François Lieutier

Induced Responses in Phenolic Metabolism in Two Norway Spruce Clones after Wounding and Inoculations with Ophiostoma polonicum, a Bark Beetle-Associated Fungus.

Two Norway spruce (Picea abies Karst.) clones, one resistant and the other susceptible to mass inoculation with Ophiostoma polonicum Siem., were compared with regard to their phenolic compositions and chalcone synthase (CHS) and stilbene synthase activities of their phloem before and at 6 and 12 d after artificial inoculation with sterile malt agar or O. polonicum. In unwounded phloem, the resistant clone differed from the susceptible clone by the presence of taxifolin glycoside, lower concentrations of stilbene glycosides, and higher CHS activity. After inoculation, (+)-catechin concentration and CHS activity dramatically increased around the wound, particularly in the resistant clone. Stilbene synthase activity also increased, but more slowly and to a lower level, whereas the concentrations of stilbenes remained stable. Tanning ability decreased in the susceptible clone, whereas it remained stable in the resistant one. It is proposed that the induced phenolic response of Norway spruce phloem consists of an activation of the phenolic pathway, finally leading to tannins and insoluble polymers. It is suggested that resistance to O. polonicum depends on the ability of the tree to easily activate the flavonoid pathway.

Mechanisms and deployment of resistance in trees to insects.

Preface. Acknowledgement. 1. Resistance in trees to insects - An overview of mechanisms and interactions S. Larson. 2. Mechanisms of resistance in conifers and bark beetle attack strategies F. Lieutier. 3. Mechanisms of resistance in trees to defoliators K. Clancy. 4. Mechanisms of Resistance in Conifers against shoot infesting insects. The case of the white pine weevil Pissodes strobi (Peck): (Coleoptera: Curculionidae) R.I. Alfaro, et al. 5. Host tree resistance to wood-boring insects T.D. Paine. 6. Plant resistance against gall-forming insects: the role of hypersensititiy T.G. Cornelissen, et al. 7. The resistance of hybrid willows to specialist and generalist herbivores and pathogens: the potential role of secondary chemistry and parent host plant status J. Hjalten, P. Hallgren. 8. Deploying pest resistance in genetically-limited forest plantations: developing ecologically-based strategies for managing risk D.J. Robinson. 9. Deployment of tree resistance to insects in short-rotation Populus D.R. Coyle, et al. 10. Strategies for Deployment of Insect Resistant Ornamental Plants D.A. Herms. 11. Possibilities to utilize tree resistance to insects in forest pest management in central and western Europe C.H. Heidger, F. Lieutier. 12. Deployment of tree resistance to pests in Asia N. Kamata. 13. Using Resistance in Tropical Forest Plantations J.D. Nichols, et al. Index.

Stimulation of tree defenses by Ophiostomatoid fungi can explain attack success of bark beetles on conifers

