Phoretic, trophic and fungal associations of Histiogaster arborsignis (Acari: Acaridae) with Ips typographus: a study case from the Czech Republic

Ips typographusL. (Coleoptera: Curculionidae) is one of the serious pests causing mass destruction of European spruce forests, with a substantial economic impact. Symbiotic microbes associated with bark beetles often play a definitive role in accomplishing their physiological and ecological functions by detoxifying chemicals, inhibiting pathogens, and offering nutrients. Although a few research works explored the microbes associated withI. typographus, much is yet to be studied to understand their adaptive ecology as holobionts comprehensively. The present study examined bacterial and fungal communities of larvae, adults, and feeding galleries from Austria and Czech Republic using high-throughput sequencing that elucidated the influence of geographic location, host, and life stage on the microbial assemblage in Eurasian spruce bark beetle,I. typographus. The most abundant bacterial genera inI. typographusincludedPseudoxanthomonas,Spiroplasma,Pseudomonas,Cellulomonas,Tyzzerella,Bacillus, andMycobacterium. Alternatively,Wickerhamomyces,Nakazawaea,Aspergillus,Ophiostoma,Cryptococcus,Rhexographium,Yamadazyma,Talaromyces, andKuraishiawere highly dominant fungal genera. Significant differences in bacterial and fungal community richness and diversity were detected among the tested samples. LEfSe analysis revealed species-specific bacterial and fungal biomarkers from different locations in the larvae, adults, and feeding gallery samples. PICRUSt2 and FUNGuild analysis documented putative roles of the bacterial and fungal communities in beetle holobiont and provided a foundation for downstream functional analyses. The current findings further enhanced our understanding of bark beetles as holobionts.

The European spruce bark beetle is a major disturbance agent in Norway spruce forests in Europe, and with a changing climate it is predicted that damage will increase. To prevent the bark beetle population buildup, and to limit further spread during outbreaks, it is crucial to detect attacked trees early. In this study, we utilize Sentinel-2 data in combination with a risk map, created from geodata and forestry data, to detect trees predisposed to and attacked by the European spruce bark beetle. Random forest models were trained over two tiles (90 × 90 km) in southern Sweden for all dates with a sufficient number of cloud-free Sentinel-2 pixels during the period May–September in 2017 and 2018. The pixels were classified into attacked and healthy to study how detection accuracy changed with time after bark beetle swarming and to find which Sentinel-2 bands are more important for detecting bark beetle attacked trees. Random forest models were trained with (1) single-date data, (2) temporal features (1-year difference), (3) single-date and temporal features combined, and (4) Sentinel-2 data and a risk map combined. We also included a spatial variability metric. The results show that detection accuracy was high already before the trees were attacked in May 2018, indicating that the Sentinel-2 data detect predisposed trees and that the early signs of attack are low for trees at high risk of being attacked. For single-date models, the accuracy ranged from 63 to 79% and 84 to 94% for the two tiles. For temporal features, accuracy ranged from 65 to 81% and 81 to 92%. When the single-date and temporal features were combined, the accuracy ranged from 70 to 84% and 90 to 96% for the two tiles, and with the risk map included, the accuracy ranged from 83 to 91% and 92 to 97%, showing that remote sensing data and geodata can be combined to increase detection accuracy. The differences in accuracy between the two tiles indicate that local differences can influence accuracy, suggesting that geographically weighted methods should be applied. For the single-date models, the SWIR, red-edge, and blue bands were generally more important, and the SWIR bands were more important after the attack, suggesting that they are most suitable for detecting the early signs of a bark beetle attack. For the temporal features, the SWIR and blue bands were more important, and for the variability metric, the green band was generally more important.

Spruce bark beetle phenological modelling and drought risk within framework of TANABBO II model

