Spatial and temporal patterns of wound periderm development in Cryptomeria japonica bark

Limited investigations have been carried out on the physiological and growth responses of bark to wounding, even though wound periderms play crucial roles in tree defenses. To understand the mechanisms of wound periderm formation, we studied the growth responses and structural changes of wounded bark of three Cryptomeria japonica individuals. We observed the developmental time frame and morphology of wound periderms around mechanically induced wounds in summer. The wound responses included discoloration, lignification, and suberization in tissues present at the time of wounding, followed by wound periderm formation and secondary metabolite deposition. The trees had developed wound periderms approximately 4 weeks after wounding. The wound periderms were within 3 mm in the axial directions and within 1 mm in the lateral directions from the wound surfaces. The distinct patterns of wound periderm formation in the axial and lateral regions resulted from the arrangement and anatomical features of the cells adjacent to the wounds. The wound phellem cells were tangentially narrower and axially shorter in the side and upper/lower regions, respectively, of the wounds. Therefore, the cell division frequencies in the planes parallel to the wound surface may be greater than those in the other directions. Wound reactions in bark might initially be triggered by microenvironmental changes, such as the spread of desiccation, which depends directly on the morphology of phloem cell complexes.

Tree resin flow dynamics during an experimentally induced attack by Ips avulsus, I. calligraphus, and I. grandicollis

The success of tree colonization by bark beetles depends on their ability to overcome host tree defenses, including resin exudation and toxic chemicals, which deter bark beetle colonization. Resin defenses during insect outbreaks are challenging to study in situ, as outbreaks are stochastic events that progress quickly and thus preclude the establishment of baseline observations of non-infested controls. We use synthetic aggregation pheromones to demonstrate that confined Ips bark beetle herbivory can be successfully initiated to provide opportunities for studying interactions between bark beetles and their hosts, including the dynamics of constitutive and induced resin exudation. In Pinus taeda L. plantations between 12 and 19 years old in North and South Carolina, U.S., trees were affixed with pheromone lures, monitored for evidence of bark beetle attacks, and resin samples were collected throughout the growing season. Baiting increased beetle herbivory to an extent sufficient to produce an induced resin response. Attacked trees exuded about three times more resin at some time than control trees. This supports previous work that demonstrated that information on constitutive resin dynamics alone provides an incomplete view of a host tree’s resistance to bark beetle attack.

Differential drivers of chemical defenses in three widespread pine species along a 1500-m elevational gradient: a test of the elevational gradient in plant defense hypothesis

1. The elevational gradient in plant defense (EGPD) hypothesis posits that natural enemy pressures increase positively alongside temperature across elevational climatic gradients, thereby selecting for enhanced defenses at lower elevations while leaving plants less defended at higher elevations. Phylogenetically constrained tests of this hypothesis in tree populations are exceedingly rare. Nevertheless, the presumed presence of poorly-defended trees has been invoked as an important driver of recent pest outbreaks at higher elevations than historically common. 2. Tree age, growth rate, and size have all been correlated with levels of tree defenses. Thus, we sought to disentangle the interacting influences of these properties from possible elevational climatic effects on monoterpene composition, concentrations, and diversity of constitutive resin within three widespread pines (Pinus contorta, Pinus ponderosa, and Pinus flexilis) across a 1532 m elevational transect in the Rocky Mountains, Colorado, USA. 3. Collectively, elevation and tree properties were relatively weak predictors of defenses in P. ponderosa and P. flexilis, but explained ~ 75% of variation in monoterpene concentration and ~ 50% of diversity in P. contorta. 4. Increasing tree age had the greatest positive influence on monoterpene concentration and diversity in P. contorta, while increasing tree size had a negative influence suggesting a potential lifelong tradeoff between tree growth and defense. Elevation had a significant, negative influence on monoterpene concentration but little to no effect on monoterpene diversity in P. contorta. 5. Overall, we found some support for the EGPD hypothesis within P. contorta, but no support within P. ponderosa or P. flexilis. Our results suggest the presence of divergent conifer defense allocation strategies and drivers even among congeners growing in shared environments. An improved understanding of the controls on tree defenses, particularly possible influences of climate-based drivers, is necessary for predicting forest pest dynamics under global change scenarios.