Abstract• Our aim is to present why the hypothesis, that Ophiostomatoid fungi play an important role in the establishment of most bark beetle species on living conifers, is valuable.• After summarizing knowledge about the relationships of bark beetles with conifers and fungi, we conclude that controversy results from misinterpretations when using fungal pathogenicity to demonstrate the role of Ophiostomatoid fungi in beetle establishment on host trees.• We demonstrate that fungal pathogenicity is not the right parameter to appreciate the role of fungus in beetle establishment on host trees. We argue that artificial low density inoculations that allow the appreciation of fungus ability to stimulate tree defenses and thus to help beetles in overcoming tree resistance must be used in complement to mass inoculations. In both cases, results must be expressed in terms of tree defense stimulation rather than in terms of tree killing.(i)Fungal species stimulating tree defenses are generally not those that grow the best in the sapwood.(ii)We argue that beetle development in the phloem, fungal invasion of the sapwood and phloem, and tree death, occur after tree defenses are exhausted, and that any fungus present in the beetle gallery could thus potentially invade the sapwood after defense exhaustion.• We conclude that stimulation of the tree defense reactions in both the phloem and the superficial sapwood is a real benefit brought by fungi to the beetles during the first phase of establishment (overcoming tree resistance).• Considering the origin of the bark beetle fungus associations attacking living trees and their general functioning based on stimulation of tree defenses, we develop three hypotheses:(i)any beetle species would be helped in its establishment in a given tree species by developing an association, even loosely, with a fungus species belonging to the Ophiostomatoid flora of that tree species;(ii)the necessity of a considerably low level of tree resistance for fungus extension into the tree is the selection pressure that has led fungi to develop their intrinsic ability to stimulate tree defenses, through their ability to grow into the phloem. This association can be completed by antagonistic fungal species controlling extension of the previous fungal species in the tree tissues;(iii)Beetle species using the strategy of overcoming tree resistance are associated with a fungal complex, of which species could assume three roles regarding relationships between beetles and trees: 1- to stimulate tree defenses in the phloem and superficial sapwood, 2- to grow into the sapwood after tree resistance is overcome, and 3- to control phloem extension of the first other two categories. Bringing nutrients to the beetle progeny can be a fourth role.• We propose that bark beetle — Ophiostomatoid associations can be categorized, based on associations’ frequency and complexity while taking into account beetle aggressiveness. We show that a close correspondence exists between beetles’ aggressiveness and the ability of their main associated fungal species to stimulate the defenses of their host tree.• We conclude with suggesting that most sapwood invading fungi might be “cheaters” which have taken advantage of the efficiency of the relationship between beetles and fungi that stimulate tree defenses.Résumé• Notre objectif est de présenter les raisons de la validité de l’hypothèse selon laquelle les champignons jouent un rôle important dans l’installation de la plupart des espèces de Scolytes sur conifères vivants.• Après avoir résumé les connaissances sur les relations des Scolytes avec les conifères et les champignons, nous concluons que la controverse résulte d’interprétations erronées lorsque l’on utilise le pouvoir pathogène des champignons pour démontrer le rôle des Ophiostomatoïdes dans l’installation des insectes sur les arbres hôtes.• Nous démontrons que le pouvoir pathogène n’est pas le paramètre correct pour apprécier le rôle du champignon dans l’installation des Scolytes sur les arbres hôtes. Nous soutenons que des inoculations artificielles à faible densité, qui permettent d’apprécier la capacité du champignon à stimuler les défenses de l’arbre et à ainsi aider le Scolyte à surmonter la résistance de celui-ci, doivent être utilisées en complément des inoculations massives. Dans les deux cas, les résultats doivent être exprimés en termes de stimulation des défenses de l’arbre plutôt qu’en termes de mortalité de l’arbre.(i)les espèces de champignons qui stimulent les défenses de l’arbre ne sont généralement pas celles qui présentent la meilleure croissance dans l’aubier.(ii)nous soutenons que le développement de l’insecte dans le phloème, l’invasion de l’aubier et du phloème par le champignon, et la mort de l’arbre, interviennent après épuisement des défenses de l’arbre, et que tout champignon présent dans les galeries de l’insecte pourrait donc potentiellement envahir l’aubier après épuisement de ces défenses.• Nous concluons que la stimulation des réactions de défense de l’arbre à la fois dans le phloème et l’aubier superficiel représente un bénéfice réel apporté par les champignons aux Scolytes pendant la première phase de leur installation (surmonter la résistance de l’arbre).• En ce qui concerne l’origine des associations Scolytes — champignons attaquant les arbres vivants et considérant leur fonctionnement général basé sur une stimulation des défenses de l’arbre, nous développons trois hypothèses :(i)toute espèce de Scolyte serait aidée dans son installation sur une espèce d’arbre donnée en développant une association, même lâche, avec une espèce de champignon appartenant à la flore Ophiostomatoïde de cette espèce d’arbre ;(ii)la nécessité d’un très faible niveau de résistance de l’arbre pour autoriser l’extension fongique dans le végétal est la pression de sélection qui a conduit les champignons à développer leur capacité intrinsèque de stimulation des défenses de l’arbre, à travers leur capacité à croître dans le phloème. Cette association peut être complétée par des espèces fongiques antagonistes contrôlant l’extension des espèces précédentes dans les tissus de l’arbre ;(iii)les espèces de Scolytes utilisant la stratégie de surmonter la résistance de l’arbre sont associées à un complexe fongique dont les espèces assurent trois fonctions eu égard aux relations entre les Scolytes et les arbres : 1— stimuler les défenses de l’arbre dans le phloème et l’aubier superficiel, 2— croître dans l’aubier après que la résistance de l’arbre ait été vaincue, and 3— contrôler l’extension des deux catégories précédentes dans le phloème. L’apport de nutriments à la progéniture du Scolyte peut représenter une quatrième fonction.• Nous proposons que les associations Scolytes — Ophiostomatoïdes puissent être classées, en se basant sur la fréquence et la complexité de l’association et en prenant en compte l’agressivité de l’insecte. Nous montrons qu’il existe une étroite correspondance entre l’agressivité des insectes et la capacité de leur principale espèce fongique associée à stimuler les défenses de l’arbre hôte.• Nous concluons en suggérant que la plupart des espèces de champignons envahissant l’aubier pourraient être des “tricheurs” qui ont profité de l’efficacité des relations entre les Scolytes et les espèces fongiques stimulatrices des défenses de l’arbre.

