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The S-adenosyl-L-methionine:pinosylvin-O-methyltransferase (PMT) gene was sequenced from Scots pine (Pinus sylvestris). The open reading frame is arranged in two exons spaced by one 102-bp intron. Promoter regulatory elements such as two "CAAT" boxes and one "TATA" box were identified. Several cis-regulatory elements were recognized: stress-responsive elements (Myb-responsive elements) as well as G, H, and GC boxes. Moreover, elicitor-responsive elements (W boxes) and a sequence resembling the simian virus 40 enhancer core were found. In phloem and needles of control trees, the transcripts of stilbene synthase (STS) and PMT were hardly detectable. Increased ozone fumigation up to 0.3 microL L(-1) enhanced the transcript level of STS and PMT in needles but not in healthy phloem. Wounding, e.g. mock inoculation, of stem-phloem was characterized by a transient increase in STS and PMT transcripts, which was more pronounced in the case of fungal inoculation. Combination of fungal-challenge or mock treatment with ozone resulted in a positive interaction at 0.3 microL L(-1). Scots pine stilbene formation appeared to be induced via STS and PMT gene expression upon ozone and fungal stress as well as wounding. The broad stress-responsiveness is in agreement with the range of various cis-acting elements detected in the STS and PMT promoters.

Les Ophiostomatales associ?es ? lps acuminatus ont ?t? isol?es des insectes, de leurs galeries et de l'aubier des pins sylvestres attaqu?s, dans le Sud-Est de la France (d?partement du Var), pendant l'?pid?mie de 1987 et 1988. Des isolements ont aussi ?t? r?alis?s dans la m?me r?gion en 1987 sur des populations end?miques d'Ips sexdentatus, et dans 2 autres r?gions (l'Orl?anais (1988) et le Mont Ventoux (1989)) sur des populations end?miques d'I acuminatus. Les vitesses de croissance des champignons varois ont ?t? compar?es au laboratoire sur malt-agar. Leur agressivit? a ?t? appr?ci?e par inoculations artificielles ? des pins sylvestres, et mesur?e par l'importance du d?veloppement de la r?action r?sinique induite et par l'importance de l'extension des champignons dans cette r?action. Le cort?ge fongique associ? ? I acuminatus dans le Sud-Est de la France comprend 4 esp?ces d'Ophiostomatales et diff?re profond?ment de celui associ? aux populations scandinaves et allemandes du m?me insecte. Il diff?re ?galement de celui associ? ? I sexdentatus, m?me quand les 2 scolytides ?voluent sur les m?mes arbres, bien que 2 esp?ces de champignons (Ophiostoma brunneo-ciliatum et Ophiostoma ips) soient communes aux 2 insectes. Dans le Var, la fr?quence d'association avec I acuminatus et le taux de fructification des champignons dans les galeries sont nettement sup?rieurs en 1987 ? ceux de 1988. Cette derni?re ann?e, aucune diff?rence n'est observ?e entre le Var et les r?gions o? les populations sont end?miques. La fr?quence d'association est aussi significativement plus faible chez les insectes essaimants que chez ceux dont l'installation a r?ussi. Des 4 esp?ces associ?es ? / acuminatus, O ips et O brunneo-ciliatum se montrent les plus agressives apr?s inoculation des arbres. Apr?s l'attaque d'I acuminatus dans la moiti? sup?rieure de l'arbre, le bleuissement de l'aubier semble s'?tendre vers la partie inf?rieure en envahissant d'abord les zones les plus profondes de ce tissu. Ophiostomatales (Ascomycetes) associated with lps acuminatus Gyll (Coleoptera: Scolytidae) in Scots pine (Pinus sylvestris L) in south-eastern France, and comparison with lps sexdentatus Boern. Ophiostomatales associated with lps acuminatus were isolated from beetles, their galleries and the sapwood of the attacked Scots pines, in south-eastern France (department of the Var), during outbreaks in 1987 and 1988. In 2 other regions, (Orl?anais and Mont Ventoux), fungi were isolated from endemic populations of the same beetles in 1988 and 1989. Fungi were also isolated from endemic populations of lps sexdentatus in the department of the Var in 1987. The growth of the fungi isolated from the Var samples was analysed on malt-agar medium. Their agressiveness was determined after artificial inoculation of Scots pines, by measuring the development of the induced defense reaction (length and resin quantity) and the extension of the fungi within this reaction. The Ophiostomatales associated with I acuminatus in south-eastern France included 4 species that differed greatly from the Scandinavian and German flora associated with the same insect. They also differed from the flora associated with I sexdentatus, even when the 2 beetles occurred in the same trees, and although the 2 fungi (Ophiostoma lps and Ophiostoma brunneo-ciliatum) were both associated with the 2 insect species (table I). In the Var region, the frequency of the association with I acuminatus, and the fructification rate of the fungi in the galleries were higher in 1987 than in 1988. In 1988, there were no differences between the Var and regions where the beetle populations were endemic. The frequency of association was also significantly lower in the swarming beetles than in the successful attacks (table I). Among the 4 fungus species associated with I acuminatus, O ips and O brunneo-ciliatum were the most aggressive after inoculation of trees (fig 2). After the successful attacks of I acuminatus in the upper half of the trees, the blue-stain invaded the lower half, beginning with the innermost zones of the sapwood.

