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There are more than one million described insect species. This species richness is reflected in the diversity of insect metabolic processes. In particular, biosynthesis of secondary metabolites, such as defensive compounds and chemical signals, encompasses an extraordinarily wide range of chemicals that are generally unparalleled among natural products from other organisms. Insect genomes, transcriptomes and proteomes thus offer a valuable resource for discovery of novel enzymes with potential for biotechnological applications. Here, we focus on fatty acid (FA) metabolism-related enzymes, notably the fatty acyl desaturases and fatty acyl reductases involved in the biosynthesis of FA-derived pheromones. Research on insect pheromone-biosynthetic enzymes, which exhibit diverse enzymatic properties, has the potential to broaden the understanding of enzyme specificity determinants and contribute to engineering of enzymes with desired properties for biotechnological production of FA derivatives. Additionally, the application of such pheromone-biosynthetic enzymes represents an environmentally friendly and economic alternative to the chemical synthesis of pheromones that are used in insect pest management strategies.

Males of the closely related species Bombus terrestris and Bombus lucorum attract conspecific females by completely different marking pheromones. MP of B. terrestris and B. lucorum pheromones contain mainly isoprenoid (ISP) compounds and fatty acid derivatives, respectively. Here, we studied the regulation of ISP biosynthesis in both bumblebees. RNA-seq and qRT-PCR analyses indicated that acetoacetyl-CoA thiolase (AACT), 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR), and farnesyl diphosphate synthase (FPPS) transcripts are abundant in the B. terrestris labial gland. Maximal abundance of these transcripts correlated well with AACT enzymatic activity detected in the LG extracts. In contrast, transcript abundances of AACT, HMGR, and FPPS in B. lucorum were low, and AACT activity was not detected in LGs. These results suggest that transcriptional regulation plays a key role in the control of ISP biosynthetic gene expression and ISP pheromone biosynthesis in bumblebee males.

Buff-tailed bumblebees, Bombus terrestris, use a male sex pheromone for premating communication. Its main component is a sesquiterpene, 2,3-dihydrofarnesol. This paper reports the isolation of a thiolase (acetyl-CoA thiolase, AACT_BT), the first enzyme involved in the biosynthetic pathway leading to formation of isoprenoids in the B. terrestris male sex pheromone. Characterisation of AACT_BT might contribute to a better understanding of pheromonogenesis in the labial gland of B. terrestris males. The protein was purified to apparent homogeneity by column chromatography with subsequent stepwise treatment. AACT_BT showed optimum acetyltransferase activity at pH 7.1 and was strongly inhibited by iodoacetamide. The enzyme migrated as a band with an apparent mass of 42.9 kDa on SDS-PAGE. MS analysis of an AACT_BT tryptic digest revealed high homology to representatives of the thiolase family. AACT_BT has 96 % amino acid sequence identity with the previously reported Bombus impatiens thiolase.

Two-dimensional comprehensive gas chromatography (GC×GC) coupled with mass detection was used as a tool for biosynthetic studies of bumblebee pheromones. Prior to biosynthetic experiments, the chromatographic behaviour of isotopically modified esters in the GC×GC system as well as their behaviour in mass detection was studied. The male marking pheromones of Bombus lucorum, Bombus lapidarius and Bombus terrestris were investigated. Main pheromonal components are ethyl tetradec-9-enoate (53 %) and ethyl dodecanoate (6 %) in B. lucorum, hexadec-9-en-1-ol (52 %) and hexadecan-1-ol (31 %) in B. lapidarius, and 2,3-dihydrofarnesol (58 %) and ethyl dodecanoate (15 %) in B. terrestris. The research strategy was based on 1) in vivo incubation of isotopically (2H, 13C) modified fatty acids (FAs) and analysis of their metabolites and 2) feeding experiments with 2H- and 13C-labelled FAs mixed with food. It was observed that labelled FAs were modified into the most abundant aliphatic compounds present in labial gland secretions. In feeding experiments, the labelled FAs were transformed into pheromone components. Transport of the FA precursors from the fat body through haemolymph was confirmed. The results show that FAs, stored in the form of triacylglycerols in the fat body, are likely to participate in the biosynthesis of some aliphatic pheromone components.

Sex-specific chemical secretions have been widely used as diagnostic characters in chemotaxonomy. The taxonomically confused group of bumblebees has reaped the benefit of this approach through the analyses of cephalic labial gland secretions (CLGS). Most of currently available CLGS descriptions concern species from the West-Palearctic region but few from the New World. Here, the CLGS of four East-Palearctic species Bombus deuteronymus, B. filchnerae, B. humilis, and B. exil (subgenus Thoracobombus) are analysed. Our results show high levels of variability in the major compounds in B. exil. In contrast, we describe a low differentiation in CLGS compounds between B. filchnerae and its phylogenetically closely related taxon B. muscorum. Moreover, the chemical profiles of B. filchnerae and B. muscorum are characterized by low concentrations of the C16 component, which is found in higher concentrations in the other Thoracobombus species. This raises the possibility that courtship behavior as well as environmental constraints could affect the role of the bumblebee males' CLGS.

Triacylglycerols (TGs) stored in the fat bodies of bumblebee males have a species-specific composition. The striking structural similarities between TG fatty acids (FAs) and components of the male marking pheromone in certain species led to the hypothesis that FAs may serve as precursors in pheromone biosynthesis. Here, we analysed TGs from B. ruderatus, B. bohemicus, and B. campestris. Nonadec-9-ene and icos-15-en-1-ol are the main components of B. ruderatus labial gland secretion, forming up to 92% of the gland extract. The corresponding icos-11-enic and icos-15-enic acids were found in TGs at levels higher than usual for bumblebee species. We found similar relationships in B. campestris and B. bohemicus. These results suggest that FAs might be precursors of aliphatic compounds in the male pheromones. Furthermore, we report for the first time the pheromone structure of B. ruderatus males.

