Aggregation pheromones have a non-linear effect on oviposition behavior in Drosophila melanogaster.

Female fruit flies (Drosophila melanogaster) oviposit at communal sites where the larvae may cooperate or compete for resources depending on group size. This offers a model system to determine how females assess quantitative social information. We show that the concentration of pheromones found on a substrate increases linearly with the number of adult flies that have visited that site. Females prefer oviposition sites with pheromone concentrations corresponding to an intermediate number of previous visitors, whereas sites with low or high concentrations are unattractive. This dose-dependent decision is based on a blend of 11-cis-Vaccenyl Acetate (cVA) indicating the number of previous visitors and heptanal (a novel pheromone deriving from the oxidation of 7-Tricosene), which acts as a dose-independent co-factor. This response is mediated by detection of cVA by odorant receptor neurons Or67d and Or65a, and at least five different odorant receptor neurons for heptanal. Our results identify a mechanism allowing individuals to transform a linear increase of pheromones into a non-linear behavioral response.

Dynamic changes to signal allocation rules in response to variable social environments in house mice.

Urine marking is central to mouse social behavior. Males use depletable and costly urine marks in intrasexual competition and mate attraction. We investigate how males alter signaling decisions across variable social landscapes using thermal imaging to capture spatiotemporal marking data. Thermal recording reveals fine-scale adjustments in urinary motor patterns in response to competition and social odors. Males demonstrate striking winner-loser effects in scent mark allocation effort and timing. Competitive experience primes temporal features of marking and modulates responses to scent familiarity. Males adjust signaling effort, mark latency, and marking rhythm, depending on the scent identities in the environment. Notably, recent contest outcome affects how males respond to familiar and unfamiliar urine. Winners increase marking effort toward unfamiliar relative to familiar male scents, whereas losers reduce marking effort to unfamiliar but increase to familiar rival scents. All males adjust their scent mark timing after a contest regardless of fight outcome, and deposit marks in more rapid bursts during marking bouts. In contrast to this dynamism, initial signal investment predicts aspects of scent marking days later, revealing the possibility of alternative marking strategies among competitive males. These data show that mice flexibly update their signaling decisions in response to changing social landscapes.

Ozone exposure disrupts insect sexual communication.

Insect sexual communication often relies upon sex pheromones. Most insect pheromones, however, contain carbon-carbon double bonds and potentially degrade by oxidation. Here, we show that frequently reported increased levels of Anthropocenic ozone can oxidize all described male-specific pheromones of Drosophila melanogaster, resulting in reduced amounts of pheromones such as cis-Vaccenyl Acetate and (Z)-7-Tricosene. At the same time female acceptance of ozone-exposed males is significantly delayed. Interestingly, groups of ozone-exposed males also exhibit significantly increased levels of male-male courtship behaviour. When repeating similar experiments with nine other drosophilid species, we observe pheromone degradation and/or disrupted sex recognition in eight of them. Our data suggest that Anthropocenic levels of ozone can extensively oxidize double bonds in a variety of insect pheromones, thereby leading to deviations in sexual recognition.

Geosmin suppresses defensive behaviour and elicits unusual neural responses in honey bees.

Geosmin is an odorant produced by bacteria in moist soil. It has been found to be extraordinarily relevant to some insects, but the reasons for this are not yet fully understood. Here we report the first tests of the effect of geosmin on honey bees. A stinging assay showed that the defensive behaviour elicited by the bee's alarm pheromone component isoamyl acetate (IAA) is strongly suppressed by geosmin. Surprisingly, the suppression is, however, only present at very low geosmin concentrations, and disappears at higher concentrations. We investigated the underlying mechanisms at the level of the olfactory receptor neurons by means of electroantennography, finding the responses to mixtures of geosmin and IAA to be lower than to pure IAA, suggesting an interaction of both compounds at the olfactory receptor level. Calcium imaging of the antennal lobe (AL) revealed that neuronal responses to geosmin decreased with increasing concentration, correlating well with the observed behaviour. Computational modelling of odour transduction and coding in the AL suggests that a broader activation of olfactory receptor types by geosmin in combination with lateral inhibition could lead to the observed non-monotonic increasing-decreasing responses to geosmin and thus underlie the specificity of the behavioural response to low geosmin concentrations.

Nematode Pheromones: Structures and Functions.

