Klaus H. Hoffmann

A Family of Neuropeptides That Inhibit Juvenile Hormone Biosynthesis in the Cricket, Gryllus bimaculatus

Four nonapeptides that inhibit juvenile hormone synthesis have been isolated by four high performance liquid chromatographic steps from extracts of the brain of the field cricket, Gryllus bimaculatus. The primary structures of these peptides were assigned by Edman degradation and mass spectrometry as Gly-Trp-Gln-Asp-Leu-Asn-Gly-Gly-Trp-NH2 (Grb-AST B1), Gly-Trp-Arg-Asp-Leu-Asn-Gly-Gly-Trp-NH2 (Grb-AST B2), Ala-Trp-Arg-Asp-Leu-Ser-Gly-Gly-Trp-NH2 (Grb-AST B3), and Ala-Trp-Glu-Arg-Phe-His-Gly-Ser-Trp-NH2 (Grb-AST B4). Each of the peptides shows high sequence similarity to the locustamyoinhibiting peptide (Lom-MIP), but is structurally different from all the allatostatins so far identified. The synthetic allatostatins Grb-AST B1-4 are potent inhibitors (50% inhibition at 10 to 7 10M) of juvenile hormone III biosynthesis by corpora allata from 3-day-old virgin females of G. bimaculatus using an in vitro bioassay. At 10M, Grb-AST B1 also strongly inhibits juvenile hormone III biosynthesis by corpora allata from 2-day-old adult males and 1-day-old (males and females) and 4-day-old (females) last instar larvae of G. bimaculatus. The inhibitory effect of Grb-AST B1 was also evident on corpora allata from a related species, Acheta domesticus. Inhibition of juvenile hormone synthesis by Grb-AST B1-4 is reversible.

Identification of two allatostatins from the cricket, Gryllus bimaculatus de Geer (Ensifera, Gryllidae): additional members of a family of neuropeptides inhibiting juvenile hormone biosynthesis.

Two peptide inhibitors of juvenile hormone biosynthesis, designated G. bimaculatus allatostatins A1 and A2, have been purified from extracts of the brain of the field cricket Gryllus bimaculatus. The primary structures of these peptides were assigned as Ala-Gln-His-Gln-Tyr-Ser-Phe-Gly-Leu-NH2 (Grb-AST A1) and Ala-Gly-Gly-Arg-Gln-Tyr-Gly-Phe-Gly-Leu-NH2 (Grb-AST A2). Each of the peptides shows C-terminal amino acid sequence similarity to cockroach allatostatins and blowfly callatostatins. The two peptides are potent inhibitors of in vitro juvenile hormone production by corpora allata from virgin females of G. bimaculatus.

Alternative sites for ecdysteroid production in insects

Summary Several evidences have been obtained in various insect species demonstrating that, besides prothoracic glands and ovaries, other tissues could be alternative sites of moulting hormone production. After a detailed review on the various methods of investigation and criteria required to validate such observations, the nature of these sites, namely oenocytes, epidermis and testes, is discussed. Their possible involvement in moulting and/or reproduction is analyzed, giving the opportunity to put forward several new hypotheses. In particular, autocrine and paracrine secretions of ecdysteroids could play a role in localized developmental events, more difficult to control from endocrine glands possibly mitosis, meiosis, reprogramming, regeneration or early steps of embryogenesis).

Age-specific patterns in honeydew production and honeydew composition in the aphid Metopeurum fuscoviride : implications for ant-attendance

The intensity of the mutualistic relationship between aphids and ants depends mainly on the composition and amount of honeydew. We used the model system Tanacetum vulgare-Metopeurum fuscoviride to study age-related differences in honeydew production and composition and its effect on the mutualism between M. fuscoviride and the ant Lasius niger. First and second instar larvae of M. fuscoviride produced only half of the amount of honeydew as older larvae or adults. There were, however, no differences between age classes in the total honeydew sugar concentration, which averaged approx. 80 &mgr;g sugar/&mgr;l honeydew. Honeydew sugar composition also did not differ between age classes, and melezitose was the dominant sugar (59% in all classes). The amino acid concentration, by contrast, increased significantly with aphid age, reaching 22.6 nmol per &mgr;l honeydew in adult M. fuscoviride. This increase was mainly caused by asparagine and glutamine, while there were no differences in the concentrations of the five other regularly detected amino acids and cystine, respectively. The intensity of ant-attendance was significantly lower in colonies of first and second instar larvae than in colonies of older age classes. Ant-attendance correlated with the amount of honeydew produced, and not with the total amino acid concentration.