A serine protease was isolated from midguts of the bumblebee male Bombus terrestris by a combination of precipitation procedures with column chromatography. The purified enzyme exhibited two bands with molecular masses of 25 and 26 kDa as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. These bands showed a proteolytic activity in zymography assay. Midgut enzymes showed optimum proteolytic activity at pH 9 and 35°C using N-succinyl-L-alanyl-L-alanyl-L-prolyl-L-phenyl-alanine 4-nitroanilide as a substrate. The Michaelis constant (Km) and maximum reaction rate (Vmax) were 0.55±0.042 mM and 0.714±0.056 μmol p-nitroalanine produced min(-1) mg protein(-1) , respectively. Inhibition was affected by trypsin inhibitor, but not by phenylmethylsulfonyl fluoride and N-tosyl-L-phenylalanine chloromethyl ketone, which indicated the trypsin-like but not chymotrypsin-like specificity. The identity of the serine protease was confirmed by nanoliquid-tandem mass spectrometry. Eleven unique peptides of the B. terrestris serine protease were found. It shows high homology to a previously reported B. ignitus serine protease covering more than 65% of the protein amino acid sequence.

Instrumental human scent analysis is undoubtedly desirable for many forensic as well medical applications. Most of the previous human scent studies were focused on volatile organic compounds (VOCs) which were analysed by head space solid phase micro-extraction gas chromatography/mass spectrometry (HS-SPME-GC/MS). This method is, however, significantly less sensitive to "heavier" less volatile compounds emitted from the human skin. These less volatile organic scent molecules probably create the basis of the individual human scent signature, and therefore, our attention is focused mainly on these "heavier" compounds. The human scent was adsorbed onto purified glass beads and samples were prepared as hexane solutions obtained by extraction from the sampled glass beads. To resolve a lot of very similar molecules, the comprehensive two-dimensional gas chromatography with time-of-flight mass spectrometer (GCxGC-TOFMS) was used to analyse the hexane scent solutions. Using this technique, more than 137 less volatile molecules including organic fatty acids, ketones, aldehydes, simple esters, alcohols, and especially various fatty acid esters with different carbon chains were identified. A considerable number of these molecules were identified in the scent samples for the first time.

Three kinds of turpentine (a waste by-product of industrial wood processing) of various monoterpene compositions were transformed by Picea abies cells and the product yields monitored in relation to the initial turpentine concentration. The major products obtained were trans-verbenol, trans-pinocarveol, myrtenol, α -terpineol, and p-cymen-8-ol depending on the substrate composition. The absolute quantitative values of the major products were evaluated for a substrate concentration of 0.86 g L 1 . The concentration of trans-verbenol and trans-pinocarveol after twelve days of biotransformation was 768 and 388 mg L 1 , respectively. The substrate was uptaken by Picea abies cells within the fi rst three days and the majority of products released in fi ve days. Although not all the starting material was consumed, the Picea abies suspension culture was able to convert concentrations of turpentine as high as 4.3 g L 1 into valuable products. By precise selection of the substrate concentration and time course, favourable conversion to products could be achieved.

Human cerumen was separated by column chromatography into the following groups of compounds: hydrocarbons, squalene, wax esters and cholesterol esters, triacylglycerols, free fatty acids, free fatty alcohols, monoacylglycerols, free cholesterol, free sterols, and free hydroxy acids. The groups of compounds obtained were examined in detail by gas chromatography and gas chromatography-mass spectrometry. In total, about one thousand compounds have been identified.

Males of Bombus terrestris (L.) adopt a patrolling behaviour during their nuptial parade using cephalic labial gland (CLG) secretions containing sexual pheromones to attract conspecific virgin queens. The changes in chemical composition of their CLG secretions with age are quite well known. In this study, we investigate the evolution of CLG secretions with age in greater detail and compare behavioural reactions of conspecific virgin queens to the secretions. We show that compounds of CLG secretions follow two profiles. Most of the compounds increase from the first day after emergence until the bees are 15-days-old and then decrease. Others are less abundant in 1 to 15-day-old males and then increase (e.g. tricosane, tricosene, henicosane, tetradecanoic acid, pentacosene, pentacosane, heptacosene, heptacosane, nonacosene and geranylcitronellyl tetradecanoate). Differences in secretion composition lead to preferences of virgin queens for males according to the male’s age. Virgin queens prefer the pheromonal gland secretions of bees of the following ages in decreasing order; 1 day = 3 days < 7 days = 30 days < 15 days < 10 days. The virgin queens are strongly attracted by secretions containing high amounts of 2,3-dihydrofarnesol, 2,3-dihydrofarnesal, ethyl dodecanoate and hexadecanol. On the contrary, geranylcitronellol is more abundant in 30-day-old males.

Within the complex network of chemical signals used by termites, trail pheromones and sex pheromones are among the best known. Numerous recent papers map the chemical identity and glandular origin of these pheromones in nearly all major isopteran taxa. In this study, we aimed to describe the sex pheromone and the trail pheromone of a poorly known sand termite, Psammotermes hybostoma. We identified (3Z,6Z,8E)-dodeca-3,6,8-trien-1-ol (dodecatrienol) as the sex pheromone released by tergal and sternal glands of female imagos and, at the same time, as the trail pheromone secreted from the sternal gland of workers. We conclude that chemical communication in Psammotermes does not differ from that of most other Rhinotermitidae, such as Reticulitermes, despite the presence of a diterpene as a major component of the trail pheromone of Prorhinotermes to which Psammotermes is presumed to be phylogenetically close. Our findings underline once again the conservative nature of chemical communication in termites, with dodecatrienol being a frequent component of pheromonal signals in trail following and sex attraction and, at the same time, a tight evolutionary relationship between the trail following of working castes and the sex attraction of imagos.