Pheromones are chemical signals secreted by one individual that can affect the behaviors of other individuals within the same species. Ascaroside is an evolutionarily conserved family of nematode pheromones that play an integral role in the development, lifespan, propagation, and stress response of nematodes. Their general structure comprises the dideoxysugar ascarylose and fatty-acid-like side chains. Ascarosides can vary structurally and functionally according to the lengths of their side chains and how they are derivatized with different moieties. In this review, we mainly describe the chemical structures of ascarosides and their different effects on the development, mating, and aggregation of nematodes, as well as how they are synthesized and regulated. In addition, we discuss their influences on other species in various aspects. This review provides a reference for the functions and structures of ascarosides and enables their better application.

Post-fumigation sub-lethal activities of phosphine and ethyl formate on survivorship, fertility and female sex pheromone production of Callosobruchus chinensis (L.).

Phosphine (PH3) and ethyl formate (EF), the two popular fumigant disinfectants of stored product insect pests, are primarily evaluated for their knock down effects without considering their post-fumigation sub-lethal activities. The sub-lethal activities (adult survivorship, fecundity, sterility and female sex pheromone production) of the fumigants were evaluated on a field-to-storage insect pest adzuki bean beetle, Callosobruchus chinensis (L.). The adults' survivorship and female fecundity, both were dose-dependently affected by sub-lethal PH3 and EF fumigation exposures. Hatchability of the eggs laid by fumigated female adults were also significantly affected. Gas-chromatography mass-spectrometry analysis of solid-phase micro-extraction from virgin fumigated C. cinensis females revealed that the PH3 LC25 (the lethal concentration required to kill the 25% of the population) fumigated female C. chinensis released significantly less amount of the pheromone components. In contrast, EF LC25 exposure did not affect the pheromone release. This study unveils the facts that the EF and PH3 fumigation have detrimental bioactivities against C. chinensis. Notably, this suggests to consider the sub-lethal EF and PH3 fumigation rather than the dose required to instantly kill all the C. chinensis individuals for disinfestation of stored adzuki bean.

Human scent signature on cartridge case survives gun being fired: A preliminary study on a potential of scent residues as an identification tool.

This paper focuses on a chemical analysis of human scent samples that were obtained from cartridge cases after being fired and their comparison with scent samples collected under laboratory conditions. Scent samples were analyzed by comprehensive two-dimensional gas chromatography coupled with the time-of-flight mass spectrometer. The results obtained from the chemical analyzes confirmed the desired stability of the human scent evidence and outlined the possible application for forensic purposes. The qualitative results of the study converge with the findings of previous studies on the composition of human scent and the chemical composition of human fingerprints. Furthermore, statistical analyzes were performed employing similarity algorithms such as Pearson's and Spearman's correlations, or Kendall's tau. The resulting comparison of the scent samples secured on fired cartridge cases compared with those samples collected under laboratory conditions yielded ten out of ten correct identifications of the scent inflictor.

Meta-analytic evidence that allelopathy may increase the success and impact of invasive grasses.

Background: In the grass family, a disproportionate number of species have been designated as being invasive. Various growth traits have been proposed to explain the invasiveness of grasses; however, the possibility that allelopathy gives invasive grasses a competitive advantage has attracted relatively little attention. Recent research has isolated plant allelochemicals that are mostly specific to the grass family that can breakdown into relatively stable, toxic byproducts. Methods: We conducted a meta-analysis of studies on grass allelopathy to test three prominent hypotheses from invasion biology and competition theory: (1) on native recipients, non-native grasses will have a significantly more negative effect compared to native grasses (Novel Weapons Hypothesis); (2) among native grasses, their effect on non-native recipients will be significantly more negative compared to their effect on native recipients (Biotic Resistance Hypothesis); and (3) allelopathic impacts will increase with phylogenetic distance (Phylogenetic Distance Hypothesis). From 23 studies, we gathered a dataset of 524 observed effect sizes (delta log response ratios) measuring the allelopathic impact of grasses on growth and germination of recipient species, and we used non-linear mixed-effects Bayesian modeling to test the hypotheses. Results: We found support for the Novel Weapons Hypothesis: on native recipients, non-native grasses were twice as suppressive as native grasses (22% vs 11%, respectively). The Phylogenetic Distance Hypothesis was supported by our finding of a significant correlation between phylogenetic distance and allelopathic impact. The Biotic Resistance Hypothesis was not supported. Overall, this meta-analysis adds to the evidence that allelochemicals may commonly contribute to successful or high impact invasions in the grass family. Increased awareness of the role of allelopathy in soil legacy effects associated with grass invasions may improve restoration outcomes through implementation of allelopathy-informed restoration practices. Examples of allelopathy-informed practices, and the knowledge needed to utilize them effectively, are discussed, including the use of activated carbon to neutralize allelochemicals and modify the soil microbial community.

