Changes in volatile terpene and diterpene resin acid composition of resistant and susceptible white spruce leaders exposed to simulated white pine weevil damage

Induced (traumatic) resin in white spruce (Picea glauca (Moench) Voss) leaders resistant or susceptible to the white pine weevil (Pissodes strobi Peck) was analyzed for volatile terpenes and diterpene resin acids after simulated white pine weevil damage. Leaders from 331 trees were wounded just below the apical bud with a 1-mm diameter drill, coinciding with the natural time of weevil oviposition in the spring. Leaders were removed in the fall, and the bark and xylem from the upper and lower regions of the leader extracted and analyzed by gas chromatography. Unwounded trees had low amounts of resin in xylem compared with bark. In response to wounding, volatile terpenes and diterpene resin acids increased in the upper xylem (area of wounding), with resistant trees showing a greater increase than susceptible trees. Wounding caused monoterpenes in particular to decrease in the lower region of the leader (away from the drilled area) in greater amounts in susceptible trees than in resistant trees. In response to wounding, the proportion of monoterpene to resin acid increased in the upper and lower xylem of resistant trees, and slightly increased in the upper xylem of susceptible trees. Monoterpene-enriched resin is more fluid than constitutive resin, and probably flows more readily into oviposition cavities and larval mines, where it may kill immature weevils. Loss of resin components in the lower xylem suggested catabolism and transport of these materials to the site of wounding; however, energetic and regulatory data are necessary to confirm this hypothesis. This study provides a basis for measuring the ability of a tree to undergo traumatic resinosis that could be used to screen for resistance to white pine weevil.     

Inhibiting factors of symptom development in several Japanese red pine (<Emphasis Type="Italic">Pinus densiflora</Emphasis>) families selected as resistant to pine wilt

Pinus densiflora and P. thunbergii, native to Japan, are highly susceptible to pine wilt disease caused by infection with a pine wood nematode (Bursaphelenchus xylophilus). Trees of these susceptible species have occasionally been found surviving in forests that are extensively damaged by this disease. Seedlings from a part of surviving trees that were selected as resistant families indicate lower mortality rates after the infection. The factors that prevent the symptoms from developing in resistant families of a susceptible species, P. densiflora, as based on the analysis of the pathogens behavior in the tree tissue and the anatomy of the resistant families, are presented in this paper. Nematode populations remained lower in the stems of seedlings from resistant families of P. densiflora than in the stems of non-resistant families. Areas dysfunctional in water transport developed in the stems of resistant families, but did not reach a size large enough to seriously block the ascent of sap. These results suggest that there are systems within the seedlings that prevent nematode migration and reproduction. The 2-year-old seedlings from resistant families of P. densiflora, however, did not suppress the pathogen activity. Numerous branches are a visible characteristic in the seedlings of some resistant families. The arrangement of the resin canals, the only channels in the tree to the pathogen migration, was disoriented at the joints between the branches and the main stem. Such a structure may be effective as a barrier to nematode dispersal.     

Resin canal traits relevant for constitutive resistance of Norway spruce against bark beetles: environmental and genetic variability

The influence of genetic determination and environment on the variability of secondary resin canal traits was investigated on 15–19-year-old Norway spruce (Picea abies (L.) Karst.) clones grown in Lower Austria (Pressbaum and Ulmerfeld) and southern Sweden (Knutstorp and Hermanstorp). Eleven and 20 clones were present on the two Austrian and the two Swedish sites, respectively. The sites differed in their water availability, as indicated by different annual precipitation and soil type. Resin canal traits measured were the number of epithelial cells per canal, the number of resin canals per unit tangential wood surface area, the mean resin canal area and the total resin canal area per unit tangential wood surface area. The latter three traits are known to be related to the constitutive resin flow of Norway spruce.