Two mass spectrometric methods for analysing triacylglycerols (HPLC/APCI-MS and MALDI-MS) were used and compared in terms of the relevance of the data for further biostatistical evaluation. While MALDI-MS is simpler and significantly faster, the time-consuming and labour-intensive HPLC/APCI-MS provides more complete information about the lipid components. However, both methods provide well-comparable results concerning the grouping of specimens belonging to different species when evaluated with multivariate exploratory approaches. The compositions of triacylglycerols in the fat bodies of males in 11 bumblebee species (Bombus terrestris, B. lucorum, B. lapidarius, B. pratorum, B. sylvarum, B. ruderatus, B. pomorum, B. subterraneus, B. campestris, B. bohemicus, and B. rupestris) were found to be species-specific.

A detailed analysis of Rhaponticum carthamoides (Willd.) Iljin root essential oil was carried out by GC, GC–MS and GC–FTIR techniques. In total, 30 components were identified, accounting for 98.0% of total volatiles. A norsesquiterpene 13-norcypera-1(5),11(12)-diene (22.6%), followed by aplotaxene (21.2%) and cyperene (17.9%), were isolated and their structures confirmed by 1D and 2D-NMR spectra (COSY, ROESY, HSQC, HMBC and INADEQUATE). Selinene type sesquiterpenes and aliphatic hydrocarbons were among minor constituents of the essential oil. The oil exhibited antimicrobial activity against 5 of 9 strains of bacteria and yeast, when tested using broth micro-dilution method. Minimum inhibitory concentrations ranged between 32 and 256 μg/ml.

The knowledge of the molecular basis of communication in bumblebee communities is limited. None of the enzymes that participate in pheromone production have been characterized. Here, we cloned the gene encoding the Delta(9) desaturase from cDNA prepared from the total RNA of the pheromone gland and fat bodies of Bombus lucorum male. Functional expression of BlucNPVE desaturase in Saccharomyces cerevisiae and GC-MS analyses revealed its preference for C(18) saturated fatty acids. This suggests that Delta(9) desaturase is involved in the desaturation of metabolic fatty acids stored in triacylglyceroles (TAGs), because oleic acid is the most abundant fatty acid bound in TAG in B. lucorum and it is present in low concentration in the pheromone blend. The incubation of pheromone precursors with a dissected labial gland as well as direct injection of labelled pheromone substrates into B. lucorum males revealed that esterification of pheromone products occurs in the labial gland. These results support both the biosynthesis of pheromones from common lipids and the de novo synthesis of unsaturated pheromones in the labial gland.

Extracts of three different glands (mandibular, labial, and Dufour's) of virgin Bombus terrestris queens at ten different ages (1-8, 12, and 18 days) were analyzed for chemical composition. One hundred and twenty-seven compounds were identified in the extracts. The mandibular and labial glands contained previously reported electroantennogram-active compounds (3-hydroxydecanoic acid, fatty acids of different chain lengths, their esters, and heptacosene). These compounds reached a maximum concentration in 3- to 7-d-old queens. Geranylcitronellol was found in both labial and Dufour's glands. Its amount was inversely correlated to age of queens.

Unusual fatty acids with 24, 26, and 28 carbon atoms were found in triacylglycerols (TAGs) isolated from fat body tissue of bumblebee Bombus pratorum. The most abundant one was (Z,Z)-9,19-hexacosadienoic acid. Its structure was determined by mass spectrometry after derivatization with dimethyl disulfide and by infrared spectroscopy. ECL (equivalent chain length) values of its methyl ester were determined on both DB-1 and DB-WAX capillary columns. (Z,Z)-9,19-Hexacosadienoic acid is quite rare in nature. So far it has been identified only in marine sponges, and this work is the first evidence of its occurrence in a terrestrial organism. HPLC/MS analysis of the bumblebee TAGs showed that (Z,Z)-9,19-hexacosadienoic acid is present in one third of all TAG molecular species. As it was found in all sn-TAG positions, it is likely that (Z,Z)-9,19-hexacosadienoic acid is transported to tissues. Interestingly, labial gland secretion of B. pratorum was found to contain (Z,Z)-7,17-pentacosadiene, a hydrocarbon with markedly similar double bond positions and geometry. Possible biosynthetic relationships between these two compounds are discussed.

In bumblebees, the male secretion of the cephalic labial gland is species-specific. It is highly involved in the nuptial behavior, acting as a sexual attracting pheromone. Therefore, it is also used to accurately identify the species. In contrast to this common scheme, the secretions of Bombus mesomelas are found to be strongly reduced and do not include the most volatile compounds that are present in the secretions of all the other studied bumblebee species. These secretions correspond to cuticular hydrocarbons that can be found in all bumblebee species. This was also the case for another bumblebee species from the same Rhodobombus subgenus: Bombus pomorum. This atypical composition of the male cephalic labial gland secretions seems to indicate that, at least for these two species of Rhodobombus, these secretions are not used to attract virgin females from a long distance, as it is the case for all the other bumblebee species studied.

