Aleksandra Urbanek

Composition and antimicrobial activity of fatty acids detected in the hygroscopic secretion collected from the secretory setae of larvae of the biting midge Forcipomyia nigra (Diptera: Ceratopogonidae)

The hygroscopic secretion produced by the secretory setae of terrestrial larvae of the biting midge Forcipomyia nigra (Winnertz) was analysed using gas chromatography coupled with mass spectrometry (GC-MS). The viscous secretion is stored at the top of each seta and absorbs water from moist air. GC-MS analyses (four independent tests) showed that the secretion contained 12 free fatty acids, the most abundant of which were oleic (18:1), palmitic (16:0), palmitoleic (16:1) and linoleic (18:2). Other acids identified were valeric (5:0), enanthic (7:0), caprylic (8:0), pelargonic (9:0), capric (10:0), lauric (12:0), myristic (14:0) and stearic (18:0). Two other compounds, glycerol and pyroglutamic acid, were also found. The antibacterial activity of the fatty acids and pyroglutamic acid was tested using the agar disc diffusion method and targeted Gram positive (Bacillus cereus, Bacillus subtilis, Enterococcus faecalis) and Gram negative bacterial strains (Citrobacter freundii, Pseudomonas aeruginosa, Pseudomonas fluorescens). The antifungal activity was tested by determining minimal inhibitory concentration (MIC) of examined compounds. Fatty acids were tested against enthomopathogenic fungi (Paecilomyces lilacinus, Paecilomyces fumosoroseus, Lecanicillium lecanii, Metarhizium anisopliae, Beauveria bassiana (Tve-N39), Beauveria bassiana (Dv-1/07)). The most effective acids against bacterial and fungal growth were C(9:0), C(10:0) and C(16:1), whereas C(14:0), C(16:0,) C(18:0) and C(18:1) demonstrated rather poor antifungal activity and did not inhibit the growth of bacteria. The antimicrobial assay investigated mixtures of fatty and pyroglutamic acids (corresponding to the results of each GC-MS test): they were found to be active against almost all the bacteria except P. fluorescens and also demonstrated certain fungistatic activity against enthomopathogenic fungi. The hygroscopic secretion facilitates cuticular respiration and plays an important role in the antimicrobial protection of F. nigra larvae living in moist terrestrial habitats.

The antifungal activity of fatty acids of all stages of Sarcophaga carnaria L. (Diptera: Sarcophagidae).

Fatty acids as components of cuticular lipids of insects play a significant role in antifungal in protection against fungal infection. The chemical composition of cuticular and internal extracts obtained from all developmental stages of flesh flies Sarcophaga carnaria was identified. The fatty acids were detected using gas chromatography coupled with mass spectrometry and the most abundant for all examined stages were: 18:1 > 16:0 > 16:1 > 18:0 > 18:2. Polyunsaturated fatty acids (PUFA) C20 were found in both, cuticular and internal extracts. GC-MS analysis showed higher relative content of PUFA in adults than in preimaginal stages. Fatty acids alone as well as their cuticular and internal extracts obtained from larvae, pupae male and female of S. carnaria were tested according to their potential antimicrobial activity against entomopathogenic fungi: Paecilomyces lilacinus, Paecilomyces fumosoroseus, Lecanicillium lecanii, Metarhizium anisopliae, Beauveria bassiana (Tve-N39) and B. bassiana (Dv-1/07). FA presented diverse antimicrobial activity depending on the length of the chain and the presence of unsaturated bonds. Short chain and unsaturated FA (6:0, 11:0, 13:0) have shown significantly stronger activity against fungi but they were detected in lower concentrations. PUFA inhibit fungal growth more effectively than unsaturated long chain fatty acids. Cuticular and internal extracts of all living forms of S. carnaria exhibited approximately equal activity against tested entomopathogenic fungi. We presumed that the most abundant saturated long chain FA and additionally PUFA founded in our analysis are involved in protecting the flies against fungal infection.

Antimicrobial activity of untypical lipid compounds in the cuticular and internal lipids of four fly species.

