Characterization of microsatellite loci in the fungus, Grosmannia clavigera, a pine pathogen associated with the mountain pine beetle

The largest forest pest epidemic in Canadian history caused by the mountain pine beetle (MPB) and its fungal associates has killed over 15 million hectares of forest. Sixty simple sequence repeat regions were identified from Grosmannia clavigera, an MPB associated fungus. Eight loci genotyped in 53 isolates from two populations in British Columbia, Canada revealed three to 10 alleles per locus and gene diversities of 0 to 0.79. All but two of these loci showed length polymorphism in Leptographium longiclavatum, a related MPB fungal associate. These microsatellites will be useful in population genetic studies of these fungi.     

Population Structure of Mountain Pine Beetle Symbiont Leptographium longiclavatum and the Implication on the Multipartite Beetle-Fungi Relationships

Over 18 million ha of forests have been destroyed in the past decade in Canada by the mountain pine beetle (MPB) and its fungal symbionts. Understanding their population dynamics is critical to improving modeling of beetle epidemics and providing potential clues to predict population expansion. Leptographium longiclavatum and Grosmannia clavigera are fungal symbionts of MPB that aid the beetle to colonize and kill their pine hosts. We investigated the genetic structure and demographic expansion of L. longiclavatum in populations established within the historic distribution range and in the newly colonized regions. We identified three genetic clusters/populations that coincide with independent geographic locations. The genetic profiles of the recently established populations in northern British Columbia (BC) and Alberta suggest that they originated from central and southern BC. Approximate Bayesian Computation supports the scenario that this recent expansion represents an admixture of individuals originating from BC and the Rocky Mountains. Highly significant correlations were found among genetic distance matrices of L. longiclavatum, G. clavigera, and MPB. This highlights the concordance of demographic processes in these interacting organisms sharing a highly specialized niche and supports the hypothesis of long-term multipartite beetle-fungus co-evolutionary history and mutualistic relationships.     

Target-specific PCR primers can detect and differentiate ophiostomatoid fungi from microbial communities associated with the mountain pine beetle Dendroctonus ponderosae

The aim of this study was to develop DNA probes that could identify the major fungal species associated with mountain pine beetles (MPB). The beetles are closely associated with fungal species that include ophiostomatoid fungi that can be difficult to differentiate morphologically. The most frequently isolated associates are the pine pathogens Grosmannia clavigera and Leptographium longiclavatum, the less pathogenic Ophiostoma montium, and an undescribed Ceratocystiopsis species (Cop. sp.). Because growing, isolating and extracting DNA from fungi vectored by MPB can be time and labour intensive, we designed three rDNA primer sets that specifically amplify short rDNA amplicons from O. montium, Cop. sp. and the pine Leptographium clade. We also designed two primer sets on a gene of unknown function that can differentiate G. clavigera and L. longiclavatum. We tested the primers on 76 fungal isolates that included MPB associates. The primers reliably identified their targets from DNA obtained from pure fungal cultures, pulverized beetles, beetle galleries, and tree phloem inoculated with G. clavigera. The primers will facilitate large-scale work on the ecology of the MPB-fungal-lodgepole pine ecosystem, as well as phytosanitary/quarantine sample screening.     

Gene genealogies reveal cryptic species and host preferences for the pine fungal pathogen Grosmannia clavigera

Grosmannia clavigera is a fungal pathogen of pine forests in western North America and a symbiotic associate of two sister bark beetles: Dendroctonus ponderosae and D. jeffreyi. This fungus and its beetle associate D. ponderosae are expanding in large epidemics in western North America. Using the fungal genome sequence and gene annotations, we assessed whether fungal isolates from the two beetles inhabiting different species of pine in epidemic regions of western Canada and the USA, as well as in localized populations outside of the current epidemic, represent different genetic lineages. We characterized nucleotide variations in 67 genomic regions and selected 15 for the phylogenetic analysis. Using concordance of gene genealogies and distinct ecological characteristics, we identified two sibling phylogenetic species: Gc and Gs. Where the closely related Pinus ponderosa and P. jeffreyi are infested by localized populations of their respective beetles, Gc is present. In contrast, Gs is an exclusive associate of D. ponderosae mainly present on its primary host‐tree P. contorta; however, in the current epidemic areas, it is also found in other pine species. These results suggest that the host‐tree species and the beetle population dynamics may be important factors associated with the genetic divergence and diversity of fungal partners in the beetle‐tree ecosystems. Gc represents the original G. clavigera holotype, and Gs should be described as a new species.     