Phylogeographic structure and past history of the circum-Mediterranean species Tomicus destruens Woll. (Coleoptera: Scolytinae).

Phylogeographic studies are often focused on temperate European species with relict footholds in the Mediterranean region. Past climatic oscillations usually induced range contractions and expansions from refugial areas located in southern Europe, and spatial distribution of genetic diversity show that northward expansions were usually pioneer‐like. Actually, few studies have focused on circum‐Mediterranean species, which probably were not influenced in the same way by climatic oscillations. We present the phylogeography of the bark beetle Tomicus destruens, which is restricted to the whole Mediterranean basin and the Atlantic coasts of North Africa and Portugal. We systematically sequenced 617 bp of the mitochondrial genes COI and COII for 42 populations (N = 219). Analysis revealed 53 haplotypes geographically structured in two clades, namely eastern and western clades, that diverged during the Pleistocene. A contact zone was identified along the Adriatic coast of Italy. Interestingly, we found contrasting levels of genetic structure within each clade. The eastern group was characterized by a significant phylogeographic pattern and low levels of gene flow, whereas the western group barely showed a spatial structure in haplotype distribution. Moreover, the main pine hosts were different between groups, with the Aleppo‐brutia complex in the east and the maritime pine in the west. Potential roles of host species, climatic parameters and geographical barriers are discussed and the phylogeographic patterns are compared to classical models of postglacial recolonization in Europe.

Population genetic structure of Tomicus piniperda L. (Curculionidae: Scolytinae) on different pine species and validation of T. destruens (Woll.)

Genetic diversity and population structure of Tomicus piniperda was assessed using mitochondrial sequences on 16 populations sampled on 6 pine species in France. Amplifications of Internal transcribed space 1 (ITS1) were also performed. Our goals were to determine the taxonomic status of the Mediterranean ecotype T. piniperda destruens, and to test for host plant or geographical isolation effect on population genetic structure. We showed that T. piniperda clusters in two mtDNA haplotypic groups. Clade A corresponds to insects sampled in continental France on Pinus sylvestris, P. pinaster and P. uncinata, whereas clade B gathers the individuals sampled in Corsica on P. pinaster and P. radiata and in continental France on P. pinea and P. halepensis. Insects belonging to clade A and clade B also consistently differ in the length of ITS1. Individuals belonging to both clades were found once in sympatry on P. pinaster. Genetic distances between clades are similar to those measured between distinct species of Tomicus. We concluded that clade B actually corresponds to the destruens ecotype and forms a good species, T. destruens. Analyses of molecular variance (amova) were conducted separately on T. destruens and T. piniperda to test for an effect of either geographical isolation or host species. Interestingly, the effect of host plant was significant for T. piniperda only, while the effect of geographical isolation was not. Pine species therefore seems to act as a significant barrier to gene flow, even if host race formation is not observed. These results still need to be confirmed by nuclear markers.

Novel insect-tree associations resulting from accidental and intentional biological 'invasions': a meta-analysis of effects on insect fitness.

The translocation of species beyond their native range is a major threat to biodiversity. Invasions by tree-feeding insects attacking native trees and the colonization of introduced trees by native insects result in new insect-tree relationships. To date there is uncertainty about the key factors that influence the outcome of these novel interactions. We report the results of a meta-analysis of 346 pairwise comparisons of forest insect fitness on novel and ancient host tree species from 31 publications. Host specificity of insects and phylogenetic relatedness between ancient and novel host trees emerged as key factors influencing insect fitness. Overall, fitness was significantly lower on novel host species than on ancient hosts. However, in some cases, fitness increased on novel hosts, mainly in polyphagous insects or when close relatives of ancient host trees were colonized. Our synthesis enables greatly improved impact prediction and risk assessment of biological invasions.