Environment had an influence on the variability of resin canal traits but the most important factor for the variability was the tree’s genetic disposition. Within countries, clones from the drier sites (Pressbaum and Hermanstorp) showed significantly smaller resin canals. Trees from Pressbaum also had smaller total resin canal areas than trees from Ulmerfeld. The number of epithelial cells and the number of canals did not differ between sites.

Resin canal traits had wide genetic variation and high broad sense heritabilities (H²), with values between 0.28 and 0.82. Highest heritability values were reached for the number of epithelial cells and the number of canals (H² > 0.8). Genotypic correlations across trials were high for the resin canal traits and approached 1 in both the Austrian and the Swedish trials, indicating that there was little genotype by environment interaction for these traits and thus the ranking of clones was very similar in the different environments.

The number of epithelial cells, the mean area and the total resin canal area showed either moderately significant positive genotypic correlations with tree growth traits or none at all.

In a breeding context, our results are encouraging and indicate that high constitutive defence potential against bark beetles, such as Ips typographus, is not aligned with low volume growth. But it should be taken into consideration that environment and forestry practices can also have an impact on the resin reservoir provided by the radial resin canals.     

Breeding eastern white pine: A world-wide perspective

Eastern white pine (Pinus strobus L.) has wide genetic variability and high productivity in plantations in North America, Europe and the Far East. In regions where neither white pine blister rust nor the white pine weevil are a problem, as in the central states of the U.S.A. and the warmer parts of Europe, primary attention is given to selection and breeding for vigor. Estimates for trees up to age 18 indicate that a first-generation genetic gain in volume of over 20% can be obtained from open-pollinated family selection within stands. Volume gains of 50% or more over local genotypes have been realized at ages 15–20 through the use of seed from selected southern Appalachian stands and in some individual tree crosses of eastern white pine with blue (Himalayan) pine. In the cool, humid regions of eastern North America and Europe, where blister rust resistance is important, breeding strategies include cloning of resistant P. strobus individuals, use of blue and Balkan pines in breeding programs, and exploitation of low-level rust resistance in low-risk localities. Eastern white pine losses from blister rust can be avoided in Japan and Korea by good plantation management without genetic selection. In some regions where the white pine weevil is the major problem, western white pine is useful both for hybridization and as an alternative, more weevil-resistant species. Analysis of relative photosynthesis of SO₂- and O₃-fumigated clones may be useful as a method of screening for pollution tolerance in regions where air pollution is likely to intensify. Efficient techniques for cloning by rooting of cuttings are now available. Efforts are under way to develop technology for micropropagation for mass production of superior clones. Breeding strategies for the major P. strobus plantation regions of the world are summarized.     

Variation in Elatobium abietinum Attack on Picea glauca and its Relation to Pissodes strobi Resistance

White spruce (Picea glauca (Moench) Voss) is host to several pests, including the white pine weevil (Pissodes strobi (Peck)) and the green spruce aphid (Elatobium abietinum (Walker)). The larvae of the white pine weevil damage spruce leaders by consuming the cortex while the green spruce aphid is a defoliator. White spruce emblings (seedlings produced by culturing tissues from seed embryos) from 18 families previously ranked for resistance to the white pine weevil were defoliated to varying degrees by the green spruce aphid in a natural outbreak that developed within a holding shadehouse. A strong relationship was shown between damage caused by the aphids and weevil resistance. Emblings ranked as highly weevil - resistant sustained significantly less aphid defoliation.     

Variation in damage by terminal shoot insects among jack pine families

The susceptibility of jack pine, Pinus banksiana Lamb., to damage by the white pine weevil, Pissodes strobi (Peck) and the eastern pine shoot borer, Eucosma gloriola Heinrich, was examined over a three-year period (1994–1996) in a 400-family genetic test near New Liskeard, Ontario. Of the 7180 trees examined, white pine weevil damaged 1041 and eastern pine shoot borer damaged 1913 trees, yielding damage frequencies of 14.5% and 26.6%, respectively. Thirty families of jack pine never sustained weevil damage during the study period, while only one family escaped damage by the shoot borer. The expected, and observed, damage rates were significantly different for the weevil, but not for the shoot borer. There was no discernible spatial pattern in the incidence of shoot damage by either insect species, based on the geographic location of the parent trees. The data suggest that there is a basis for pursuing further studies to evaluate genetic resistance in tree improvement programs.     