Honeydew amino acids in relation to sugars and their role in the establishment of ant‐attendance hierarchy in eight species of aphids feeding on tansy (Tanacetum vulgare)

Abstract.  The ratio of the concentration of honeydew total amino acids to total sugars in the honeydew of eight species of aphids, all feeding on tansy, Tanacetum vulgare (L.), was determined and correlated with honeydew production and ant‐attendance. The honeydew of the five ant‐attended aphid species [Metopeurum fuscoviride (Stroyan), Trama troglodytes (v. Hayd), Aphis vandergooti (Börner), Brachycardus cardui (L.), Aphis fabae (Scopoli)] was rich in total amino acids, ranging from 12.9 to 20.8 nmol µL−1 compared with the unattended aphid Macrosiphoniella tanacetaria (Kalt.) with only 3 nmol µL−1. Asparagine, glutamine, glutamic acid and serine (all nonessential amino acids) were the predominant amino acids in the honeydew of all species. The total concentration of amino acids in the phloem sap of tansy was much higher (78.7 nmol µL−1) then in the honeydew samples, and the predominant amino acids were glutamate (34.3%) and threonine (17.7%). A somewhat unexpected result was the finding that those aphid species with the highest total amino acid concentration in the honeydew always had the highest concentration of sugars. The lowest amino acid–sugar combined value was 104–28.8 nmol µL−1 in the non ant‐attended species M. tanacetaria, and the highest value was an average of 270–89.9 nmol µL−1 for the three most intensely attended aphid species M. fuscoviride, A. vandergooti and T. troglodytes. There is no evidence that any single amino acid or group of amino acids in the honeydew acted as an attractant for ant‐attendance in these eight aphid species. The richness of the honeydew (rate of secretion × total concentration of sugars), along with the presence of the attractant sugar melezitose, comprised the critical factors determining the extent of ant‐attendance of the aphids feeding on T. vulgare. The high total amino acid concentration in sugar‐rich honeydews can be explained by the high flow‐through of nutrients in aphids that are particularly well attended by ants.

Recent Advances in Hormones in Insect Pest Control

New approaches to the development of insect control agents have been revealed through the description of natural and synthetic compounds capable of interfering with the processes of development and reproduction of the target insects. The review presents information on novel insecticides that mimic the action of two insect growth and developmental hormone classes, the ecdysteroids and the juvenile hormones. Neuropeptide structures, their biogenesis, action, and metabolism also offer the opportunity to exploit novel control agents.

Molecular characterisation of cDNAs from the fall armyworm Spodoptera frugiperda encoding Manduca sexta allatotropin and allatostatin preprohormone peptides.

Allatotropin (AT) is a 13-residue amidated neuropeptide, first isolated from pharate adult heads of the tobacco hornworm, Manduca sexta (Manse-AT), which strongly stimulates the biosynthesis of juvenile hormones (JH) in the corpora allata (CA) of adult moths. In Spodoptera frugiperda, a cDNA that encodes 134 amino acids, including an AT peptide, has been cloned. The S. frugiperda allatotropin mature peptide (Spofr-AT) [GFKNVEMMTARGFa] is identical to that isolated from M. sexta. The basic organization of the Spofr-AT precursor is similar to that of Agrius convolvuli, M. sexta, Pseudaletia unipuncta, and Bombyx mori with 83-93% amino acid sequence identity. The Spofr-AT gene is expressed in at least three mRNA isoforms with 134, 171 and 200 amino acids, differing from each other by alternative splicing. All allatostatins (AS) have an inhibitory action on the JH biosynthesis in the CA. A cDNA that encodes 125 amino acid residues including one copy of the Manse-AS peptide has been cloned from S. frugiperda (Spofr-AS; QVRFRQCYFNPISCF). The basic organization of the Spofr-AS precursor is similar to that of P. unipuncta with 85% amino acid sequence identity. Using one step RT-PCR for semi-quantification of the gene expression, we showed that the three mRNAs of the Spofr-AT gene and the Spofr-AS gene are expressed in brains of last instar larvae, prepupae, pupae, and adults of both sexes of S. frugiperda with variable intensity.

The effects of octopamine, dopamine and serotonin on juvenile hormone synthesis, In vitro, in the cricket, Gryllus bimaculatus