Breach of the northern Rocky Mountain geoclimatic barrier: initiation of range expansion by the mountain pine beetle

Aim Our aim is to examine the historical breach of the geoclimatic barrier of the Rocky Mountains by the mountain pine beetle (Dendroctonus ponderosae Hopkins). This recent range expansion from west of the North American continental divide into the eastern boreal forest threatens to provide a conduit to naïve pine hosts in eastern North America. We examine the initial expansion events and determine potential mechanism(s) of spread by comparing spread patterns in consecutive years to various dispersal hypotheses such as: (1) meso‐scale atmospheric dispersal of insects from source populations south‐west of the Rocky Mountains in British Columbia (i.e. their historical range), (2) anthropogenic transport of infested plant material, and (3) spread of insect populations across adjacent stands via corridors of suitable habitat. Location British Columbia, Canada. Methods We explore potential mechanism(s) of invasion of the mountain pine beetle using spatial point process models for the initial 3 years of landscape‐level data collection, 2004–2006. Specifically, we examine observed patterns of infestation relative to covariates reflecting various dispersal hypotheses. We select the most parsimonious models for each of the initial 3 years of invasion using information criteria statistics. Results The initial range expansion and invasion of the beetle was characterized by aerial deposition along a strong north‐west to south‐east gradient, with additional aerial deposition and localized dispersal from persisting populations in following years. Main conclusions Following deposition of a wave front of mountain pine beetles parallel to the Rocky Mountains via meso‐scale atmospheric dispersal, the areas of highest intensity of infestations advanced up to 25 km north‐east towards jack pine (Pinus banksiana) habitat in a single year. There appeared to be no association between putative anthropogenic movement of infested materials and initial range expansion of the mountain pine beetle across the continental divide.     

Spatial–temporal analysis of species range expansion: the case of the mountain pine beetle, Dendroctonus ponderosae

Aim The spatial extent of western Canada’s current epidemic of mountain pine beetle, Dendroctonus ponderosae Hopkins (Coleoptera: Curculionidae, Scolytinae), is increasing. The roles of the various dispersal processes acting as drivers of range expansion are poorly understood for most species. The aim of this paper is to characterize the movement patterns of the mountain pine beetle in areas where range expansion is occurring, in order to describe the fine‐scale spatial dynamics of processes associated with mountain pine beetle range expansion. Location Three regions of Canada’s Rocky Mountains: Kicking Horse Pass, Yellowhead Pass and Pine Pass. Methods Data on locations of mountain pine beetle‐attacked trees of predominantly lodgepole pine (Pinus contorta var. latifolia) were obtained from annual fixed‐wing aircraft surveys of forest health and helicopter‐based GPS surveys of mountain pine beetle‐damaged areas in British Columbia and Alberta. The annual (1999–2005) spatial extents of outbreak ranges were delineated from these data. Spatial analysis was conducted using the spatial–temporal analysis of moving polygons (STAMP), a recently developed pattern‐based approach. Results We found that distant dispersal patterns (spot infestations) were most often associated with marginal increases in the areal size of mountain pine beetle range polygons. When the mountain pine beetle range size increased rapidly relative to the years examined, local dispersal patterns (adjacent infestation) were more common. In Pine Pass, long‐range dispersal (> 2 km) markedly extended the north‐east border of the mountain pine beetle range. In Yellowhead Pass and Kicking Horse Pass, the extension of the range occurred incrementally via ground‐based spread. Main conclusions Dispersal of mountain pine beetle varies with geography as well as with host and beetle population dynamics. Although colonization is mediated by habitat connectivity, during periods of low overall habitat expansion, dispersal to new distant locations is common, whereas during periods of rapid invasion, locally connected spread is the dominant mode of dispersal. The propensity for long‐range transport to establish new beetle populations, and thus to be considered a driver of range expansion, is likely to be determined by regional weather patterns, and influenced by local topography. We conclude that STAMP appears to be a useful approach for examining changes in biogeograpical ranges, with the potential to reveal both fine‐ and large‐scale patterns.     