The characteristics of oleoresin exudation amount from the trunks of pine trees

The amounts of oleoresin exudation from artificially-marked wounds on the trunks of pine trees were measured, and several characteristics of the oleoresin exudation amount (OEA) were elucidated. OEA showed no regular diurnal change and had no relation to both heights and directions of the wounds marked on the trunks, but differed between individual trees and between locations on the same tree. The exudation of oleoresin from each wound stopped after several hours. Mean OEA per wound decreased with increase in the number of wounds per given area. When the exudation of oleoresin from a wound was exhausted, further oleoresin was scarcely exuded even from a new wound made close to the initial one. In response to wounding, therefore, primary oleoresin exudation seems to occur by releasing the resin stocked in the resin canal.     

European larch and eastern white pine respond similarly during three years of partial defoliation

To test whether trees with different leaf life spans respond differently to defoliation, eastern white pine (Pinus strobus L.) and European larch (Larix decidua Mill.) trees (9 years old in 1991) were partially defoliated by hand between July 1 and 10 in 1989, 1990 and 1991. At the end of 1991, trees of both species had received either 0, 1, 2 or 3 years of defoliation. Trees that received only 1 year of defoliation were defoliated in 1989. Variables measured included photosynthesis, twig water potential, leaf mass per area and leaf nitrogen concentration. There were few significant responses to defoliation in any of the three years of treatment in either species, and only the current-year defoliation treatments caused significant responses. Both species had reduced photosynthetic rates and less negative twig water potentials in response to defoliation in 1989. In 1990 and 1991, the defoliation treatments had no significant effect on any of the parameters measured in European larch. In 1990, there was a significant reduction in foliar nitrogen concentration in eastern white pine in response to defoliation in 1990. In 1991, eastern white pine had significantly less negative twig water potentials in response to defoliation in 1991. Leaf mass per area was not affected by defoliation in either species. We conclude that, for European larch and eastern white pine, differences in leaf life span have no effect on leaf- and twig-level responses to defoliation.     

Limited response of ponderosa pine bole defenses to wounding and fungi

Tree defense against bark beetles (Curculionidae: Scolytinae) and their associated fungi generally comprises some combination of constitutive (primary) and induced (secondary) defenses. In pines, the primary constitutive defense against bark beetles consists of preformed resin stored in resin ducts. Induced defenses at the wound site (point of beetle entry) in pines may consist of an increase in resin flow and necrotic lesion formation. The quantity and quality of both induced and constitutive defenses can vary by species and season. The inducible defense response in ponderosa pine is not well understood. Our study examined the inducible defense response in ponderosa pine using traumatic mechanical wounding, and wounding with and without fungal inoculations with two different bark beetle-associated fungi (Ophiostoma minus and Grosmannia clavigera). Resin flow did not significantly increase in response to any treatment. In addition, necrotic lesion formation on the bole after fungal inoculation was minimal. Stand thinning, which has been shown to increase water availability, had no, or inconsistent, effects on inducible tree defense. Our results suggest that ponderosa pine bole defense against bark beetles and their associated fungi is primarily constitutive and not induced.     