Three-dimensional continuous picking path planning based on ant colony optimization algorithm.

Fruit-picking robots are one of the important means to promote agricultural modernization and improve agricultural efficiency. With the development of artificial intelligence technology, people are demanding higher picking efficiency from fruit-picking robots. And a good fruit-picking path determines the efficiency of fruit-picking. Currently, most picking path planning is a point-to-point approach, which means that the path needs to be re-planned after each completed path planning. If the picking path planning method of the fruit-picking robot is changed from a point-to-point approach to a continuous picking method, it will significantly improve its picking efficiency. The optimal sequential ant colony optimization algorithm(OSACO) is proposed for the path planning problem of continuous fruit-picking. The algorithm adopts a new pheromone update method. It introduces a reward and punishment mechanism and a pheromone volatility factor adaptive adjustment mechanism to ensure the global search capability of the algorithm, while solving the premature and local convergence problems in the solution process. And the multi-variable bit adaptive genetic algorithm is used to optimize its initial parameters so that the parameter selection does not depend on empirical and the combination of parameters can be intelligently adjusted according to different scales, thus bringing out the best performance of the ant colony algorithm. The results show that OSACO algorithms have better global search capability, higher quality of convergence to the optimal solution, shorter generated path lengths, and greater robustness than other variants of the ant colony algorithm.

Discovery of novel anti-cyanobacterial allelochemicals by multi-conformational QSAR approach.

Microcystis aeruginosa causes cyanobacterial harmful algal blooms (cHABs) in various freshwater environments. Due to global climate change, the cHABs have even spread to estuaries and coasts. Plant-derived flavones have been reported as allelochemicals that efficiently inhibit the growth of M. aeruginosa. Quantitative structure-activity relationship (QSAR) was applied to investigate the factors affecting the M. aeruginosa inhibitory activity of flavones, and to discover novel allelochemicals against M. aeruginosa. We constructed 2D and 3D-QSAR models based on the half maximum inhibitory concentration (IC50) of 22 flavones against M. aeruginosa, using molecular descriptors from multiple stable conformations. Both models showed satisfactory performances (2D-QSAR: r2=0.899, q2=0.596, rtest2=0.801; 3D-QSAR: r2=0.810, q2=0.516, rtest2=0.897). The 2D-QSAR model indicates that the anti-cyanobacterial activity is positively correlated with minimum and maximum surface electrostatic potential, and negatively correlated with polarity index and polar surface area. Through the 3D-QSAR approach, electronegative hydroxyl groups in 5- and 4'-position were favorable for the anti-cyanobacterial activity. In addition, we selected six untested flavones that fit the "activity-favorable" pattern of the visualized 3D-QSAR model. Five of the external flavones exhibited significant cyanobacterial inhibitory ability at their predicted IC50 by the 3D-QSAR model. In particular, diosmetin achieved an inhibition rate of 70.50±4.74%, which was much higher than expected. The flavones screened by the 3D-QSAR model are novel cyanobacterial inhibitors and should be fully exploited to mitigate cHABs.

The monodominance of Acuri in Pantanal formations: allelochemical effects of its leaves and the presence of Acurizais.

The presence of monodominant vegetative formations almost exclusively composed of Acuri palm trees (Attalea phalerata) stands out in some regions of the Pantanal Sul-Mato-Grossense. These formations are generally associated with anthropic, edaphic and/or hydrological factors. However, little is known about the effect of allelopathy on the formation and maintenance of these areas. Herein, we investigated the chemical composition of A. phalerata aqueous leaf extract and the potential allelopathic effects on germination and growth of target L. sativa, L. esculentum and S. obtusifolia species. Thus, extracts at concentrations of 0, 2.5, 5, 10, 15, and 20% were used for germination and growth bioassays with a completely randomised design in a germination chamber and greenhouse. The results showed that the A. phalerata extracts negatively affected the germination speed index and mean germination time of the target species and positively affected seedling length under controlled conditions and were also stimulated in the greenhouse. Thus, the formation of Acurizals can be related to the presence of secondary metabolites in the leaves, in addition to other environmental factors.

Development of a New Sex Attractant via the Peripheral Coding of Pheromones in Mythimna loreyi.