International audience; The evolution of signals and reproductive traits involved in the pre-mating recognition has been in focus of abundant research in several model species, such as bumblebees (genus Bombus). However, the most-studied bumblebee reproductive trait, the male cephalic labial gland secretions (CLGS), remains unknown among bumblebee species from South America. In this study, the CLGS of five South American bumblebees of the subgenera Thoracobombus (Bombus excellens and B. atratus) and Cullumanobombus (B. rubicundus, B. hortulanus, and B. melaleucus) were investigated, by comparing the chemical compositions of their secretions to those of closely related European species. The results showed an obvious interspecific differentiation in both subgenera. The interspecific differentiation among the species of the Thoracobombus subgenus involved different compounds present at high contents (main compounds), while those of the Cullumanobombus subgenus shared the same main components. This suggests that among the species of the Cullumanobombus subgenus, the differentiation in minor components could lead to species discrimination.

The cephalic labial gland secretion of Bombus semenoviellus males was analyzed, and its chemical composition is reported for the first time. The secretion functions as sex or marking pheromone. Eighty compounds were identified in the secretion, the main one being all-trans-geranylgeranyl acetate (48%). The same compound was shown to form 87% of the labial gland secretion of B. cullumanus males. Both species are closely related and belong to the subgenus Cullumanobombus.

International audience; The diterpene neocembrene A or (1E,5E,9E,12R)-1,5,9-trimethyl-12-(1-methylethenyl)-1,5,9-cyclotetradecatriene, known as the trail-following pheromone of the advanced Termitidae Nasutitermitinae Nasutitermes exitiosus and Trinervitermes bettonianus, has been identified after SPME-GC/MS as the major component of the trail-following pheromone of the Rhinotermitidae Prorhinotermitinae, Prorhinotermes canalifrons and P. simplex. In all the other Rhinotermitidae studied until now, the major component of their trail pheromones is dodecatrienol ((3Z,6Z,8E)-dodeca-3,6,8-trien-1-ol). This biochemical data further add to the anatomical and molecular characteristics that give a special status to the taxon Prorhinotermes among Rhinotermitidae. In Prorhinotermes canalifrons and P. simplex, neocembrene A was the only secretory compound specific to the sternal gland surface that could be detected after SPME. It elicited orientation as well as recruitment behavioral effects. However, the comparison of the respective biological activities triggered by neocembrene A and by sternal gland secretion suggests that minor components of the latter are acting in synergy with neocembrene A.

Labial gland secretions of males of six cuckoo bumblebee species (Bombus vestalis, Bombus rupestris, Bombus campestris, Bombus sylvestris, Bombus bohemicus, and Bombus maxillosus) were analysed and components of their extracts were identified. The specimens of two species collected in the Czech Republic had a significantly different composition of the labial gland secretions than those described earlier in Scandinavia (B. rupestris, B. campestris). Czech populations of three other species studied (B. vestalis, B. sylvestris, and B. bohemicus) produced similar components as those known from the literature. Marking pheromone of B. maxillosus is here reported for the first time. This paper describes detailed analyses of the labial gland secretions including minor and trace components and a determination of the double bond positions in unsaturated compounds.

The biosynthetic pathway of the formation of males'marking pheromones of two bumblebee species, Bombus lucorum and B. lapidarius, was studied for the first time. After applications of deuterium labelled palmitic acid ([2H31]-hexadecanoic acid) into either the head capsules or abdomens the corresponding deuterium labelled pheromone components were detected in the labial gland (LG) extracts of males of both species. In B. lucorum, ethyl [2H29]-hexadec-9-enoate and ethyl [2H31]-hexadecanoate were identified. No deuterium-labelled ethyl tetradecenoate, that would correspond to shortening of the carbon chain, was detected. In B. B. lapidarius, [2H31]-hexadecan-1-ol and [2H29]-hexadec-9-en-1-ol were found in the LG. Furthermore, the deuterium labelled precursor was incorporated into triacylglycerols (TAG) of the fat bodies of males. In vitro incubation of the labial glands of B. lucorum males with deuterium-and 13C-labelled palmitic acid lead to the formation of saturated ethyl esters only.

Enantiomerically pure (S)-(+)-linalool was the main constituent in the extracts of the cephalic secretions of virgin females, mated females, freshly emerged males, and patrolling males of the solitary bee Colletes cunicularius. After copulation, the content of (S)-(+)-linalool emitted by the female was strongly reduced. Electrophysiological experiments revealed that both enantiomers of linalool elicited responses from the antennae of the males. Field tests using the pure enantiomers and the racemate of linalool showed that the number of male bees attracted was highest for (S)-(+)-linalool. The search flight activity in the mating flight area increased dramatically when patrolling males were presented with (S)-(+)-linalool vs (R)-(-)-linalool. Taken together, these data indicate a mate attractant pheromone function of (S)-(+)-linalool.

Insects’ fat bodies are responsible for nutrient storage and for a significant part of intermediary metabolism. Thus, it can be expected that the structure and content of the fat body will adaptively change, if an insect is going through different life stages. Bumblebee queens belong to such insects as they dramatically change their physiology several times over their lives in relation to their solitary overwintering, independent colony foundation stage, and during the colony life-cycle ending in the senescent stage. Here, we report on changes in the ultrastructure and lipid composition of the peripheral fat body of Bombus terrestris queens in relation to seasonal changes in the queens’ activity. Six life stages are defined and evaluated in particular: pharate, callow, before and after hibernation, egg-laying, and senescence. Transmission electron microscopy revealed that the fat body contained two main cell types–adipocytes and oenocytes. Only adipocytes reveal important changes related to the life phase, and mostly the ration between inclusion and cytoplasm volume varies among particular stages. Both electron microscopy and chemical analyses of lipids highlighted seasonal variability in the quantity of the stored lipids, which peaked prior to hibernation. Triacylglycerols appeared to be the main energy source during hibernation, while the amount of glycogen before and after hibernation remained unchanged. In addition, we observed that the representation of some fatty acids within the triacylglycerols change during the queen’s life. Last but not least, we show that fat body cell membranes do not undergo substantial changes concerning phospholipid composition in relation to overwintering. This finding supports the hypothesis that the cold-adaptation strategy of bumblebee queens is more likely to be based on polyol accumulation than on the restructuring of lipid membranes.