Histopathology of primary needles and mortality associated with white pine blister rust in resistant and susceptible Pinus strobus

White pine blister rust caused by Cronartium ribicola is a damaging non‐native disease of five‐needled pines in North America. Efforts to control the disease and mitigate damage to date have been only somewhat effective. Recent efforts to improve the health of eastern white pine and reestablish the tree as a dominant species in the North Central United States have focused on identification and propagation of disease‐free eastern white pine (Pinus strobus) growing in areas with a high incidence of blister rust. Many of these selections have been shown to resist infection following artificial inoculation with C. ribicola. In this study, 13 eastern white pine families derived from controlled pollination of selections previously determined to possess putative resistance as well as susceptible selections were inoculated with C. ribicola. Mortality data from inoculation studies show superior survivability in three families with over 60% of seedlings able to survive the 52 week post‐inoculation monitoring period compared to 0–10% survival of the most susceptible families. Primary needles were collected for histological analysis from all inoculated families 4 weeks after inoculation and from selected families 6.5 weeks and 38 weeks after inoculation. Histological observations of infection sites show distinct resistance reactions in the families more likely to survive infection based on mortality data. Analysis of the reactions in susceptible families revealed extensive hyphal colonization of the vascular bundle and adjacent mesophyll cells that appear uninhibited by tree responses. In resistant families, collapsed cells adjacent to infection sites, heavy deposition of phenolic compounds and abnormal cell growth were documented more frequently and appear to play an integral role in the ability of these eastern white pine families to impede growth of C. ribicola in primary needle tissue.     

Comparison of histological responses and tissue damage expansion between resistant and susceptible Pinus thunbergii infected with pine wood nematode Bursaphelenchus xylophilus

Pine wilt disease caused by the pine wood nematode (PWN), Bursaphelenchus xylophilus, has been epidemic and has had disastrous impacts on pine forests and forest ecosystems in eastern Asia. Many pine species in this area are susceptible to this disease. Pinus thunbergii is particularly susceptible. In Japan, tree breeders have selected surviving trees from severely damaged forests as resistant candidates, and have finally established several resistant varieties of P. thunbergii. However, this breeding procedure requires much time and effort due to the lack of physiological and phenotypical information about resistance. To investigate the resistance mechanisms of selected P. thunbergii, we compared histochemical responses, tissue damage expansion, and PWN distribution in resistant and susceptible clones of P. thunbergii after PWN inoculation. The results suggested that the mechanisms of resistance are as follows: damage expansion in the cortex, cambium, and xylem axial resin canals are retarded in resistant trees soon after inoculation, probably due to the induction of wall protein-based defenses. Suppression of PWN reproduction was particularly caused by inhibition of damage expansion in the cambium. The slow expansion of damage in each tissue provides time for the host to complete the biosynthesis of lignin in the walls of cells that surround the damaged regions. This lignification of cell walls is assumed to effectively inhibit the migration and reproduction of the PWNs. The mechanism of initial damage retardation is presumed to be a key for resistance.     

Migration of Bursaphelenchus xylophilus in cortical and xylem axial resin canals of resistant pines

Migration of the pine wood nematode (PWN), Bursaphelenchus xylophilus, in susceptible and resistant pines was investigated at the tissue level. PWN was inoculated onto the top cross‐cut surface of 20‐cm stem cuttings of susceptible Pinus thunbergii and resistant pines (P. strobus, P. rigida and P. thunbergii of a resistant family Namikata‐(t)‐73 (half‐sib)). PWNs were mainly distributed in cortical resin canals of susceptible P. thunbergii down to 15 cm from the inoculated surface by 6 h after inoculation (HAI) and all tissues (including cortical and xylem resin canals) down to the bottom at 192 HAI. In P. strobus, P. rigida and P. thunbergii family Namikata‐(t)‐73 (half‐sib), PWN was distributed in cortical resin canals down to 5 cm by 6 HAI and down to the base at 192 HAI. However, the distribution of PWN in xylem resin canals of the resistant pines was restricted near inoculated surfaces down to 5 cm, even at 192 HAI. These results demonstrated that migration of PWN in resistant pines was slowed in cortical resin canals and restricted in xylem axial resin canals, features which may be associated with the resistance.     