Spatial genetic structure of the mountain pine beetle (Dendroctonus ponderosae) outbreak in western Canada: historical patterns and contemporary dispersal

Environmental change has a wide range of ecological consequences, including species extinction and range expansion. Many studies have shown that insect species respond rapidly to climatic change. A mountain pine beetle epidemic of record size in North America has led to unprecedented mortality of lodgepole pine, and a significant range expansion to the northeast of its historic range. Our goal was to determine the spatial genetic variation found among outbreak population from which genetic structure, and dispersal patterns may be inferred. Beetles from 49 sampling locations throughout the outbreak area in western Canada were analysed at 13 microsatellite loci. We found significant north-south population structure as evidenced by: (i) Bayesian-based analyses, (ii) north-south genetic relationships and diversity gradients; and (iii) a lack of isolation-by-distance in the northernmost cluster. The north-south structure is proposed to have arisen from the processes of postglacial colonization as well as recent climate-driven changes in population dynamics. Our data support the hypothesis of multiple sources of origin for the outbreak and point to the need for population specific information to improve our understanding and management of outbreaks. The recent range expansion across the Rocky Mountains into the jack/lodgepole hybrid and pure jack pine zones of northern Alberta is consistent with a northern British Columbia origin. We detected no loss of genetic variability in these populations, indicating that the evolutionary potential of mountain pine beetle to adapt has not been reduced by founder events. This study illustrates a rapid range-wide response to the removal of climatic constraints, and the potential for range expansion of a regional population.     

A population survey of Beauveria bassiana in the microhabitat of the red turpentine beetle,Dendroctonus valens,in Chinese pine forests

Field survey of the entomopathogenic fungus Beauveria bassiana in association with the red turpentine beetle, Dendroctonus valens, was undertaken in three pine plantations in Northern China. In total, 88 strains of B. bassiana sensu lato were isolated from the soil, bark, beetle frass, living adult and cadaver samples and soil was proved to be an important inoculum reservoir for fungal entomopathogens. Of these, 77 isolates were included for genetic diversity analysis by PCR for inter-simple sequence repeats (ISSR). Genetic diversity and population structure analysis of the isolates from three sites and five niches demonstrated high genetic diversity and heterogeneity between and/or within populations. Wright's statistics revealed a high gene flow rate (4.529) among the three populations, especially among the soil-derived isolate subpopulations. Low variation was mainly caused (94.8%) by variation among different substrates, suggesting the importance of microhabitat substrates on genetic diversity of B. bassiana. Phylogenetic variation was not associated with geographic distance.     

Phylogeography of spruce beetles (Dendroctonus rufipennis Kirby) (Curculionidae: Scolytinae) in North America

Tree-feeding insects that are widespread in north temperate regions are excellent models for studying how past glaciations have impacted differentiation and speciation. We used mitochondrial DNA (mtDNA) sequences and allele frequencies at nine microsatellite loci to examine genetic population structure across the current range of the spruce beetle (Dendroctonus rufipennis), an economically important insect in North America. Two major haplotype groups occur across northern North America, from Newfoundland to Alaska, on white spruce (Picea glauca), and a third distinctive haplotype group occurs throughout the Rocky Mountains on Engelmann spruce (Picea engelmannii). The two mtDNA lineages found in northern populations are 3-4% divergent from each other and from the lineages found in the Rocky Mountains. Analyses of microsatellite data also suggest the existence of major population groupings associated with different geographical regions. In the Pacific Northwest, concordant contact zones for genetically distinct populations of spruce beetles and their principal hosts appear to reflect recent secondary contact. Although we could detect no evidence of historical mtDNA gene flow between allopatric population groups, patterns of variation in the Pacific Northwest suggest recent hybridization and introgression. Together with the pollen record for spruce, they also suggest that beetles have spread from at least three glacial refugia. A minimum estimate of divergence time between the Rocky Mountain and northern populations was 1.7 Myr (million years), presumably reflecting the combined effects of isolation during multiple glacial cycles.     