BackgroundIps typographus (European spruce bark beetle) is the most destructive pest of spruce forests in Europe. As for other animals, it has been proposed that the microbiome plays important roles in the biology of bark beetles. About the bacteriome, there still are many uncertainties regarding the taxonomical composition, insect-bacteriome interactions, and their potential roles in the beetle ecology. Here, we aim to deep into the ecological functions and taxonomical composition of I. typographus associated bacteria.ResultsWe assessed the metabolic potential of a collection of isolates obtained from different life stages of I. typographus beetles. All strains showed the capacity to hydrolyse one or more complex polysaccharides into simpler molecules, which may provide an additional carbon source to its host. Also, 83.9% of the strains isolated showed antagonistic effect against one or more entomopathogenic fungi, which could assist the beetle in its fight against this pathogenic threat. Using culture-dependent and -independent techniques, we present a taxonomical analysis of the bacteriome associated with the I. typographus beetle during its different life stages. We have observed an evolution of its bacteriome, which is diverse at the larval phase, substantially diminished in pupae, greater in the teneral adult phase, and similar to that of the larval stage in mature adults. Our results suggest that taxa belonging to the Erwiniaceae family, and the Pseudoxanthomonas and Pseudomonas genera, as well as an undescribed genus within the Enterobactereaceae family, are part of the core microbiome and may perform vital roles in maintaining beetle fitness.ConclusionOur results indicate that isolates within the bacteriome of I. typographus beetle have the metabolic potential to increase beetle fitness by proving additional and assimilable carbon sources for the beetle, and by antagonizing fungi entomopathogens. Furthermore, we observed that isolates from adult beetles are more likely to have these capacities but those obtained from larvae showed strongest antifungal activity. Our taxonomical analysis showed that Erwinia typographi, Pseudomonas bohemica, and Pseudomonas typographi species along with Pseudoxanthomonas genus, and putative new taxa belonging to the Erwiniaceae and Enterobacterales group are repeatedly present within the bacteriome of I. typographus beetles, indicating that these species might be part of the core microbiome. In addition to Pseudomonas and Erwinia group, Staphylococcus, Acinetobacter, Curtobacterium, Streptomyces, and Bacillus genera seem to also have interesting metabolic capacities but are present in a lower frequency. Future studies involving bacterial-insect interactions or analysing other potential roles would provide more insights into the bacteriome capacity to be beneficial to the beetle.

In recent years, carbon sequestration in European forests has decreased due to extensive droughts and bark beetle outbreaks brought on by climate change. The Eurasian spruce bark beetle (Ips typographus (L.)) is a natural disturbance agent in forests where Norway spruce (Picea abies (Karst)) is dominant. Through mass attacks the beetle can overwhelm defences of healthy trees. Drought stress can further reduce tree resistance and predispose the trees to bark beetle attack. The aim of this thesis was to generate knowledge and investigate both the region-specific seasonal phenology of the spruce bark beetle in southern Sweden and the resistance of its host, the Norway spruce. Pheromone traps and felled trees that were colonised were used to determine flight activity and required thermal sums (degree-days, dd >5°C) for flight start, reemergence of parental beetles, and the emergence of filial beetles. The influence of weather, phenology, and soil moisture conditions, and lag effects following a severe drought event in 2018, on spruce tree resistance was quantified by inoculating a blue stain fungus associated with bark beetles on three sites in southern Sweden on four occasions during the 2019 and 2021 growing seasons. Fungal growth was expected to show susceptibility to bark beetle attack. Flight activity started at the end of April (47 dd after 1 January). Re-emergence of parental beetles from the first brood started at the end of May (122 dd after colonisation), and continued flight showed that sister broods were frequent. The new generation started to emerge at the end of June (437 dd after colonisation) to initiate a second generation. Tree resistance was lower in the early season (June) and differed with both precipitation levels and local soil moisture conditions. Resistance was higher in 2021 than 2019, indicating a recovery. In both years, there was a significant correlation between lesion size (tree resistance) and water availability in the autumn of the previous year. In the current climate in southern Sweden, tree resistance is lowest during tree attacks from sister brood flights (June). Understanding thresholds in both bark beetle development and tree drought stress variables is crucial for predicting the impact of future bark beetle outbreaks in a climate change context.

На території Ботанічного саду Харківського національного університету (ХНУ) ім. В. М. Каразіна на хвойних породах визначено 4 види короїдів: короїда-типографа (Ips typographus L.), гравера звичайного (Pityogenes chalcographus L.), соснового короїда-крихітку (Crypturgus cinereus Herb.) та тайгового короїда-крихітку (C. subcribrosus Eg.). Відзначено, що з роду Abies лише п’ять видів не мали ознак заселення, а саме: A. balsamia, A. cephalonica, А. concolor, A. grandis, А. violacea. Picea koraiensis та P. alba не були заселені короїдами, але на стовбурі помічено спроби заселення. Встановлено, що форму P. obovata «Glauca» заселяли всі визначені види короїдів. Найбільш поширеним на ялині серед інших видів короїдів виявився Ips typographus. Серед 198 обстежених дерев визначено: живих із відсутністю ознак заселення короїдами – 27,3 %, зі спробами заселення – 9,1 %, заселених короїдами – 63,6 %.

