS was more efficient at the lowest selection intensity. An optimization point, which maximized E(g) and GD relative to each other, was calculated for each selection method. In all cases the optimization point indicated that both E(g) and GD would be favorably high when optimized relative to each other. The implications for volume gain, genetic diversity and potential inbreeding in red pine, a species with inherently low levels of genetic variation, are discussed.

Genetic gain and gene diversity following thinning in a half-sib plantation

Abstract Status number, gene diversity, inbreeding coefficient and genetic gain were calculated following phenotypic rogueing of different intensities in a half-sib progeny plantation of Scots pine (Pinus sylvestris L.). Across most selection intensities the status number, gene diversity and genetic gain remained favorably high while the inbreeding coefficient was remarkably stable. It is suggested that phenotypic selection in half-sib progeny tests or plantations with unknown pedigrees, can be used to manage seed collection areas or as a component in low-input breeding without a fast build up of coancestry or inbreeding, provided the initial number of progenies of unrelated parents is sufficiently high and that a high number of these families are retained with a few individuals per family.