The compositions of 23 monoterpene hydrocarbons of six pine species (Pinus sylvestris, P. yunnanensis, P. armandii, P. tropicalis, P. cubensis and P. caribaea) were compared, using multivariate data analysis. Four of the six species were clearly different from the other species in a Principal Components Analysis (PCA) model, based on the relative amounts (selective normalization) of the monoterpenes. The correlation coefficients between constituents were determined separately for each species and the strongest correlations were found between (+)-α-pinene and (+)-camphene and between the corresponding (—)-enantiomers, in all species. This pattern, i.e. a good correlation in all species, was neither shown by the correlation of the structurally more similar (+)-α/β-pinenes, nor by the (—)-α/β-pinenes or within the enantiomeric pairs of α-pinene and β-pinene. For these pairs of monoterpenes, good correlations were found in some species. None of the species showed good correlations in all the investigated monoterpene pairs presented here. Correlations between monoterpenes in insect-attacked trees (P. cubensis and P. caribaea, attacked by Dioryctria horneana, and P. yunnanensis, attacked by Tomicus piniperda) were also determined. The results are discussed from chemotaxonomic and biosynthetic points of view.

The release of volatile compounds from a flower of Hydrosme rivieri was recorded during the whole flowering period (7 days). The quantities of six odour components (dimethyl disulphide, dimethyl trisulphide, n -alkanes C 10 , C 11 , C 12 and C 13 ) forming the main part of emanating volatiles were plotted versus time. n -Alkanes started to emanate 3 days before the release of dimethyl disulphide and dimethyl trisulphide (the components with a rotting meat odour).

(S)-(−)- and (R)-(+)-limonene was transformed to carvone via corresponding cis- and trans-carveol using Solanum aviculare and Dioscorea deltoidea plant cells. Both carveols and carvone formed were racemic in all biotransformations.

Premating behavior in the species Bombus confusus Schenck, 1859 was studied. The marking habits of bumblebee males were revealed and the use of their labial gland secretion in scent marks was proved chemically. Identical compounds were present both in the labial gland secretion and on male-marked objects. This chemical proof is reported for the first time from natural conditions of a B. confusus locality. Males' flight activities, site tenacity, and male–male interactions are described in detail. Contrary to literature reports, male B. confusus behavior does not basically differ from that of already known perching bumblebee species.

Sixtee n belonging to the family Apiaceae were examined for their acceptance as host plants by Trioza apicalis in non-choice tests. The number of eggs laid per day varied from 18 on Daucus carota subsp, sativus to zero on Aegopodium podagraria. In a comparative test, the psyllids landed significantly faster and started egg-laying earlier on carrots (D. carota subsp, sativus) than on dill (Anethum graveolens). In a study of shelter plants for T. apicalis, the psyllids survived on spruce, Picea abies, for more than 30weeks, but they died on grass, Phleum pratense and Brachypodium sylvaticum, within 2 weeks. The length of the day is one of the factors regulating hibernation. At 20°C, light regimes shorter than 17 h induced hibernation, while longer light periods led to reproduction. Both the shelter plants and the summer hosts, including the Apiaceae species, released large amounts of monoterpene hydrocarbons, with great differences among the species. The most preferred host plants (carrots) were found to contain (+)- and (-)-c~-pinene and (+)-sabinene, while large amounts ( > 20%) of either (+) - or ( - ) -limonene were released by the species of low preference. ( - ).-c~- Pinene, ( - ) -J3-pinene, and (-) -limonene were the main components of the spruce odour. A multivariate data analysis of the chemical results showed clearly the significant similarities of the summer host and shelter plants (Daucus carota, Picea abies, and Juniperus communis). © 1997 Elsevier Science Ltd

BACKGROUND: In addition to their general role in the hydrolysis of storage lipids, bumblebee lipases can participate in the biosynthesis of fatty acids that serve as precursors of pheromones used for sexual communication. RESULTS: We studied the temporal dynamics of lipolytic activity in crude extracts from the cephalic part of Bombus terrestris labial glands. Extracts from 3-day-old males displayed the highest lipolytic activity. The highest lipase gene expression level was observed in freshly emerged bumblebees, and both gene expression and lipase activity were lower in bumblebees older than 3 days. Lipase was purified from labial glands, further characterized and named as BT-1. The B. terrestris orthologue shares 88% sequence identity with B. impatiens lipase HA. The molecular weight of B. terrestris lipase BT-1 was approximately 30 kDa, the pH optimum was 8.3, and the temperature optimum was 50°C. Lipase BT-1 showed a notable preference for C8-C10 p-nitrophenyl esters, with the highest activity toward p-nitrophenyl caprylate (C8). The Michaelis constant (Km) and maximum reaction rate (Vmax) for p-nitrophenyl laurate hydrolysis were Km = 0.0011 mM and Vmax = 0.15 U/mg. CONCLUSION: This is the first report describing neutral lipase from the labial gland of B. terrestris. Our findings help increase understanding of its possible function in the labial gland.