Genetic study of the forest pest Tomicus piniperda (Col., Scolytinae) in Yunnan province (China) compared to Europe: new insights for the systematics and evolution of the genus Tomicus

The pine shoot beetle Tomicus piniperda is present throughout Eurasia. In Europe, it is considered as a secondary pest that rarely causes tree mortality, while heavy damage is observed in Yunnan Province (China) where it exhibits a novel aggregative behaviour during shoot attack. To understand why the ecological characteristics of the European and Chinese populations differ so strongly, we conducted an analysis of population genetic structure on 12 populations in Yunnan and one in JiLin using mitochondrial (COI-COII) and nuclear (ITS2 and 28S rDNA) DNA sequences, and compared the results to those obtained in France. We showed that the Yunnan populations differed markedly from French and JiLin populations. For all three markers, the genetic distances measured between the Tomicus from Yunnan and those from France were similar to distances previously observed between species. Similar distances were found between Yunnan and JiLin populations. Conversely, the distances between French and JiLin individuals were substantially lower, falling in the intraspecific range. We concluded that the individuals sampled in Yunnan belong to a new, undescribed species (Tomicus sp. nov.). We also showed that some individuals belong to the species T. brevipilosus that had never been recorded from this region before. Evolution of the genus Tomicus is discussed in the light of these new results.

High intercontinental migration rates and population admixture in the sapstain fungus Ophiostoma ips

Ophiostoma ips is a common fungal associate of various conifer‐infesting bark beetles in their native ranges and has been introduced into non‐native pine plantations in the Southern Hemisphere. In this study, we used 10 microsatellite markers to investigate the population biology of O. ips in native (Cuba, France, Morocco and USA) and non‐native (Australia, Chile and South Africa) areas to characterize host specificity, reproductive behaviour, and the potential origin as well as patterns of spread of the fungus and its insect vectors. The markers resolved a total of 41 alleles and 75 haplotypes. Higher genetic diversity was found in the native populations than in the introduced populations. Based on the origin of the insect vectors, the populations of O. ips in Australia would be expected to reflect a North American origin, and those in Chile and South Africa to reflect a European origin. However, most alleles observed in the native European population were also found in the native North American population; only the allele frequencies among the populations varied. This admixture made it impossible to confirm the origin of the introduced Southern Hemisphere (SH) populations of O. ips. There was also no evidence for specificity of the fungus to particular bark beetle vectors or hosts. Although O. ips is thought to be mainly self‐fertilizing, evidence for recombination was found in the four native populations surveyed. The higher genetic diversity in the North American than in the European population suggests that North America could be the possible source region of O. ips.

A new Leptographium species associated with Tomicus piniperda in south-western China.

Tomicus species (Coleoptera: Scolytidae) are serious pests of pines with a wide distribution in Europe, Asia and America. In Yunnan, south-western China,T. piniperda has destroyed more than 0.5 million ha ofPinus yunnanensis in the past 15 years. A blue stain fungus belonging to the genusLeptographium is associated with both the shoot-feeding and trunk-attacking stages of the beetles life cycle. The fungus is morphologically similar to the anamorph ofOphiostoma crassivaginatum and toL. pyrinum, which are both characterised by short robust conidiophores and hyphae covered by a granular layer. Both these species have been isolated from conifers and are associated with insects. After comparing the fungus fromT. piniperda with similarLeptographium species, using light and scanning electron microscopy, we concluded that it represents a new taxon, which is described here asL. yunnanense sp. nov.

The effects of forest age on saproxylic beetle biodiversity: implications of shortened and extended rotation lengths in a French oak high forest

In French oak high forests, current silvicultural trends include two seemingly opposed practices: shortening the forestry cycle, which contributes to forestry intensification, and lengthening rotations in temporary set‐aside stands, called ‘ageing islands’, to favour biodiversity and high‐quality large‐diameter tree production. To derive the potential effects of these two trends, we studied habitat structure and saproxylic beetles biodiversity along an age gradient in a French oak high forest. Four age classes were surveyed: premature (i) 160/180 years and (ii) 180/200 years, (iii) mature 200/220 years, and (iv) overmature 300‐year‐old stands. Structural features were noted: deadwood volumes, density of large trees with or without microhabitats, number of cavities, presence of dead large canopy branches, sap droppings and sporophores of saproxylic fungi. Results showed that beetle species richness was positively related to stand age. Globally, overmature stands differed significantly from younger premature and mature forests in species composition and structure. Younger stands tended to show both fewer structural features and lower levels of saproxylic biodiversity. As the forest aged, the overall structural complexity and saproxylic biodiversity increased. However, no individual stand characteristic influenced preferentially biodiversity, and stand age was the best explaining factor. In conclusion, we discuss how (i) shorter rotations in the high forest production cycle and (ii) temporary set‐aside forest islands affect forest structure and deadwood‐associated assemblages. Shortening rotation length in oak high forests may negatively impact saproxylic biodiversity, whereas temporary set‐asides may play a key role for biodiversity conservation in a managed forest matrix.