James S. Buckner

THE COMPOSITION OF THE CUTICULAR LIPIDS FROM NYMPHS AND EXUVIAE OF THE SILVERLEAF WHITEFLY, BEMISIA ARGENTIFOLII

Abstract The surface lipids of Bemisia argentifolii Bellows and Perring (Homoptera: Aleyrodidae) nymphs and exuviae were characterized. The identification and distribution of lipids were determined by capillary gas chromatography (CGC) and CGC-mass spectrometry (CGC-MS). The quantity of lipid from exuviae was 80% greater than that from nymphs. The composition for the lipid classes and the chain-length distribution of each lipid class were similar for both nymphs and exuviae: wax esters (86%), long-chain aldehydes (7%), hydrocarbons (3–4%) and long-chain alcohols (3–4%). The wax esters were composed of even-numbered-carbon compounds ranging from C 38 to C 64 . Structural analyses of the wax esters by CGC-MS with single ion monitoring revealed that the carbon numbers for the major acid and alcohol moieties ranged from C 14 –C 28 and C 24 –C 36 , respectively, and the major wax ester constituents were the C 52 ester, dotriacontanyl icosanoate (27%) and the C 54 ester, tetratriacontanyl icosanoate (14%). The major aldehyde was dotriacontanal (C 32 ), and the major alcohols were near equal amounts of dotriacontan-1-ol (C 32 ) and tetratriacontan-1-ol (C 34 ). The major hydrocarbons for both nymphs and exuviae were odd-carbon-numbered C 25 –C 35 n-alkanes. Lipids extracted from the exuviae from laboratory-reared sweetpotato whiteflies, Bemisia tabaci (Gennadius), were essentially the same as the lipids from B. argentifolii .

The lipid composition of the wax particles from adult whiteflies, Bemisia tabaci and Trialeurodes vaporariorum

Abstract Adult whiteflies are characterized by the presence of copious amounts of wax particles covering all surfaces of the body except the eyes. The lipid composition was determined for wax particles removed from the surfaces of the sweetpotato whitefly, Bemisia tabaci (Gennadius), and the greenhouse whitefly, Trialeurodes vaporariorum (Westwood). The lipid components in the wax particles of both species were mostly mixtures of long-chain aldehydes and long-chain primary alcohols. The major wax particle components for B. tabaci were C 34 aldehyde and C 34 alcohol and small amounts of C 32 aldhyde and alcohol. For the wax particles from T. vaporariorum , C 32 aldehyde and C 32 alcohol were the major components with lesser amounts of the C 30 components. These findings were compared to the surface lipids of fully-waxed B. tabaci and T. vaporariorum adults that contained, in addition to the major amounts of long-chain aldehydes and alcohols, quantities of long-chain wax esters. Wax esters were not present in lipid extracts from the surface of B. tabaci whiteflies at the time of adult emergence (prior to deposition of wax particles). Thus, the appearance of wax esters on the cuticular surfaces occurred during the period of deposition of wax particles. The quantities of wax esters in the surface lipid extracts of wing tissues separated from the bodies of adult whiteflies indicated that the wing surfaces were a major site of wax ester deposition.

Cuticular lipid composition of Heliothis virescens and Helicoverpa zea pupae

Abstract The cuticular lipids of laboratory-reared tobacco budworms, Heliothis virescens (Fabricius), and corn earworms, Helicoverpa zea (Boddie), pupae were characterized. Quantities of cuticular lipid were greater for pupae in diapause than for non-diapausing pupae. Major cuticular lipids of H. virescens pupae were longchain n-alcohols (42%) and aldehydes (41%) with chain lengths of mainly C26, C28 and C24 in decreasing order of abundance. Lesser amounts (2–5%) of wax esters and hydrocarbons were identified. The wax ester fraction was a complex mixture of C32-C52 components composed mainly of C24-C28 n-alcohol and saturated, monounsaturated and diunsaturated fatty acid (C16-C22) moieties. For H. zea, the major fractions were wax esters (38%), hydrocarbons (19%), n-alcohols (17%), diols (16%) and lesser amounts (2–5%) of aldehydes, acetate esters of alcohols and oxoalcohols. Triacontanyl hexadecanoate comprised 85% of the wax ester fraction. The H. zea alcohols, alcohol ester moieties and aldehydes consisted of a homologous series of C32-C52 compounds with the 30 carbon chain length as major constituents. The H. zea diols were C30-C36 even-chain n-alcohols with hydroxyl groups on carbon numbers 11, 12, 13, 14 or 15. Mass spectral analysis indicated the presence of unsaturation in the alkyl chain of the major diol components.