Bumblebee males (Hymenoptera) produce species-specific labial gland secretions called marking pheromones (MPs). MPs generally consist of terpenoids and fatty-acid-derived aliphatic compounds with various chain lengths predominantly containing one or no double bonds. The unsaturated fatty-acid-derived MP components were hypothesized to be produced by fatty acid desaturases (FADs) that exhibit diverse substrate specificities. To address this hypothesis, we isolated and functionally characterized FADs from three bumblebee species: Bombus lucorum, Bombus terrestris, and Bombus lapidarius. By employing RNA sequencing of the male labial glands and fat bodies of B. lucorum and B. terrestris, we identified five paralogous FAD-like sequences but only two FAD lineages were abundant and differentially expressed in the labial glands. We found that abundant FAD lineages were also expressed in the labial gland and fat body of Bombus lapidarius. Functional characterization of FADs in a yeast expression system confirmed that Δ4-FADs exhibited a unique Δ4-desaturase activity exclusively on 14-carbon fatty acyls and Δ9-FADs displayed Δ9-desaturase activity on 14- to 18-carbon fatty acyls. These results indicate that Δ9-FADs are involved in the biosynthesis of major unsaturated components of MPs in B. lucorum and B. lapidarius despite the diverse MP composition of these bumblebee species. The contribution of lipases, acyltransferases, esterases, and fatty acid reductases to production of the species-specific MP composition is also discussed in light of the transcriptomic data obtained in this study.

The floral fragrance of the shrub Daphne mezereum in central Sweden was collected by means of the head-space technique and investigated by GC-MS and multi-dimensional GC. ( S )-(+)-Linalool was the main constituent (95%) of the flower fragrance and its enantiomeric purity exceeded 99% in the samples. The (2 S , 5 S )- and (2 R , 5 S )-furanoid and the (3 R , 6 S )- and (3 S , 6 S )-pyranoid linalool oxide isomers constituted 2–5% of the fragrance. The elution order of these compounds on a permethylated β-cyclodextrin column is reported. A fragrance sample of D. mezereum as well as ( S )-(+)-linalool attracted males of the vernal solitary bee species Colletes cunicularius and Andrena cinerea . A racemic mixture of the two enantiomeric pairs of known furanoid linalool oxides was only weakly attractive to the bees. The role of the fragrance in the pollination specialization of the plant is discussed.

Labial glands of the cuckoo-bumblebee males of the species Psithyrus vestalis were extracted and the components of their secretions were identified. Chemical composition of the males' signal of Psithyrus vestalis has not yet been described in the literature. We found geranycitronellyl acetate to be the main component (48%). Geranylcitronellol and (Z)-15-eicosen-1-ol were present in the secretion in lower amounts. Long-chain aldehydes, acetates, and hydrocarbons formed only minor components of the mixture. The identification of minor components was based on GC-MS, while the more abundant compounds were isolated and fully characterized by spectral and chemical methods. For the structure confirmation, geranylcitronellol was prepared from geranyl bromide via a three-step synthesis.

Populations on islands often exhibit lower levels of genetic variation and ecomorphological divergence compared to their mainland relatives. While phenotypic differentiation in characters, such as size or shape among insular organisms, has been well studied, insular differentiation in quantitative reproductive traits involved in chemical communication has received very little attention to date. Here, we investigated the impact of insularity on two syntopic bumblebee species pairs: one including species that are phylogenetically related (Bombus terrestris and B. lucorum), and the other including species that interact ecologically (B. terrestris and its specific nest inquiline B. vestalis). For each bumblebee species, we characterized the patterns of variation and differentiation of insular (Corsican) vs. mainland (European) populations (i) with four genes (nuclear and mitochondrial, 3781 bp) and (ii) in the chemical composition of male marking secretions (MMS), a key trait for mate attraction in bumblebees, by gas chromatography-mass spectrometry (GC-MS). Our results provide evidence for genetic differentiation in Corsican bumblebees and show that, contrary to theoretical expectations, island populations of bumblebees exhibit levels of genetic variation similar to the mainland populations. Likewise, our comparative chemical analyses of MMS indicate that Corsican populations of bumblebees are significantly differentiated from the mainland yet they hold comparative levels of within-population MMS variability compared to the mainland. Therefore, insularity has led Corsican populations to diverge both genetically and chemically from their mainland relatives, presumably through genetic drift, but without a decrease of genetic diversity in island populations. We hypothesize that MMS divergence in Corsican bumblebees was driven by a persistent lack of gene flow with mainland populations and reinforced by the preference of Corsican females for sympatric (Corsican) MMS. The impoverished Corsican bumblebee fauna has not led to relaxation of stabilizing selection on MMS but to consistent differentiation chemical reproductive traits on the island. © 2013 Lecocq et al., SCOPUS: ar.j, info:eu-repo/semantics/published

Monoterpene hydrocarbons from the xylem oleoresins of Pinus tropicalis, P. caribaea var. caribaea, P. cubensis and P. maestrensis growing in Cuba were investigated. A large number of oleoresin samples from trees, attacked and unattacked by the moth Dioryctria horneana , as well as from trees growing in different localities, were analyzed. In each sample, the relative amounts of the volatiles and the enantiomeric compositions of six chiral monoterpene hydrocarbons were determined using a two-dimensional gas chromatographic (2D-GC) system. The GC-data were evaluated using two methods for multivariate data analysis, PCA (principal components analysis) and PLS-DA (projections to latent structures-discriminant analysis). PC-plots based on the relative amounts of the monoterpenes present in the volatile oleoresin fractions analyzed, showed that P. tropicalis and P. cubensis formed one group, separated from P. caribaea , while pines belonging to the species P. maestrensis were found in both groups. The species P. tropicalis and P. maestrensis , both of which are resistant to D. horneana , and the subgroups within the species P. caribaea and P. cubensis , that had not been attacked by D. horneana , showed no signs of constituting a chemical subgroup. No significant difference was found between the attacked and the unattacked P. cubensis trees. In P. caribaea , on the other hand, the composition of volatiles in attacked trees was significantly different from that in unattacked trees. However, an influence of monoterpenes on the oviposition of D. horneana was not proved.