Dietary benefits of fungal associates to an eruptive herbivore: potential implications of multiple associates on host population dynamics

We used the mountain pine beetle (Dendroctonus ponderosae Hopkins) and its two fungal associates, Grosmannia clavigera and Ophiostoma montium, to study potential nutritional benefits of fungi to bark beetles. We tested for potential effects of feeding on phloem colonized by fungi on beetle performance in field and laboratory studies. The fungi increased nitrogen levels in the phloem of attacked trees by 40%, indicating that it may be an important source of dietary nitrogen for mountain pine beetles. However, nitrogen levels of phloem inoculated with fungi in the laboratory were similar to uncolonized phloem, indicating that the fungi may redistribute nitrogen from the sapwood to the phloem rather than increase absolute levels of nitrogen. Beetles emerging from attacked trees carrying G. clavigera were larger than beetles carrying O. montium, which in turn were larger than beetles lacking fungi. Results of experimental laboratory studies varied, likely because of differences in the growth and sporulation of fungi under artificial conditions. Results indicate that the two fungi may offer complementary benefits to the mountain pine beetle because larvae preferentially fed on phloem colonized by both fungi together over phloem colonized by one fungus or uncolonized phloem. Teneral adults preemergence fed on spores in pupal chambers when they were produced and consumed little phloem before emerging. Teneral adults mined extensively in the phloem before emerging when spores were not produced in the pupal chamber. Our results provide evidence for a nutritional role of fungi in the diet of bark beetles and show that multiple associates may differentially affect beetle performance, which could have important implications for bark beetle population dynamics.     

Mountain pine beetle host-range expansion threatens the boreal forest

The current epidemic of the mountain pine beetle (MPB), an indigenous pest of western North American pine, has resulted in significant losses of lodgepole pine. The leading edge has reached Alberta where forest composition shifts from lodgepole to jack pine through a hybrid zone. The susceptibility of jack pine to MPB is a major concern, but there has been no evidence of host-range expansion, in part due to the difficulty in distinguishing the parentals and their hybrids. We tested the utility of a panel of microsatellite loci optimized for both species to classify lodgepole pine, jack pine and their hybrids using simulated data. We were able to accurately classify simulated individuals, and hence applied these markers to identify the ancestry of attacked trees. Here we show for the first time successful MPB attack in natural jack pine stands at the leading edge of the epidemic. This once unsuitable habitat is now a novel environment for MPB to exploit, a potential risk which could be exacerbated by further climate change. The consequences of host-range expansion for the vast boreal ecosystem could be significant.     

The Relative Abundance of Mountain Pine Beetle Fungal Associates Through the Beetle Life Cycle in Pine Trees

The mountain pine beetle (MPB) is a native bark beetle of western North America that attacks pine tree species, particularly lodgepole pine. It is closely associated with the ophiostomatoid ascomycetes Grosmannia clavigera, Leptographium longiclavatum, Ophiostoma montium, and Ceratocystiopsis sp.1, with which it is symbiotically associated. To develop a better understanding of interactions between beetles, fungi, and host trees, we used target-specific DNA primers with qPCR to assess the changes in fungal associate abundance over the stages of the MPB life cycle that occur in galleries under the bark of pine trees. Multivariate analysis of covariance identified statistically significant changes in the relative abundance of the fungi over the life cycle of the MPB. Univariate analysis of covariance identified a statistically significant increase in the abundance of Ceratocystiopsis sp.1 through the beetle life cycle, and pair-wise analysis showed that this increase occurs after the larval stage. In contrast, the abundance of O. montium and Leptographium species (G. clavigera, L. longiclavatum) did not change significantly through the MPB life cycle. From these results, the only fungus showing a significant increase in relative abundance has not been formally described and has been largely ignored by other MPB studies. Although our results were from only one site, in previous studies we have shown that the fungi described were all present in at least ten sites in British Columbia. We suggest that the role of Ceratocystiopsis sp.1 in the MPB system should be explored, particularly its potential as a source of nutrients for teneral adults.     

