Evolution of moth sex pheromone composition by a single amino acid substitution in a fatty acid desaturase.

For sexual communication, moths primarily use blends of fatty acid derivatives containing one or more double bonds in various positions and configurations, called sex pheromones (SPs). To study the molecular basis of novel SP component (SPC) acquisition, we used the tobacco hornworm (Manduca sexta), which uses a blend of mono-, di-, and uncommon triunsaturated fatty acid (3UFA) derivatives as SP. We identified pheromone-biosynthetic fatty acid desaturases (FADs) MsexD3, MsexD5, and MsexD6 abundantly expressed in the M. sexta female pheromone gland. Their functional characterization and in vivo application of FAD substrates indicated that MsexD3 and MsexD5 biosynthesize 3UFAs via E/Z14 desaturation from diunsaturated fatty acids produced by previously characterized Z11-desaturase/conjugase MsexD2. Site-directed mutagenesis of sequentially highly similar MsexD3 and MsexD2 demonstrated that swapping of a single amino acid in the fatty acyl substrate binding tunnel introduces E/Z14-desaturase specificity to mutated MsexD2. Reconstruction of FAD gene phylogeny indicates that MsexD3 was recruited for biosynthesis of 3UFA SPCs in M. sexta lineage via gene duplication and neofunctionalization, whereas MsexD5 representing an alternative 3UFA-producing FAD has been acquired via activation of a presumably inactive ancestral MsexD5. Our results demonstrate that a change as small as a single amino acid substitution in a FAD enzyme might result in the acquisition of new SP compounds.

The role of desaturases in the biosynthesis of marking pheromones in bumblebee males.

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.

Total synthesis, proof of absolute configuration, and biosynthetic origin of stylopsal, the first isolated sex pheromone of strepsiptera.

The asymmetric total synthesis of the diastereomers of stylopsal establishes the absolute configuration of the first reported sex pheromone of the twisted-wing parasite Stylops muelleri as (3R,5R,9R)-trimethyldodecanal. The key steps for the diastereo- and enantiodivergent introduction of the methyl groups are two different types of asymmetric conjugate addition reactions of organocopper reagents to α,ß-unsaturated esters, whereas the dodecanal skeleton is assembled by Wittig reactions. The structure of the natural product was confirmed by chiral gas chromatography (GC) techniques, GC/MS and GC/electroantennography (EAD) as well as field tests. An investigation into the biosynthesis of the pheromone revealed that it is likely to be produced by decarboxylation of a 4,6,10-trimethyltridecanoic acid derivative, which was found in substantial amounts in the fat body of the female, but not in the host bee Andrena vaga. This triple-branched fatty acid precursor thus seems to be biosynthesized de novo through a polyketide pathway with two consecutive propionate-propionate-acetate assemblies to form the complete skeleton. The simplified, motionless and fully host-dependent female exploits a remarkable strategy to maximize its reproductive success by employing a relatively complex and potent sex pheromone.

Stylopsal: the first identified female-produced sex pheromone of strepsiptera.

A female-produced sex pheromone of Stylops muelleri was identified as an unusually branched saturated aldehyde (9R)-3,5-syn-3,5,9-trimethyldodecanal. We named it stylopsal. Its structure was established by using mass spectrometry, infrared spectroscopy, and organic synthesis of candidate compounds. The synthetic standard of (9R)-3,5-syn-3,5,9-trimethyldodecanal gave identical chromatographic and mass spectrometric data as the natural pheromone and also was active in electroantennographic and behavioral assays. The female fat body lipids contained the corresponding fatty acid, indicating a possible link between lipid metabolism and the pheromone biosynthesis.

Total spondylectomy of C2: report of three cases and review of the literature.

STUDY DESIGN: A report on 3 patients undergoing total spondylectomy of the C2 vertebra for tumor and the technique for C1-3 reconstruction. OBJECTIVE: To illustrate the feasibility of complete resection of the C2 vertebra with preservation of the vertebral arteries and cervical nerve roots. BACKGROUND: Total spondylectomy provides improved progression free survival in many patients with locally aggressive spinal tumors. However, the perceived technical demands of effectively preserving both vertebral arteries, maintaining cervical nerve roots, and biomechanical reconstruction of the cranial-cervical junction often dissuades surgeons from carrying out total spondylectomy of the C2 vertebra. METHODS: A review of 3 patients undergoing total C2 spondylectomy for tumor (thyroid adenocarcinoma, chordoma, and solitary plasmocytoma) was done. The surgical procedure that was undertaken and the technique used are described. RESULTS: Postoperatively, all 3 patients had uneventful postoperative recovery with gradual improvement in their neurologic functions. CONCLUSION: Preservation of bilateral vertebral arteries and all cervical nerve roots is feasible when carrying out intralesional total spondylectomy in patients with C2 vertebral body tumors and should be considered in patients thought to benefit from total C2 vertebra excision. In an attempt to augment construct stability and provide anterior column load sharing, we have used mesh cage and iliac crest graft between C1 and C3 held in place with a short cervical plate without complications.

Atlantoaxial fixation using the polyaxial screw-rod system.

The aim of this study is to evaluate the first results of the atlantoaxial fixation using polyaxial screw-rod system. Twenty-eight patients followed-up 12-29 months (average 17.1 months) were included in this study. The average age was 59.5 years (range 23-89 years). The atlantoaxial fusion was employed in 20 patients for an acute injury to the upper cervical spine, in 1 patient with rheumatoid arthritis for atlantoaxial vertical instability, in 1 patient for C1-C2 osteoarthritis, in 2 patients for malunion of the fractured dens. Temporary fixation was applied in two patients for type III displaced fractures of the dens and in two patients for the atlantoaxial rotatory dislocation. Retrospectively, we evaluated operative time, intraoperative bleeding and the interval of X-ray exposure. The resulting condition was subjectively evaluated by patients. We evaluated also the placement, direction and length of the screws. Fusion or stability in the temporary fixation was evaluated on radiographs taken at 3, 6, 12 weeks and 6 and 12 months after the surgery. As concerns complications, intraoperatively we monitored injury of the nerve structures and the vertebral artery. Monitoring of postoperative complications was focused on delayed healing of the wound, breaking or loosening of screws and development of malunion. Operative time ranged from 35 to 155 min, (average 83 min). Intraoperative blood loss ranged from 50 to 1,500 ml (average 540 ml). The image intensifier was used for a period of 24 s to 2 min 36 s (average 1 min 6 s). Within the postoperative evaluation, four patients complained of paresthesia in the region innervated by the greater occipital nerve. A total of 56 screws were inserted into C1, their length ranged from 26 to 34 mm (average, 30.8 mm). All screws were positioned correctly in the C1 lateral mass. Another 56 screws were inserted into C2. Their length ranged from 28 to 36 mm (average 31.4 mm). Three screws were malpositioned: one screw perforated the spinal canal and two screws protruded into the vertebral artery canal. C1-C2 stability was achieved in all patients 12 weeks after the surgery. No clinically manifested injury of the vertebral artery or nerve structures was observed in any of these cases. As for postoperative complications, we recorded wound dehiscence in one patient. The Harms C1-C2 fixation is a very effective method of stabilizing the atlantoaxial complex. The possibility of a temporary fixation without damage to the atlantoaxial joints and of reduction after the screws and rods had been inserted is quite unique.