Differential drivers of chemical defenses in three widespread pine species along a 1500-m elevational gradient: a test of the elevational gradient in plant defense hypothesis

1. The elevational gradient in plant defense (EGPD) hypothesis posits that natural enemy pressures increase positively alongside temperature across elevational climatic gradients, thereby selecting for enhanced defenses at lower elevations while leaving plants less defended at higher elevations. Phylogenetically constrained tests of this hypothesis in tree populations are exceedingly rare. Nevertheless, the presumed presence of poorly-defended trees has been invoked as an important driver of recent pest outbreaks at higher elevations than historically common. 2. Tree age, growth rate, and size have all been correlated with levels of tree defenses. Thus, we sought to disentangle the interacting influences of these properties from possible elevational climatic effects on monoterpene composition, concentrations, and diversity of constitutive resin within three widespread pines (Pinus contorta, Pinus ponderosa, and Pinus flexilis) across a 1532 m elevational transect in the Rocky Mountains, Colorado, USA. 3. Collectively, elevation and tree properties were relatively weak predictors of defenses in P. ponderosa and P. flexilis, but explained ~ 75% of variation in monoterpene concentration and ~ 50% of diversity in P. contorta. 4. Increasing tree age had the greatest positive influence on monoterpene concentration and diversity in P. contorta, while increasing tree size had a negative influence suggesting a potential lifelong tradeoff between tree growth and defense. Elevation had a significant, negative influence on monoterpene concentration but little to no effect on monoterpene diversity in P. contorta. 5. Overall, we found some support for the EGPD hypothesis within P. contorta, but no support within P. ponderosa or P. flexilis. Our results suggest the presence of divergent conifer defense allocation strategies and drivers even among congeners growing in shared environments. An improved understanding of the controls on tree defenses, particularly possible influences of climate-based drivers, is necessary for predicting forest pest dynamics under global change scenarios.

Mountain Pine Beetles Colonizing Historical and Naïve Host Trees Are Associated with a Bacterial Community Highly Enriched in Genes Contributing to Terpene Metabolism

ABSTRACTThe mountain pine beetle,Dendroctonus ponderosae, is a subcortical herbivore native to western North America that can kill healthy conifers by overcoming host tree defenses, which consist largely of high terpene concentrations. The mechanisms by which these beetles contend with toxic compounds are not well understood. Here, we explore a component of the hypothesis that beetle-associated bacterial symbionts contribute to the ability ofD. ponderosaeto overcome tree defenses by assisting with terpene detoxification. Such symbionts may facilitate host tree transitions during range expansions currently being driven by climate change. For example, this insect has recently breached the historical geophysical barrier of the Canadian Rocky Mountains, providing access to näive tree hosts and unprecedented connectivity to eastern forests. We use culture-independent techniques to describe the bacterial community associated withD. ponderosaebeetles and their galleries from their historical host,Pinus contorta, and their more recent host, hybridP. contorta-Pinus banksiana. We show that these communities are enriched with genes involved in terpene degradation compared with other plant biomass-processing microbial communities. These pine beetle microbial communities are dominated by members of the generaPseudomonas,Rahnella,Serratia, andBurkholderia, and the majority of genes involved in terpene degradation belong to these genera. Our work provides the first metagenome of bacterial communities associated with a bark beetle and is consistent with a potential microbial contribution to detoxification of tree defenses needed to survive the subcortical environment.

Use of Loop-Mediated Isothermal Amplification for Detection of Ophiostoma clavatum, the Primary Blue Stain Fungus Associated with Ips acuminatus

ABSTRACTLoop-mediated isothermal amplification (LAMP) is an alternative amplification technology which is highly sensitive and less time-consuming than conventional PCR-based methods. Three LAMP assays were developed, two for detection of species of symbiotic blue stain fungi associated withIps acuminatus, a bark beetle infesting Scots pine (Pinus sylvestris), and an additional assay specific toI. acuminatusitself for use as a control. In common with most bark beetles,I. acuminatusis associated with phytopathogenic blue stain fungi involved in the process of exhausting tree defenses, which is a necessary step for the colonization of the plant by the insect. However, the identity of the main blue stain fungus vectored byI. acuminatuswas still uncertain, as well as its frequency of association withI. acuminatusunder outbreak and non-outbreak conditions. In this study, we employed LAMP technology to survey six populations ofI. acuminatussampled from the Southern Alps.Ophiostoma clavatumwas detected at all sampling sites, whileOphiostoma brunneo-ciliatum, reported in part of the literature as the main blue stain fungus associated withI. acuminatus, was not detected on any of the samples. These results are consistent with the hypothesis thatO. clavatumis the main blue stain fungus associated withI. acuminatusin the Southern Alps. The method developed in the course of this work provides a molecular tool by which it will be easy to screen populations and derive important data regarding the ecology of the species involved.