This article describes the qualitative and quantitative analyses of untypical compounds in the cuticular and internal lipids of four dipteran species. For isolated compounds, antimicrobial activity against 18 reference strains of bacteria and fungi was determined.

Sensilla coeloconica ringed by microtrichia in host-seeking biting midges.

The distribution and morphology of antennal sensilla coeloconica in parasitic and predaceous biting midges were studied in females of Forcipomyia (feeding on the blood of frogs), Atrichopogon (feeding on haemolymph), Austroconops, Culicoides (feeding on the blood of birds and mammals) and Brachypogon (feeding on haemolymph and dissolved tissues of insects) (all: Diptera: Ceratopogonidae). A Lower Cretaceous female of Archiculicoides (Diptera: Ceratopogonidae) from Lebanese amber, which fed on the blood of unknown vertebrates, was also examined. In sensilla coeloconica ringed by microtrichia, the peg is grooved longitudinally and protrudes distinctly from the pit. We suggest that the microtrichia encircling the protruding peg form a structure resembling a picket fence in order to maintain a higher level of humidity, which facilitates the capture and transport of odour molecules through the channels in the peg wall. Sensilla coeloconica ringed by microtrichia function as very effective chemoreceptors in host‐ and prey‐seeking activity. During the evolution of Ceratopogonidae, sensilla coeloconica with a fence of microtrichia have evolved twice in groups feeding on the blood of vertebrates (i.e. in the basal lineage: Lower Cretaceous or earlier) and in the subgenus Lasiohelea of Forcipomyia (Palaeogene). Sensilla coeloconica ringed by microtrichia are described for the first time in the relict genus Austroconops.

Non-biting midges of the tribe Tanytarsini in Eocene amber from the Rovno region (Ukraine): a pioneer systematic study with notes on the phylogeny (Diptera: Chironomidae)

The first detailed systematic study on the tanytarsine chironomids recorded in the Eocene amber from the Rovno region (Ukraine) revealed seven new taxa. Archistempellina gen. nov., represented by A. bifurca sp. nov. (type for the genus, male) and A. falcifera sp. nov. (male), displays characters similar to those found in Stempellina Thienemann et Bause, and is recognized as one of probable basal lineages in the subtribe Stempellinina Shilova. Corneliola gen. nov., represented by C. avia sp. nov. (male, female), combines features known from several genera of the tribe Tanytarsini, and tentatively is regarded as the closest relative of Constempellina Brundin. Rheotanytarsus alliciens sp. nov. (male), featuring a long hypopygial digitus, is considered to be the oldest species of the genus. Tanytarsus congregabilis sp. nov. (male) is the first known Eocene representative of the lugens systematic species group. A complemented description of the male of Tanytarsus serafini Giłka is provided as well.

Morphology and histology of secretory setae in terrestrial larvae of biting midges of the genus Forcipomyia (Diptera: Ceratopogonidae).

Apneustic larvae of the genus Forcipomyia possess unique secretory setae located on the dorsal surface along the body in two rows, one pair on each thoracic and abdominal segment and two pairs on the head. Morphological and histological studies of secretory setae in fourth instar larvae of Forcipomyia nigra (Winnertz) and Forcipomyia nigrans Remm indicate they are modified mechanoreceptors (sensilla trichodea) in which the trichogen cell is a glandular cell producing a hygroscopic secretion. The cytoplasm of the glandular trichogen cell fills the lumen of a secretory seta, which shows one or more pores on the apex. The cytoplasm contains numerous microtubules responsible for transportation of proteinaceous vesicles, and an extremely large polyploid nucleus typical of gland cells. The main role of the hygroscopic secretion is to moist the body and thus facilitate cuticular respiration.

Identification and antifungal activity of novel organic compounds found in cuticular and internal lipids of medically important flies.