Sex pheromones play an essential role when moths are searching for mates. Male olfactory receptor neurons (ORNs) are the primary determinant during peripheral pheromone recognition. Here, we identified the sex pheromones of a global agricultural pest, Mythimna loreyi, using gas chromatography coupled with mass spectrometry and electroantennographic detection. Nine pheromone components were identified, including (Z)-9-tetradecen-1-yl acetate (Z9-14:OAc), (Z)-7-dodecen-1-yl acetate (Z7-12:OAc), and (Z)-11-hexadecen-1-yl acetate (Z11-16:OAc), and the first two elicited electrophysiological activities in the male antennae. Trichoid sensilla were classified into four functional types on the basis of neuronal responses to pheromones by single sensillum recording. Five functional ORNs were involved in recognizing pheromones and pheromone analogues. Finally, a field bioassay revealed that a blend of Z9-14:OAc, Z7-12:OAc, and Z11-16:OAc at a ratio of 100:8.8:19.7 was highly efficient for trapping males. Our results uncover the pheromone recognition mechanism in M. loreyi and provide a novel angle for developing efficient sex attractants of pests on the basis of screening the peripheral olfactory neurons.

Scent mark signal investment predicts fight dynamics in house mice.

Signals mediate competitive interactions by allowing rival assessment, yet are often energetically expensive to produce. One of the key mechanisms maintaining signal reliability is social costs. While the social costs of over-signalling are well known, the social costs of under-signalling are underexplored, particularly for dynamic signals. In this study, we investigate a dynamic and olfactory-mediated signalling system that is ubiquitous among mammals: scent marking. Male house mice territorially scent mark their environment with metabolically costly urine marks. Competitive male mice are thought to deposit abundant scent marks in the environment. However, we recently identified a cohort of low-marking males that win fights. We hypothesized that there may be social costs imposed on individuals who under-invest in signalling. Here we find that scent mark investment predicts fight dynamics. Winning males that produce fewer scent marks prior to a fight engage in more intense fights that take longer to resolve. This effect appears to be driven by an unwillingness among losers to acquiesce to weakly signalling winners. We, therefore, find evidence for rival assessment of scent marks as well as social costs to under-signalling. This supports existing hypotheses for the importance of social punishment in maintaining optimal signalling equilibria. Our results further highlight the possibility of diverse signalling strategies in house mice.

Septin Defects Favour Symmetric Inheritance of the Budding Yeast Deceptive Courtship Memory.

Mnemons are prion-like elements that encode cellular memories of past cellular adaptations and do not spread to progenies during cell divisions. During the deceptive courtship in budding yeast, the Whi3 mnemon (Whi3mnem) condenses into a super-assembly to encode a mating pheromone refractory state established in the mother cell. Whi3mnem is confined to the mother cell such that their daughter cells have the ability to respond to the mating pheromone. Confinement of Whi3mnem involves its association with the endoplasmic reticulum membranes and the compartmentalization of these membranes by the lateral membrane diffusion barrier at the bud neck, the limit between the mother cell and the bud. However, during the first cell division after the establishment of the pheromone refractory state, this adaptation is more likely to be inherited by the daughter cell than in subsequent cell divisions. Here, we show that the first cell division is associated with larger daughter cells and cytokinesis defects, traits that are not observed in subsequent cell divisions. The cytoskeletal septin protein shows aberrant localisation in these divisions and the septin-dependent endoplasmic reticulum membrane diffusion barrier is weakened. Overall, these data suggest that cytokinesis defects associated with prolonged cell division can alter the confinement and inheritance pattern of a cellular memory.

Ecotoxic side-effects of allelochemicals on submerged plant and its associated microfloras effectively relieved by sustained-release microspheres.