Waxes and lipids associated with the external waxy structures of nymphs and pupae of the giant whitefly, Aleurodicus dugesii

The nymphs and pupae of the giant whitefly, Aleurodicus dugesii, produce large quantities of external lipids, both as waxy particles and as waxy filaments. The nymphs and pupae extrude filaments from two dorsal rows of five pores each. Filaments can attain lengths of 5-8 cm. The external lipids of nymphs and pupae consist largely of long-chain aldehydes, alcohols, acetate esters and wax esters. Hydrocarbons are minor components. Soon after hatching, the nymph produced an unidentified waxy fringe extruded laterally from its margin. After molting to the second instar, long, hollow, waxy filaments were produced by the immature stages. The major lipid class associated with the filaments was saturated wax esters (89%), mainly C44, C46 and C60. Associated with formation of the filaments were waxy particles in the shape of curls, which peeled off of the extruding filaments. Similar but more tubular-shaped curls were also produced by numerous lateral pores so that, eventually, the curls completely camouflaged the nymph. The major lipid class of the curls was wax esters (50%), mainly C44 and C46. The cuticular surface lipids of the nymphs were mainly long-chain aldehydes (43%) and wax esters (27%). Unsaturated fatty acid moieties constituted 2 and 19% of the wax esters of curls and nymph cuticular surface lipids, respectively. The major lipid classes of pupae and of their palisade were long-chain aldehydes and alcohols. No unsaturated wax esters were detected in the filaments, but 30% of pupal and 21% of palisade surface wax esters were unsaturated in their fatty acid moieties, 16:1, 18:1 and 20:1.

External lipids of adults of the giant whitefly, Aleurodicus dugesii

Abstract The external lipids of male and female adults of the giant whitefly, Aleurodicus dugesii , were found in three forms; as cuticular lipids, as waxy particles, and only on the females, as waxy filaments. Collectively, the external lipids consisted of long-chain aldehydes, long-chain alcohols and wax esters. Cuticular lipids consisted mainly of wax esters. The major wax esters were C44 (tetracosanyl icosanoate and docosanyl docosanoate) and C46 (mainly triacontanyl hexadecanoate and tetracosanyl docosanoate). Hydrocarbons, largely n -alkanes, were minor components of the cuticular surface lipids. Both sexes produced waxy particles soon after eclosion by breaking off extruding strands (or ribbons) from anterior wax plates on their abdomens; three pair on males and two pair on females. These particles coated the adults and their surroundings. The waxy particles produced by the anterior wax plates of the adults were a mixture of aldehydes and alcohols, predominantly C30. The wax esters were not part of the waxy particles despite being the major lipid class on the cuticular surface. In addition, the adult female has two pair of posterior abdominal wax plates which produce waxy filaments that are broken off during oviposition and form spiral trails on the leaf. These trails appear to function to camouflage the eggs which are laid horizontally in the waxy trail. The lipid obtained from the waxy filaments from the posterior abdominal wax plates of the female was largely wax esters. The predominant wax ester was C46 (mainly triacontanyl hexadecanoate).

Uric acid levels during last larval instar of Manduca sexta, an abrupt transition from excretion to storage in fat body

Abstract Levels of uric acid in the whole body of the tobacco hornworm, Manduca sexta increased steadily for the 9 days of the fifth instar. However, concentrations in the haemolymph were lowest during the transition from the feeding stage to the wandering stage (days 3, 4), the time when there was a switch from uric acid excretion by the Malpighian tubule-hindgut system to storage in the fat body. Haemolymph volumes, determined for larvae between 2 and 6 days into the fifth instar by isotope dilution with [ 14 C]-inulin, were used to calculate rates of incorporation of uric acid into Malpighian tubules and fat body of larvae injected with [ 14 C]-uric acid. These labelling studies indicated that the Malpighian tubules ceased to remove uric acid from the haemolymph some time between the last 6 hr of day 3 of the fifth instar and the first 18 hr of day 4. At the same period, fat body removed significant quantities of uric acid from the haemolymph. The times of initial decreases and increases in levels of uric acid in haemolymph and fat body, respectively, indicated that storage in the fat body started before cessation of elimination via the Malpighian tubule-hindgut system.