A 3 saisons diff?rentes de l'ann?e, 5 esp?ces de champignons associ?es ? I sexdentatus et T piniperda ont ?t? inocul?es ? des pins sylvestres. Des rondins fra?chement coup?s ont aussi ?t? inocul?s au laboratoire, ? 1 seule saison. Le suivi pendant 60 j de la cin?tique de croissance des r?actions de d?fense, estim?es par leur longueur ou par la quantit? de r?sine (quantit? totale et concentration), ainsi que la cin?tique de croissance des champignons ? l'int?rieur des r?actions, ont permis d'avancer diverses hypoth?ses quant au fonctionnement des relations pin sylvestre-scolytides-champignons. Le r?le potentiel de chacun des champignons dans l'induction des r?actions de d?fense, ou dans la mort des arbres lorsque ceux-ci sont attaqu?s par les scolytes, est discut?. La r?action aurait pour r?le de stopper ou de fortement ralentir la croissance de ces champignons, en attendant que le bourrelet cicatriciel isole compl?tement les tissus agress?s. La longueur de la r?action dans le phlo?me serait conditionn?e par les capacit?s de diffusion de l'?liciteur, alors que la quantit? de r?sine accumul?e d?pendrait de la quantit? d'?liciteur. L'arr?t de l'accumulation de la r?sine en un lieu donn? ne serait pas d? ? la saturation des tissus, mais ? un arr?t de l'?licitation en ce lieu. Apr?s la fin de la croissance de la r?action, il y aurait homog?n?isation de la r?partition de la r?sine ? l'int?rieur des tissus concern?s, par diffusion de la r?sine des zones ? concentration ?lev?e vers les zones ? concentration plus faible et, dans certains cas, par continuation de la synth?se r?sinique. Il est aussi propos? de d?finir 3 param?tres repr?sentatifs de la r?action : rapidit?, intensit?, efficacit?. Kinetics of growth of fungi associated with lps sexdentatus and Tomicus piniperda (Coleoptera: Scolytidae) and of the defence reaction of the inoculated scots pines. At 3 different seasons in the year, 5 species of fungi associated with I sexdentatus and T piniperda were inoculated into scots pine trees. Logs were also inoculated once during the year in the lab. The reactions were estimated by their length and by the quantity of resin (both total and concentration). The percentage of sieve cells filled with resin was appreciated by histological sections made in each reaction, at 10 and 30 mm from the point of inoculation and in the middle between this point and the end of the reaction. The kinetics of growth of the fungi and of the defence reactions (length, resin, histology) were followed for 60 days. The temperature and defence reactions were not sufficient to explain the differences in the kinetics of growth between fungi and between seasons (figs 1 and 7). Factors related to intrinsic characteristics of the fungi and to physiological state of the tree may play a role. The possible role of each fungus is discussed according to the induction of the reaction and to tree death when attacked by bark beetle populations. The wound periderm does not seem to be directly responsible for stopping the fungal growth. In return, the considerable slowing down of this growth, followed by a slowing down of the development of the reaction length (fig 3) may give sufficient time for the wound periderm to be built, to insulate the wounded tissues completely and to contain the aggressors. The parallel between the kinetics of growth of the fungi and the kinetics of resin accumulation suggests that the quantity of resin would depend upon the quantity of elicitor produced during the fungal growth. The reaction length in the phloem would depend on the diffusion ability of the elicitor. The end of the resin accumulation at a given place would not depend upon tissue saturation, but rather on the end of the elicitation at this place (compare figs 3, 5 and table III). After the end of the reaction growth, an homogenization process of the resin distribution may take place inside the reaction, by a diffusion of the resin from the highly concentrated zones to the less concentrated zones and, in some cases, by a continuation of the resin synthesis. A definition of 3 parameters representative for the defence reaction is proposed: speed, intensity, efficiency.