Exploring complex pheromone biosynthetic processes in the bumblebee male labial gland by RNA sequencing.

Male marking pheromones (MPs) are used by the majority of bumblebee species (Hymenoptera: Apidae), including a commercially important greenhouse pollinator, the buff-tailed bumblebee (Bombus terrestris), to attract conspecific females. MP biosynthetic processes in the cephalic part of the bumblebee male labial gland (LG) are of extraordinary complexity, involving enzymes of fatty acid and isoprenoid biosynthesis, which jointly produce more than 50 compounds. We employed a differential transcriptomic approach to identify candidate genes involved in MP biosynthesis by sequencing Bombus terrestris LG and fat body (FB) transcriptomes. We identified 12 454 abundantly expressed gene products (reads per kilobase of exon model per million mapped reads value > 1) that had significant hits in the GenBank nonredundant database. Of these, 876 were upregulated in the LG (> 4-fold difference). We identified more than 140 candidate genes potentially involved in MP biosynthesis, including esterases, fatty acid reductases, lipases, enzymes involved in limited fatty acid chain shortening, neuropeptide receptors and enzymes involved in biosynthesis of triacylglycerols, isoprenoids and fatty acids. For selected candidates, we confirmed their abundant expression in LG using quantitative real-time reverse transcription-PCR (qRT-PCR). Our study shows that the Bombus terrestris LG transcriptome reflects both fatty acid and isoprenoid MP biosynthetic processes and identifies rational gene targets for future studies to disentangle the molecular basis of MP biosynthesis. Additionally, LG and FB transcriptomes enrich the available transcriptomic resources for Bombus terrestris.

Gene expression responses in larvae of the fleshfly Sarcophaga bullata after immune stimulation.

Insect larvae develop in decaying organic matter and their defence against various microorganisms must therefore be highly efficient. In the present study, we explored the transcriptional kinetics and induction levels of eight genes in Sarcophaga bullata larvae after infection or aseptic injury. Using real-time PCR, we studied the time-dependent immune response of larvae of the fleshfly S. bullata. We compared the mRNA levels of eight selected genes in induced and non-induced larvae. The third-instar larvae of S. bullata were induced by injecting a bacterial suspension of Escherichia coli, Staphylococcus aureus or Pseudomonas aeruginosa, or by simple aseptic injury with an entomological pin. We used intact larvae as a control for basal mRNA expression. Total RNA was isolated from the whole body, fat body and haemocytes. We determined the mRNA levels of genes encoding sapecin, transferrin, prophenoloxidase 1 and 2, storage-binding protein, cathe psin L, sarcocystatin, and 26/29 kDa protease. We found that there was massive up-regulation of genes encoding the fleshfly peptide sapecin, as well as the protein transferrin. We also detected down-regulation of, or no change in, the expression of genes that encode prophenoloxidase 1 and 2, storage-binding protein, cathepsin L, sarcocystatin, and 26/29 kDa protease.

Composition of the labial gland secretion of the bumblebee males Bombus pomorum.

Labial gland secretions of 22 males of the bumblebee Bombus pomorum, collected in the Czech Republic, were analysed separately for each individual. The secretions contained 70 compounds among which saturated and unsaturated hydrocarbons strongly dominated. The proportion of hydrocarbons in the secretion was unusually high (85-100%) compared to other bumblebee species studied so far (3-15%). Methyl and ethyl esters of fatty acids, known from many other bumblebee species, formed only minor components (less than 1% in sum) of the secretions of several B. pomorum individuals. No terpenic compounds, typical for males' marking secretion of many bumblebee species, were detected in B. pomorum. The absolute quantities of hydrocarbons present in the labial gland extracts were comparable with those usually present in other species. The composition of hydrocarbons found in the labial glands was different from the profile of the cuticular hydrocarbons. Despite our expectations in species exhibiting a regular patrolling and scent-marking behaviour, the labial gland extracts obtained from B. pomorum males were unusually low concentrated and their chemical composition was atypical with respect of the proportions of hydrocarbons when compared with other patrolling species. This is the first report on the analysis of the labial gland secretion of the B. pomorum males.

Sex pheromone of horse-chestnut leafminer Camneraria ohridella and its use in a pheromone-based monitoring system.

Gas chromatography combined with electroantennographic detection (GC-EAD), electroantennography (EAG), and wind-tunnel and field experiments were used to reinvestigate the composition of Cameraria ohridella (Lepidoptera, Gracillariidae, Lithocolletinae) sex pheromone. The GC-EAD experiments showed one EAD-active area corresponding to the major pheromone component. (8E,10Z)-tetradeca-8,10-dienal. The EAG experiments proved that (9E)-tetracedecenal and stereoisomers of (8E,10Z)-tetradeca-8,10-dienal exhibited significant electrophysiological activity and could, therefore, be considered as possible minor pheromone components. However, wind-tunnel and field experiments demonstrated that none of these compounds affect the efficacy of the main pheromone component. A monitoring system based on (8E,10Z)tetradeca-8,10-dienal was developed and used to study the flight activity of C. ohridella.