Pathogenicity and induced resistance in Larix kaempferi and Larix olgensis inoculated with Endoconidiophora fujiensis.

With climate warming and economic globalization, insect-microbe assemblages are becoming increasingly responsible for various devastating forest diseases worldwide. Japanese larch (Larix kaempferi) is extensively cultivated in China because of its high survival rate, rapid maturation and robust mechanical properties. Endoconidiophora fujiensis, an ophiostomatoid fungus associated with Ips subelongatus, has been identified as a lethal pathogen of L. kaempferi in Japan. However, there is a dearth of research on the pathogenicity of E. fujiensis in larches in China. Therefore, we investigated the pathogenicity of E. fujiensis in introduced L. kaempferi and indigenous larch (Larix olgensis) trees and compared the induced resistance responses to the pathogen in both tree species in terms of physiology and gene expression. Five-year-old saplings and 25-year-old adult trees of L. olgensis and L. kaempferi were inoculated in parallel during the same growing season. Endoconidiophora fujiensis exhibited high pathogenicity in both larch species, but particularly in L. kaempferi compared with L. olgensis adult trees; adult L. olgensis was more resistant to E. fujiensis than adult L. kaempferi, which was reflected in higher accumulation of defensive monoterpenes, such as myrcene, 3-carene and limonene and the earlier induction of defense genes catalase (CAT) and pathogenesis-related protein 1 (PR1). This study contributes to our understanding of the interactions between bark beetle-associated ophiostomatoid fungi and host larches, from phenotypic responses to alterations in secondary metabolites via defense- and metabolism-related gene activation, providing a valuable foundation for the management of larch diseases and pests in the future.

SEM analysis of sensilla on the mouthparts and antennae of Asian larch bark beetle Ips subelongatus.

The Asian larch bark beetle, Ips subelongatus, is a severe pest of larches in Northeastern China. The gustatory and olfactory systems of I. subelongatus play important roles in host location, mating, and feeding. In this study, we examined the types, distributions, and abundances of various sensilla associated with the mouthparts and antennae of I. subelongatus using scanning electron microscopy (SEM). On the mouthparts, five types of sensilla are present: sensilla trichodea (S.t.1-3), sensilla chaetica (S.c.1-3), sensilla basiconica (S.b.1-2), sensilla twig basiconica (S.tb.1-3), and sensilla placodea (S.p). S.t.3 are the most abundant sensilla subtype on the mouthparts in both sexes, while S.b.1 are the least abundant. Most sensilla on the mouthparts are located on the maxillae and labium, and the apex of each maxillary and labial palp carry the same sensilla subtypes (S.b.2 and S.tb.1-3). However, the total number of sensilla on the apex of each maxillary palp is higher than that on the labial palp. On the antennae, five types of sensilla are present: sensilla trichodea (S.t.1-3), sensilla chaetica (S.c.1-2), sensilla basiconica (S.b.1-3), Böhm bristles (B.b), and sensilla coeloconica (S.co). Antennal sensilla are mostly situated on the anterior surface of the antennal club, particularly on the two dense sensory bands. S.b.1 are the most abundant sensilla subtype on the antennae in both sexes, while S.t.1 are the least abundant. No sexual dimorphism in sensilla type or distribution on the mouthparts or antennae is observed between the sexes of I. subelongatus. However, S.t.3 (on mouthparts) and S.c.1 (on antennae) were significantly more abundant in males than in females, while more S.t.1 (on mouthparts) were observed in females than in males. Finally, the putative functions of each kind of sensilla with respect to their fine structures, distributions, and abundances on the mouthparts and antennae are discussed.

Chemical signal interactions of the bark beetle with fungal symbionts, and host/non-host trees.