Northernmost European spruce bark beetle Ips typographus outbreak: Modelling tree mortality using remote sensing and climate data

Individuals across various animal species communicate their presence to conspecifics. Especially phytophagous and parasitoid insects with their brood developing on limited resources rely on chemical cues, such as host-marking pheromones, to reduce intraspecific competition. Bark beetles are phytophagous insects with some species being economically and ecologically relevant forest pests. Several of them use the volatile compound verbenone to inhibit attraction and reduce intraspecific competition. However, in the Eurasian spruce bark beetle, Ips typographus (L.), temporal emission patterns did so far not quite support the putative function of verbenone as an indicator of densely colonised host trees. More importantly, it is currently unclear how well verbenone emission is actually related to colonisation density and thus intraspecific competition. Here, we inoculated Norway spruce logs with I. typographus at two defined colonisation densities in the greenhouse and measured the emission of verbenone and its precursors α-pinene and verbenol over time. Verbenone emission was 3–7 times greater from colonised logs compared to decaying logs without beetles during the major part of larval development. Furthermore, our data supports the quantitative hypothesis, that the termination of attack on a tree is mediated by a cessation of the release of verbenol and continuous emission of verbenone. The latter is most likely a passively produced host-marking cue reflecting the actual density of conspecifics since per-beetle emission was unaffected by colonisation density. These findings shed new light on the regulation of bark beetle mass aggregations, which are currently causing previously unseen economic damages in temperate forests.

The Eurasian spruce bark beetle (ESBB), Ips typographus, has recently caused catastrophic damage to Norway spruce (Picea abies) forests in Europe, resulting in the loss of more than 100 million cubic meters of wood. Traditional forest management strategies have failed to constrain the growing infestation rate; hence, novel measures must be deployed. A better understanding of ESBB physiology and adaptation to host allelochemicals may provide a platform for future management strategies using molecular tools such as RNA interference. To understand ESBB physiology and adaptation, the current study unraveled the gene expression dynamics of ESBB in different life stages and tissues. We obtained ESBB transcriptomes for different life stages [larvae (L1, L2, and L3), pupa, callow, and sclerotized adult] and male/female tissues (gut, fat body, and head) from callow and sclerotized adult beetles. Differential gene expression analysis (DGE) identified multiple gene families related to detoxification, digestion, resistance, and transport in different life stages and tissues of the beetle. Gene Ontology (GO) enrichment revealed 61 critical metabolic pathways enriched across all DGE comparisons. DGE analysis further pinpointed the differential expression of essential genes involved in detoxification, digestion, transport, and defense in various tissues and life stages. RT-qPCR experiments and enzymatic assays corroborated the findings further. The catalogue of differentially expressed genes identified in ESBB could aid better understanding of ESBB physiology and adaptation to hosts and serve as targets for future RNAi-based ESBB management.

Tree-killing bark beetles cause great economic and ecological damage worldwide. Several long-legged fly species of the genus Medetera are natural enemies of tree-killing bark beetles and are of interest as potential biological control agents. However, flies within the Medetera genus have been poorly studied to date, partly due to very difficult species identification that often requires studying the fine structures of male genital morphology. In this thesis, morphological analysis and barcoding were applied in species level identification of different Medetera specimens collected from the bark of Norway spruce trees in Southern Sweden infested with the Eurasian spruce bark beetle, Ips typographus. Adult Medetera flies use olfaction to locate bark beetle-infested trees. Olfactory cues (odours) are detected by various types of hair-like structures called sensilla distributed on the fly antennae and maxillary palps. This thesis shows that the number and subtypes of sensilla differs between M. signaticornis and M. infumata, the two most common species found at the study sites. Therefore, it is possible that these two species respond to different odours, or process the same odours in different ways. Further analysis showed that male and female M. signaticornis were able to detect more than 20 compounds emitted from Norway spruce trees infested with I. typographus. These compounds included metabolites produced by the trees and also compounds produced by bark beetles and associated microorganisms. Such detection of odours from different trophic levels by Medetera flies may facilitate the location of infested trees throughout a bark beetle attack. Multiple comparisons of synthetic blends containing different combinations of these compounds demonstrated that some compounds (e.g. (– )-cis-verbenol, ipsdienol, myrtenol, α- and γ-terpinene; limonene, camphor, terpinen-4- ol and borneol) were more important than others in the attraction of Medetera species. Overall, the findings presented in this thesis can facilitate identification and monitoring of Medetera flies and might help to improve management of I. typographus in the future.