Analysis of bark proteins in blister rust-resistant and susceptible western white pine (Pinus monticola)

We compared bark proteins from four contrasting (blister rust-resistant versus susceptible) half-sib seedling pairs of western white pine (Pinus monticola D. Don). Pooled proteins from resistant and susceptible groups (four trees per group) were separated by two-dimensional gel electrophoresis, silver stained, and analyzed with the aid of a laser scanner interfaced with a computerized gel documentation system. Qualitative and quantitative protein differences were observed between resistant and susceptible groups. The number of proteins unique to a group was greater in the susceptible category than in the resistant category. Biosynthesis of some common proteins was enhanced near lesioned areas of susceptible seedlings. Many proteins shared similar charge and mass characteristics with those of pathogenesis-related (PR) proteins. Two protein bands were isolated and partially characterized by N-terminal amino acid sequencing: a 10.6-kDa band that was selectively enriched in all resistant individuals, and a 26.0-kDa band that was enriched in some susceptible individuals. The significance of these protein differences and the possible use of selected proteins as disease or resistance markers are discussed.     

The effects of herbaceous and woody competition on planted white pine in a clearcut site

We investigated the effects of herbaceous and woody vegetation control on the survival and growth of planted eastern white pine (Pinus strobus L.) seedlings through six growing seasons. Herbaceous vegetation control involved the suppression of grasses, forbs, ferns, and low-shrubs, and was maintained for 0, 2, or 4 years after white pine seedlings were planted. Woody control involved the removal of all tall-shrub and deciduous trees, and was conducted at the time of planting, at the end of the second or fifth growing seasons, or not at all. Seedling height and basal diameter responded positively and proportionally to duration of herbaceous vegetation control. Gains associated with woody control were generally not significant unless some degree of herbaceous vegetation control was also conducted. Only herbaceous control increased pine crown closure and rate of crown closure. Herbaceous control and the presence of 5000–15,000 stems per ha of young overtopping aspen were associated with reduced weevil (Pissodes strobi Peck.) injury and increased pine height growth. The study suggests that white pine restoration strategies on clearcut sites should focus on the proactive, early management of understory vegetation and the gradual reduction of overtopping cover from woody vegetation to create a seedling light environment that supports acceptable growth with minimal weevil damage.     

Pattern of parenchyma and canal resin composition in softwoods and hardwoods

There is a general pattern of resin chemistry for parenchyma cells and secretory tissue. In sapwood the parenchyma resin is composed of fats, steryl esters, and occasionally waxes. Secretory tissue (most often resin canals) contains terpenes, terpenoids, and polyisoprenes. Other types of canal resin occur in a few hardwood families. The pattern is valid for softwoods and hardwoods. It has been defined from a comparison of available information on wood chemistry, wood anatomy, and the chemistry of oleoresin exudates from trees. This is a short overview with the most important references of two detailed reports.     

Traumatic resin canals as markers of infection events in Douglas‐fir roots infected with Armillaria root disease

In response to an infection, traumatic resin canals (TRCs) are formed in the roots of many conifers, which may be used to determine the timing and sequence of infection events essential for epidemiological studies of root diseases. Juvenile Douglas‐fir (Pseudotsuga menziesii) tree roots at coastal and interior sites in British Columbia were wounded at various times of the year or were inoculated with an isolate of Armillaria ostoyae, and root sections were taken to determine the timing and extent of TRC formation. Naturally infected Douglas‐fir were also examined to determine the extent of the TRCs in infected and uninfected roots on infected trees and in the lower stem. Wounds made in March and October had poor or no TRC formation while the summer wounds responded strongly and were associated with resin soaking. Roots wounded in October did not respond until the following year in all trees except one. Trees produced TRCs and resin soaked tracheids at all times in response to the fungal inoculations. The most striking difference between wounding or fungal inoculation was the multiple bands of TRCs produced in response to the fungus. TRCs at natural A. ostoyae infections were found 92% of time in roots at the stem junction and 74% of the time in the stem at soil line. TRCs were produced in uninfected roots on infected trees but disappeared with increasing distance from the initiating lesion. TRCs can be used to time yearly and seasonal root infections when they can be traced from an identified lesion.     