Genetic variation within the cosmopolitan aquatic fungus Lignincola laevis (Microascales, Ascomycota)

Recent estimates suggest that there are as many as 10,000 marine fungal species, although the current number stands at just over 500. Previous studies were predominantly surveys of marine fungal diversity on various substrata in Europe, north American and Asia, while little research exists on the possible processes leading to their current distribution. Therefore, this study was initiated to assess the genetic structure and geographical distribution of the cosmopolitan aquatic fungus Lignincola laevis. The internal transcribed spacers and partial 28S region of the rRNA gene clusters, and the partial MCM7 protein gene were sequenced and analysed using maximum parsimony and Bayesian likelihood methods. Three distinct lineages were present in the sampled L. laevis isolates with a mean pairwise distance between the groups >10 %. However, no geographical assemblages could be identified in the phylogeny. The high genetic divergence suggests that Lignincola laevis is a species complex that consists of a group of closely related species with subtle morphological differences. The lack of geographical structure in the sampled isolates of L. laevis may suggest a high dispersal capacity for L. laevis.     

Fire, fuels and restoration of ponderosa pine–Douglas fir forests in the Rocky Mountains, USA

Aim Forest restoration in ponderosa pine and mixed ponderosa pine–Douglas fir forests in the US Rocky Mountains has been highly influenced by a historical model of frequent, low‐severity surface fires developed for the ponderosa pine forests of the Southwestern USA. A restoration model, based on this low‐severity fire model, focuses on thinning and prescribed burning to restore historical forest structure. However, in the US Rocky Mountains, research on fire history and forest structure, and early historical reports, suggest the low‐severity model may only apply in limited geographical areas. The aim of this article is to elaborate a new variable‐severity fire model and evaluate the applicability of this model, along with the low‐severity model, for the ponderosa pine–Douglas fir forests of the Rocky Mountains. Location Rocky Mountains, USA. Methods The geographical applicability of the two fire models is evaluated using historical records, fire histories and forest age‐structure analyses. Results Historical sources and tree‐ring reconstructions document that, near or before ad 1900, the low‐severity model may apply in dry, low‐elevation settings, but that fires naturally varied in severity in most of these forests. Low‐severity fires were common, but high‐severity fires also burned thousands of hectares. Tree regeneration increased after these high‐severity fires, and often attained densities much greater than those reconstructed for Southwestern ponderosa pine forests. Main conclusions Exclusion of fire has not clearly and uniformly increased fuels or shifted the fire type from low‐ to high‐severity fires. However, logging and livestock grazing have increased tree densities and risk of high‐severity fires in some areas. Restoration is likely to be most effective which seeks to (1) restore variability of fire, (2) reverse changes brought about by livestock grazing and logging, and (3) modify these land uses so that degradation is not repeated.     

Maintenance of divergent lineages of the Rice Blast Fungus Pyricularia oryzae through niche separation,loss of sex and post-mating genetic incompatibilities