Novel organic compounds found in the cuticular and internal lipids of medically important flies were identified. Uracil, 9-tricosene, 1-oleoyl glycerol, dimethyl suberate and butyl stearate were tested for their potential antifungal activity. Minimal inhibitory concentrations of the compounds against reference strains of fungi were determined. Uracil and dimethyl suberate slightly inhibited the growth of entomopathogenic fungi. The cuticular and internal lipids of Calliphora vicina, Calliphora vomitoria, Sarcophaga carnaria and Musca domestica were studied by gas chromatography (GC) combined with mass spectrometry (GC/MS). A comparison of the lipid extracts between the preimaginal and mature stages showed adults flies contained a higher total content of the identified components. Furthermore, their amounts distinctly predominated in the internal lipids of all the species. The amount of 9-tricosene was the highest in adults of C. vicina, while the larvae and pupae had a definitively lower amount of this compound. Uracil was found to be the most abundant component in extracts obtained from C. vomitoria especially in the internal lipids of adults. 1-oleoyl glycerol was detected in all of the examined species of flies. It was most abundant in the internal extracts isolated from the larvae of C. vicina and the pupae of C. vomitoria. Suberic acid dimethyl ester was found in the larval and pupal internal lipids of C. vicina and S. carnaria in low amounts. Butyl stearate was identified only in the internal lipids of the larvae and adults of houseflies.

Adipokinetic hormone induces changes in the fat body lipid composition of the beetle Zophobas atratus

In insects, neuropeptide adipokinetic hormone (AKH) released from the corpora cardiaca mobilizes lipids and carbohydrates in the fat body. We examined the developmental differences in the action of Tenmo-AKH, a bioanalogue belonging to the adipokinetic/hypertrahelosemic family (AKH/HrTH), on the lipid composition of larval and pupal fat bodies in the beetle Zophobas atratus. Tenmo-AKH was administered to the beetle larvae and pupae either as a single dose or as two doses of 20 pmol during a 24h interval. Extracts of fat bodies were used to analyse the lipid composition by gas chromatography (GC) combined with mass spectrometry (GC-MS). Control extracts were analyzed using the same method. Fatty acids (FA) and fatty acid methyl esters (FAME) were the most abundant compounds in the fat bodies from both developmental stages. We observed significant differences in their concentrations following hormonal treatment. Tenmo-AKH also induced a distinct increase in larval sterols, fatty alcohols and benzoic acid.

Antennal and mouthpart sensilla of Acanthoscelides obtectus say (Coleoptera: Bruchidae)

Acanthoscelides obtectus, a serious pest with a worldwide distribution, damages stored seeds of Phaseolus sp. To acquire a better understanding of the sensory orientation of bean weevils, their antennal and palpal sensilla were investigated. The antennae and palps possess several types of sensory hairs: blunt‐tipped sensilla chaetica and trichoidea and also sharp‐tipped sensilla trichoidea, these last being the most numerous. Sensilla basiconica are present only on the flagellomeres, but Böhm bristles occur on the basal sclerite and scape of the antennae. The terminal segments of the maxillary and labial palps bear a group of sensilla styloconica with single apical pores. Sensilla coeloconica are located only on the terminal palpal segments. They were unable to detect any distinct morphological differences between males and females. The possible chemosensory, mechanosensory, and thermo‐/hygroreceptive functions of the sensilla are discussed.

Ultrastructural studies on the midgut of biting midge Forcipomyia nigra (Winnertz) (Diptera: Ceratopogonidae).

Biting midges belonging to the genus Forcipomyia are known to be hematophagous, predatory or saprophagous. Different stages of Forcipomyia nigra midges were investigated to provide a description of midgut ultrastructure. Larvae feeding on decaying organic matter possess simple, straight alimentary tracts whose middle regions are the longest. TEM studies of the larval midgut epithelium reveal that digestive cells show different ultrastructure depending on their age. The older cells with electron-dense cytoplasm degenerate while the younger ones with electron-lucent cytoplasm remain active in digestion. In saprophagous females, the ultrastructure of midgut epithelium changes according to the age of flies. Oogenesis induces degeneration of digestive cells and utilization of reserve material accumulated by them. The midgut epithelia of male midges consist of digestive and regenerative cells that show no evidence of cell degeneration as observed in females. Our results demonstrate differences between midgut digestive cells of males and females.