Abstract Maximum inhibition of juvenile hormone (JH) III synthesis induced by octopamine in isolated corpora allata (CA) of female and male adult Gryllus bimaculatus occurred at 10−4 M. Though control rates of JH III synthesis rise to a peak on day 3 and decline over the next 6 days, the rate of JH III synthesis resulting from octopamine inhibition was constant at 6–8.5 pmol/pair CA/h. The JH synthesis was therefore never completely inhibited by octopamine at any age in either males or females. Elevated Ca ion (25 mM) in the medium tripled JH III synthesis to about 60 pmol/pair CA/h and at this Ca ion concentration the normal inhibition of JH synthesis by octopamine or synthetic allatostatin-1 (Dip AS-1) was overcome. This suggests that the Ca ion is important in transducing the octopamine inhibition. The potent octopamine-2 antagonists, phentolamine and gramine, did not block the octopamine inhibition of JH III synthesis, which indicates that in G. bimaculatus the octopamine type-2 receptors are not involved, but rather receptors involved in the Ca ion flux may be more important in octopamine inhibition. There was no CA-inhibiting action of dopamine or serotonin in adult G. bimaculatus, and there was no CA-inhibiting action of dopamine, serotonin or octopamine in 1-h-old males which normally have a very low JH synthese rate. Addition of farnesol to the incubation medium had no effect on JH synthesis, but induced a large accumulation of methyl farnesoate in the CA. Octopamine added with farnesol only slightly reduced the methyl farnesoate accumulation, however, the addition of farnesol along with octopamine to the medium overcame the inhibitory effect of octopamine and allatostatin-1 of JH III synthesis, which indicates that these CA-inhibiting factors were blocking an early step in JH III synthesis.

RNA interference with the allatoregulating neuropeptide genes from the fall armyworm Spodoptera frugiperda and its effects on the JH titer in the hemolymph

The juvenile hormone (JH) titer was measured by liquid chromatography-mass spectrometry (LC-MS) with electrospray ionization (ESI). Three JH homologs, the JH I-III were detected in various amounts in larvae, prepupae and virgin adult females of Spodoptera frugiperda. In penultimate larvae, the JH II and III titers were relatively high, but decreased continuously during the 3 days of that stage, whereas JH I was detectable at low amounts only on the first 2 days. At the beginning of the last larval stage almost no JH could be detected but thereafter, a consistent low amount of JH III was present until the prepupal stage. In adult virgins, the JH titer peaked on the 2nd and 6th day after the imaginal molt. The measured hormone titers well agree with general lepidopteran physiology, because in larvae the JH titer should be high to prevent premature metamorphosis, but decrease in last instar larvae before pupation, whereas in adults JH returns to control various aspects of reproduction. JH biosynthesis is thought to be the main factor influencing the JH titer in the hemolymph and there is evidence that neuropeptides either act stimulatory (allatotropins) or inhibitory (allatostatins) on this process. After silencing of either the allatostatin AS-C-type (Spofr/Manse-AS) or the allatotropin AT 2 (Spofr-AT 2) gene the transcript level was reduced in brain and gut of last instar larvae as well as of adult S. frugiperda. This suppression led to an increased JH titer in larvae, suggesting an allatostatic activity of both the peptides in this stage. As a result of the elevated hormone titer, the last larval stage was prolonged. In prepupae, the JH titer was decreased, but the animals pupated and molted normally. In adult female virgin moths the effect on the JH titer was inversely dependent on the age of the moths and varied among the JH homologs, indicating that the peptides act either allatostatic or allatotropic. For both peptides, gene silencing clearly reduced the oviposition rates of adult females.

Adult nutrition and butterfly fitness: effects of diet quality on reproductive output, egg composition, and egg hatching success

BackgroundIn the Lepidoptera it was historically believed that adult butterflies rely primarily on larval-derived nutrients for reproduction and somatic maintenance. However, recent studies highlight the complex interactions between storage reserves and adult income, and that the latter may contribute significantly to reproduction. Effects of adult diet were commonly assessed by determining the number and/or size of the eggs produced, whilst its consequences for egg composition and offspring viability were largely neglected (as is generally true for insects). We here specifically focus on these latter issues by using the fruit-feeding tropical butterfly Bicyclus anynana, which is highly dependent on adult-derived carbohydrates for reproduction.ResultsAdult diet of female B. anynana had pronounced effects on fecundity, egg composition and egg hatching success, with butterflies feeding on the complex nutrition of banana fruit performing best. Adding vitamins and minerals to a sucrose-based diet increased fecundity, but not offspring viability. All other groups (plain sucrose solution, sucrose solution enriched with lipids or yeast) had a substantially lower fecundity and egg hatching success compared to the banana group. Differences were particularly pronounced later in life, presumably indicating the depletion of essential nutrients in sucrose-fed females. Effects of adult diet on egg composition were not straightforward, indicating complex interactions among specific compounds. There was some evidence that total egg energy and water content were related to hatching success, while egg protein, lipid, glycogen and free carbohydrate content did not seem to limit successful development.ConclusionThe patterns shown here exemplify the complexity of reproductive resource allocation in B. anynana, and the need to consider egg composition and offspring viability when trying to estimate the effects of adult nutrition on fitness in this butterfly and other insects.