Canker on Bark of Populus spp. Caused by Cytospora tritici, a New Disease in China

Species of Cytospora Ehrenb. and associated teleomorphs cause dieback and canker on over 85 species of angiosperm and gymnosperm plants throughout the world (2). Cytospora tritici Punith. was first observed on Triticum asetivum in Germany in 1980 but may also affect many hardwoods (3). During a survey of landscape trees in 2007, Populus spp. with cankers were found in Fushun, Baoxing, and Luding counties and Chengdu city in Sichuan Province. In these trees, bark canker pathogens discolored the sapwood. During damp weather, conidia were pushed out and formed orange spore horns. Conidiomatal stromata were immersed in bark, prominent, and 1.53 ± 0.33 mm in diameter (n = 10). Discs were white to grey, circular, oval, and 0.59 ± 0.14 mm in diameter (n = 10), with one ostiole per disc. Ostioles were dark grey. Locules were multi-chambered, chambers irregular. Conidia were lelongate-allantoid shaped, hyaline, aseptate, 5.04 ± 0.65 μm long (n = 50), and 1.22 ± 0.13 μm wide (n = 50). Fragments (5 × 5 mm2) of the junction of diseased and healthy tissues were surface sterilized with 1% NaOCl for 30 s and then rinsed twice in sterile distilled water. The pieces were placed on potato dextrose agar (PDA) plates and incubated at 25°C for 7 days. The obtained isolates were cultured on PDA at 25°C in diffuse fluorescent light for 30 days. Upon isolation, the mycelium grew at a rate of 3 to 5 mm per day at 25°C, forming pale white-to-pure white flat colonies. Conidiomata never formed on PDA. ITS1-5.8S-ITS2 sequences were amplified via PCR from genomic DNA obtained from mycelia using universal primers ITS1 and ITS4 (4). The amplification products showed 100% sequence homology with C. tritici isolate DQ243812 from the GenBank database. The ITS sequences were submitted to GenBank (Accession No. JQ277333 to JQ277336). Pathogenicity was confirmed by inoculating 20 disinfected (70% ethanol) Populus tomentosa cuttings. Cuttings were incubated at 25°C for 30 days. Another two cuttings were treated with water agar as controls. In 18 of the 20 cuttings, the cambium developed a brown color and appeared water soaked 15 days later, whereas controls did not develop any symptoms. C. tritici was reisolated from symptomatic tissues. To our knowledge, this is the first report of C. tritici in China causing canker on Populus spp. Cytospora canker is common in practically all countries where poplar are grown. Canker expansion increases when tree defenses are compromised, usually by seasonal dormancy but also by drought, cold injury of wood, sun scald of bark, flooding of root, hail, freezing, or other stress (1). Future spread of C. tritici to western China is considered highly likely. References: (1) G. C. Adams et al. Stud. Mycol. 52:1, 2005. (2) D. F. Farr and A. Y. Rossman. Fungal Databases. Systematic Mycology and Microbiology Laboratory, ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/ March 25, 2012. (3) E. Punithalingam. Nova Hedwigia 32:585, 1980. (4) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al., eds. Academic Press, San Diego, 1990.

Why does longleaf pine have low susceptibility to southern pine beetle?

Pine forests throughout the world are subject to disturbance from tree-killing bark beetles, but pine species differ in their susceptibilities. In the southeastern United States, Pinus palustris Mill. suffers far less mortality from the southern pine beetle, Dendroctonus frontalis Zimmermann, than do its sympatric congeners. We tested the commonly invoked hypothesis that P. palustris has relatively low susceptibility because it has higher oleoresin flow than other pines, especially Pinus taeda L. However, seven studies in three states over 6 years refuted the hypothesis that P. palustris and P. taeda differ in their constitutive resin flow or in their capacity to replace resin depleted by either experimental wounding or natural beetle attacks. Additionally, surveys of natural beetle attacks revealed that P. taeda and P. palustris were equally likely to be attacked and killed when they cooccurred in front of growing infestations. Thus, the relative susceptibility of these two species changes with the spatial scale at which they are mixed, and the strong landscape-scale pattern of low mortality in P. palustris is not because individual trees are physiologically less susceptible. Ultimately, the conspicuous differential impact of D. frontalis on P. taeda and P. palustris may be the product of coevolution between tree defenses and beetle behavior.