Harmful algae bloom caused by water eutrophication is a burning question worldwide. Allelochemicals sustained-release microspheres (ACs-SMs) exhibited remarkable inhibition effect on algae, however, few studies have focused on the ecotoxic side-effects of ACs-SMs on submerged plant and its associated microfloras. Herein the effects of different exposure situations including single high-concentration ACs (15 mg/L, SH-ACs), repeated low-concentration ACs (3 × 5 mg/L, RL-ACs) and ACs-SMs containing 15 mg/L ACs on morphological indexes, chlorophyll content, lipid peroxidation, enzymatic activity, and chlorophyll fluorescence indexes of submerged plant Vallisneria natans and the richness and diversity of its associated microfloras (epibiotic microbes and sediment microbes) were studied. The results showed that pure ACs (RL-ACs and SH-ACs groups) had negative effects on plant height, mean leaf number and area of V. natans, but promoted the increase of mean leaf length. In addition, pure ACs caused lipid peroxidation, activated the antioxidant defense system, decreased chlorophyll content, and damaged photosynthetic system in leaves. Interestingly, ACs-SMs not only had barely negative effects on above indexes of V. natans, but had certain positive effects at the later experiment stage (days 50-60). Pure ACs and ACs-SMs all reduced the richness and diversity of microfloras in each group, and promoted the increase of relative abundance of dominant bacteria Pseudomonas, leading to a simpler community structure. Significantly, V. natans leaves diminished the effects of pure ACs and ACs-SMs on epibiotic microbes, and the plant rhizosphere was beneficial to the increase of dominant bacteria that promoted plant growth. Thus, sustained-release microspherification technology can effectively relieve the ecotoxic side-effects of pure ACs on submerged plant and its associated microfloras. This study fills the gap on the ecological safety knowledge of ACs-SMs and provides primary data for evaluating the feasibility and commercialization prospects of ACs-SMs as algae inhibitor in aquatic ecosystem.

Intrasexual cuticular hydrocarbon dimorphism in a wasp sheds light on hydrocarbon biosynthesis genes in Hymenoptera.

Cuticular hydrocarbons (CHCs) cover the cuticle of insects and serve as desiccation barrier and as semiochemicals. While the main enzymatic steps of CHC biosynthesis are well understood, few of the underlying genes have been identified. Here we show how exploitation of intrasexual CHC dimorphism in a mason wasp, Odynerus spinipes, in combination with whole-genome sequencing and comparative transcriptomics facilitated identification of such genes. RNAi-mediated knockdown of twelve candidate gene orthologs in the honey bee, Apis mellifera, confirmed nine genes impacting CHC profile composition. Most of them have predicted functions consistent with current knowledge of CHC metabolism. However, we found first-time evidence for a fatty acid amide hydrolase also influencing CHC profile composition. In situ hybridization experiments furthermore suggest trophocytes participating in CHC biosynthesis. Our results set the base for experimental CHC profile manipulation in Hymenoptera and imply that the evolutionary origin of CHC biosynthesis predates the arthropods' colonization of land.

The influence of manual semen collection in male trained dogs (Canis familiaris), in the presence or absence of a female in estrus, on the concentrations of cortisol, oxytocin, prolactin and testosterone.

Sex pheromones are chemical substances secreted into the environment that affect the physiology and behavior of recipients. Females use these compounds during oestrus to attract males, which leads to attempts of mating. This study evaluates the influence of manual semen collection in male dogs, in the presence or absence of a female in estrus, on the blood concentrations of cortisol (CRT), oxytocin (OXT), prolactin (PRL) and testosterone (T), as hormones involved both in the physiology of reproduction and stress. Ten male dogs were used in Experiment 1 to measure the serum and plasma concentrations of the aforementioned hormones in the absence of semen collection. Subsequently in the same animals, the concentrations of these hormones were evaluated before and after semen collection in the presence (Exp. 2) or in absence of a female in estrus (Exp. 3). No significant changes in hormone concentration caused by the semen collection were found, either with, or without the presence of female in estrus. Obtained results suggest that the procedure of manual semen collection in dogs, probably due to its passive character, does not stimulate endocrine glands to secrete hormones, and the process of ejaculation is probably controlled by neural pathway. The lack of effect of semiochemical stimulation to the CRT, PRL, OXT and T level, could be caused by a short contact with female during semen collection. Further studies on involvement of the hormones during the process of natural mating, especially preceded by long courtships, similar to that observed under natural conditions, should shed a light on the physiology of mating and the connection between the endocrine system and semiochemical stimulation in dogs.

Brown bear skin-borne secretions display evidence of individuality and age-sex variation.