De novo biosynthesis of male sex pheromone from two bumblebee species (Bombus terrestris and Bombus lucorum) was studied by using in vitro incubations of labial glands (LGs) with radioactive [1,2-(14)C]acetate and deuterated [D(3)]acetate. The labeled substrate was incorporated into several types of compounds, such as terpenic alcohols, fatty acids, esters, and hydrocarbons. A similar incubation of [1,2-(14)C]acetate with fat bodies (FB) led to the formation of fatty acids, triacylglycerols (TAG), and hydrocarbons. To support the results from in vitro incubations, PCR analysis of fatty acid synthase (FAS) transcripts in LG and FB was performed. Relative quantification of FAS transcription levels revealed that the abundance of mRNA from the FAS gene is a function of the age of B. terrestris males. A comparison of the relative FAS mRNA gene transcription level in FB and LGs of B. terrestris and B. lucorum males proved that high biosynthetic activity takes place in the LGs of both species. Together, these results indicate that pheromone components are synthesized de novo in the LG.

In this study, we analyzed the chemical composition of volatile oils hydrodistilled from seeds of Consolida regalis, Delphinium elatum, Nigella hispanica, and N. nigellastrum using GC and GC/MS. In C. regalis, octadecenoic (77.79%) and hexadecanoic acid (8.34%) were the main constituents. Similarly, the oils from D. elatum and N. hispanica seeds consisted chiefly of octadecadienoic (42.83 and 35.58%, resp.), hexadecanoic (23.87 and 28.59%, resp.), and octadecenoic acid (21.67 and 19.76%, resp.). Contrastingly, the monoterpene hydrocarbons α-pinene (34.67%) and β-pinene (36.42%) were the main components of N. nigellastrum essential oil. Our results confirm the presence of essential oils in the family Ranunculaceae and suggest chemotaxonomical relationships within the representatives of the genera Consolida, Delphinium, and Nigella. In addition, the presence of various bioactive constituents such as linoleic acid, (−)-β-pinene, squalene, or carotol in seeds of D. elatum, N. hispanica, and N. nigellastrum indicates a possible industrial use of these plants.

The influence of different immobilization methods on the biotransformation of verbenol bySolanum aviculare plant cells has been studied. The biotransformation course was compared using free and immobilized cells. Immobilization techniques used included entrapment in alginate, pectate and carrageenane gels, in polyurethane foam, and on the surface of polyphenylenoxide.

A great diversity of defensive chemicals has been described in termite soldiers equipped with a unique defensive organ, the frontal gland. Along with the functional diversity of these compounds, reflecting the evolutionary history of particular lineages and their defensive strategies, a considerable degree of chemical variability often occurs among species and populations. Thus, the chemistry of termite defense may provide information on the phylogeny and geographic dispersal of species and populations. In this paper, we report on the anatomy of the frontal gland and on the diversity of soldier defensive chemicals in the sand termite, Psammotermes hybostoma, from nine colonies and five different localities in Egypt. Using gas chromatography–mass spectrometry, a total of 30 sesquiterpene hydrocarbons, or their oxygenated derivatives, were detected, and the chemical identity of most of them identified. In addition, a ketone, an ester, and a diterpene were identified in some colonies. Within colonies, the chemical composition was stable and did not differ among soldier size categories. However, there were pronounced quantitative and qualitative differences in frontal gland chemicals among colonies and geographic locations. The findings are discussed in a broader comparison with other termite taxa.

Volatile compounds from flowers, buds and leaves of Aegopodium podagraria, Anthriscus sylvestris, Carum carvi, Heracleum sibiricum, Laserpitium latifolium and Pastinaca sativa were collected on PorapakQ and TenaxGC and identified by GC-MS. The enantiomeric compositions of the main chiral monoterpenes were determined by multidimensional gas chromatography. The characteristics of the floral fragrances were found to originate in differences in the proportions of the monoterpene hydrocarbons, among which α-pinene, β-pinene, cis-β-ocimene, trans-β-ocimene, limonene, sabinene and myrcene dominated. Heracleum sibiricum and P. sativa had very similar fragrances. Pure (+)-3-carene was found in H. sibiricum and C. carvi, and (-)-sabinene was the main compound in L. latifolium and A. sylvestris. Nitrogenous compounds, some aliphatic short-chain esters and linalool were found exclusively in the flower scents. The possible biological functions of the volatile compounds are discussed.

Frontal gland secretions of termite soldiers of the speciesNasutitermes costalis, N. ephratae, N. nigriceps, N. rippertii, andN. hubbardii were analyzed. The chiral compositions of the monoterpene hydrocarbons were investigated by means of two-dimensional gas chromatography. Two of them, 3-carene andβ-pinene were present in the form of (−)-enantiomers of high purity in the majority of the samples. The other monoterpenes,α-pinene, camphene, sabinene, limonene, andβ-phellandrene, exhibited a great variation in the enantiopurity between different species.α-Pinene and limonene varied significantly even between different colonies within the species.N. costalis was the only species with an invariable composition of the secretion, independent of the locality or origin.