The symbiosis between the bark beetle (Ips subelongatus) and its fungal symbiont (Endoconidiophora fujiensis) poses a serious threat to larch forests. However, the signaling pathways between these symbiotic partners and their host/non-host trees are not fully understood. Inoculation of the host larch (Larix principis-rupprechtii) with two strains of E. fujiensis induced a rapid and long-term release of monoterpenes. Although the fungi had a level of tolerance to these compounds, many monoterpenes inhibited fungal growth in culture. Moreover, monoterpenes with stronger inhibitory effects on fungal growth exhibited weaker synergistic effects on the attraction of I. subelongatus to aggregation pheromone. Surprisingly, individual isomers of aggregation pheromone components promoted fungal symbiont growth in a culture medium. Non-host volatiles (NHVs) were tested and shown to completely inhibit the growth of fungal symbionts in culture but had no effects on beetle responses to aggregation pheromone, with the exception of (Z)-3-hexen-1-ol. These results reveal convergence and mutualism patterns in the evolution of I. subelongatus and E. fujiensis with respect to host tree volatiles but not in response to NHVs. Ultimately, we put forward a hypothesis that host plants are ecological and evolutionary determinants of bark beetle-fungus symbioses in terms of their complex signaling interactions.

Forecasting Spring Flight of Ips subelongatus (Coleoptera: Curculionidae: Scolytinae) in Japanese larch Larix kaempferi (Pinales: Pinaceae) Forests in the Republic of Korea.

Ips subelongatus is a species of bark beetle experiencing population outbreaks in Korea. In this study, a predictive model and empirical prediction were used to forecast the spring flight of these beetles in Japanese larch forests. The number of beetles caught in pheromone traps was investigated in larch forests thinned in 2009, 2010, 2012, or 2013. Data from the sites thinned in 2009, 2010, or 2012 were used in the predictive model based on a degree-day model that was validated using data from the site thinned in 2013. The lower threshold temperature for flight (LTF) and a thermal sum for the spring flight of I. subelongatus were estimated. The empirical prediction that beetles initiate their flight when daily maximum temperatures reach 16 or 20°C was tested using daily maximum temperature and the beetles caught. The LTF was estimated as 5.97°C, with 42.95 degree-days required for initiation of spring flight. The median flight dates were estimated with a discrepancy from 1 to 3 d by the predictive model. Using the empirical prediction, differences between the day when daily maximum temperature reached 16 or 20°C and flight peak days ranged from 4 to 45 d. These results demonstrate that the predictive model is more suitable than the empirical prediction for predicting the spring flight of I. subelongatus. Overwintering I. subelongatus adults seem to need to gain a determined thermal sum before initiating spring flight rather than merely waiting for the daily maximum temperature to exceed a critical temperature.

Population divergence of aggregation pheromone responses in Ips subelongatus in northeastern China.

The Asian larch bark beetle, Ips subelongatus, is considered to be the major pest of larch within its natural range. We investigated the electrophysiological and behavioral characteristics as well as mitochondrial DNA cytochrome oxidase subunit I sequences of I. subelongatus from 13 geographic populations throughout northeastern China in order to explore population divergence of aggregation pheromone responses and the extent of potential genetic divergence. Electrophysiological analyses showed that antennae of I. subelongatus from all the six tested populations responded strongly to (S)-(-)-ipsenol (100% detection; 0.35-0.73 mV) in gas chromatography (GC)-electroantennographic detection (EAD) analyses, while its antipode, (R)-(+)-ipsenol was antennally inactive. I. subelongatus populations varied in their responses to (R)-(-)- and (S)-(+)-ipsdienol in GC-EAD analyses. Behavioral bioassays demonstrated that (S)-(-)-ipsenol alone was significantly attractive at all the tested sites, supporting its status as a key pheromone component of I. subelongatus, whereas (S)-(+)-ipsdienol was inactive alone. Adding (S)-(+)-ipsdienol to (S)-(-)-ipsenol did not have any effect on the trap catches from some populations in Inner Mongolia. However, (S)-(+)-ipsdienol showed a strong synergistic effect on (S)-(-)-ipsenol from several populations in Jilin and Liaoning Provinces, and a weak synergistic effect from some transition populations in Heilongjiang Province. Furthermore, 27 mitochondrial haplotypes were found among the 13 populations (intraspecific nucleotide divergence, 0.1%-1.1%). Analyses of molecular variance and haplotype networks indicated that different geographic populations have developed some genetic variation but did not form completely independent groups. From an applied point of view, a universal synthetic binary blend of racemic ipsenol and (S)-(+)-ipsdienol might have a potential for monitoring or even mass-trapping of I. subelongatus across northeastern China, even though some populations only use (S)-(-)-ipsenol alone as their active pheromone component.