The incidence of salvage felling is a significant indicator of stands‘ health and stability. Health is mainly indicated by biotic and anthropogenic factors, while abiotic effects are primarily an indicator of a stand’s stability. All these factors influence each other and subsequently they can result in salvage felling. For the Czech Republic, there have been relevant data for the period since 1964. Actually, recent data cover approximately 70% of the Czech Republic´s area in the twenty-year period assessed in this text. During this period, the volume of salvage felling amounted to 89.2 million m3 which represents 28.4% of total felling in this period. The largest share is caused by abiotic effects (18.6%), next by biotic agents (9.6%) and anthropogenic are only responsible for 0.2%. In the last two years, the volume of salvage felling caused by biotic agents was higher than the volume of salvage felling caused by abiotic and anthropogenic agents for the first time. In terms of biotic agents, almost the whole volume is represented by bark beetle wood as a result of spruce stands infestation by the European spruce bark beetle – Ips typographus (L.) and double-spined bark beetle – Ips duplicatus (Sahl.), and to a small extent also by other species of bark beetles on spruce, pine and occasionally other wood tree species. In the last three years, mainly the incidence of the Ips typographus L., has concentrated in North Moravia and Silesia. Currently, it is also spreading in South Moravia and Bohemia and in districts along the state borders with Austria and Germany, with the most serious situation in this region being the one in the Bohemian-Moravian Highlands.

4 - The Eurasian spruce bark beetle in a warming climate: Phenology, behavior, and biotic interactions

Storms, temperature maxima and the Eurasian spruce bark beetle Ips typographus—An infernal trio in Norway spruce forests of the Central European High Tatra Mountains

Eurasian spruce bark beetle, Ips typographus is a destructive pest of the Norway spruce (Picea abies). Recent outbreaks in Europe have been attributed to global warming and other anthropogenic impacts. Bark beetles are guided by multiple complex olfactory cues throughout their life cycle. Male-produced aggregation pheromones, comprising 2-methyl-3-buten-2-ol and cis-verbenol, have been identified as the most powerful attractants for dispersing conspecifics. In addition to host trees, bark beetles interact with multiple organisms, including symbiotic ophiostomatoid fungi, which may promote beetle colonization success and offspring development. Previously, in a short-distance laboratory assay, we demonstrated that I. typographus adults are attracted to the volatile organic compounds (VOCs) produced by three symbiotic fungi: Grosmannia penicillata, Endoconidiophora polonica, and Leptographium europhioides. Furthermore, the abundant fusel alcohols and their acetates were found to be the most attractive odorants in the fungal VOC profile. In this study, using a long-distance field-trapping experiment, we analyzed the role of fungal VOCs as attractants for dispersing I. typographus. Two types of fungal lures were tested in combination with pheromones in traps: (1) live cultures of fungi grown on potato dextrose agar (PDA) and (2) dispensers containing synthetic fusel alcohols and their acetates in equal proportions. Subsequently, the composition of VOCs emitted from live fungal lures were analyzed. We found that the symbiotic fungi synergistically increased the attraction of beetles to pheromones in field traps and the attractiveness of live fungal lures depended on the fungal load. While one Petri dish with E. polonica, when combined with pheromones synergistically increased trapping efficiency, three Petri dishes with L. europhioides were required to achieve the same. The synthetic mix of fungal fusel alcohols and acetates improved the catch efficiency of pheromones only at a low tested dose. VOC analysis of fungal cultures revealed that all the three fungi produced fusel alcohols and acetates but in variable composition and amounts. Collectively, the results of this study show that, in addition to pheromones, bark beetles might also use volatile cues from their symbiotic fungi to improve tree colonization and reproductive success in their breeding and feeding sites.

A Festschrift for Stephen L. Wood