Many species of fungal plant pathogens coexist as multiple lineages on the same host, but the factors underlying the origin and maintenance of population structure remain largely unknown. The rice blast fungus Pyricularia oryzae is a widespread model plant pathogen displaying population subdivision. However, most studies of natural variation in P. oryzae have been limited in genomic or geographic resolution, and host adaptation is the only factor that has been investigated extensively as a contributor to population subdivision. In an effort to complement previous studies, we analyzed genetic and phenotypic diversity in isolates of the rice blast fungus covering a broad geographical range. Using single-nucleotide polymorphism genotyping data for 886 isolates sampled from 152 sites in 51 countries, we showed that population subdivision of P. oryzae in one recombining and three clonal lineages with broad distributions persisted with deeper sampling. We also extended previous findings by showing further population subdivision of the recombining lineage into one international and three Asian clusters, and by providing evidence that the three clonal lineages of P. oryzae were found in areas with different prevailing environmental conditions, indicating niche separation. Pathogenicity tests and bioinformatic analyses using an extended set of isolates and rice varieties indicated that partial specialization to rice subgroups contributed to niche separation between lineages, and differences in repertoires of putative virulence effectors were consistent with differences in host range. Experimental crosses revealed that female sterility and early post-mating genetic incompatibilities acted as strong additional barriers to gene flow between clonal lineages. Our results demonstrate that the spread of a fungal pathogen across heterogeneous habitats and divergent populations of a crop species can lead to niche separation and reproductive isolation between distinct, widely distributed, lineages.     

Broadscale Specificity in a Bark Beetle–Fungal Symbiosis: a Spatio-temporal Analysis of the Mycangial Fungi of the Western Pine Beetle

Whether and how mutualisms are maintained through ecological and evolutionary time is a seldom studied aspect of bark beetle–fungal symbioses. All bark beetles are associated with fungi and some species have evolved structures for transporting their symbiotic partners. However, the fungal assemblages and specificity in these symbioses are not well known. To determine the distribution of fungi associated with the mycangia of the western pine beetle (Dendroctonus brevicomis), we collected beetles from across the insect’s geographic range including multiple genetically distinct populations. Two fungi, Entomocorticium sp. B and Ceratocystiopsis brevicomi, were isolated from the mycangia of beetles from all locations. Repeated sampling at two sites in Montana found that Entomocorticium sp. B was the most prevalent fungus throughout the beetle’s flight season, and that females carrying that fungus were on average larger than females carrying C. brevicomi. We present evidence that throughout the flight season, over broad geographic distances, and among genetically distinct populations of beetle, the western pine beetle is associated with the same two species of fungi. In addition, we provide evidence that one fungal species is associated with larger adult beetles and therefore might provide greater benefit during beetle development. The importance and maintenance of this bark beetle–fungus interaction is discussed.     

Mountain pine beetle selectivity in old‐growth ponderosa pine forests,Montana, USA

A historically unprecedented mountain pine beetle (MPB) outbreak affected western Montana during the past decade. We examined radial growth rates (AD 1860–2007/8) of co‐occurring mature healthy and MPB‐infected ponderosa pine trees collected at two sites (Cabin Gulch and Kitchen Gulch) in western Montana and: (1) compared basal area increment (BAI) values within populations and between sites; (2) used carbon isotope analysis to calculate intrinsic water‐use efficiency (iWUE) at Cabin Gulch; and (3) compared climate‐growth responses using a suite of monthly climatic variables. BAI values within populations and between sites were similar until the last 20–30 years, at which point the visually healthy populations had consistently higher BAI values (22–34%) than the MPB‐infected trees. These results suggest that growth rates two–three decades prior to the current outbreak diverged between our selected populations, with the slower‐growing trees being more vulnerable to beetle infestation. Both samples from Cabin Gulch experienced upward trends in iWUE, with significant regime shifts toward higher iWUE beginning in 1955–59 for the visually healthy trees and 1960–64 for the MPB‐infected trees. Drought tolerance also varied between the two populations with the visually healthy trees having higher growth rates than MPB‐infected trees prior to infection during a multi‐decadal period of drying summertime conditions. Intrinsic water‐use efficiency significantly increased for both populations during the past 150 years, but there were no significant differences between the visually healthy and MPB‐infected chronologies.     