Scent originates from excretions and secretions, and its chemical complexity in mammals translates into a diverse mode of signalling. Identifying how information is encoded can help to establish the mechanisms of olfactory communication and the use of odours as chemical signals. Building upon existing behavioural and histological literature, we examined the chemical profile of secretions used for scent marking by a solitary, non-territorial carnivore, the brown bear (Ursus arctos). We investigated the incidence, abundance, and uniqueness of volatile organic compounds (VOCs) from cutaneous glandular secretions of 12 wild brown bears collected during late and post-breeding season, and assessed whether age-sex class, body site, and individual identity explained profile variation. VOC profiles varied in the average number of compounds, compound incidence, and compound abundance by age-sex class and individual identity (when individuals were grouped by sex), but not by body site. Mature males differed from other age-sex classes, secreting fewer compounds on average with the least variance between individuals. Compound uniqueness varied by body site and age for both males and females and across individuals. Our results indicate that brown bear skin-borne secretions may facilitate age-sex class and individual recognition, which can contribute towards further understanding of mating systems and social behaviour.

The Alarm Pheromone and Alarm Response of the Clonal Raider Ant.

Ants communicate via an arsenal of different pheromones produced in a variety of exocrine glands. For example, ants release alarm pheromones in response to danger to alert their nestmates and to trigger behavioral alarm responses. Here we characterize the alarm pheromone and the alarm response of the clonal raider ant Ooceraea biroi, a species that is amenable to laboratory studies but for which no pheromones have been identified. During an alarm response, ants quickly become unsettled, leave their nest pile, and are sometimes initially attracted to the source of alarm, but ultimately move away from it. We find that the alarm pheromone is released from the head of the ant and identify the putative alarm pheromone as a blend of two compounds found in the head, 4-methyl-3-heptanone and 4-methyl-3-heptanol. These compounds are sufficient to induce alarm behavior alone and in combination. They elicit similar, though slightly different behavioral features of the alarm response, with 4-methyl-3-heptanone being immediately repulsive and 4-methyl-3-heptanol being initially attractive before causing ants to move away. The behavioral response to these compounds in combination is dose-dependent, with ants becoming unsettled and attracted to the source of alarm pheromone at low concentrations and repulsed at high concentrations. While 4-methyl-3-heptanone and 4-methyl-3-heptanol are known alarm pheromones in other more distantly related ant species, this is the first report of the chemical identity of a pheromone in O. biroi, and the first alarm pheromone identified in the genus Ooceraea. Identification of a pheromone that triggers a robust, consistent, and conserved behavior, like the alarm pheromone, provides an avenue to dissect the behavioral and neuronal mechanisms underpinning chemical communication.

Conifer-killing bark beetles locate fungal symbionts by detecting volatile fungal metabolites of host tree resin monoterpenes.

Outbreaks of the Eurasian spruce bark beetle (Ips typographus) have decimated millions of hectares of conifer forests in Europe in recent years. The ability of these 4.0 to 5.5 mm long insects to kill mature trees over a short period has been sometimes ascribed to two main factors: (1) mass attacks on the host tree to overcome tree defenses and (2) the presence of fungal symbionts that support successful beetle development in the tree. While the role of pheromones in coordinating mass attacks has been well studied, the role of chemical communication in maintaining the fungal symbiosis is poorly understood. Previous evidence indicates that I. typographus can distinguish fungal symbionts of the genera Grosmannia, Endoconidiophora, and Ophiostoma by their de novo synthesized volatile compounds. Here, we hypothesize that the fungal symbionts of this bark beetle species metabolize spruce resin monoterpenes of the beetle's host tree, Norway spruce (Picea abies), and that the volatile products are used as cues by beetles for locating breeding sites with beneficial symbionts. We show that Grosmannia penicillata and other fungal symbionts alter the profile of spruce bark volatiles by converting the major monoterpenes into an attractive blend of oxygenated derivatives. Bornyl acetate was metabolized to camphor, and α- and ß-pinene to trans-4-thujanol and other oxygenated products. Electrophysiological measurements showed that I. typographus possesses dedicated olfactory sensory neurons for oxygenated metabolites. Both camphor and trans-4-thujanol attracted beetles at specific doses in walking olfactometer experiments, and the presence of symbiotic fungi enhanced attraction of females to pheromones. Another co-occurring nonbeneficial fungus (Trichoderma sp.) also produced oxygenated monoterpenes, but these were not attractive to I. typographus. Finally, we show that colonization of fungal symbionts on spruce bark diet stimulated beetles to make tunnels into the diet. Collectively, our study suggests that the blends of oxygenated metabolites of conifer monoterpenes produced by fungal symbionts are used by walking bark beetles as attractive or repellent cues to locate breeding or feeding sites containing beneficial microbial symbionts. The oxygenated metabolites may aid beetles in assessing the presence of the fungus, the defense status of the host tree and the density of conspecifics at potential feeding and breeding sites.