The composition of external lipids from adult whiteflies, Bemisia tabaci and Trialeurodes vaporariorum

Abstract The total surface lipids, including the wax particles, of the adult whiteflies of Bemisia tabaci and Trialeurodes vaporariorum were characterized. At eclosion, there were similar amounts of long-chain hydrocarbons, aldehydes, alcohols and wax esters. Within a few hours post-eclosion, long-chain aldehydes and long-chain alcohols were the dominant surface lipid components, C 34 on B. tabaci and C 32 on T. vaporariorum . Hydrocarbons, mainly n -alkanes, were minor components of the surface lipids. The major wax esters were C 46 on B. tabaci and C 42 on T. vaporariorum . The major acid and alcohol moieties in the wax esters of B. tabaci were C 20 and C 26 , respectively, and of T. vaporariorum were C 20 and C 22 , respectively. Both B. tabaci and T. vaporariorum had a minor wax ester composed of the fatty acid C 18:1 esterified to the major alcohols, C 34 and C 32 , respectively. Bemisia were readily distinguished from Trialeurodes based on the composition of their wax particles and/or their wax esters; however, no differentiating surface lipid components were detected between biotypes A and B of B. tabaci .

Subcellular localization of uric acid storage in the fat body of Manduca sexta during the larval-pupal transformation

Abstract The fat body of the tobacco hornworm, Manduca sexta , serves as the major site for uric acid storage during metamorphosis. Light and electron microscopic examinations of fat body stained with reduced silver to show the location of stored uric acid have revealed that most, if not all, fat body cells store uric acid. The extent of specific staining is proportional to the increase in uric acid concentration in fat body during the initial stages of metamorphosis. Storage is associated with discrete membrane-bound structures, designated as uric acid storage vacuoles. In larval fat body, the structures are round or elliptical-shaped vacuoles with electron-dense fibrous interiors and are about the size of observed mitocondria (0.5–1.0 μm). During the larval-pupal transformation, the storage vacuoles double in size and appear as fibrous cores with spaces between the cores and the surrounding membranes. Before pupal ecdysis, the storage vacuoles are concentrated around the nucleus of each cell but after that event they are more uniformly distributed within fat body cells.

Long-chain and very long-chain methyl-branched alcohols and their acetate esters in pupae of the tobacco hornworm, Manduca sexta

Abstract Four homologous series of very long-chain methyl-branched alcohols (VLMA, C38 to >C44) were found in the internal lipids of developing male pupae of the tobacco hornworm, Manduca sexta, both as free alcohols and as acetate esters. The four major homologous series, with carbon chain backbones of 36–44 carbon atoms, consisted of a monomethyl, two dimethyls and a trimethyl-branched alcohol series. The major alcohol of each homologous series (with the corresponding alkane obtained by reduction in parentheses) was identified as 24-methyltetracontan-1-ol (17-methyltetracontane), 24,28-dimethyltetracontan-1-ol (13,17-dimethyltetracontane), 22,34-dimethyloctatriacontan-1-ol (5,17-dimethyloctatriacontane) and 22,26,34-trimethyloctatriacontan-1-ol (5,13,17-trimethyloctatriacontane). The minor components of the VLMA had backbones with an odd number of carbon atoms (37, 39, 41 and 43). Methyl branches of the minor components were identified on the 18- and 14,18-positions when numbered from the alkyl end of the chain. Also identified were minor amounts of long-chain methyl-branched alcohols (LMA, C25 to C32). The major components in the “wax ester” TLC fraction were acetate esters of the LMA and VLMA.

Uric acid excretion in larval Manduca sexta.

Abstract The characteristic coating of frass of last-instar tobacco hornworms, Manduca sexta, reared on artificial diet, proved to be uric acid. Results indicated that uric acid is the major nitrogenous excretory product; during most of the larval feeding stage, 5–7% of the excreta (dry weight basis) was uric acid; only minute quantities of allantoic acid were present. The rate of uric acid excretion was linear for the periods when coated pellets were observed. Abrupt increases in uric acid resulted from delays in pellet expulsion associated with delays in feeding activity. A distinctive coating was not generated by penultimate instar larvae, but abrupt changes in uric acid content did occur, which suggests that the phenomenon of coated frass is directly related to a differential in uric acid concentration. The source of uric acid in the frass was the Malpighian tubule system. The transition period between feeding and the wandering stage was a time of rapid decrease in uric acid excretion; there were only low levels in the last fecal pellets and none in Malpighian ampullae of wandering-stage larvae. Since the first appearance of coated fecal pellets preceded the release of ecdysone by about 24 hr, the involvement of this hormone was not indicated.