The age-dependent changes in the composition of triacylglycerols (TAG) in the fat bodies of bumblebee males were studied using HPLC/MS. Two related species (Bombus terrestris and B. lucorum) were compared, with the age of the males being 0–30 days. The total amount of TAG in B. lucorum was about 2.7 times higher than that in B. terrestris for all of the ages studied. One to three-day-old males had the highest content of TAG in their fat bodies (1.6–2.3 mg/individual in B. terrestris and 3.8–4.2 mg/individual in B. lucorum). The analytical data show different patterns in both species. The qualitative composition of fatty acids in TAG was similar, but the mean relative abundance between B. terrestris and B. lucorum differed: 14:0, 7 and 14%; 16:0, 20 and 44%; 18:3, 62 and 23%; 18:1, 3 and 8%, respectively (the data is based on a GC/MS integration). A statistical evaluation of the dynamic changes in the TAG composition revealed that in B. terrestris different age classes were well separated according to their TAG composition while in B. lucorum the TAG did not change substantially during the male’s life. The TAG analyses provide more precise information on the differences between the classes studied than the FA composition alone.

Labial glands of the cuckoo-bumblebee males of the species Psithyrus vestalis were extracted and the components of their secretions were identified. Chemical composition of the males' signal of Psithyrus vestalis has not yet been described in the literature. We found geranycitronellyl acetate to be the main component (48%). Geranylcitronellol and ( Z )-15-eicosen-1-ol were present in the secretion in lower amounts. Long-chain aldehydes, acetates, and hydrocarbons formed only minor components of the mixture. The identification of minor components was based on GC-MS, while the more abundant compounds were isolated and fully characterized by spectral and chemical methods. For the structure confirmation, geranylcitronellol was prepared from geranyl bromide via a three-step synthesis.

Quantitative real-time polymerase chain reaction (PCR) is an accurate and sensitive technique for gene expression analysis. However, it requires data normalization using reference genes. Here we assessed the stability of eight reference genes in the labial gland and fat body of the bumblebees Bombus terrestris and Bombus lucorum of different ages. To date, no reference genes have been identified for these species. Our data show that arginine kinase (AK) and phospholipase A2 (PLA2) are the most stable genes in both tissues of B. terrestris. The most stable genes for the labial gland and fat body of B. lucorum were found to be elongation factor 1alpha (EEF1A) and PLA2.

Age-related changes of antennal-active components of male labial gland extracts were studied in two closely related bumblebee species, Bombus terrestris and B. lucorum. In B. terrestris, compounds eliciting electroantennogram (EAG) responses of virgin queens were ethyl dodecanoate, 2,3-dihydrofarnesal, 2,3-dihydrofarnesol, hexadecan-1-ol, octadeca-9,12,15-trien-1-ol, and geranylcitronellol. Compounds that elicited EAG responses from queens of B. lucorum were ethyl dodecanoate, ethyl tetradec-7-enoate, ethyl tetradec-9-enoate, ethyl hexadec-9-enoate, hexadecan-1-ol, hexadec-7-enal, octadeca-9,12-dien-1-ol, octadeca-9,12,15-trien-1-ol, and octadecan-1-ol. Quantities of these compounds in the labial glands changed significantly over the lifetime of the respective males of the two species. In both species, concentrations of the respective compounds reached their maximum within seven days after eclosion. Subsequently, a rapid decrease in the amount of EAG-active compounds occurred in B. terrestris, whereas in B. lucorum the amount of active compounds stayed approximately constant or decreased at a slow rate. Microscopy showed that in B. terrestris secretory cells of the labial glands undergo apoptosis from the fifth to the tenth day of life, whilst in B. lucorum labial gland cells remain unchanged throughout the life of the males.

Analytical determination by HPLC of five taxanes—paclitaxel, cephalomannine, 10-deacetylpaclitaxel, 10-deacetylcephalomannine and taxcultine—in extracts from the bark of Taxus baccata indicates that taxane concentrations vary seasonally. The highest concentration was found in October and the lowest in January for all taxanes examined. Copyright © 1999 John Wiley & Sons, Ltd.

b Variations of secretions of the cephalic part of the labial glands from four different subspecies of Bombus terrestris, B. t. terrestris, B. t. lusitanicus, B. t. sassaricus, and B. t. dalmatinus, were investigated. 95 compounds were detected in the whole data set: 54 in B. t. terrestris ,5 4 inB. t. lusitanicus, 48 in B. t. sassaricus, and 44 in B. t. dalmatinus. The (E)-2,3-dihydrofarnesol is the main compound in B. t. dalmatinus and B. t. sassaricus, while it is dihydrofarnesyl dodecanoate in B. t. terrestris and B. t. lusitanicus. A principal component analysis produced a pattern showing three well distinct groups corresponding to dalmatinus, sassaricus, and terrestris þ lusitanicus.

Volatiles released by seedlings of Norway spruce infested with the aphid Cinara pilicornis were analyzed using SPME-GC-MS. Among the stress-induced compounds released by the host plant, citronellol, cis-trans-nepetalactone and cis-trans-nepetalactol was found. These compounds originated from the aphids and they were assumed to be pheromone components for this aphid species. To determine the relative stereochemistry of the nepetalactone, a diagnostic method was developed. The method was based on multivariate analysis of tabulated relative intensities of mass fragments of the four nepetalactone diastereomers. In the practical method described, a few pairs of fragments in the mass spectra were compared and, in combination with the Kovat's index, were used to unambiguously identify the relative stereochemistry of the nepetalactone.