Epicuticular wax content of western white pine is involved in Cronartium ribicola resistance

The introduction of Cronartium ribicola, the causative agent of white pine blister rust, has been devastating for white pines in North America, including western white pine. Among the observed partial resistance responses to white pine blister rust is a mechanism referred to as difficult-to-infect (DI), which is characterized by lower frequency of infection spots on needles and a lower incidence of branch and stem cankers than susceptible host plants. Parents with the DI trait were selected and bred to produce nine full-sib families. Progeny were propagated and cloned using in vitro techniques in controlled environmental conditions. Explants were inoculated with a single strain of C. ribicola and characterized by using a disease assessment index (DAI), which was used to compare DI full-sib families to several other full-sib families (from Idaho) selected for other partial resistance traits. Most DI families had significantly lower DAI scores and higher epicuticular wax content. When the wax was removed from the surface of needles, the DAI of all full-sib families increased. Scanning electron microscopy revealed that stomata in DI full-sib families are mostly occluded by epicuticular wax. The results of this study suggest that the DI resistance mechanism of western white pine is related to needle surface epicuticular wax, which likely hinders the penetration of stomata by fungal hyphae. The occurrence and magnitude of this trait in the breeding population remains unknown, and special assessment technique for large-scale screening will have to be developed.     

Effects of mass inoculation on induced oleoresin response in intensively managed loblolly pine

Oleoresin flow is an important factor in the resistance of pines to attack by southern pine beetle, Dendroctonus frontalis Zimm., and its associated fungi. Abiotic factors, such as nutrient supply and water relations, have the potential to modify this plant-insect-fungus interaction; however, little is known of the effects of inoculation with beetle-associated fungi on oleoresin flow. We observed that constitutive and induced resin yield in loblolly pine, Pinus taeda L., were affected by either fungal inoculation (with the southern pine beetle-associated fungus Ophiostoma minus (Hedgcock) H. & P. Sydow) or silvicultural treatment. The effects of mass wounding (400 wounds m(-2)) and mass wounding and inoculation with O. minus were assessed by comparison with untreated (control) trees. The treatments were applied to trees in a 2 x 2 factorial combination of fertilizer and irrigation treatments. Fertilization did not significantly affect constitutive resin yield. Even as long as 105 days post-treatment, however, mass-inoculated trees produced higher induced resin yields than control or wounded-only trees, indicating a localized induced response to fungal inoculation. We noted no systemic induction of host defenses against fungal colonization. Although beetles attacking previously attacked trees face a greater resinous response from their host than beetles attacking trees that had not been previously attacked, the effect of an earlier attack may not last more than one flight season. Despite mass inoculations, O. minus did not kill the host trees, suggesting that this fungus is not a virulent plant pathogen.     

Resin flow responses to fertilization, wounding and fungal inoculation in loblolly pine (Pinus taeda) in North Carolina

Resin flow is the primary means of natural defense against southern pine beetle (Dendroctonus frontalis Zimm.), the most important insect pest of Pinus spp. in the southern United States. As a result, factors affecting resin flow are of interest to researchers and forest managers. We examined the influence of fertilization, artificial wounding and fungal inoculation on resin flow in 6- and 12-year-old stands of loblolly pine (Pinus taeda L.) and determined the extent of that influence within and above the wounded stem area and through time. Fertilization increased constitutive resin flow, but only the younger trees sustained increased resin flow after wounding and inoculation treatments. An induced resin flow response occurred between 1 and 30 days after wounding and inoculation treatments. Wounding with inoculation resulted in greater resin flow than wounding alone, but increasing amounts of inoculum did not increase resin flow. Increased resin flow (relative to controls) lasted for at least 90 days after wounding and inoculation. This increase appeared to be limited to the area of treatment, at least in younger trees. The long-lasting effects of fungal inoculation on resin flow, as well as the response to fertilization, suggest that acquired resistance through induced resin flow aids in decreasing susceptibility of loblolly pine to southern pine beetle.