Single‐nucleotide polymorphism discovery in Leptographium longiclavatum,a mountain pine beetle‐associated symbiotic fungus,using whole‐genome resequencing

Single‐nucleotide polymorphisms (SNPs) are rapidly becoming the standard markers in population genomics studies; however, their use in nonmodel organisms is limited due to the lack of cost‐effective approaches to uncover genome‐wide variation, and the large number of individuals needed in the screening process to reduce ascertainment bias. To discover SNPs for population genomics studies in the fungal symbionts of the mountain pine beetle (MPB), we developed a road map to discover SNPs and to produce a genotyping platform. We undertook a whole‐genome sequencing approach of Leptographium longiclavatum in combination with available genomics resources of another MPB symbiont, Grosmannia clavigera. We sequenced 71 individuals pooled into four groups using the Illumina sequencing technology. We generated between 27 and 30 million reads of 75 bp that resulted in a total of 1, 181 contigs longer than 2 kb and an assembled genome size of 28.9 Mb (N₅₀ = 48 kb, average depth = 125x). A total of 9052 proteins were annotated, and between 9531 and 17 266 SNPs were identified in the four pools. A subset of 206 genes (containing 574 SNPs, 11% false positives) was used to develop a genotyping platform for this species. Using this roadmap, we developed a genotyping assay with a total of 147 SNPs located in 121 genes using the Illumina^® Sequenom iPLEX Gold. Our preliminary genotyping (success rate = 85%) of 304 individuals from 36 populations supports the utility of this approach for population genomics studies in other MPB fungal symbionts and other fungal nonmodel species.     

Tree‐mediated interactions between the jack pine budworm and a mountain pine beetle fungal associate

1. Coniferous trees deploy a combination of constitutive (pre‐existing) and induced (post‐invasion), structural and biochemical defences against invaders. Induced responses can also alter host suitability for other organisms sharing the same host, which may result in indirect, plant‐mediated interactions between different species of attacking organisms. 2. Current range and host expansion of the mountain pine beetle (Dendroctonus ponderosae Hopkins; MPB) from lodgepole pine‐dominated forests to the jack pine‐dominated boreal forests provides a unique opportunity to investigate whether the colonisation of jack pine (Pinus banksiana Lamb.) by MPB will be affected by induced responses of jack pine to a native herbaceous insect species: the jack pine budworm (Choristoneura pinus pinus Freeman; JPBW). 3. We simulated MPB attacks with one of its fungal associates, Grosmannia clavigera Robinson‐Jeffrey & Davidson, and tested induction of either herbivory by JPBW or inoculation with the fungus followed by a challenge treatment with the other organism on jack pine seedlings and measured and compared monoterpene responses in needles. 4. There was clear evidence of an increase in jack pine resistance to G. clavigera with previous herbivory, indicated by smaller lesions in response to fungal inoculations. In contrast, although needle monoterpenes greatly increased after G. clavigera inoculation and continued to increase during the herbivory challenge, JPBW growth was not affected, but JPBW increased the feeding rate to possibly compensate for altered host quality. 5. Jack pine responses varied greatly and depended on whether seedlings were treated with single or multiple organisms, and their order of damage.     

Geographic patterns of microsatellite variation in Boechera stricta, a close relative of Arabidopsis

The genus Boechera is a widespread North American group with great potential for studies of ecology and evolution: Boechera is closely related to Arabidopsis and exhibits different ecological and reproductive strategies. Boechera stricta (previously Arabis drummondii) is a morphologically and genetically well-defined, perennial crucifer species. Fifteen natural populations of diploid individuals from the Rocky Mountains were analysed using 21 microsatellite loci. In accordance with our expectation for this predominately inbreeding species, a high F IS value (0.89) was observed. Furthermore, populations of B. stricta were highly differentiated, as indicated by F ST = 0.56. Three clusters were identified using structure- the majority of populations belonged to either the Northern or Southern cluster. Together, the north-south partitioning and evenness of genetic variation across the two clusters suggested multiple refugia for this perennial herb in the Rocky Mountains. Pleistocene glaciation, together with the topographically and climatologically heterogeneous cordillera, has profoundly influenced the genetic architecture of B. stricta. Genetic population structure was also influenced by relatively recent genome admixture at two levels: within species (involving individuals from the Northern and Southern clusters) and between species (with the hybridization of B. stricta and Boechera holboellii). This complexity of population structure at presumably neutral microsatellite loci located throughout the genome in B. stricta provides a baseline against which to test whether functional genetic variation is undergoing local adaptive evolution throughout the natural species range.